Practical Problems in Assisted Conception [1st ed.] 9781108149891

Table of contents :
Practical Problems in Assisted Conception......Page 1
Half Title......Page 3
Title Page......Page 5
Copyright......Page 6
Contents......Page 7
Contributors......Page 9
Chapter 1 A Lifetime Observations and Reflections as an IVF Doctor......Page 13
References......Page 16
Chapter 2 The Patient Perspective of Assisted Reproduction......Page 17
Data from Published Systematic Reviews......Page 19
Significance of Size of Fibroids......Page 20
Conclusions and a Practical Approach to Management in Clinical Practice......Page 21
References......Page 22
Which Follicle-Stimulating Hormone to Use......Page 23
What Dosage to Give......Page 24
References......Page 25
Predictive Tests of Poor Response......Page 26
Stimulation Protocols Used in Poor Responders......Page 27
Adjuvant Therapies for Poor Responder Patients......Page 28
Counseling......Page 29
References......Page 30
Background......Page 31
Considerations for Patient Selection......Page 32
Assessment of Risk to Fertility (Table 6.2)......Page 33
Suggested Documentation......Page 34
References......Page 35
Gonadotropin Hormone-Releasing Hormone Agonists......Page 36
Cryopreservation of Unfertilized or Fertilized Oocytes......Page 37
References......Page 38
Diagnosis......Page 40
Surgical Treatment......Page 41
Suggested Audits......Page 42
References......Page 43
Sperm Quality in Older Men......Page 44
Risks to Child Health and Development......Page 45
Suggested Standard Operating Protocol (SOP)......Page 46
References......Page 47
Discussion on Endometrioma-Specific Treatment Options......Page 48
Discussion on Stimulation Protocols and Treatment-Adverse Effects......Page 49
Further Reading......Page 50
References......Page 51
Ultrasound......Page 52
Treatment Hysteroscopic Surgery......Page 53
Foley Catheter......Page 54
Antibiotics......Page 55
References......Page 56
In Vitro Research: Biologically Plausible Mechanisms......Page 58
In Vivo Research: Clinical Trials on Acupuncture and CHM Acupuncture......Page 59
CAM Interventions during IVF......Page 60
References......Page 61
How Does the 揚roblem? Relate to IVF and Vice Versa?......Page 63
Strategies/Solutions......Page 64
References......Page 65
The Patient Who Has a Uterine Abnormality Diagnosed at Tubal Assessment......Page 67
Methods of Tubal Assessment Hysterosalpingography (HSG)......Page 68
Laparoscopy and Dye......Page 70
Fertiloscopy or Transvaginal Hydrolaparoscopy......Page 71
Chlamydia Testing......Page 72
Suggested Audit......Page 73
References......Page 74
Which One Is Better? Single Step or Sequential?......Page 75
Embryo Manipulation and Culture Approaches......Page 76
References......Page 77
How to Set Up and Manage Key Performance Indicators (KPI)......Page 78
Processes......Page 82
Laboratory Equipment......Page 83
Conclusion......Page 84
References......Page 85
Clinical Significance of EMT and Endometrial Pattern Clinical Significance of EMT......Page 86
Clinical Significance of Endometrial Pattern......Page 87
Managing Thin Endometrium in IVF......Page 88
Therapies to Improve Endometrial Perfusion......Page 89
References......Page 90
Pathophysiology......Page 91
Management......Page 93
References......Page 95
Progesterone Preparations and Routes of Administration and Duration of Luteal Support......Page 98
Protocols......Page 101
References......Page 102
Controlled Ovarian Stimulation: Protocols and Regime......Page 104
Pharmacologic Interventions......Page 106
References......Page 107
Cause of Subfertility......Page 109
Anesthetic Management......Page 110
References......Page 112
Preliminary Steps......Page 114
Movement of Aspiration Needle......Page 115
Follicle/Blood Vessel......Page 116
References......Page 117
The Setup......Page 119
Issues Relating to Machine/System Setup......Page 120
Reference......Page 121
Adequate Follicular Recruitment and Development......Page 122
Reduced Human Chorionic Gonadotrophin or Luteinizing Hormone/Progesterone Bioavailability......Page 123
Suggested Audit......Page 124
References......Page 125
Technique for Egg Collection......Page 126
Early Complications......Page 127
References......Page 129
Cervical Mucus and Blood......Page 130
Bladder Filling......Page 131
Cervical Dilatation......Page 132
Avoiding a Difficult Embryo Transfer......Page 133
References......Page 134
Normal Erectile Function and Ejaculation......Page 135
Conservative Measures......Page 136
Surgical Sperm Recovery (SSR)......Page 137
References......Page 138
Identification......Page 139
Identification......Page 140
Identification......Page 141
Resolution......Page 142
References......Page 143
Operator and Protocol......Page 144
Human Factors......Page 145
Patient Demographics......Page 146
Corrective Actions and Preventative Measures......Page 147
Reference......Page 149
Moving Oocytes and Embryos between Dishes......Page 150
Conclusion......Page 153
Acknowledgements......Page 154
Sources of Infection......Page 155
Identification......Page 156
Prevention......Page 157
References......Page 159
Polar Body Biopsy......Page 160
Platform for Couples with Genetic Disorders......Page 161
Mosaicism......Page 162
Should PGD Always Be Coupled with PGS?......Page 163
References......Page 164
Endometrium and Sub-Endometrium......Page 165
Congenital Anomalies......Page 166
Patency......Page 167
Follicle Tracking......Page 168
Conclusion......Page 169
References......Page 170
Lifestyle Changes......Page 171
Improving Embryo Quality and Selection......Page 172
Intra-Cavity Lesion......Page 173
Clinical Trials......Page 174
References......Page 175
What Is Fertilization Failure?......Page 177
Optimizing the Laboratory Environment......Page 178
Conclusion......Page 179
References......Page 180
What Is Poor Embryo Development?......Page 181
Oocytes: Cause of Aneuploidy and Influence on Embryo Development......Page 182
Laboratory: In Vitro Culture and Reprotoxicity......Page 183
Will Poor-Quality Embryos in One Cycle Mean Poor-Quality Embryos in the Subsequent Cycles?......Page 184
References......Page 185
What Is a Quality Management System and Why Have It?......Page 186
The Crucial Elements of the QMS Personnel......Page 187
Document Control......Page 188
Validation......Page 190
Conclusion......Page 191
References......Page 192
Human Performance Is Variable......Page 193
Teamwork......Page 194
Checks and Checklists......Page 195
References......Page 196
Suggestions for Further Reading......Page 197
Positioning and Branding......Page 198
How Are We Going to Get There? Develop a Marketing Plan......Page 199
References......Page 202
Study Setup......Page 203
Data Collection......Page 204
The Referral Process......Page 205
Summary......Page 207
References......Page 208
How to Get Information to Develop a Business Plan......Page 209
Understanding a Business Plan for an IVF Unit Setup......Page 210
Understanding the Business Economics: Profit and Loss Statement......Page 211
Revenue and Costs for an IVF/ Infertility Clinic Revenue from Services......Page 213
Cost of Running an Infertility Center......Page 214
Introduction to a Balance Sheet......Page 215
Which Measure to Use While Setting Up a Business......Page 216
Operational/Strategic Choices (Options) while Setting Up an IVF Business......Page 217
For Further Reading......Page 218
Chapter 42 The Essentials for Building a Great Fertility Team Take Five with Fiona Pringle......Page 219
ART 3.0......Page 221
References......Page 223
Conversation Questions......Page 224
Index......Page 227

Citation preview

Practical Problems in Assisted Conception

Practical Problems in Assisted Conception Edited by

Ying Cheong University of Southampton

Togas Tulandi McGill University

Tin-Chiu Li Chinese University of Hong Kong

University Printing House, Cambridge CB2 8BS, United Kingdom One Liberty Plaza, 20th Floor, New York, NY 10006, USA 477 Williamstown Road, Port Melbourne, VIC 3207, Australia 4843/24, 2nd Floor, Ansari Road, Daryaganj, Delhi – 110002, India 79 Anson Road, #06–04/06, Singapore 079906 Cambridge University Press is part of the University of Cambridge. It furthers the University’s mission by disseminating knowledge in the pursuit of education, learning, and research at the highest international levels of excellence. www.cambridge.org Information on this title: www.cambridge.org/9781316645185 DOI: 10.1017/9781108149891 © Cambridge University Press 2018 This publication is in copyright. Subject to statutory exception and to the provisions of relevant collective licensing agreements, no reproduction of any part may take place without the written permission of Cambridge University Press. First published 2018 Printed in the United Kingdom by TJ International Ltd. Padstow Cornwall A catalogue record for this publication is available from the British Library. ISBN 978-1-316-64518-5 Paperback Cambridge University Press has no responsibility for the persistence or accuracy of URLs for external or third-party internet websites referred to in this publication and does not guarantee that any content on such websites is, or will remain, accurate or appropriate. Every effort has been made in preparing this book to provide accurate and up-to-date information that is in accord with accepted standards and practice at the time of publication. Although case histories are drawn from actual cases, every effort has been made to disguise the identities of the individuals involved. Nevertheless, the authors, editors, and publishers can make no warranties that the information contained herein is totally free from error, not least because clinical standards are constantly changing through research and regulation. The authors, editors, and publishers therefore disclaim all liability for direct or consequential damages resulting from the use of material contained in this book. Readers are strongly advised to pay careful attention to information provided by the manufacturer of any drugs or equipment that they plan to use.

Contents List of Contributors

vii

1 A Lifetime’s Observations and Reflections as an IVF Doctor 1 Ian D. Cooke 2 The Patient’s Perspective of Assisted Reproduction 5 Louise Nicola Burton 3 How to Manage Intramural Fibroids before an IVF Cycle 7 Ertan Saridogan 4 Ovarian Stimulation Protocols: Horses for Courses 11 Ying Cheong 5 The Consultation with the Poor Responder 14 Paul Atkinson and William L. Ledger 6 The Consultation with a Patient Who Wishes to Have Ovarian Tissue Cryopreservation 19 Richard A. Anderson 7 How to Manage Patients with Cancer Who Need Fast-Track Treatment 24 Kirsten Tryde Macklon and Michael von Wolff 8 How to Manage Patients with Adenomyosis before an IVF Cycle 28 Stephan Gordts 9 The Consultation with Older Men Seeking ART Treatment 32 Allan Pacey 10 How to Manage Endometrioma in the IVF Patient 36 Mukhri Hamdan and Ying Cheong 11 How to Manage IVF Patients with Intrauterine Adhesions 40 Jacqueline Chung Pui Wah and Tin-Chiu Li

12 What Is the Role of Complementary and Alternative Medicine during IVF? A Review of the Evidence 46 Trevor Wing and Andrew Flower 13 How to Manage the Patient with Hydrosalpinges before an IVF Cycle 51 Arri Coomarasamy and Justin Chu 14 Tubal Assessment in the IVF Patient Ka Ying Bonnie Ng and Ying Cheong

55

15 Which Culture Media to Use 63 Necati Findikli and Mustafa Bahceci 16 How to Set Up and Manage Key Performance Indicators: What Are Red-Alerts and What to Do When They Occur? 66 Rachel Cutting and Caitriona Meaney 17 How to Manage the Patient with Thin Endometrium 74 Tse Yeun Tan and Heng Hao Tan 18 How to Manage the Patient with Fluid in the Endometrium Prior to Embryo Transfer 79 Christian M. Becker and Monica Mittal 19 Practical Tips on Personalizing Luteal Phase Support 86 Srividya Seshadri and Sesh Kamal Sunkara 20 How to Avoid Ovarian Hyperstimulation Syndrome: Best Management Strategies Nikolaos Tsampras and Raj Mathur 21 Anesthetic Choices in IVF Practice 97 Singaraselvan Nagarajan and Eileen Lew 22 Tips and Tricks of Transvaginal Oocyte Retrieval 102 Durga G. Rao, Sujatha Vellanki and Ying Cheong

92

Contents

23 What to Do When the Electric Oocyte Aspiration Pump Stops Working 107 Ying Cheong

34 How to Draw Up a Management Plan When an IVF Cycle Has Failed 159 Carol Coughlan

24 How to Manage the Patient Who Has No Egg Retrieval 110 Nikoletta Panagiotopoulou and Abha Maheshwari

35 When Fertilization Fails 165 Tanya Milachich and Atanas Shterev

25 Managing Unusual Complications in IVF Relating to Transvaginal Oocyte Retrieval 114 Ying Cheong

36 What Does Poor Embryo Development Mean, and How Does It Influence Subsequent Cycles? 169 Emma S. Adolfsson 37 The ABCs of Quality Management in IVF Oksana Stidston

26 Tips and Tricks on the Management of a Difficult Embryo Transfer 118 Edward Coats and Marco Gaudoin

38 Human Factors in IVF Practice Rachel J. Broadley

27 How to Manage the Patient Who Fails to Produce a Semen Sample 123 Alpha K. Gebeh and Mostafa Metwally

39 Customer Service and Marketing in IVF Practice 186 Helen Kennard

28 Prevention and Management of Incubator Failure 127 Charissa Watchorn and Julia Paget

40 The Role of the Research Nurse in an IVF Center 191 Susan Wellstead and Jane Forbes

29 Troubleshooting Poor Survival after Oocyte and Embryo Cryopreservation 132 Victoria Ryder and Dawn Yell

41 Financial Models in an IVF Practice Kiran Gedela

30 The “Lost” Embryo in the Laboratory Virginia N. Bolton

138

31 What to Do When There Is a Suspected Infection in the IVF Laboratory 143 Philippa Tucker and Julia Paget 32 Dealing with Practical Problems Encountered in PGD and PGS 148 Junhao Yan and Zi-Jiang Chen 33 Tips and Tricks in Performing 3-D Ultrasound before an IVF Cycle 153 Sotiris H. Saravelos

174

181

197

42 The Essentials for Building a Great Fertility Team: Take Five with Fiona Pringle 207 Fiona Pringle and Ying Cheong 43 The Future of IVF: ART 3.0 Nicholas S. Macklon

209

44 Peering into the Future of the Fertility Business: A Conversation with Sue Channon, CEO, Virtus Health 212 Sue Channon and Ying Cheong

Index

215

Contributors

Emma S. Adolfsson MSc Department of Pathology, University Hospital of Örebro, Örebro, Sweden Richard A. Anderson MD PhD FRCOG FRCPEd MRC Centre for Reproductive Health, University of Edinburgh, Edinburgh, UK Paul Atkinson FRANZCOG CREI University of New South Wales Royal Hospital for Women, Sydney, Australia Mustafa Bahceci MD Bahceci Fulya IVF Center, Istanbul, Turkey Christian M. Becker MD Nuffield Department of Obstetrics & Gynaecology, University of Oxford, John Radcliffe Hospital, Oxford, UK

Justin Chu PhD MRCOG University Department of Obstetrics & Gynaecology, Birmingham Women’s Hospital, Birmingham, UK Edward Coats MRCOG BSc LLM DFRSH DHMSA East Midlands Fertility Centre, Nottingham, UK Ian D. Cooke FRCOG University Department of Obstetrics & Gynaecology, The Royal Hallamshire Hospital, Sheffield, UK Arri Coomarasamy FRCOG Institute of Metabolism & Systems Research, University of Birmingham, Birmingham, UK Carol Coughlan MD FRCPI FRCOG IVI Middle East, Dubai, UAE

Virginia N. Bolton MA PhD Assisted Conception Unit, Guy’s Hospital, London, UK

Rachel Cutting MBE Jessop Fertility, The Royal Hallamshire Hospital, Sheffield, UK

Rachel J. Broadley FRCA Southampton NHS Treatment Centre, Southampton, UK

Necati Findikli MSc PhD Bahceci Fulya IVF Center, Istanbul, Turkey

Louise Nicola Burton BSc PgC PgD Derby University, UK Sue Channon RN Virtus Health, Greenwich, NSW, Australia Zi-Jiang Chen MD PhD Center for Reproductive Medicine, Shandong University, Jinan, China Ying Cheong MA MD FRCOG Complete Fertility Centre, University Department of Obstetrics & Gynaecology, Princess Anne Hospital, Southampton, UK

Andrew Flower PhD MRCHM MBAcC Primary Care & Population Sciences, University of Southampton Aldermoor Health Centre, Southampton, UK Jane Forbes MRes, BSc hons, RM, RN Complete Fertility Centre, Princess Anne Hospital, Southampton, UK Marco Gaudoin MD FRCOG GRCM Ltd, Glasgow, UK Alpha K. Gebeh PhD MRCOG Jessop Fertility, The Royal Hallamshire Hospital, Sheffield, UK

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List of Contributors

Kiran Gedela MD Oasis Centre for Reproductive Medicine, Hyderabad, India

Caitriona Meaney BSc (Hons), MSc Jessop Fertility, The Royal Hallamshire Hospital, Sheffield, UK

Stephan Gordts MD Life Expert Center, Leuven, Belgium

Mostafa Metwally MD FRCOG Jessop Fertility, The Royal Hallamshire Hospital, Sheffield, UK

Mukhri Hamdan PhD MObGyn Fertility Unit, University of Malaya, Kuala Lumpur, Malaysia Chung Pui Wah, Jacqueline MbCHB, MRCOG, FHKCOG, FHKAM (O&G), CERT HKCOG (Reprod Med) The Chinese University of Hong Kong, Prince of Wales Hospital, Shatin, New Territories, Hong Kong SAR Helen Kennard MA MDip BSc Complete Fertility Centre, Princess Anne Hospital, Southampton, UK William L. Ledger MA DPhil FRCOG FRANZCOG University of New South Wales Royal Hospital for Women, Sydney, Australia Eileen Lew MBBS MMed (Anaes) Department of Anesthesiology, KK Women’s & Children’s Hospital, Singapore Tin-Chiu Li MD PhD MRCP FRCOG Department of Obstetrics & Gynecology, Chinese University of Hong Kong, Prince of Wales Hospital, Hong Kong, China Philippa Lowen PhD Complete Fertility Centre, Princess Anne Hospital, Southampton, UK Kirsten Tryde Macklon MD PhD Fertility Clinic, University Hospital Copenhagen, Copenhagen, Denmark Nicholas S. Macklon MD FRCOG University Department of Obstetrics & Gynaecology, Princess Anne Hospital, Southampton, UK

Tanya Milachich PhD Dr Shterev IVF Unit, Sofia, Bulgaria Monica Mittal BSc, MB BS, MRCOG Oxford University Hospital NHS Foundation Trust, John Radcliffe Hospital, Oxford Singaraselvan Nagarajan MBBS FCARCSI Department of Anesthesiology, KK Women’s & Children’s Hospital, Singapore Ka Ying Bonnie Ng MBChB BMedSci University Department of Obstetrics & Gynaecology, Princess Anne Hospital, Southampton, UK Allan Pacey MBE PhD FRCOG Academic Unit of Reproductive & Developmental Medicine, University of Sheffield, Sheffield, UK Julia Paget BSc DipClinEmbryol Complete Fertility Centre, Princess Anne Hospital, Southampton, UK Nikoletta Panagiotopoulou MSc MRCOG Aberdeen Fertility Centre, Aberdeen, UK Durga G. Rao MRCOG Oasis Centre for Reproductive Medicine, Hyderabad, India Victoria Ryder BSc MMedSci Complete Fertility Centre, Princess Anne Hospital, Southampton, UK

Abha Maheshwari MD FRCOG Department of Obstetrics & Gynaecology, Aberdeen Maternity Hospital, Aberdeen, UK

Sotiris H. Saravelos MD MRCOG Assisted Reproductive Technology Unit, Department of Obstetrics & Gynaecology, Prince of Wales Hospital, Hong Kong, China

Raj Mathur MD FRCOG Department of Reproductive Medicine, Manchester Academic Health Sciences Centre, Manchester, UK

Ertan Saridogan MD PhD FRCOG Institute for Women’s Health, University College London Hospitals, London, UK

List of Contributors

Srividya Seshadri MD MSc MRCOG Centre for Reproductive & Genetic Health, London, UK Atanas Shterev PhD Dr Shterev IVF Unit, Sofia, Bulgaria Oksana Stidston M.P.A. Complete Fertility Centre, Princess Anne Hospital, Southampton, UK Sesh Kamal Sunkara MD MRCOG Assisted Conception Unit, Guy’s Hospital, London, UK Heng Hao Tan MMed MRCOG Department of Reproductive Medicine, KK Women’s & Children’s Hospital, Singapore Tse Yeun Tan FRANZCOG Department of Reproductive Medicine, KK Women’s & Children’s Hospital, Singapore Nikolaos Tsampras MBBS Department of Reproductive Medicine, Manchester Academic Health Sciences Centre, Manchester, UK Togas Tulandi MD MHCM Department of Obstetrics and Gynecology, Milton Leong Chair in Reproductive Medicine, McGill University, Montreal, Canada

Venkata Sujatha Vellanki MS MICOG FMAS Clinical Head Oasis–Centre for Reproductive Medicine,Vijayawada Michael von Wolff MD PhD Department of Gynaecological Endocrinology & Reproductive Medicine, Bern University Hospital, Bern, Switzerland Charissa Watchorn MSc Complete Fertility Centre, Princess Anne Hospital, Southampton, UK Susan Wellstead Complete Fertility Centre, Princess Anne Hospital, Southampton, UK Trevor Wing BSc MSc PhD The Women’s Natural Health Clinic, Twickenham, UK Junhao Yan MD PhD Center for Reproductive Medicine, Shandong University, Jinan, China Dawn Yell MSc Complete Fertility Centre, Princess Anne Hospital, Southampton, UK

ix

Chapter

1

A Lifetime’s Observations and Reflections as an IVF Doctor Ian D. Cooke

Considering life as an in vitro fertilization (IVF) doctor leads one to think that IVF has always been readily available, but this is not so. I did my first infertility clinic in 1959 as a Senior House Officer in Australia; it had already been differentiated from the gynecology clinic because of the number of patients. A hysterosalpingogram provided evidence for tubal macrosurgery, semen analysis was not standardized and there was no endocrine test for ovulation nor any means to stimulate it. A patient went abroad for donor insemination. After extensive clinical training and research in endocrinology, I ran an infertility clinic in Wales in 1969, although tubal surgery was still the only treatment used. Urinary total estrogen and pregnanediol assays in 24-hour urine samples became possible and clomiphene and human menopausal gonadotropin (hMG) were introduced. Multiple pregnancies followed in spite of urinary estrogen monitoring that used the postal service to send samples to a distant lab, the results being phoned through the following day. In 1973 I established my infertility clinic in Sheffield, by which time it was possible to measure serum hormones. Prolactin measurement led to a search for pituitary microadenomas, and bromocriptine was extensively used before it was fully appreciated that prolactin was also a stress hormone. Laparoscopy was developed into an important diagnostic tool and became essential to define peritubal and periovarian adhesions, better dealt with by microsurgery, as was tubocornual obstruction. Sperm cryopreservation using slow freezing became practical after earlier use in animal husbandry and sperm donation then became feasible. Sperm banks developed, so donors were screened, interviewed and counseled, as were the couples; it was recognized that counselors had a role and needed specific training. Sperm banks made donations available for sale to other clinics and trade flourished. During this time the World Health Organization (WHO) developed its Task Force on the Diagnosis

and Treatment of Infertility, began optimizing semen analysis and structured a formal evaluation of both female and male history, physical examination and investigation. The 1978 protocol was used in 23 countries and the project recruited about 10,000 patients. Features required for diagnosis of each cause were identified and resulted in the publication of the Laboratory Manual for the Examination of Human Semen and Semen-Cervical Mucus Interaction and the Manual for the Standardized Investigation and Diagnosis of the Infertile Couple, which helped to regularize the clinical management. The former volume progressed to its fifth edition in 2010 and is the world reference standard for semen analysis, incorporating the major changes in evaluation of sperm morphology using strict criteria. Ultrasound, which had been mostly used for obstetric measurement, began to be used for assessment of the nonpregnant pelvic organs and follicle growth was charted. Ovulation was timed more accurately by first urinary and then serum luteinizing hormone (LH) assays. For 10 years Edwards and Steptoe had been researching human IVF. Edwards predicted that success would raise all sorts of ethical questions – and he was roundly ignored. Finally in 1978, they succeeded. Louise Brown was born after a non-stimulated cycle and laparoscopic oocyte retrieval and the world was changed. Developments came slowly. In view of the poor results, ovarian stimulation was introduced and more oocytes were obtained. The Melbourne group refined the stimulation schedule, leading to more embryos being replaced and the inevitable rise in multiple births and associated prematurity causing pressure on neonatal units. Laparoscopic egg retrieval was replaced by transurethral and then transvaginal aspiration under ultrasound control and aspiration pressures were reduced when syringes were replaced by low-pressure pumps. Human menopausal gonadotropin with its 1:1 ratio of

1

Ian D. Cooke

follicle-stimulating hormone (FSH) and LH (with added human chorionic gonadotropin [hCG] that was not highlighted) were gradually replaced by preparations containing more FSH until recombinant FSH was introduced with the claim that LH was unnecessary. Around the same time, embryo cryopreservation, still with slow freezing, was introduced and seen as an answer to excess egg/embryo production. In the United Kingdom, social and political discussion led the Medical Research Council to set up a group to advise on research policy and Parliament to form the Warnock Committee of Inquiry into Human Fertilisation and Embryology, which recommended creating a Statutory Licensing Authority to regulate research and services. As little happened, the Medical Research Council and the Royal College of Obstetricians and Gynaecologists established a Voluntary (later Interim) Licensing Authority with a view to creating standards of clinical practice and as a stimulus to government to implement the Warnock recommendations. The Department of Health held discussions about the meaning of “proper counseling” and how it could be developed and practiced. After an unprecedented two white papers describing the government legislative intentions and then their revised proposals following public consultation, the Human Fertilisation and Embryology Act was passed in 1990. The Human Fertilisation and Embryology Authority was established in 1991 and quickly formulated its Code of Practice, instituting site visits and dialogue with practitioners and publishing its annual reports. At the same time intrauterine insemination began to be used as a treatment for selected couples prior to moving on to IVF, particularly for those couples with a mild male factor, mild endometriosis or unexplained infertility. Endometriosis was increasingly being diagnosed with more rigorous application of the revised American Fertility Society classification. Laparoscopic salpingostomy began to replace microsurgery and there was debate about the optimal way to manage endometriotic cysts. The European Society of Human Reproduction and Embryology (ESHRE) held its first annual meeting in Bonn, Germany, in 1985, and its journal, Human Reproduction, was published in 1986, climbing to be the principal one in its field as evidenced by its impact factor. In Italy the law forbade embryo cryopreservation, so efforts were directed to oocyte freezing. Data collection began to be taken seriously and national data were aggregated into European IVF-monitoring (EIM) program reports, leading to

the development of the International Committee for Monitoring Assisted Reproduction (ICMART) and its efforts to collate world data. Capri workshops of international experts provided an opportunity to synthesize knowledge in infertility. The British Infertility Counselling Association was founded to promote professional standards and somewhat later, embryologists formed the Association of Clinical Embryologists in the United Kingdom to standardize practice, training and certification and develop continuing education. Although WHO had published a technical bulletin in 1990 reviewing the field of assisted reproduction and putting forward a series of suggestions for research, it evinced little further interest until a large meeting was convened in Geneva in 2001 on “Current Practices and Controversies in Assisted Reproduction.” A global perspective was framed and the importance of low-cost methods was emphasized. As well as the scientific and clinical perspectives, attention was given to social and psychological issues, ethical aspects of infertility and assisted reproductive technology (ART) and national and international data surveillance. In 1998 the International Federation of Fertility Societies (IFFS) had begun its surveillance of laws and guidelines relating to ART and has published data relating at its peak to 102 countries (2010), those with laws, guidelines, both or neither. Its documentation extended to the status of conception, embodying religious and ethical dimensions, and its triennial publications have continued [1]. These aspects were explored progressively from 2001 in a series of publications by ESHRE’s Task Force on Law and Ethics [2]. The novel intracytoplasmic sperm injection (ICSI) allowed poor-quality semen to be used, but its indications were soon distorted and it has been used more widely, particularly in some parts of the Middle East, as the routine approach. The Cochrane Collaboration, founded in 1993 and developed from its initial reviews in perinatal medicine, has embraced ART. An examination of the 59 reviews of randomized controlled trials covering many areas of ART practice (up to July 2015) was produced using A Measurement Tool to Assess Systematic Reviews (AMSTAR) criteria and it concluded that most were of a high standard, although there was evidence of publication bias [3]. Anxiety about the high frequency of multiple births soon emerged and single embryo transfer

Observations and Reflections as an IVF Doctor

(SET) was pioneered in Scandinavia; it was shown that live birth rates could be sustained and this has led to an effort in many parts of the world to reduce the number of embryos transferred. In Belgium regulation was introduced so that patient reimbursement was dependent on SET under defined circumstances, including maternal age and the number of attempts. A steady improvement in laboratory standards and competence has also helped lead to a reduction in the frequency of multiple births. Later, in vitro culture of embryos to blastocyst helped the selection for transfer and time-lapse imaging has improved understanding of the variety of embryological stages, but improvement in “take home baby” rates remains to be demonstrated. Although embryo cryopreservation has been essential to the support of the SET concept, it was realized that too many embryos were being produced using the gonadotropin-releasing hormone (GnRH) agonist protocols and longer-term storage was becoming a problem. Gentler stimulation regimes were proposed to reduce the number of oocytes and embryos with some evidence that this led to betterquality embryos. Ovarian hyperstimulation syndrome (OHSS) was also occurring too frequently and antagonists caused less impact on the woman and markedly reduced the frequency of OHSS. Freezing was extended to oocytes, particularly in Italy, which created laws based on religious dogma in order to restrict the number of oocytes fertilized and mandated their replacement, although these were ultimately struck down. The slow cryopreservation technique has largely been replaced by vitrification, although more extensive longer-term data on ovarian tissue preservation are awaited. Surrogacy was introduced, leading to cross-border activity, complicating the ethical debate and raising legal issues about the adoption process. Some countries have responded by restricting its practice within their borders. The Royal College of Physicians and the Universities of Leeds and York published their Effective Health Care Bulletin on the Management of Subfertility based on systematic reviews in 1992. This was followed by the Royal College of Obstetricians and Gynaecologists’ series of evidence-based Guidelines for the management of infertility in primary, secondary and tertiary care. The ESHRE Capri Workshop set out its Guidelines on Prevalence, Diagnosis and Management of

Infertility and the National Institute for Health and Care Excellence (NICE) issued its Fertility Assessment and Treatment for People with Fertility Problems extensively using systematic reviews and publishing its evidence base. Those recommendations were subsequently reviewed and have significantly influenced national criteria for funding within the National Health Service. WHO is currently finalizing its Guidelines. It defined the questions for systematic reviews using the Patient/ Problem/Population; Intervention; Comparison; Outcome and Setting [4] (PICOS) system using the Grading of Recommendations Assessment, Development and Evaluation (GRADE) system to determine the level of the Recommendations [5] and these seem likely to determine the standard of future Guidelines. The impacts of the procedures and monitoring on patients have both physical and mental dimensions, with anxiety and stress playing a significant role. Recognition has led to efforts to improve the procedures and designate “patient-friendly” IVF with emphasis on communication and support. ESHRE has recently published its Guideline on Psychosocial Care in Infertility and Medically Assisted Conception and has issued a pocket guideline for use by professionals in their routine care delivery [6]. Although the isolation of embryonic stem cells, derived from the inner cell mass of blastocysts grown for research, has stimulated the new field of organ replacement therapy, the ethical constraints have helped to push research into pluripotent cell derivatives of somatic cell lines. This research field has emphasized the segregation of opinion driven by religious principles, a problem recognized early on in reproductive medicine by Dr. Mary (later Baroness) Warnock. IFFS Surveillance publications regularly underscore this, identifying those countries where specific practices, such as donor gametes, are forbidden by law. Costa Rica was the only country to pass a law prohibiting IVF, but an appeal by citizens to the Inter-American Court of Human Rights resulted in a judgment supporting them and instructing the government to repeal the law and provide IVF. The Court gave a robust refutation of the biological and philosophical premises underlying the law, providing reassurance that science and rationality can lead to progress [7]. Huge advances have been made in the science and clinical practice of reproductive medicine and the latest

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Ian D. Cooke

evidence is presented in this volume. However, to make further progress, such as by the creation of artificial gametes, there will need to be high-quality communication in extending the public understanding not only of science but also of the ethical basis of both the science and its clinical application. At the other end of the spectrum, more attention will need to be given to the politics of health and the financial aspects of clinical service. Access may seem a problem in the developed world, but this problem has hardly been touched in low-resource economies, where appropriate forms of ARTs have not been available for the general population. They really have to be part of the WHO approach of universal health coverage [8] at the same time as greater efforts must be made to reduce the massive burden of tubal damage from unsafe abortion and lack of obstetric care [9]. It is a privilege to have been part of the revolution in fertility care. These selective memories highlight the fact that the discussions have been on a much broader stage than clinical science alone. We need to keep that perspective. The technology has not reached most people in the world. It remains a challenge to deliver to them affordable reproductive health care and extend the gains made in the delivery of contraception, so that fertility care truly covers the life course for everyone [10].

References 1. IFFS Surveillance. 2013. Ed. S. J. Ory. https://c.ymcdn .com/sites/iffs.site-ym.com/resource/resmgr/iffs_sur veillance_09-19-13.pdf

2. Statements from the Task Force on Ethics and Law (ESHRE). www.eshre.eu/Specialty-groups/SpecialInterest-Groups/Ethics-and-Law/Documents-of-theTask-Force-Ethics-Law.aspx 3. C. Farquhar, J. R. Rishworth, J. Brown, W. L. D. M. Nelen and J. Marjoribanks. Assisted reproductive technology: An overview of Cochrane Reviews (Review). Cochrane Database of Systematic Reviews 2015, Issue 7. Art. No.: CD010537. http://onli nelibrary.wiley.com/doi/10.1002/14651858.CD010537 .pub3/full 4. PICOS System to Define Questions for Systematic Reviews. https://consortiumlibrary.org/aml/research aids/handouts/PICOSworksheet.pdf 5. The GRADE Classification System. http://clinicale vidence.bmj.com/x/set/static/ebm/learn/665072 .html 6. Psychosocial Care in Infertility and MAR, ESHRE. www.eshre.eu/Guidelines-and-Legal/Guidelines/Psyc hosocial-care-guideline.aspx 7. Inter-American Court of Human Rights Report No. 1/ 15 Case 12.798 Artavia Murillo et al. (“In Vitro Fertilization”) v. Costa Rica https://iachr .lls.edu/sites/iachr.lls.edu/files/iachr/Cases/Artavia_M urillo_et_al_v_Costa_Rica/peterson_artavia_muril lo_et_al_v._costa_rica.pdf. pp. 1360–1. 8. Universal Health Care. www.who.int/mediacentre/fac tsheets/fs395/en/ 9. World Disability Survey. 2011. www.who.int/disabil ities/world_report/2011/ 10. The Life Course Approach in Sexual and Reproductive Health. www.euro.who.int/__data/assets/pdf_file/ 0017/292202/Life-Course-Approach-in-SRH.pdf? ua=1

Chapter

2

The Patient’s Perspective of Assisted Reproduction Louise Nicola Burton

Working in health care, it is natural to be in “professional mode,” and sometimes we forget about the most important person – the patient. While in vitro fertilization (IVF) and everything associated with it might be second nature, it is important to remember that it might be the first time that the patient has encountered this new and intimidating world. I have been on both sides of the coin. By the time the patient is sitting in your clinic, she may be further down the investigative line and know what the issues are. The discovery of an issue and the need to have investigations and intervention can come as quite a shock. The devastating effect that this news can have should not be underestimated. As a patient, it can be a scary time and the feeling of isolation simply adds to emotions. After having the investigations in our local hospital and learning that we would need IVF, we looked into local clinics and went along to an open evening. We found this very informative. All the staff came across as professional and, just as important, friendly and approachable. You might be a professor in your field, but if you appear standoffish, that is likely to put some people off. It was also interesting to hear from all the different specialties (the lab team, the nurses and the counselor) and see how they worked as a team and genuinely seemed to respect each other. Combined with the tour, this made us feel so sure we were in good hands, we actually didn’t bother going to any other clinics. Subsequently, I have been lucky enough to go along and talk for a few minutes at these open evenings as I feel that clinics will easily be able to “big themselves up,” but it helped to hear from someone who had been in their shoes and actually been through it. It’s all well and good that you have the most up-todate lab or an embryoscope, but what’s it like to have to go through the ups and downs, or to inject yourself? I also felt reassured that we weren’t the only couple going through this.

It’s a strange stage to be at, with a mix of trepidation and excitement that you have finally started on the journey. At all the consultations with both the consultants and the nursing team we had everything explained to us in a language and at a level we understood. This is another important point from a patient’s point of view. We weren’t being spoken down to or impressed by big, fancy words. Explaining all the options to patients and making them part of the decision-making process can give them some element of feeling in control. When we were told something, it was made clear and explained to us, and we were frequently asked if we had any questions before we moved on. We were often reminded that we could call the clinic at any point if we had any questions. The day that the drugs arrived was a bit of a reality check. I remember opening the box and seeing the different drugs, syringes and needles (so many needles …). I must admit I had a bit of a wobble, but it wasn’t long until our first ultrasound scan and a meeting with our lovely, reassuring, friendly nurses. During all our scans, the results and how things were progressing were discussed. One of the most important things to me, and to others that I’ve spoken to, is that the partner is included. They may not be going through the actual interventions, but they are emotionally invested in the procedure and are a very important part, especially to the patient. The day of egg collection can be another mixed bag of emotions. It is another step forward, but there can also be disappointment if fewer eggs than expected are collected. On one occasion, my left ovary was inaccessible, which obviously had an effect. The staff members were upbeat, though, and we left with a promise of a phone call from the embryologists the next day. We have been through IVF more than once and had both fresh and frozen cycles. Waiting for that call doesn’t get any easier. Embryo transfer day was made to feel as relaxed as possible for us. We got to see the embryo and were

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given a picture to take home. This may seem like a small thing, but it was huge for us, and, if successful, many people have a baby photograph that early! The time between transfer and getting the results was perhaps the strangest time of all. After having injections every day and scans every other day, suddenly nothing. Luckily we had been warned about this, and once again the offer of help and support being only a phone call away was reiterated. We have had the best and the worst outcomes through IVF. Poor egg numbers, poor embryo quality, positive and negative pregnancy blood test results, the first scans with a heartbeat, the loss of twins, the birth of our rainbow baby. Throughout all of this our clinic was there to support us. Counseling was included in the package, and having a specialist that dealt with these highs and lows every day was invaluable. We have been blessed with our clinic, but I know others have not been so lucky. The technology involved is impressive, but the majority of patients I’ve spoken to are more interested in something far more basic – being treated as an individual and not just another patient on a conveyor belt. Due to the way that the

National Health Service is funded, there are likely to be a significant number of patients who are paying privately. IVF isn’t cheap, understandably so, but some patients won’t have money to splash around and a significant amount of savings will be necessary. They may be able to afford only one attempt. There are so many extra add-ons available out there that patients can find out about, thanks to Dr. Google. It is important that what is being offered at clinics is through evidence-based medicine and not seen as either trying to get more money or offering false hope to people who are often in a desperate situation. These may sound like simple points, but I think in the world of health care, with time pressures and targets to hit, we often forget the basics and it does no harm to be reminded of them on occasions. To sum the whole experience up in one word would be difficult, but I’d say “emotional.” Remembering that the patient may be experiencing excitement to fear, and everything in between, is important. Whether she is going through this for the first or fifth time, the patient should always be at the center of the decisions made.

Chapter

3

How to Manage Intramural Fibroids before an IVF Cycle Ertan Saridogan

Fibroids are frequently encountered prior to or during in vitro fertilization (IVF) cycles due to their high prevalence in the female population. The estimated cumulative incidence of fibroids by age 50 years is >80 percent in black women and almost 70 percent in white women [1]. However, many fibroids are completely harmless and have no clinical relevance. When clinically relevant fibroids are taken into account to include uteri nine weeks gestation size or larger, at least one submucosal fibroid or at least one fibroid of ≥4 cm, the prevalence is 10–15 percent in white women and 30–40 percent in black women between the ages of 35 and 39 years. In contrast, 35 percent of white women and 50 percent of black women aged 40–50 years have clinically relevant fibroids [1]. It is, however, debatable how clinically relevant are the inclusion criteria used to produce these figures. The impact of fibroids on IVF outcome is very controversial; a number of published systematic reviews and meta-analyses in the past decade have come up with different conclusions [2–6]. This is probably a reflection of the differences in the methodology of reviews and how stringent are the inclusion/ exclusion criteria that have been used. Furthermore, and probably more importantly, the differences may stem from the fact that the number, size, shape, location and consistency of fibroids vary and their impact on reproduction would be almost impossible to stratify. In general, there is consensus that submucosal fibroids or those that distort the uterine cavity do have a detrimental impact on fertility outcome. However, the quality of evidence to support this is weak and the significance of benefit has been brought into question in a recent Cochrane review [7]. An additional issue is that uterine cavity distortion is not restricted to submucosal fibroids and some intramural fibroids do cause significant distortion to the uterine cavity. Studies that set out to examine the

impact of intramural fibroids go to great lengths to ensure that uterine cavity distortion is excluded with a high-quality or reliable test. This may have resulted in exclusion of a subgroup of women who have intramural fibroids with cavity distortion, and the published systematic reviews do not provide a clear outcome analysis for this group. In this chapter, the evidence from published literature is critically analyzed to attempt to provide guidance to clinicians as to how intramural fibroids can be managed in women undergoing IVF treatment.

Data from Published Systematic Reviews Several major reviews have been published on the impact of fibroids on reproductive outcomes in the past decade [2–6]. Three of these reviews included studies that looked at the impact of all types of fibroids on both spontaneous pregnancy and IVF treatment outcomes [2–4], whereas the other two specifically looked at studies that analyzed the impact of intramural fibroids not distorting the uterine cavity on the outcome of IVF treatment [5, 6]. Somigliana et al. [2] reviewed the published literature related to fibroids and reproduction. In one of their analyses, they carried out a meta-analysis of 15 articles on IVF outcome and fibroids. Seven of these articles reported IVF outcome separately for intramural fibroids and the meta-analysis showed a small but significant detrimental impact of intramural fibroids on conception (OR 0.8, 95% CI 0.6–0.9) and delivery (OR 0.7, 95% CI 0.5–0.8) rates following IVF/ intracytoplasmic sperm injection (ICSI) treatment. They noted that the mean or median diameter of fibroids in the included studies was rarely above 3 cm and that the detrimental impact emerging from the published articles may have been an underestimation of the real impact. The latter opinion was based on the finding that the negative impact was seen in women

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with fibroids > 4 cm [8]. Somigliana et al. also made reference to a nonrandomized comparative study by Bulletti et al. [9], who found higher cumulative clinical pregnancy (33 percent versus 15 percent) and delivery (25 percent versus 12 percent) rates after one to three cycles of IVF treatment in women who underwent myomectomy for intramural fibroids > 5 cm compared to those who decided against myomectomy. Klatsky et al. [3] carried out a similar metaanalysis of 19 studies that were mostly included in the previous systematic review. These studies compared the IVF outcome in women with intramural fibroids of 1–8 cm with those controls without fibroids. Most studies included women with relatively small fibroids of 2–3 cm. The meta-analysis showed a significant decrease in implantation (OR 0.79, 95% CI 0.71–0.88) and clinical pregnancy rates (OR 0.84, 95% CI 0.74–0.95) and an increase in miscarriage rates (OR 1.82, 95% CI 1.43–2.30). The authors urged caution in interpreting the results as they pointed out that meta-analysis may replicate and amplify biases in each study. Pritts et al. [4] analyzed 23 studies that mostly gave IVF/ICSI-related outcomes. Twelve of these studies included outcomes related to intramural fibroids. These studies showed lower clinical pregnancy rates (OR 0.81, 95% CI 0.70–0.94), ongoing pregnancy/live birth rates (OR 0.70, 95% CI 0.58–0.85) and implantation rates (OR 0.68, 95% CI 0.59–0.80) and higher miscarriage rates (OR 1.75, 95% CI 1.23–2.49) compared to control women without fibroids. When only prospective studies or studies that assessed the uterine cavity distortion with hysteroscopy or sonohysterography were examined, clinical pregnancy rates were no longer significantly different, while the implantation rates remained significantly lower in women with intramural fibroids. Two studies that assessed the clinical pregnancy rates and one that gave the ongoing/live pregnancy rates showed that myomectomy for intramural fibroids did not improve the outcomes compared to controls with in situ fibroids. Sunkara et al. [5] published an analysis of 19 studies on the impact of non-cavity-distorting intramural fibroids on IVF outcome. They found significant reductions in live birth (OR 0.79, 95% CI 0.70–0.88) and clinical pregnancy (OR 0.85, 95% CI 0.77–0.0.94) rates in women with fibroids compared to women without fibroids. Implantation and miscarriage rates were not statistically different. The studies included in this

article had analyzed data from women with fibroids of 0.4–8.0 cm, the majority being less than 5 cm. Metwally et al. [6] carried out a further analysis of the published studies on the effect of intramural fibroids on assisted reproduction technology (ART) treatment using stricter criteria. Inclusion criteria were presence of a control group, analysis of intramural fibroids separately (not grouping them together with subserosal fibroids) and exclusion of submucosal fibroids by assessing the endometrial cavity with an objective method (hysteroscopy, hysterography, ultrasonography and sonohysterography). With this approach they included only 10 studies from a similar period of publication year to the previous four systematic reviews. The analysis of nine studies that gave the outcome of ART treatment showed no differences in live birth and miscarriage rates, but demonstrated lower clinical pregnancy rates in women with fibroids (OR 0.60, 95% CI 0.42–0.87). When further sensitivity analyses were carried out to include only the studies where age was not a confounding factor and/or studies that used a high-quality method (hysteroscopy or sonohysterography) to exclude cavity involvement, no differences in the live birth, clinical pregnancy or miscarriage rates were found between women with and without fibroids. Importantly, four studies that gave the size of fibroids included women with fibroid size of 5 cm or less. It appears that, despite some degree of differences in the conclusions of these systematic reviews, the common finding is that the presence of fibroids probably has a detrimental impact on the outcome of IVF.

Significance of Size of Fibroids As mentioned earlier, a common feature in these reviews is that the majority of studies included only women with relatively small intramural fibroids, probably because women with larger fibroids underwent a myomectomy. Hence, the published literature may be underestimating the impact of intramural fibroids, particularly the larger ones. Only a few studies attempted to assess the impact of fibroid size. Oliviera et al. [8] found significantly lower clinical pregnancy rates after IVF/ICSI in women with intramural or subserosal fibroids of 4.1–6.9 cm compared to women with no fibroids or fibroids ≤ 4 cm. There was no difference in pregnancy rates between the control group and women with fibroids ≤ 4 cm. Women with fibroids of ≥ 7 cm were excluded.

How to Manage Intramural Fibroids before an IVF Cycle

Another retrospective study of impact of fibroids not distorting the cavity found that delivery rates were lower in the presence of fibroids > 2.85 cm whilst there was no detrimental impact in the presence of smaller fibroids [10].

Mechanism of Action The actual mechanism of how intramural fibroids may affect ART outcome is not known. It is possible that the presence of fibroids affects uterine contractility, intrauterine environment or endometrial receptivity through endocrine, paracrine mechanisms or inflammatory pathways [2]. In addition, larger fibroids may affect ovarian accessibility for the purpose of egg collection. This latter point is less well recognized in the published literature, but is an important clinical challenge in the presence of some fibroids. This may result in a lower number of collected oocytes and may occasionally force clinicians to perform transabdominal egg collections.

Impact of Myomectomy Evidence on the potential benefit of myomectomy prior to ART for women with intramural fibroids is scarce. Only one comparative nonrandomized study assessed the potential benefit of myomectomy prior to IVF [9]. One hundred sixty-eight women with at least one fibroid > 5 cm were allowed to choose between myomectomy and expectant management prior to IVF. Submucosal fibroids were excluded, but it is likely that some women had subserosal fibroids. In the 84 women who had a myomectomy, clinical pregnancy (33 percent versus 15 percent, 0.05 < p) and delivery (25 percent versus 12 percent, p < 0.05) rates were significantly lower compared to the other 84 women who did not have surgery after one to three cycles of IVF treatment. Myomectomy is a relatively frequently performed procedure, particularly in the presence of symptomatic fibroids. However, questions remain as to its effect on fertility and outcome of ART. While the potential harm of postoperative pelvic adhesions on spontaneous conceptions is well recognized, the impact of myometrial trauma or intrauterine adhesions after myomectomy on IVF is less recognized. Potential benefits of laparoscopy against laparotomy for myomectomy have been well established in a number of randomized controlled trials. In comparison to traditional open myomectomy, the

laparoscopic approach is associated with less postoperative pain and fever, and shorter hospital stay at the expense of longer operating times [11]. Other potential advantages of the laparoscopic approach include a shorter recovery time with a quicker return to activities of daily living [12]. Nevertheless, myomectomy is still a major operation and is associated with significant morbidity. Furthermore, the women are usually advised to avoid a pregnancy for at least three months postoperatively, resulting in delays in the planned IVF treatment. This may potentially be an issue for older women, particularly for those with reduced ovarian reserve.

Conclusions and a Practical Approach to Management in Clinical Practice The published evidence on the impact of intramural fibroids on IVF outcome is suggestive of a detrimental impact; however, this is based on relatively lowquality studies that show significant variability in inclusion/exclusion criteria and outcome parameters. This is hardly surprising, considering that fibroids come in different numbers, size and consistency. While there is a need to perform prospective randomized studies in this field, this is likely to be extremely challenging due to a high number of confounding factors. The majority of published studies included women with relatively small intramural fibroids, hence there is a significant possibility that the detected impact in the systematic reviews is an underestimation. Currently there is very little evidence from controlled studies on the benefit of myomectomy for intramural fibroids prior to IVF treatment, although the procedure is relatively frequently performed. It is quite likely that numbers needed to treat (NNT) for this purpose would be very high for small fibroids, while the NNT would be lower for larger fibroids. This point would need to be taken into account when decisions are made on myomectomy, and potential benefits should be weighed against the associated morbidity, cost and delay in treatment. In our practice we take a number of factors into account when we counsel our patients who have intramural fibroids that do not distort the cavity prior to IVF treatment. These include the age of the woman, her ovarian reserve, the number and size of fibroids, the overall size of the uterus, history of previous surgery and ovarian accessibility. We try to avoid

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surgery in the presence of fibroids < 5 cm when the uterine cavity is regular. We tend to offer surgery first to women with fibroids ≥ 7 cm but proceed with IVF treatment without surgery in the presence of fibroids of 5–6 cm in the first IVF attempt. We usually offer surgery for fibroids of 5–6 cm, if the woman had one or two failed IVF attempts. With this approach we aim to keep the NNT as low as possible per additional pregnancy achieved. If there are difficulties with ovarian accessibility due to fibroids, we prefer surgery before IVF. We usually wait for three months before proceeding with IVF postoperatively but in older women with reduced ovarian reserve, we proceed with IVF earlier and freeze embryos for delayed transfer.

Suggested Standard Operating Protocol (SOP) –

Management of women with subfertility and fibroids

Suggested Patient Information Sheet –

Subfertility and fibroids

References 1. D. D. Baird, D. B. Dunson, M. C. Hill, D. Cousins and J. M. Schectman. High cumulative incidence of uterine leiomyoma in black and white women: Ultrasound evidence. Am. J. Obstet. Gynecol. 2003;188:100–7. 2. E. Somigliana, P. Vercellini, R. Daguati, R. Pasin, O. De Giorgi and P. G. Crosignani. Fibroids and female reproduction: A critical analysis of the evidence. Hum. Reprod. Update 2007;13:465–76. 3. P. C. Klatsky, N. D. Tran, A. B. Caughey and V. Y. Fujimoto. Fibroids and reproductive outcomes: A systematic literature review from conception to delivery. Am. J. Obstet. Gynecol. 2008;198:357–66.

4. E. A. Pritts, W. H. Parker and D. L. Olive. Fibroids and infertility: An updated systematic review of the evidence. Fertil. Steril. 2009;91:1215–23. 5. S. K. Sunkara, M. Khairy, T. El-Toukhy, Y. Khalaf and A. Coomarasamy. The effect of intramural fibroids without uterine cavity involvement on the outcome of IVF treatment: A systematic review and meta-analysis. Hum. Reprod. 2010;25:418–29. 6. M. Metwally, C. M. Farquhar and T. C. Li. Is another meta-analysis on the effects of intramural fibroids on reproductive outcomes needed? Reprod. Biomed. Online 2011;23:2–14. 7. J. Bosteels, J. Kasius, S. Weyers, F. J. Broekmans, B. W. Mol, and T. M. D’Hooghe. Hysteroscopy for treating subfertility associated with suspected major uterine cavity abnormalities. Cochrane Database Syst. Rev. 2015;21:CD009461. 8. F. G. Oliveira, V. G. Abdelmassih, M. P. Diamond, D. Dozortsev, N. R. Melo and R. Abdelmassih. Impact of subserosal and intramural uterine fibroids that do not distort the endometrial cavity on the outcome of in vitro fertilization intracytoplasmic sperm injection. Fertil. Steril. 2004;81:582–7. 9. C. Bulletti, D. De Ziegler, P. L. Setti, E. Cicinelli, V. Polli and M. Stefenatti. Myomas, pregnancy outcome, and in vitro fertilization. Ann. N. Y. Acad. Sci. 2004;1034:84–92. 10. L. Yan, L. Ding, C. H. Li, Y. Wang, R. Tang and Z. J. Chen. Effect of fibroids not distorting the endometrial cavity on the outcome of IVF treatment: A retrospective cohort study. Fertil. Steril. 2014;101: 716–21. 11. P. Bhave Chittawar, S. Franik, A. W. Pouwer and C. Farquhar. Minimally invasive surgical techniques versus open myomectomy for uterine fibroids. Cochrane Database Syst. Rev. 2014;10: CD004638. 12. T. Tulandi and H. Youseff. Laparoscopy-assisted myomectomy of large uterine myomas. Gynaecological Endoscopy. 1997 Apr. 1;6:105–8.

Chapter

4

Ovarian Stimulation Protocols Horses for Courses Ying Cheong

Introduction Many in vitro fertilization (IVF) centers in the United Kingdom have protocols they predominately follow for several reasons. First, with a standardized protocol, the clinics develop clinical, organizational and administrative memory over a shorter period of time. This cumulative experience with protocol troubleshooting and issues relating to protocol deviation is immensely valuable in the context of treatment outcomes and patient journey as well as operational efficiency. Second, any changes made to a single standardized protocol can be introduced in a single step-wise manner, reducing the potential for confusion and error. Third, it simplifies other, nonclinical aspects of clinic management, including finance, accounting and auditing. However, many other IVF centers in the United Kingdom and worldwide are led by an individual clinician, each with his own clinical protocol variation, which the patient and clinic nurses follow. Hence, from a clinic management perspective, there needs to be a good balance between diversification of protocols and streamlining, embedded within the appropriate management and robust governance oversight.

Summary of the Current Evidence Long-Agonist or Antagonist Protocol? Long-agonist protocol: • Yields more eggs and a higher pregnancy rate than the short-agonist protocol (but the shortantagonist protocol is no longer commonly practiced). • Allows advanced, predictable scheduling of IVF cycles. • Commences during the luteal phase of the previous cycle, with longer duration and more side effects for patients. Antagonist protocol:

•

•

•

Recent randomized studies comparing the gonadotropin-releasing hormone (GnRH) longagonist protocol and the GnRH antagonist protocol have shown no significant differences in live birth rates. Associated with a significantly lower probability of ovarian hyperstimulation syndrome (OHSS) resulting in morbidity and hospital admission compared to the long-agonist protocol. Allows for GnRH agonist ovulation trigger in those at risk of OHSS.

Which Follicle-Stimulating Hormone to Use Follicle-stimulating hormones (FSH) activate the FSH receptors within the follicles to stimulate follicular growth. Different formulated brands can vary in terms of the recombinant preparations (Follitropin alfa, beta or delta) or purity (purified versus highly purified FSH). With the expiry of patents, the marketplace is now flooded with different biosimilar brands. These may differ in strength and purity and contain different composition of isoforms and/or various glycosylation profiles. Some clinicians have expressed concern over potential alterations in clinical efficacy or safety. However, one does need to bear in mind that the introduction of drugs into most developed countries, including the United Kingdom, is rigorously regulated. In practice, the aforementioned small differences do not affect bioactivity compared to natural hFSH and are thus unlikely to result in significant clinical differences. The more important factors are how familiar the clinic is with the product, if the method and mode of administration of the drug are patient friendly and if the price is economically attractive. Indeed, practical issues influencing the market such as ease of product availability and streamlining of distribution and

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product support available from the companies may be more influential than any subtle clinical variations. Regardless, the evidence currently shows: • In long-agonist protocol, urinary human menopausal gonadotrophin (hMG) versus recombinant FSH (rFSH), the live birth rate was significantly higher with the hMG group. • In antagonist treatment, no difference appeared in live birth rates between hMG and rFSH.

Which Protocol to Choose for the Patient at a Consultation The choice of protocol during a consultation depends on several factors: 1) Has the patient had previous treatment(s) and what was her response? Varying the protocol from previous protocols that may not have worked as well can be useful, although it does not guarantee success. 2) Did the patient suffer any side effects from previous treatment? If she had significant side effects from downregulation, then changing to the antagonist protocol will reduce this risk. 3) In a patient who has never received ovarian stimulation before but has endometriosis, the use of downregulation for three to six months can improve her pregnancy rates, although the evidence for this practice is based on studies with small sample size. 4) In a patient who has low ovarian reserve, no one protocol will reliably fix this challenge, and the patient has to be counseled accordingly with careful management of expectations. The evidence based around more egg yield using the long protocol was not based on a study population with low ovarian reserve. 5) In the gold standard patient having her first IVF treatment, follow the individual clinic’s preferred protocol. This will generally mean a smoother, streamlined pathway for the patient, as the clinic will have systems in place to deal with all aspects of this treatment protocol.

What Dosage to Give A proficient clinic will have a standard operating procedure (SOP) to guide clinical staff on dosing regimes. The SOP will set out dosing range for the good

prognosis patients, poor prognosis patients and for those at risk of OHSS. The dosage will be graduated from low doses, e.g. FSH of 112.5 IU daily, to that of around 300 IU daily. Note there is no value in increasing the FSH dosage much above 300 IU daily as studies have shown that beyond this dose, the egg yield plateaus and the associated increased in drug expenditure comes without the necessary clinical benefit. Using GnRH as an ovulation trigger: One of the most significant and high-risk side effects of ovarian stimulation is OHSS. Coasting and cryopreservation are the two methods currently used to prevent OHSS. Recently, Cabergoline used in low doses for eight days from the day of human chorionic gonadotropin (hCG) administration was shown to reduce the rate of OHSS compared with placebo. Studies revealed the incidence, but not the severity, of OHSS was reduced by Cabergoline treatment, without reducing pregnancy rate. Many centers worldwide have also used buserelin as an ovulation trigger to reduce the rate of OHSS. Gonadotrophin-releasing hormone agonist can be used as an alternative trigger to hCG cycles that have been suppressed by GnRH antagonist action. Administration of a GnRH agonist displaces the GnRH antagonist in the pituitary gland, activating GnRH receptors and resulting in a surge of gonadotrophins similar to that of a natural cycle or an hCG trigger. However, the luteinizing hormone (LH) surge in a natural cycle has a total duration of around 48 hours. The LH surge duration using a traditional hCG trigger is up to five days, but that of a GnRH agonist trigger only 24–36 hours. Gonadotrophin-releasing hormone agonist triggers therefore have the advantage of a lower rate of OHSS. This advantage has been clearly demonstrated in oocyte donation cycles, where OHSS is not a complication in the oocyte donors, while the pregnancy rates of oocyte recipients were equivalent to those with an hCG trigger. While the shorter duration of a GnRH trigger does not have a detrimental impact on oocyte function, the reduced level of endogenous LH during the early luteal phase has an unfavorable impact on corpus luteal function and the endometrium’s ability to sustain early implantation. In order to overcome the latter issue, two solutions exist. First, adopt a “freeze all” policy for these patients and replace their embryos in a frozen embryo transfer cycle, although this is potentially more costly for self-paying patients. Second,

Ovarian Stimulation Protocols: Horses for Courses

supplement the lower LH concentrations with lowdose hCG injections (e.g., 1500 IU). Several studies have reported success with this strategy although it does not completely eliminate the risk of OHSS.

Conclusion The choice of ovarian stimulation protocol is indeed horses for courses. Individualization is crucial and clinicians also need to remember that at the end of the day, the patient’s experience throughout her IVF journey is as important and sometimes arguably matters more to the patient than the clinical outcome.

Suggested Standard Operating Protocol (SOP) –

Ovarian stimulation protocol

Suggested Audit –

Ovarian stimulation protocol deviation

4. E. M. Kolibianakis, J. Collins, B. C. Tarlatzis, P. Devroey, K. Diedrich and G. Griesinger. Among patients treated for IVF with gonadotrophins and GnRH analogues, is the probability of live birth dependent on the type of analogue used? A systematic review and meta-analysis. Human Reproduction Update 2006;12(6):651–71. 5. H. G. Al-Inany, M. A. Youssef, M. Aboulghar, F. Broekmans, M. Sterrenburg, J. Smit et al. Gonadotrophin-releasing hormone antagonists for assisted reproductive technology. Cochrane Database Syst. Rev. 2011(5):CD001750. 6. S. Kol. Embryo implantation and GnRH antagonists: GnRH antagonists in ART: Lower embryo implantation? Hum. Reprod. 2000;15(9): 1881–2. 7. K. Zikopoulos, E. M. Kolibianakis, M. Camus, H. Tournaye, E. Van den Abbeel, H. Joris et al. Duration of gonadotropin-releasing hormone antagonist administration does not affect the outcome of subsequent frozen-thawed cycles. Fertility and Sterility 2004;81(2):473–5.

1. S. Daya. Gonadotropin releasing hormone agonist protocols for pituitary desensitization in in vitro fertilization and gamete intrafallopian transfer cycles. Cochrane Database Syst. Rev. 2000(2):CD001299.

8. P. Devroey, M. Aboulghar, J. Garcia-Velasco, G. Griesinger, P. Humaidan, E. Kolibianakis et al. Improving the patient’s experience of IVF/ICSI: A proposal for an ovarian stimulation protocol with GnRH antagonist co-treatment. Hum. Reprod. 2009;24 (4):764–74.

2. H. G. Al-Inany, A. M. Abou-Setta, M. A. Aboulghar, R. T. Mansour and G. I. Serour. Efficacy and safety of human menopausal gonadotrophins versus recombinant FSH: A meta analysis. Reproductive Biomedicine Online 2008;16(1):81–8.

9. M. van Wely, I. Kwan, A. L. Burt, J. Thomas, A. Vail, F. Van der Veen et al. Recombinant versus urinary gonadotrophin for ovarian stimulation in assisted reproductive technology cycles. Cochrane Database Syst. Rev. 2011(2):CD005354.

References

3. G. Griesinger, R. Felberbaum and K. Diedrich. GnRH antagonists in ovarian stimulation: A treatment regimen of clinicians’ second choice? Data from the German national IVF registry. Hum. Reprod. 2005;20(9):2373–5.

10. P. Humaidan. Luteal phase rescue in high-risk OHSS patients by GnRHa triggering in combination with low-dose HCG: A pilot study. Reproductive Biomedicine Online 2009;18(5):630–4.

13

Chapter

5

The Consultation with the Poor Responder Paul Atkinson and William L. Ledger

Introduction Poor response to gonadotropin therapy for superovulation carries with it a lower chance of pregnancy after in vitro fertilization (IVF) and significant distress for the patient/couple concerned. By definition, the consultation with the “poor responder” patient will occur after at least one and frequently several cycles of gonadotropin superovulation and must be handled with sensitivity and empathy while maintaining an objective yet sympathetic approach. The term “poor responder” itself is pejorative and is often best avoided, along with labels such as “inadequate,” “failure” and “insufficiency.” The prevalence of poor responders is reported to be between 5.6 and 35.1 percent [1]. The variation in reported prevalence is largely due to differences in the definition of a “poor responder.” A review in 2011 identified no fewer than 41 different definitions used in 47 trials [2]. In response to this lack of clarity, a European Society of Human Reproduction and Embryology (ESHRE) working group devised a common definition for “poor responder” that represents the first necessary step to harmonize research leading to development of evidenced-based treatment strategies for these patients [3]. The Bologna criteria are summarized in Table 5.1. It is important to note that the Bologna criteria were primarily developed to give uniformity to inclusion criteria for clinical trials so that valid conclusions could be extrapolated from research, rather than for use in clinical practice. The criteria have been criticized due to their pooling of a number of heterogeneous groups of patients, both young and old, with potentially different prognoses that require different treatment strategies. In general, poor responders have much lower pregnancy rates compared to normal responders, 14.8 percent versus 34.5 percent in one systematic review [1]. However, the same review highlighted that poor responders are not a homogeneous group. Older poor responders have lower pregnancy rates than younger (1.5–12.7

percent versus 13.0–35 percent), reflecting the importance of age-related oocyte quality as a prognostic factor.

Etiology and Heterogeneity The common and obvious cause of ovaries responding poorly to superovulation is their having a reduced number of follicle-stimulating hormone (FSH)sensitive antral follicles, or reduced ovarian reserve. However, in some patients it may be due to suboptimal exposure to gonadotropins, especially in overweight women, or the follicles being less sensitive to exogenous FSH due to receptor subtypes. The definition of poor responders should more accurately be divided into those patients who have an expected poor response, as predicted by a low AMH and AFC, and less so by age; and those that have an unexpected poor response, i.e. the younger patient with normal ovarian reserve tests who fails to achieve more than four oocytes during stimulation. The etiologies are different and the prognosis for the latter group is more favorable.

Predictive Tests of Poor Response Other than age, a number of tests are useful in the prediction of poor response. The most clinically useful are AMH, AFC and day 3 FSH coupled with estrogen. Inhibin B, clomiphene challenge tests and ovarian volume are not useful. AFC and AMH have the best sensitivity and specificity, but it is important to note that the false positive rate for the prediction of poor response is 10–20 percent [4]. The results of these tests should not be used initially to exclude patients from access to stimulation cycles but as tools for advising on likely response and to help in gonadotropin dose selection. In health systems where economic considerations are at the forefront, extreme cut-off values are preferred before denying treatment as they have higher specificity and a lower false positive rate.

The Consultation with the Poor Responder

Table 5.1 Bologna Criteria for Poor Responders

Table 5.2 Stimulation protocols and adjuvants used in poor responders

Bologna Criteria: Requires two of three 1. ≥ age 40 or other risk factor for low ovarian reserve e.g. chemotherapy 2. poor ovarian response, ≤ three oocytes with conventional stimulation 3. abnormal ovarian reserve test (AMH < 1.1 ng/mL or AFC < 7) Also defined if during at least two previous stimulation cycles using ≥ 300 IU gonadotropins, < 4 oocytes achieved

When measuring the day 3 serum FSH it is important to couple it with a measurement of estrogen so as not to be falsely reassured by a normal FSH that is in fact suppressed by elevated serum estrogen released due to early follicle recruitment, as is often seen with very diminished ovarian reserve, when the premature rising estrogen provides negative feedback and a normalizing of serum FSH. The estrogen level should be basal for the FSH to be interpretable. Pregnancy rates for poor responders with an FSH above 12.0 IU/L are significantly lower compared to those with normal levels (4.0 percent versus 14.8 percent), even after correction for age [5].

Use of Outcomes of Previous Cycles to Predict Poor Response The best test of poor ovarian response is a dynamic test, specifically the response during the first IVF cycle. However, this also needs to be looked at critically, especially when dealing with the younger patient who had an unexpected poor response. In expected poor responders, namely older women with reduced ovarian reserve, a decrease in pregnancy rate was observed in subsequent cycles after the initial poor response cycle, with rates of 7–9 percent in the second cycle and no pregnancies in the third cycle [1]. Based on these data, older patients with low ovarian reserve tests who have experienced two stimulation cycles with a poor response should be counseled that their chance of pregnancy in subsequent cycles is poor and donor options should be explored. This is in contrast to unexpected poor responders. These patients showed an increase in pregnancy rate from 11–22 percent in the second cycle to 21–25 percent in the third cycle [1]. There is a natural

Stimulation Protocols Used in Poor Responders

Adjuvants Used in Poor Responders

GnRH antagonist

DHEA

Long Down Regulation GnRH agonist

Testosterone

Short “Flare” agonist cycle

Growth Hormone

Microdose Flare

Recombinant LH

GnRH agonist stop protocol

Aspirin

Clomifene FSH protocol

Inositol

Corifollitropin alfa, HMG with Antagonist

Co-enzyme Q

“Elonva Flare”

Estradiol in the luteal phase

Late luteal FSH

variation in the ovarian response to stimulation from cycle to cycle, and with continuing cycles the number of oocytes collected will regress toward the mean. It is largely dependent on how many FSHreceptive follicles are present at the start of stimulation, and this can vary. A recent study showed that up to 75 percent of patients had a significant variation in response to the exact same gonadotropin stimulation over three separate IVF cycles and that this was not predicted by basal FSH or AFC [6].

Stimulation Protocols Used in Poor Responders The existence of a large number of different stimulation protocols (Table 5.2) for poor responder patients demonstrates the lack of consensus among fertility specialists as to how to manage this group of patients optimally. No stimulation protocol has yet been shown to be superior for the treatment of poor responders. The development of a follicle from a primordial to an ovulating follicle takes approximately 120 days, with only the last 16 days or so of follicle development being under the control of gonadotropins. If the follicle is simply not there or is not mature enough to respond, then no stimulation protocol will work. It is helpful to explain this to patients, often with the aid of a diagram.

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Paul Atkinson and William L. Ledger

If follicles are there, then the key to successful superovulation has traditionally been seen as timely and relatively high doses of FSH to maintain serum FSH concentration above the threshold needed to sustain multi-follicular growth. However several studies have shown that increasing the FSH dose much beyond 300 IU is of little benefit [7], although more may be needed in the obese patient. Some authors suggest the gonadotropin-releasing hormone (GnRH) agonist long down-regulation protocol is detrimental in poor responders as it causes excessive ovarian suppression, leading to reduced ovarian response. Trials have studied lowering or stopping the GnRH agonist once pituitary suppression has occurred or alternatively using a flare-up regimen to boost early-cycle FSH and decrease the length of time of suppression. Other clinics, including ours, prefer the use of late-follicular-phase GnRH antagonists so as not to suppress early-cycle FSH. Our research group is interested in the use of a novel stimulation protocol for poor responders. We hypothesize that the unique pharmacodynamic and pharmacokinetic properties of corifollitropin alfa when combined with a short agonist flare cycle, “Elonva Flare,” will maximize FSH exposure at the critical time of antral follicle recruitment and hence lead to a higher oocyte yield and improved pregnancy rates. Our clinical pregnancy rates are encouraging and require validation in a prospective randomized controlled trial (RCT). There are two schools of thought concerning the selection of the optimum stimulation protocol for poor responder patients. Each should be considered a viable strategy depending on the patient, the clinic setup and the overarching health system. The first school of thought is that if no stimulation protocol is superior to another, and if a natural variation exists in response that is more reflective of the ovarian reserve and not the stimulation protocol per se, and that clinical pregnancy rates are low anyway, then the protocol that is the simplest, most patient-friendly and that can be standardized to minimize clinic and health system costs should be used. In standard protocols this would be a GnRH antagonist protocol with consideration of use of corifollitropin alfa to decrease patient burden. Some research groups have gone further and explored even milder stimulation options such as clomiphene with FSH. The per-cycle pregnancy rates are lower, but the burden is less.

The second school of thought is that in patients with low chances of pregnancy per cycle, multiple cycles will usually be needed to achieve a pregnancy. There is a known large dropout rate of patients undergoing IVF, due to the stressful nature of the treatment and its implications [8]. In a patient-driven model of care, the reality is that patients will be reluctant to repeat the same treatment protocol if it initially gave a poor response. In poor responder patients who are thought to still have reasonable chances of success, a strategy is to move between GnRH antagonist, long down-regulation and flare cycles, not because one protocol is superior to another, but in order to keep the patient engaged within the treatment system, thus maximizing their chances of having a successful pregnancy.

Adjuvant Therapies for Poor Responder Patients The addition of adjuvant therapies before superovulation in poor responder patients is based on the concept of trying to increase the number of secondary follicles that enter the pathway of folliculogenesis to reach the point at which they can respond to FSH, i.e. increasing the antral follicle pool. Alternatively, adjuvant use during superovulation is aimed at improving antral follicle development by enhancing steroidogenesis or FSH receptivity. However, there is currently no adjuvant therapy that has been trialed before or during superovulation that has convincingly shown to improve pregnancy rates in poor responder patients. The evidence base for the routine use of different adjuvants, alone or in combination, for poor responder patients is inadequate. There is a lack of robust evidence for most of the adjuvants that have been promoted through clinics and websites, and large, well-designed RCTs are still needed for use of immunotherapy, vasodilators, uterine relaxants, aspirin, heparin, growth hormone and androgens, to list but a few, as adjuvants in IVF. The most promising data for improvement in response to superovulation in poor responder patients, though by no means conclusive, are for pre-cycle treatment with androgens. Androgens, primarily produced by theca cells, have a critical role in follicular steroidogenesis as the substrate for estrogens and subsequent early follicular and granulosa cell development. Androgens may also increase the FSH receptor expression and thus potentially enhance the ovarian response to gonadotropins.

The Consultation with the Poor Responder

A recent Cochrane review [9] of 17 RCTS and 1,500 patients showed that pretreatment with dehydroepiandrosterone (DHEA) or testosterone showed a significantly increased pregnancy rate in poor responders. The quality of the evidence was judged as moderate, and when a sensitivity analysis was done that removed trials at high risk of bias, the effect size was reduced and no longer significant. The addition of recombinant luteinizing hormone (LH) before or during gonadotropin stimulation has been trialed with inconclusive results. It has not been well studied in poor responders. There is no benefit in the older patient [9], but perhaps a benefit in the younger poor responder patient that warrants further research [10]. Growth hormone has been postulated to upregulate the local ovarian synthesis of IGF-1 and amplify the effect of FSH. Small and heterogeneous trials have suggested benefit in poor responders, although the evidence is not clear. A recent Australian trial, although failing to meet its target sample size, showed no improvement with the addition of growth hormone [12]. Other adjuvants including aspirin, co-enzyme Q and inositol have not been shown to improve pregnancy rates when tested in randomized trials [13].

Counseling The role of the fertility specialist during this consultation is to be the guide for the patient, and expertise in psychology and counseling is equally important as expertise in reproductive endocrinology. The key points are summarized in Table 5.3. The first step is to put aside plenty of time for the consultation, to ask open-ended questions and to let the patient speak. While listening the specialist needs to form in their own mind where the patient is on their fertility journey. Is the patient open to the use of donor oocytes or would this be completely unacceptable? It is counterproductive to the doctor–patient relationship to be insistent on donor oocytes when it is clear that this would be unacceptable or not feasible. However, if the doctor feels that this is likely to be what is eventually required, then it is important to plant this seed at an early consultation without laboring the point. It is important to remember that a patient’s attitude toward donor oocytes will often change over time as superovulation cycles produce a persistently poor response.

Table 5.3 Counseling the poor responder

Key points for the counseling of the poor responder Allow plenty of time Ensure both partners are present Use open-ended questions to discover where they are at Plant the seed for the possible need for donor oocytes Use nursing and counseling services If the prognosis is reasonable, engage the patient in the system: – change the stimulation, use adjuvants with an evidence base, enroll in a trial If the prognosis is poor, be realistic and firm Be the guide, don’t be paternalistic

If the patient is part of a couple, then it is vital that both partners are seen together. After a number of superovulation cycles, it is usually the female partner who bears the greater part of the burden of repeated poor response, and she often begins to attend appointments alone. The fertility specialist needs to encourage and at times be insistent that both partners attend. Follow-up should be arranged to discuss cycles that have ended in a poor response and the doctor should lead these discussions, rather than leaving them to nurses or clinic psychologists. Supportive nursing and counseling staff are vital in a well-run fertility unit. Patients will often find it easier to talk openly with nursing staff and counselors about their fears, anxieties and disappointments than with their specialist, and a cohesive team will provide a flow of communication across the unit’s members. The fertility specialist needs to have a clear idea about the prognosis of achieving a pregnancy and be strong in their recommendation if they believe the patient should not undertake further autologous cycles. The fertility specialist and other members of the clinic need to convey a consistent message and remain resolute, which is difficult when faced with desperate and emotional patients and, for some clinics, a financial incentive to continue. It is sometimes kinder to say no and be firm. However, we hold the strong view that the ultimate role of the fertility specialist is not just to achieve

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a pregnancy with a patient’s own gametes but to identify how best to guide a patient through their fertility journey so that they can eventually make peace with themselves. Taking financial considerations into account, it is not unreasonable for a patient to undergo a superovulation cycle when the prognosis is very poor as a psychological treatment to prove to her that pregnancy with autologous oocytes is extremely unlikely to happen. The cycle can always be cancelled prior to oocyte collection if there is poor response to stimulation, putting the patient at minimal risk. An open, generous approach is needed for the consultation for the poor responder. If there is a realistic chance of autologous pregnancy, multiple cycles are usually needed and this requires the patient to stay engaged with the system. Clear counseling and support, changing the stimulation recipe, being open to the use of adjuvants if the patient has faith in them, notwithstanding the poor evidence base, and actively recruiting patients for clinical trials help keep the patient engaged. If the chances are futile, then the fertility specialist needs to stop and listen to where the patient feels she is on her journey, and guide her and her partner toward ultimate acceptance of childlessness, adoption or donation.

Suggested Standard Operating Protocol (SOP) –

4. A. La Marca, G. Sighinolfi, D. Radi, C. Argento, E. Baraldi, A. Artenisio, G. Stabile and A. Volpe. Anti-Mullerian hormone (AMH) as a predictive marker in assisted reproductive technology (ART). Hum. Reprod. Update 2010;16:113–30. 5. J. Galey-Fontaine, I. Cedrin-Durnerin, R. Chaibi, N. Massin and J. Hugues. Age and ovarian reserve are distinct predictive factors of cycle outcome in low responders. Repro. Biomed. Online 2005;10:94–99. 6. L. Rombauts, C. Lambalk, A. Schultze-Mosgau, J. van Kuijk, P. Verweij, D. Gates, K. Gordon and G. Griesinger. Intercycle variability of the ovarian response in patients undergoing repeated stimulation with corifollitropin alfa in a gonadotropin-releasing hormone antagonist protocol. Fertil. Steril. 2015;104: 884–90. 7. M. Berkkanoglu and K. Ozgur. What is the optimum maximal gonadotropin dosage used in microdose flare-up cycles in poor responders? Fertil. Steril. 2010;94:662–5. 8. M. Verberg, M. Eijkemans, E. Heijnen, F. Broekmans, C. de Klerk, B. Fauser and N. Macklon. Why do couples drop-out from IVF treatment? A prospective cohort study. Hum. Reprod. 2008;23:2050–5. 9. H. Nagels, J. Rishworth, C. Siristattidis and B. Kroon. Androgens (dehydroepiandrosterone or testosterone) for women undergoing assisted reproduction. Cochrane Database of Systematic Reviews 2015;11: CD009749.

References

10. T. Konig, L. van der Houwen, A. Overbeek, M. Hendriks, S. Beutler-Beemsterboer, W. Kuchenbecker, C. Renckens, R. Bernardus, R. Schats, R. Homburg, P. Hompes and C. Lambalk. Recombinant LH supplementation to a standard GnRH antagonist protocol in women of 35 years or older undergoing IVF/ICSI: A randomized controlled multicentre study. Hum. Reprod. 2013;28:2804–12.

1. J. Oudendijk, F. Yarde, M. Eijkemans, F. Broekmans and S. Broer. The poor responder in IVF: Is the prognosis always poor? A systematic review. Hum. Reprod. Update 2012;18:1–11.

11. A. Ferraretti, L. Gianaroli, T. Motrenko, E. Feliciani, C. Tabanelli and M. Magli. LH pretreatment as a novel strategy for poor responders. Biomed. Res. Inter. 2014; Article ID 926172.

2. N. Polyzos and P. Devroey. A systematic review of randomized trials for the treatment of poor ovarian responders: Is there any light at the end of the tunnel? Fertil. Steril. 2011;96:1058.e7–61.e7.

12. Norman R. A randomized double blind placebo controlled study of recombinant human growth hormone (r-HGH) on live birth rates in women who are poor responders. Abstract O-082. ESHRE Helsinki 2016.

Management of Poor Responders

Suggested Audit –

Outcomes of poor responders

3. A. Ferraretti, A. La Marca, B. Fauser, B. Tarlatzis, G. Nargund and L. Gianroli. ESHRE consensus on the definition of the “poor response” to ovarian stimulation for in vitro fertilization: The Bologna criteria. Hum. Reprod. 2011;26:1616–24.

13. L. Nardo, T. El-Toukhy, J. Stewart, A. Balen and N. Potdar. British Fertility Society Policy and Practice Committee: Adjuvants in IVF: evidence for good clinical practice. Hum. Fertil. (Camb.) 2015;18:2–15.

Chapter

6

The Consultation with a Patient Who Wishes to Have Ovarian Tissue Cryopreservation Richard A. Anderson

Background

cryopreservation are multiple, but include that it is not readily integrated as part of conventional assisted reproduction practice in in vitro fertilization (IVF)based clinics, and regulatory aspects may be different. The first procedure of storing ovarian tissue for fertility preservation in a cancer patient was performed in 1993 in Edinburgh, following the successful demonstration of the technique in sheep [2]. Subsequently, the first baby born to a woman who had had ovarian tissue reimplanted was in 2004, and in recent years, the number of babies born has increased rapidly, although at the time of writing, it is still fewer than 100 [3, 4]. The key features of ovarian tissue transplantation are as follows:

Ovarian tissue cryopreservation is one of the range of options now available to girls and young women wishing to preserve their fertility in the face of treatment for cancer or other serious conditions (Figure 6.1) [1]. The growing demand for all aspects of fertility preservation has led to a rapid development of the provision of services in this field, although ovarian tissue cryopreservation remains more patchily available. It is, for the moment, regarded as an experimental procedure and while there have been calls for this to change to categorization as an established procedure in adults, it is undeniably experimental when applied to prepubertal girls. The reasons for the more limited development of this approach compared to oocyte and embryo

Figure 6.1 Options for fertility preservation based on cryopreservation, in girls and young women. Adapted from Anderson RA et al. 2015, Lancet Diabetes Endocrinol 2015;3:556–67, reprinted with permission.

Cryopreservation options for female fertility preservation FEMALE

Patient Assessment

Pre-pubertal

Intervention

Laparoscopy/ Ovarian biopsy

Post-pubertal

Ovarian stimulation

Partner/ Donor sperm

Storage

Ovarian Tissue Cryopreservation

Oocyte Cryo

Embryo Cryo

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Richard A. Anderson

• It does not require any pretreatment, thus can potentially be offered with minimal delay to the proposed treatment. • It requires an invasive surgical procedure, i.e. laparoscopy. • There is no minimum age: it is appropriate for prepubertal girls as well as adult women. • As the point is to remove a substantial part of the patient’s ovarian tissue and thus ovarian reserve, and most follicles from the stored tissue will be lost following replacement, the effect of the proposed treatment must be thought to have a greater impact on the ovary than this. • Reimplantation of ovarian tissue offers the potential for restoration of fertility without further need for assisted conception. • Reimplanted ovarian tissue will also be endocrinologically active, thus providing physiological hormone production and obviating the need for hormone replacement therapy (HRT) during the lifetime of the graft. • There is a risk of malignant contamination of the ovarian tissue and thus of recurrence of the disease after tissue replacement. • It is unclear how well it works: at present, it appears that approximately one in three to five women will conceive after ovarian tissue replacement. • The duration of graft function appears very variable, from minimal activity to several years. As ovarian tissue cryopreservation is at present regarded as experimental, then criteria for its use may be appropriate. Table 6.1 shows criteria developed in this regard. They should not be regarded as tablets of stone, but as a starting point for evidencebased development. They have been validated in an analysis of the provision of this service for girls with a new cancer diagnosis, and showed a very clear discrimination between those who subsequently did not develop premature ovarian insufficiency (POI) versus those at high risk of POI [5]. There are no validated criteria using fertility as the outcome.

Considerations for Patient Selection The considerations behind the criteria in Table 6.1 are as follows: • Age. This reflects the decline in the number of follicles within the ovary with age. At present, all reported live births have been from women in their 20s or early 30s [4, 6]. Assessment of the ovarian

Table 6.1 Factors for ovarian tissue cryopreservation consideration

• Intrinsic factors –Health status of patient –Consent (patient/parent) –Age –Assessment of ovarian reserve • Extrinsic factors –Nature of predicted treatment • (high/medium/low/uncertain risk) –Expertise/options available

reserve using either biomarkers such as antimüllerian hormone (AMH) or antral-follicle count (AFC) may supplement this information, but it should be recognized that age is the key determinant of oocyte quality. Evidence from animal studies shows that approximately two-thirds of the follicles present in the ovary will not survive re-engraftment, during the time the ovarian tissue is required to be re-vascularized. • It is clearly the case that the ideal situation will be that the patient has optimal ovarian function, i.e. that it has not been compromised by previous chemotherapy. Nevertheless, it may well be that the referral occurs at a time of relapse of disease, when a patient has already had a course of chemotherapy. Commonly, this might be for a woman with Hodgkin’s lymphoma who has been treated with the relatively non-gonadotoxic regimen of ABVD, but her relapse requires treatment with high doses of chemotherapy or radiotherapy to the pelvis. There are very substantial differences in the risks associated with various chemotherapeutic regimens, which are discussed briefly later in this chapter, but an additional consideration is the interval since the last course of chemotherapy and the proposed tissue cryopreservation procedure. The issue is the unknown degree of damage to oocytes in primordial follicles, which may be functionally compromised by treatment. In vivo this appears potentially recoverable with some chemotherapy regimens, but how much extra loss will be incurred with the additional stress of cryopreservation is unclear. There have, however, been pregnancies reported following storage and replacement of tissue in women who had already received chemotherapy; therefore, this

Ovarian Tissue Cryopreservation

•

•

•

•

needs to be considered carefully on an individual case basis. A key related criterion is consideration of the risk of loss of future fertility for the patient. A framework for assessment of risk is provided in Table 6.2 [7]. This divides risk into those factors that are intrinsic to the patients, and those that are extrinsic and particularly related to the nature of the predicted treatment. Treatments regarded as being of high risk include those involving administration of high doses of alkylating agents, and radiotherapy affecting the pelvis, including doses from spinal irradiation. There may or may not be precise information on this available at the time of consultation. It is important to remember that most girls and young women treated for cancer will, in fact, retain their fertility, as recently demonstrated in a large study of female survivors of childhood Hodgkin’s lymphoma, where the chance of parenthood was identical to that of the background population, although pelvic radiotherapy in particular clearly compromised this [8]. The patient should have a good prognosis. Whether that means the treatment should always be given with the intention of cure is debatable, as long-term survival may be the more realistic outcome. It is important to consider the potential for the contamination of the ovarian tissue by the malignancy. This has been well described in leukemia and is possible, although much less likely in solid malignancies [9]. It is imperative that a tissue biopsy is sent for a pathological examination at the time of surgery, and depending on the diagnosis it may also be possible to use specific molecular markers to check for contamination. At present, any evidence of contamination would preclude replacement, and require in vitro maturation of the follicles to allow pregnancy. This remains a long way from clinical practice at the moment, and therefore many working in the field would not offer tissue cryopreservation to patients with leukemia with evidence of active disease. Storage of ovarian tissue after initial cycles of chemotherapy has been described, but it remains unclear how safe and effective this approach is. It may also be pertinent to consider access to fertility preservation procedures in the context of access to other state-funded assisted reproduction techniques. At present, in the United Kingdom, criteria for access to assisted reproduction are

Table 6.2 Potential criteria for ovarian tissue cryopreservation

• Age < 35 years • No previous chemotherapy (or low risk) • High (> 50 percent) risk of ovarian failure – High-dose alkylating agents – Radiotherapy to pelvis • Good (> 50 percent) chance of survival

regarded as not applicable to patients seeking fertility preservation at the time of storage of gametes or gonadal tissue, but may well be applicable at the time that those tissues or cells are used. If there is likely to be an issue over this, it will be important to make the patient aware of this consideration.

Assessment of Risk to Fertility (Table 6.2) • As ovarian tissue cryopreservation requires a laparoscopy, there is the need to consider the surgical fitness of the patient. Patients may be anemic, neutropenic or thrombocytopenic, adding to the surgical risk, or there may be mediastinal involvement or posterior fossa tumors that may impact the anesthetic aspects of the risk. This should be clearly discussed with the patient, highlighting that this is an additional procedure that the patient is choosing to undergo rather than something that is absolutely necessary for her medical care. This risk may be tempered if the laparoscopy can be combined with some aspect of her cancer care, for example, insertion of a Hickman line. • This leads to consideration of issues of consent, which will be particularly pertinent for children when assessment of competence is relevant. The experimental nature of this procedure, particularly in this patient group, is again of key importance. • The relevance of the age of the patient and the potential value of assessment of the ovarian reserve have been discussed earlier in this chapter. Both AMH and AFC may be reduced in patients who are unwell compared to healthy patients of the same age. However, they may be of particular value in assessing patients who have already received chemotherapy that may have had a more marked

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Richard A. Anderson

effect on ovarian function than anticipated. Thus, demonstration of a very low AMH or AFC may make the storage of ovarian tissue much less likely to be successful in terms of subsequent return of fertility, which may inform the patient’s decision as to whether to proceed or not. • The importance of assessing the predicted treatment has been discussed in terms of its effect on ovarian function, but the need for uterine function to support a pregnancy should also be recognized. This is relevant to pelvic radiotherapy and total body irradiation. Radiotherapy to the uterus increases the risk of early and late miscarriage, prematurity, growth retardation and stillbirth [10]. These risks are dose related, and also greater in the prepubertal child as uterine growth at puberty may be significantly compromised. • Availability: at present, ovarian tissue cryopreservation is only available in a few centers, both globally and within the United Kingdom, and thus access within the finite timescale available to these patients may be limiting.

The Procedure Ovarian biopsy for cryopreservation is generally performed at laparoscopy, thus in adult women can generally be performed as a day case procedure. Centers differ in their surgical approach, with some performing a unilateral oophorectomy as routine, whereas others more commonly take ovarian cortical biopsies, usually in the form of strips of tissue that can be readily dissected from the ovary with minimal bleeding. Discussion of this will also include normal postoperative care, and chemotherapy can often be started a couple of days after the procedure. The patient should be informed as to where the tissue may be stored, and what follow-up arrangements are to be arranged. Storage of tissue under these circumstances may be for a protracted period, and therefore robust processes are needed to ensure its safekeeping.

What about Replacement? While the consultation will necessarily focus on the process of the surgical procedure to take the ovarian tissue for cryopreservation, due consideration should also be given to what might happen at the time of replacement. At present, the most successful procedure appears to be orthotopic replacement of the ovarian tissue within the pelvis, as in most instances

heterotopic replacement has been unsuccessful [3]. A further surgical procedure within the pelvis will therefore be necessary. One significant advantage of ovarian tissue cryopreservation is that it may give the opportunity for natural conception, and indeed a number of cases have been reported where women have had two or more children following tissue replacement. Any aspect of treatment that will compromise uterine (or tubal) function should therefore be considered. While surrogacy may potentially be used for those patients in whom ovarian tissue can be replaced but who do not have a uterus, there are no reports of this being undertaken successfully at present.

Suggested Documentation The following list provides a starting point: 1. Referral form, including • Pubertal status/reproductive history/current contraception • Past and proposed treatment regimens • Estimated risk to fertility 2. Patient information sheets: age-specific. This may include specific versions for children and adolescents, as well as adults. 3. Checklist covering options discussed and the issues associated with each. This may also be incorporated into the patient information sheet to allow her to take home a summary of the discussion for later consideration. 4. Appropriate consent forms (also age-specific) 5. Treatment plan: may be useful to include a checklist that all relevant aspects and parties have been included. Theatre booking Anesthetic review if necessary Tissue cryopreservation staff Research laboratory if appropriate 6. Documentation of outcome: operation note and documentation of tissue storage (where, how much, in what form, cryopreservation methodology details). 7. Letter to patient, referring oncologist/other clinician and general practitioner (GP) detailing outcome. 8. Plan for follow-up of patient 9. Laboratory plan for patient contact/need for ongoing tissue storage.

Ovarian Tissue Cryopreservation

Conclusion The patient presenting for discussion of ovarian tissue cryopreservation presents a very specific range of issues that need to be discussed in a time-pressured situation. The use of specific written information material is essential under these circumstances to allow her to consider the options and the pros and cons of proceeding after the consultation. In some cases, a telephone conversation either preceding the main consultation or to follow-up to pick up ongoing issues thereafter may be of considerable value to prevent the need for further detailed discussion on the day of proposed surgery.

Suggested Audits Completeness of referral information (e.g. future treatment plan) Characterization of patients completing tissue cryopreservation versus other fertility preservation strategies Surgical outcomes Later ovarian/reproductive function.

References 1. R. A. Anderson, R. T. Mitchell, T. W. Kelsey, N. Spears, E. E. Telfer and W. H. Wallace. Cancer treatment and gonadal function: Experimental and established strategies for fertility preservation in children and young adults. Lancet Diabetes Endocrinol. 2015;3: 556–67. 2. R. G. Gosden, D. T. Baird, J. C. Wade and R. Webb. Restoration of fertility to oophorectomized sheep by ovarian autografts stored at −196°C. Human Reproduction 1994;9:597–603.

3. J. Donnez, M. M. Dolmans, A. Pellicer, C. Diaz-Garcia, M. Sanchez Serrano and K. T. Schmidt et al. Restoration of ovarian activity and pregnancy after transplantation of cryopreserved ovarian tissue: A review of 60 cases of reimplantation. Fertil. Steril. 2013;99:1503–13. 4. H. Van der Ven, J. Liebenthron, M. Beckmann, B. Toth, M. Korell and J. Krussel et al. Ninety-five orthotopic transplantations in 74 women of ovarian tissue after cytotoxic treatment in a fertility preservation network: Tissue activity, pregnancy and delivery rates. Hum. Reprod. 2016. 5. W. H. Wallace, A. G. Smith, T. W. Kelsey, A. E. Edgar and R. A. Anderson. Fertility preservation for girls and young women with cancer: Population-based validation of criteria for ovarian tissue cryopreservation. Lancet Oncol. 2014;15:1129–36. 6. J. Donnez and M. M. Dolmans. Fertility preservation in women. Nat. Rev. Endocrinol. 2013;9:735–49. 7. W. H. Wallace, H. O. Critchley and R. A. Anderson. Optimizing reproductive outcome in children and young people with cancer. J. Clin. Oncol. 2012;30:3–5. 8. J. H. Bramswig, M. Riepenhausen and G. Schellong. Parenthood in adult female survivors treated for Hodgkin’s lymphoma during childhood and adolescence: A prospective, longitudinal study. Lancet Oncol. 2015;16:667–75. 9. M. M. Dolmans, V. Luyckx, J. Donnez, C. Y. Andersen and T. Greve. Risk of transferring malignant cells with transplanted frozen-thawed ovarian tissue. Fertil. Steril. 2013;99:1514–22. 10. L. B. Signorello, J. J. Mulvihill, D. M. Green, H. M. Munro, M. Stovall, R. E. Weathers et al. Stillbirth and neonatal death in relation to radiation exposure before conception: A retrospective cohort study. Lancet 2010;376:624–30.

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How to Manage Patients with Cancer Who Need Fast-Track Treatment Kirsten Tryde Macklon and Michael von Wolff

her marital status, the type of cancer and the local expertise in her hospital/country.

Methods to Preserve Fertility in Women Different options exist to preserve fertility in women (Figure 7.1). The key point is to be able to arrange to see them in the shortest time possible so that there is no substantial delay in their cancer treatment and yet they will have sufficient time to think about fertility preservation. The ovaries can be protected from chemotherapy or pelvic radiation, and thereby oocytes are left in situ. To protect ovaries from chemotherapy, gonadotropin hormone-releasing hormone (GnRH) agonists can be given. Protection of ovaries from radiotherapy of the pelvis requires laparoscopic transposition of the ovaries. Alternatively, oocytes are removed and cryopreserved in vitro. Oocytes are stored following ovarian stimulation and transvaginal aspiration of follicles, or ovarian tissue is removed and oocytes are cryopreserved embedded in the ovarian tissue. Whichever method is preferred depends upon several factors like the urgency to start cancer treatment, the age of the patient, her personal preferences,

After an initial gonadotrophin release, GnRH agonists bring about a downregulation of the GnRH receptor, followed by hypogonadism, resulting in a transient menopausal state. It is assumed, but has not been proven, that the ovaries are protected from chemotherapy due to a GnRH-induced decrease of follicular activity and perfusion. As the flare-up effect of the GnRH agonists takes about one week, they should be administered one week before the start of chemotherapy. The effect of the GnRH agonists should continue for at least one to two weeks after the last chemotherapy cycle. The treatment is noninvasive and simple to perform as it requires only one injection every 4–12 weeks. The fertility-preserving effect of GnRH agonists has been investigated in several studies, including prospective randomized trials. Meta analyses have shown some reduction in the risk of developing primary ovarian

Postpubertal girls/women

Prepubertal girls

Not sexually active

Sexually active

Imminent need for chemo

Ovarian tissue cryopreservation

Gonadotropin Hormone-Releasing Hormone Agonists

Ovarian stimulation with cryopreservation of oocytes

2 weeks before chemo

GnRH agonists

Pelvic radiation: Transposition of the ovaries

Figure 7.1 Flowchart of which type of fertility preservation to choose in girls and women depending on the kind of therapy, the urgency to start treatment and the age of the patient

Patients with Cancer Who Need Fast-Track Treatment

insufficiency (POI) in women co-treated with GnRH agonists during chemotherapy [1]. However, due to the heterogeneity of the study results, doubts remain concerning its effect. Therefore, other techniques should be considered in addition to this treatment. Note also that this medication only has an effect in postpubertal girls and women.

Transposition of the Ovaries Transposition is required in case of pelvic radiation. Radiotherapy with 2 Gy leads to a loss of circa 50 percent of the primordial follicles [2] and the risk of POI in women aged > 20 years is almost 100 per cent if they receive radiotherapy > 15 Gy. The mobilized ovary is usually transposed laparoscopically craniolaterally, fixed and marked with a clip in order to achieve the greatest distance possible from the main irradiated area. As loss of ovarian function can occur despite transposition due to scatter irradiation, additional cryopreservation of the ovarian tissue is recommended. According to the published literature, there is a success rate of up to 85 percent with this technique in patients with regular ovulatory cycles, and also in patients under the age of 40 after radiotherapy [3]. Unspecific postoperative abdominal discomfort has been described, which was a result of ovarian cysts or peritoneal adhesions in most cases.

Cryopreservation of Ovarian Tissue One option to cryopreserve oocytes is cryopreserving ovarian tissue. A whole ovary, a semi-ovary or even ovarian biopsies can be removed and prepared for cryopreservation. By using this method, a large number of oocytes can potentially be collected and stored for future use. The technique of cryopreserving ovarian tissue is a specialist task that cannot be performed by any fertility clinic. It requires a team of specialists trained to do it, and, in the European Union, a certified laboratory. The primordial follicle is the predominant type of follicle in the ovary, representing more than 90 percent of all follicles, and, fortunately, this is also the type of follicle that best tolerates freezing. Primordial follicles are located in the cortex of the ovary in the outermost 1–2 mms. When ovarian tissue is prepared for cryopreservation, the cortex is thus isolated, cut into smaller fragments and treated with a cryoprotectant to avoid freezing damage. Traditionally, the slow freezing protocol is used, but

the tissue can also be cryopreserved by vitrification. Collection of the tissue requires an operation in general anesthesia. It can favorably be done by laparoscopy at any given time of the cycle, so in theory it can be performed from one day to the next provided a certain flexibility of the operating theater. Cryopreservation of ovarian tissue can also be done in prepubertal girls. The risks of this procedure are low. Complications requiring a second operation have been described in around 1 of 500 cases [4]. If the woman develops POI and wants to become pregnant, the tissue is transplanted laparoscopically into the pelvis, either subperitonally, laterally to the uterus, into the ovaries or onto the remaining ovaries. It is estimated that more than 60 children are born per year after the procedure worldwide, many of them by spontaneous conception [5]. According to the two major study groups in Denmark [6] and the network FertiPROTEKT [5], the success rate in terms of deliveries per transplanted patients is around 30 percent in women up to 35 years of age. A further increase can be expected due to future improvement of the transplantation technique.

Cryopreservation of Unfertilized or Fertilized Oocytes Controlled ovarian hyperstimulation (COH) with gonadotropins will lead to maturation of several follicles. This principle is used in in vitro fertilization (IVF), in which all mature follicles are aspirated with the aim to collect the eggs and then fertilize them with husbands’ or donors’ sperm. For women with cancer, this is a widely accepted way of preserving fertility, provided there is enough time before the start of chemotherapy. Traditionally, stimulation with gonadotropins starts on the third day of the cycle and after 10–12 days the oocytes are aspirated, after which the woman, in principle, can start chemotherapy. Oocytes are either frozen unfertilized or are fertilized and then embryos are frozen two to five days after aspiration. Recently developed stimulation protocols allow the start of stimulation any day of the menstrual cycle, requiring only two weeks at a maximum for the whole stimulation and aspiration treatment [7]. To increase later pregnancy chances, it is also possible to perform two consecutive stimulations with only a few days between both stimulations or to first remove and cryopreserve ovarian tissue and directly afterward start ovarian stimulation.

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Kirsten Tryde Macklon and Michael von Wolff

Concern has been raised regarding the possible deleterious effects of elevated estrogen levels during COH for women with estrogen receptor positive breast cancer, which is why protocols that include an aromatase inhibitor such as Letrozole have been developed [8]. It has been found that Letrozole reduces the serum estrogen levels, and women having undergone IVF using a protocol with Letrozole do not experience more relapses than women not undergoing IVF. The success rate of ovarian stimulation and cryopreservation can still only be estimated. Based on the number of retrieved oocytes [9] and the data of several registries, delivery rates of 40 percent in women with cryopreserved oocytes at an age of up to 35 years have been estimated and confirmed by small cohort studies [4].

Methods to Preserve Fertility in Girls In young, prepubertal girls, the only option to preserve fertility is cryopreservation of ovarian tissue [10]. Ovarian stimulation can be performed postmenarche, but as this technique requires vaginal aspiration, it should only be considered in cases where the girl is already sexually active.

electroejaculation under general anesthesia can be considered. In younger boys, the only possibility to preserve fertility is cryopreservation of testicular tissue. This is still experimental and is only offered in research settings. Biopsies from the testes are cryopreserved in the hope that one day the tissue can be transplanted back to the individual or cultured in vitro in order to isolate mature sperm. It has to be stressed that this is experimental and has not yet been performed in humans.

Suggested Standard Operating Protocol (SOP) Clinical referral and management pathway for patients with cancer who need fertility preservation

Suggested Audits On efficiency of service: – Waiting time from referral to treatment – Number of patients screened versus uptake of service – Pregnancy outcomes from tissue transplant

References Methods to Preserve Fertility in Men Sperm can be collected by masturbation and cryopreserved before chemo- or radiation therapy. This can be done over a period of several days provided there is enough time or even just once before start of treatment. All it requires is that the man is able to produce a sperm sample by masturbation. Most fertility laboratories are able to provide this service. Before cryopreservation, the HIV and hepatitis B and C status of the man should be checked. After treatment, in case of azoospermia, the sperm can be thawed and used to inseminate the female partner or to fertilize her eggs in connection to IVF.

Methods to Preserve Fertility in Boys In pubertal/postpubertal boys, there are indications that at testicular volumes as low as 6–7 ml it is possible to find motile spermatozoa in the ejaculate [11]. Whenever the boy is able to produce a sperm sample, cryopreservation of sperm should be offered. In rare cases, where the size of the testes indicates sperm production but the boy is not yet capable of producing a sperm sample by masturbation,

1. R. R. Munhoz, A. A. Pereira, A. D. Sasse, P. M. Hoff, T. A. Traina, C. A. Hudis and R. J. Marques. Gonadotropin-releasing hormone agonists for ovarian function preservation in premenopausal women undergoing chemotherapy for early-stage breast cancer: A systematic review and meta-analysis. JAMA Oncol. 2016;2:65–73. 2. W. H. Wallace, A. B. Thomson and T. W. Kelsey. The radiosensitivity of the human oocyte. Hum. Reprod. 2003;18:117–21. 3. M. Bisharah and T. Tulandi. Laparoscopic preservation of ovarian function: An underused procedure. Am. J. Obstet. Gynecol. 2003;188:367–70. 4. B. Lawrenz, J. Jauckus, M. Kupka, T. Strowitzki and M. von Wolff. Efficacy and safety of ovarian stimulation before chemotherapy in 205 cases. Fertil. Steril. 2010;94:2871–3. 5. H. Van der Ven, J. Liebenthron, M. H. Beckmann, B. Toth, M. Korell, J. Krüssel et al. Ninety-five orthotropic transplantations of ovarian tissue after cytotoxic treatment in a fertility preservation network – tissue activity, pregnancy and delivery rates. Hum. Reprod., in press. 6. A. K. Jensen, S. G. Kristensen, K. T. Macklon, J. V. Jeppesen, J. Fedder, E. Ernst and C. Y. Andersen. Outcomes of transplantations of cryopreserved

Patients with Cancer Who Need Fast-Track Treatment

ovarian tissue to 41 women in Denmark. Hum. Reprod. 2015;30:2838–45. 7. H. Cakmak, A. Katz, M. I. Cedars and M. P. Rosen. Effective method for emergency fertility preservation: Random-start controlled ovarian stimulation. Fertil. Steril. 2013;100:1673–80. 8. K. Oktay, V. Turan, G. Bedoschi, F. S. Pacheco and F. Moy. Fertility preservation success subsequent to concurrent aromatase inhibitor treatment and ovarian stimulation in women with breast cancer. J. Clin. Oncol. 2015;33:2424–9. 9. M. von Wolff, R. Dittrich, J. Liebenthron, F. Nawroth, A. N. Schüring, T. Bruckner and A. Germeyer.

Fertility-preservation counselling and treatment for medical reasons: Data from a multinational network of over 5000 women. Reprod. Biomed. Online. 2015;31:605–12. 10. K. T. Schmidt, E. C. Larsen, C. Y. Andersen and A. N. Andersen. Risk of ovarian failure and fertility preserving methods in girls and adolescents with a malignant disease. BJOG 2010;117:163–74. 11. I. Hagenäs, N. Jørgensen, C. Rechnitzer, P. Sommer, M. Holm, K. Schmiegelow et al. Clinical and biochemical correlates of successful semen collection for cryopreservation from 12–18-year-old patients: A single-center study of 86 adolescents. Hum. Reprod. 2010;25: 2031–8.

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How to Manage Patients with Adenomyosis before an IVF Cycle

Introduction

structures and alterations in endometrial function [5]. Therefore, it is reasonable to hypothesize the existence of a relationship between adenomyosis and reproduction. However, the mechanisms by which adenomyosis may impede fertility remain unclear.

Stephan Gordts

Rokitansky was the first to describe adenomyosis, in 1860, as the presence of heterotopic endometrial glands and stroma in the myometrium, with adjacent smooth muscle hyperplasia [1]. Morphologically, adenomyosis is defined as an invasion of glands and stroma into the myometrium, which extends deeper than 2.5 mm from the endometrial junctional zone (EJZ). With the introduction and evolution of indirect imaging techniques, the diagnosis of adenomyosis moved from a histological to a clinical entity. The advent of magnetic resonance imaging (MRI) and 3-D ultrasound heralded a turning point in the appreciation of adenomyosis as an important disorder of the female reproductive tract. The systematic use of these techniques has enabled noninvasive visualization of distortions in the myometrial architecture, which has facilitated the distinction between pathology of the outer myometrium and pathology of the inner myometrium or junctional zone (JZ). Like endometriosis, adenomyosis may present in various disguises, ranging from a simple JZ thickening to a nodular, cystic or diffuse lesion, involving the entire uterine wall. There is a higher incidence of adenomyosis with advanced age; thus, with the current trend in postponing childbearing, an increasing number of patients who seek fertility treatments will be diagnosed with uterine pathology. In patients with infertility, dysmenorrhea and menorrhagia, the incidence of adenomyosis is reported to be as high as 50 percent [2]. Despite the absence of epidemiological evidence, it seems reasonable to hypothesize a causal relationship between adenomyosis and infertility. In baboons, adenomyosis was associated with lifelong infertility [3]. Moreover, a recent study reported that, among patients with adenomyosis, the clinical pregnancy rate decreased and the miscarriage rate increased after in vitro fertilization (IVF) treatments, compared to patients without adenomyosis [4]. Adenomyosis causes dysregulation of myometrial

Diagnosis Suspicion of adenomyosis should be based on the clinical examination and symptoms of the patient. Certainly, in patients with endometriosis, the presence of adenomyosis should be excluded. Any suspicion should be investigated with diagnostic imaging. Recent advances in ultrasound technology have increased its diagnostic capabilities. In the diagnosis of adenomyosis, ultrasound analyses have highaccuracy, with a mean sensitivity of 0.72 (95% CI: 0.65–0.79), and a specificity of 0.81 (95% CI: 0.77–0.85) [6]. However, ultrasound diagnostic performance is biased by the experience of the examiner. MRI also has a high diagnostic sensitivity of 0.77 (95% CI 67–85%) and specificity of 89 percent [6]. However, MRI has the advantage of less dependence on operator experience, and the diagnosis is based on objective image findings. MRI provides excellent soft tissue differentiation and clear visualization of the JZ. Image findings must be clearly described, and descriptions should take into consideration the size, the aspect and the location of lesions. Adenomyosis can appear as a JZ hyperplasia, myometrial striations irradiating from the endometrium, a focal or diffuse lesion or cystic adenomyosis. For many years, the uterus has been a neglected incubator. However, the introduction of more sophisticated, indirect imaging techniques has created new exploration possibilities. In addition to the uterine cavity, explorations should include the entire endo-myometrium. Transvaginal ultrasound can be used as a first screening method to determine the necessity of more accurate visualization with MRI.

Patients with Adenomyosis before an IVF Cycle

Adenomyosis and IVF: The Controversy Results reported on IVF outcomes in patients with adenomyosis should be interpreted with caution, due to the great heterogeneity among studies. Many studies are based on a retrospective design, and they frequently include a small number of patients. Different studies often employ different diagnostic techniques: in general, MRI data are less operatordependent than ultrasound data; thus, MRI studies provide a more reliable description of the pathology. Moreover, different studies use different stimulation protocols, and there is no validated classification system for reporting results. JZ pathology can be clearly diagnosed with MRI, but different JZ thicknesses are used as thresholds for diagnosing JZ hyperplasia [7, 8]. Currently, there are no stringent criteria to define JZ hyperplasia adenomyosis. Furthermore, it is unknown whether JZ hyperplasia is an initial stage in the development of more severe adenomyosis. It is also unknown whether diffuse, focal and cystic adenomyosis have the same impact on the uterine environment. In a recent meta-analysis [4], adenomyosis was associated with a 28 percent (95% CI, 5–45%) reduction in the likelihood of clinical pregnancy in infertile women who underwent IVF/intracytoplasmic sperm injection (ICSI) with autologous oocytes. Additionally, compared to healthy pregnancies, pregnancies with adenomyosis were associated with more than double the risk of miscarriage, and due to a higher abortion rate, a 30 percent lower live birth rate. This doubled risk of abortion was also reported among recipients with adenomyosis who participated in an oocyte donation program [9]. That finding is particularly interesting because it excluded an oocyte factor, and it highlighted the involvement of an abnormal uterine environment and potentially defective placentation. In a prospective study [8] where adenomyosis was diagnosed with MRI, the JZ thickness correlated negatively with the pregnancy rate after IVF.

Medical Treatment Like endometriosis, adenomyosis is hormonedependent. Therefore, medical treatment is based on hormone suppression. The treatment aims to control the clinical symptoms and induce regression of the adenomyotic lesion and the coexisting inflammatory environment. Intrauterine devices loaded with levonorgestrel or Danazol are rather impractical for women wishing to

conceive; consequently, gonadotrophin-releasing hormone (GnRha) is the most frequently used drug. Although data are scarce, spontaneous pregnancies have been reported after using GnRha for six months. Low pregnancy rates and high abortion rates associated with adenomyosis may be due to the increased inflammatory reactions caused by endometrial cell invasion into the myometrium and the increased production of estrogen caused by overexpression of aromatase P450 [10]. These conditions may explain why hormone suppression with GnRha can exert beneficial effects. The presence of adenomyosis did not affect pregnancy rates after IVF/ICSI among women treated with the long GnRha protocol [11, 12]. In a metaanalysis by Vercellini et al. [4], higher pregnancy rates were observed after the long GnRha ovarian stimulation protocol compared to the short GnRha protocol. Patients with adenomyosis frequently exhibit concomitant endometriosis; therefore, ovarian reserve can be impaired, due to the presence of ovarian endometrioma and/or due to effects from a previous surgery. In those cases, even treatment with the long GnRha protocol can result in a poor ovarian response. Those patients should be drawn into a discussion of whether to cryopreserve all the embryos and perform a cryotransfer at a later stage (i.e., two to three months), after the disease has been down-regulated with GnRha.

Surgical Treatment Adenomyosis is an infiltration of uterine tissue into the myometrium; thus, resection of the disease always results in the partial removal of healthy myometrium, and it carries the unavoidable risk of incomplete removal. In most cases, surgical excision is performed via laparotomy. However, in recent years, procedures have also been performed with laparoscopy. Complete resection is possible when dealing with focal lesions, like adenomyoma or cystic adenomyosis. In cases of diffuse adenomyosis, only cytoreductive surgery is feasible. Complete resection removes a critical amount of myometrium and results in a “nonfunctional” uterus. Different methods of cytoreductive surgery have been described, and most involve closing the uterine wall with overlapping flaps. In contrast, for uterine myoma, no healthy myometrial tissue is removed in the myomectomy. However, because the delineation between healthy tissue and adenomyosis is not clear, it is more challenging to close the myometrial layer, and a meticulous approach is necessary. Attention should be focused on

29

Stephan Gordts

performing adequate hemostasis and avoiding dead spaces. Although there is no general consensus about the optimal interval between surgery and a first attempt to achieve pregnancy, an interval of at least three to six months has been advocated. After surgery, the myometrial wall becomes thin, and fibrosis diminishes the tensile strength of the uterus. These factors elevate the risk of uterine rupture during pregnancy or labor. Although the true risk of uterine rupture is not well established, patients should be informed that a C-section delivery would be preferable. Cystic adenomyosis or small adenomyoma in the inner third of the intramural myometrium can be treated with hysteroscopy. Some lesions are directly recognizable in hysteroscopy, because they bulge into the endometrial cavity. Lesions localized deep in the uterine wall require ultrasound guidance during the hysteroscopic procedure. Hysteroscopic treatment consists of a complete resection with 5-Fr scissors, which allows clear dissection of the myometrial wall of the cyst from the surrounding myometrium. An alternative treatment option is an ablative technique that destroys the inner cystic wall. We believe that the ablative approach is preferable for cysts localized deep in the uterine wall. However, the hysteroscopic technique has the advantage of leaving the outer myometrium intact and avoiding an abdominal scar [13]. Treating adenomyosis with a partial resection leaves a portion of “diseased uterus,” which carries the risk of recurrence. However, this approach can preserve fertility, with a reported pregnancy rate of 50 percent [14], and it can increase pregnancy rates after IVF [14].

Conclusion Adenomyosis, a disease of the myometrium, became a clinical entity with the introduction of more sophisticated techniques for indirect imaging. Although not fully understood, adenomyosis negatively interferes with female fertility. Due to the advanced age of patients referred to our IVF programs, combined with the fact that the incidence of adenomyosis increases with age, the frequency of encountering patients with adenomyosis in our fertility clinics has increased over the years. When adenomyosis is suspected clinically, a meticulous uterine exploration must be performed, with special attention to the inner myometrium or JZ. Hormone suppression is beneficial for diseases involving JZ hyperplasia or

Table 8.1 Recommendations for managing patients with adenomyosis

• 3-D ultrasound and/or MRI enable a clinical diagnosis of adenomyosis. • MRI is not operator-dependent, which facilitates accurate descriptions of the lesions. • Lesions should clearly be described in terms of form, size, number and location. • Although the data are heterogeneous, the presence of adenomyosis appears to lower pregnancy rates and increase miscarriage rates. • GnRha-induced hormone suppression appears to have a beneficial effect on pregnancy rates after IVF. • Different options should be discussed with the patient, including postponing an IVF cycle until after three months of hormone suppression with GnRha or freezing all the embryos. • Surgery should be reserved for focal or cystic forms of adenomyosis.

diffuse forms of adenomyosis with global, and frequently asymmetric, thickening of the uterine wall. Treatment options (Table 8.1) should be discussed with the patient before starting an IVF treatment to consider postponing the IVF until after medical treatment with a GnRha analog for three months. This treatment for adenomyosis can potentially lower the inflammatory reaction and reduce the JZ thickness. Surgery should be reserved for patients with focal lesions, such as an adenomyoma or cystic adenomyosis. Several different operative techniques have been described; most recently, laparoscopic and hysteroscopic procedures have joined the repertoire of technical approaches. Nevertheless, it remains unclear which technique is most effective. Further studies, with conscientious descriptions of the lesions, are necessary to improve our understanding of how adenomyosis interferes with fertility and to determine the best possible treatment [15].

Suggested Standard Operating Protocol (SOP) Management of women with adenomyosis including definitions and diagnostic criteria

Suggested Audits Pregnancy outcomes of women with adenomyosis with IVF/ICSI

Patients with Adenomyosis before an IVF Cycle

References 1. C. Rokitansky. Über Uterusdrüsen-Neubildung in Uterus- und Ovarial- Sarcomen. Zeitschr. Gesellschaft der Aerzte in Wien 1860;16:577–81. 2. J. J. Brosens, N. M. De Souza and F. G. Barker. Uterine junctional zone: Function and disease. Lancet 1995;346: 558–60. 3. B. F. Barrier, M. J. Malinowski, E. J. Dick Jr., G. B. Hubbard and G. W. Bates. Adenomyosis in the baboon is associated with primary infertility. Fertil. Steril. 2004; 82 (Suppl. 3): 1091–4. 4. P. Vercellini, D. Consonni, D. Dridi, B. Bracco, M. P. Frattaruolo and E. Somigliana. Uterine adenomyosis and in vitro fertilization outcome: A systematic review and meta-analysis. Hum. Reprod. 2014;29(5):964–77. 5. S. Campo, V. Campo and G. Benagiano. Adenomyosis and infertility. RBMonline 2012;24(1):35–46. 6. R. Champaneria, P. Abedin, J. Daniels, M. Balogun and K. S. Khan. Ultrasound scan and magnetic resonance imaging for the diagnosis of adenomyosis: Systematic review comparing test accuracy. Acta Obstet. Gynecol. Scand. 2010;89:1374–84.

9. J. A. Martınez-Conejero, M. Morgan, M. Montesinos, S. Fortuño, M. Meseguer, C. Simon, J. A. Horcajadas and A. Pellicer. Adenomyosis does not affect implantation, but is associated with miscarriage in patients undergoing oocyte donation. Fertil. Steril. 2011;96:943–50. 10. J. Brosens, H. Verhoeven, R. Campo, L. Gianaroli, S. Gordts, J. Hazekamp, L. Hagglund, T. Mardesic, E. Varila, J. Zech et al. High endometrial aromatase P450 mRNA expression is associated with poor IVF outcome. Hum. Reprod. 2004;19: 352–6. 11. V. Mijatovic, E. Florijn, N. Halim, R. Schats and P. Hompes. Adenomyosis has no adverse effects on IVF/ICSI outcomes in women with endometriosis treated with long-term pituitary down-regulation before IVF/ICSI. Eur. J. Obstet. Gynecol. Reprod. Biol. 2010;151:62–5. 12. M. F. Costello, K. Lindsay and G. McNally. The effect of adenomyosis on in vitro fertilization and intra-cytoplasmic sperm injection treatment outcome. Eur. J. Obstet. Gynecol. Reprod. Biol. 2011;158:229–34. 13. S. Gordts and R. Campo. Brosens Hysteroscopic diagnosis and excision of myometrial cystic adenomyosis. Gynecol. Surg. 2014;11(4):273–8.

7. G. Kunz, D. Beil, P. Huppert, M. Noe, S. Kissler and G. Leyendecker. Adenomyosis in endometriosis – prevalence and impact on fertility. Evidence from magnetic resonance imaging. Hum. Reprod. 2005;20: 2309–16.

14. G. F. Grimbizis, T. Mikos and B. Tarlatzis. Uterus sparing operative treatment for adenomyoisis. Fertil. Steril. 2014;101(2):472–87.

8. A. Maubon, A. Faury, M. Kapella, M. Pouquet and P. Piver. Uterine junctional zone at magnetic resonance imaging: A predictor of in vitro fertilization implantation failure. J. Obstet. Gynaecol. Res. 2010;36(3):611–18.

15. S. Gordts, J. J. Brosens, L. Fusi, G. Benagiano and I. Brosens. Uterine adenomyosis: A need for uniform terminology and consensus classification. Reprod. Biomed. Online 2008; 17: 244–8.

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The Consultation with Older Men Seeking ART Treatment Allan Pacey

Introduction Consultations with older men seeking assisted conception are becoming increasingly common in many developed (high-resource) countries. This is undoubtedly because of the general trend for heterosexual couples in those countries to delay the age at which they decide to start a family. In addition, it is now more socially acceptable for men to start a second relationship later in life (perhaps following a divorce or after being widowed). Whatever the reason, clinicians and gynecologists increasingly need to be aware of some specific issues that relate to providing assisted conception for an older man and his partner. This chapter summarizes the current thinking in this area and highlights topics for further consideration and future research. However, before the chapter can begin, two questions should be considered. The first, is “what constitutes an older man in the context of assisted conception?” This is important to clarify at the start, as unlike the well-described agerelated decline in fertility of women, the age-related changes to male fertility are less obvious and less well documented (i.e. men don’t undergo a menopause). However, this chapter emphasizes that where measurable changes to male reproductive performance are seen, they tend to occur once men reach the age of 40 to 45 years. The second question for consideration is “what age-related changes are of concern to older fathers and therefore, as a consequence, those providing them and their partners with assisted conception?” Here it is harder to be specific, but this chapter considers the question in three broad areas: • Are there any changes to the quality or quantity of spermatozoa in the ejaculates of older men? • Is there any effect of male age on the probability of conception, either naturally or following assisted conception?

•

What are the risks (if any) to the health and development of any children born to an older father?

The following section deals with these three areas in turn.

Sperm Quality in Older Men There is now fairly convincing evidence that many aspects of semen quality decline markedly as a function of increasing male age. A recent metaanalysis of 93,839 men from a total of 90 studies examined seven aspects of ejaculate quality and reported a statistically significant age-associated decline in semen volume, percentage motility, progressive motility, the percentage of normal morphology and the proportion of sperm in the ejaculate with unfragmented DNA [1]. Interestingly, the analysis was unable to conclude that there was an age-related decline in sperm concentration, suggesting that while the quantity or speed of spermatogenesis changed little with increasing age, a much bigger effect was on the quality of spermatogenesis. The analysis is robust in that it controlled for a number of confounding factors, such as the duration of sexual abstinence, which is already known to increase with male age and therefore independently to have an effect on semen quality. However, from such data it is difficult to conclude what effect these changes may have on the probability of conception in older men, since the variables of semen quality are only a surrogate marker of male fertility and attempts to connect the two are usually associated with a large confidence interval. Moreover, the analysis does not publish the data in a format that allows the reader to determine if the changes are likely to be clinically significant (i.e., increase the probability that an older man may be more likely to have semen quality below one of the World Health Organization [WHO] threshold values). However, of the changes reported to be associated with increasing male age, it is

Consultation with Older Men Seeking ART Treatment

probably the increase in the number of ejaculated sperm with DNA fragmentation that is the most likely to have a clinical significance for older men. There is now fairly strong evidence that increasing levels of sperm DNA damage are associated with increased time to pregnancy, reduced fertility and increasing levels of early pregnancy loss. The importance of sperm DNA damage is a topic that I return to later in this chapter.

Paternity in Older Men Relatively little data are available to examine how male age affects the probability of paternity in older men who are not seeking assisted conception and are trying to conceive naturally with a partner in her fertile years. This is because most birth registries tend to ignore the age of the father, and the identity of a newborn’s father cannot be confirmed as accurately as that of the mother. However, there are some tantalizing clues to suggest that natural fertility does indeed decline with increasing male age. Perhaps the best study to examine this looked at the time to pregnancy in 7,870 fathers enrolled in the Avon Longitudinal Study of Pregnancy and Childhood (ALSPAC) [2]. This analysis found that after adjustment for partner age, men over the age of 40 years were half as likely to have sired a pregnancy within 12 months compared to men under the age of 25 years old (odds ratio (95% CI) = 0.51 (0.31–0.86)). This is very similar to an observation by Hassan and Killick, who used a questionnaire to ask 2,112 consecutive women attending an antenatal clinic about their time to pregnancy, contraceptive use, pregnancy planning etc. [3]. This found that relative to fathers < 25 years old, those who were older than 45 years were 4.6 and 12.5 times more likely to have a time to pregnancy of greater than one or two years, respectively. While a number of other studies have reached the same (or similar) conclusions, none has been able to establish an underlying reason for the observation that older men are generally less fertile. It is not clear whether or not it is due to an age-related decline in semen quality (see earlier in this chapter) or as a consequence of behavioral change (such as a decline in coital frequency) associated with increasing male age. While both of these may be contributory factors, data from assisted conception would tend to support the former. For example, a study of registry data from 59 French in vitro fertilization (IVF) centers examined

the outcome of IVF cycles from 1,938 men whose partners were totally sterile with bilateral tubal obstruction or where both Fallopian tubes were absent [4]. The study found that when the male partner was above the age of 40 years, the odds ratio of failure to conceive was 2.00 (95% confidence interval [CI]: 1.10–3.61) when his partner was 35–37 years old, 2.03 (95% CI: 1.12–3.68) when she was 38–40 years, and 5.74 (95% CI: 2.16, 15.23) when she was aged 41 years and over. These data are important because they rule out the influence of behavioral factors such as sexual frequency and suggest that a real difference exists in the quality of spermatozoa (probably sperm DNA) between younger and older men. This notion is further supported by a study of more than 17,000 cycles of intrauterine insemination and found that while there was a small decrease in the pregnancy rate with increasing male age (12.3 percent for men under 30 to 9.3 percent for men above the age of 45), there was a dramatic increase in the miscarriage rate in the partners of older men (13.7 percent in the partners of men below 30 to 32.4 percent in the partners of men over the age of 40).

Risks to Child Health and Development By the early 1990s, the British Andrology Society and the American Society for Reproductive Medicine both recommended that sperm donors should be below the age of 40 years old. This was because of the observations that there is an increasing risk of aneuploidy in the sperm from older men in addition to new mutations of the paternal genome. This is corroborated by epidemiological studies, which have shown that the children of older men (generally above the age of 40) are more likely to be born with, or have an increased risk of developing: • Dyskinetic cerebral palsy • Congenital malformations • Autism spectrum disorders • Achondroplasia • Schizophrenia • Retinoblastoma • Downs syndrome. While the relevance of the current sperm donor recruitment guidelines [6] to a medical consultation with an older man seeking assisted conception may not seem immediately clear, it is the same biological mechanisms at work in both scenarios and the

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clinicians and gynecologists would do well to familiarize themselves with this literature. Some of the studies that underpin these observations include population-based data with significant statistical power. For example, the health records of 754,330 people born in Sweden between 1973 and 1980 were linked to the records of people admitted to hospital with a diagnosis of schizophrenia or other non-affective psychosis between 1989 and 2001 [7]. This study concluded that after adjustment for important confounding factors, the hazard ratio for each 10-year increase in paternal age was 1.47 (95 CI: 1.23–1.76) for schizophrenia and 1.12 (0.98–1.29) for other non-affective psychoses. Although this study has not been without its critics, the data for autism spectrum disorders are very similar with a 10-year birth cohort linkage study of more than 1 million participants and a meta-analysis of more than 30 epidemiological studies showing an advancing paternal age association with increased risk of autism [8].

Strategies and Solutions for the Older Male Given the observations outlined earlier, and the impact that this might have on the outcome for older men seeking assisted conception with their partners, a number of strategies have been proposed. Perhaps the most outlandish was made in 2015 by bioethicist Kevin Smith from the University of Abertay (UK) who proposed a system of statesupported universal sperm banking for all young men [9], presumably so that they might have the opportunity to use sperm ejaculated at a younger age (say in their early 20s) should they wish to become a father later in life (perhaps in their mid-to-late 40s). Notwithstanding the cost and logistics of implementing such a venture, the idea is almost certainly flawed because given the relatively poor post-thaw survival of sperm, it would mean that most of the banked sperm would need to be used in IVF or intracytoplasmic sperm injection (ICSI), which introduces its own risks and costs. Therefore, before this approach could be implemented, there would need to be a comprehensive cost-benefit analysis undertaken. However, at the present time, it is unlikely to be the case that clinicians and gynecologists working in assisted conception would encounter an older man who “just happened” to have sperm banked earlier in life without any prior medical reason (e.g. a diagnosis

of cancer). At the present time, there is no advantage to offer precautionary sperm banking to an older male in order to avoid any further deterioration of sperm or genetic quality (notwithstanding that it may be required to guard against testicular failure or impotence prior to IVF). There are currently no specific strategies to separate “good” from “poor” sperm in the ejaculates of older men, over and above the kind of sperm preparation methods that are ordinarily used in assisted conception.

Conclusion It is likely that consultations with older men seeking assisted conception with their partners will become increasingly common over the next decade, particularly in high-resource countries. Unfortunately, there is strong (and growing) evidence of a significant reduction in the fertility of men above the age of 40, and this is also associated with an increased risk of miscarriage in their partners; epidemiological evidence also suggests that the children of older fathers may be at increased risk of some disorders (e.g., Downs syndrome, schizophrenia, autism, etc.) and this is probably caused by an increase in the mutational load in the sperm produced by older men [10]. Unfortunately, no specific protocols are available to improve sperm quality or to be able to select those with fewer mutations for use in assisted conception. The provision of sperm banking to younger men as a way of avoiding age-related changes in sperm quality is not recommended.

Suggested Standard Operating Protocol (SOP) No recognized protocols or information sheets have been developed for consultations with the older male. However, it is recommended that older men and their partners are informed about the relevant data concerning: • Possible reductions in semen quality; • Reduced chance of pregnancy (independent of female age); • Increased risk of miscarriage; and • Increased risk of disorders with a genetic component in any children born. A summary of the background literature is given earlier in this chapter.

Consultation with Older Men Seeking ART Treatment

Suggested Audits There are no recognized audits in this area, but any that are developed should take account of partner age during any analysis as this is likely to be a significant confounding variable (see earlier in this chapter).

Related Research Areas/Websites There are few dedicated resources specifically aimed at the older male and his partner, but the Australian website (www.yourfertility.org.au) does have a useful hub of resources (http://yourfertility.org.au/for-men/ age/) that patients may find useful.

References 1. S. L. Johnson, J. Dunleavy, N. J. Gemmell and S. Nakagawa. Consistent age-dependent declines in human semen quality: A systematic review and meta-analysis. Ageing Res. Rev. 2015;19:22–33. 2. W. C. Ford, K. North, H. Taylor, A. Farrow, M. G. Hull and J. Golding. Increasing paternal age is associated with delayed conception in a large population of fertile couples: Evidence for declining fecundity in older men. The ALSPAC Study Team (Avon Longitudinal Study of Pregnancy and Childhood). Hum. Reprod. 2000;15: 1703–8. 3. M. A. Hassan and S. R. Killick. Effect of male age on fertility: Evidence for the decline in male fertility with increasing age. Fertil. Steril. 2003;79 (Suppl. 3): 1520–7.

4. E. de La Rochebrochard, J. de Mouzon, F. Thépot and P. Thonneau. French National IVF Registry (FIVNAT) Association. Fathers over 40 and increased failure to conceive: The lessons of in vitro fertilisation in France. Fertil. Steril. 2006;85:1420–4. 5. S. Belloc, P. Cohen-Bacrie, M. Benkhalifa, M. CohenBacrie, J. De Mouzon, A. Hazout and Y. Ménézo. Effect of maternal and paternal age on pregnancy and miscarriage rates after intrauterine insemination. Reprod. Biomed. Online 2008;17:392–7. 6. Association of Biomedical Andrologists, Association of Clinical Embryologists, British Andrology Society, British Fertility Society and the Royal College of Obstetricians and Gynaecologists. UK guidelines for the medical and laboratory screening of sperm, egg and embryo donors (2008) Hum. Fertil. (Camb). 2008;11: 201–10. 7. A. Sipos, F. Rasmussen, G. Harrison, P. Tynelius, G. Lewis, D. A. Leon and D. Gunnell. Paternal age and schizophrenia: A population based cohort study. BMJ 2004;329:1070. 8. C. M. Hultman, S. Sandin, S. Z. Levine, P. Lichtenstein and A. Reichenberg. Advancing paternal age and risk of autism: New evidence from a population-based study and a meta-analysis of epidemiological studies. Mol. Psychiatry 2011;16:1203–12. 9. K. R. Smith. Paternal age bioethics. J. Med. Ethics 2015;41:775–9. 10. R. Ramasamy, K. Chiba, P. Butler and D. J. Lamb. Male biological clock: A critical analysis of advanced paternal age. Fertil. Steril. 2015;103:1402–6.

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10

How to Manage Endometrioma in the IVF Patient Mukhri Hamdan and Ying Cheong

Points to Consider on the Management of the Patient with Endometrioma during Her IVF Journey What Was the Patient’s History Prior to Presentation? Unilateral or bilateral endometrioma can be diagnosed by imaging, e.g., ultrasound scan or during laparoscopy. At presentation prior to their in vitro fertilization (IVF) treatment, some patients may already have had surgical treatment on their endometrioma/endometriosis. Endometrioma may present as a recurrence, sometimes after several operations. Other patients may have endometrioma diagnosed for the very first time. The management strategy for these various presentations differs depending on the patients’ age, ovarian reserve and, of course, their preference for surgical intervention versus more conservative treatment.

Discussion on Endometrioma-Specific Treatment Options

of ovulation, is ineffective in the treatment of subfertility [1] whether used alone or before and/or after surgery [2] in enhancing pregnancy outcomes. However, treatment with a gonadotrophin-releasing hormone (GnRH) agonist for three to six months [3] prior to treatment with IVF increases the odds of clinical pregnancy by more than fourfold. The European Society for Human Reproduction and Embryology (ESHRE) working party recommended surgical treatment of AFS/ASRM (American Society for Reproductive Medicine) stage I/II endometriosis prior to IVF, but due to the absence of good evidence, the working party did not provide firm guidance on the preferred surgical management of deep nodular lesions prior to assisted reproductive technology (ART) [1]. The National Institute of Clinical Excellence in 2013 [4] in contrast suggested that women with moderate or severe endometriosis be offered surgical treatment because it improves the chance of spontaneous pregnancy, although this guidance is not specific to women undergoing IVF/intracytoplasmic sperm injection (ICSI) who have endometrioma.

Surgery Conservative Conservative management involves starting ovarian stimulation without surgically treating the existing endometrioma. This option may be particularly suitable for those who have had previous surgery, those who have endometrioma that is smaller (usually under 3 cm diameter) and/or those with an already low ovarian reserve. Medical treatment is in general not recommended prior to IVF [1].

Medical Very often, medical treatment is used as a form of interim symptomatic control for those patients with symptoms of pelvic pain. In the context of the patient with endometriosis, it is generally recognized that hormonal manipulation, which results in suppression

The benefit of surgical treatment of endometriomas prior to IVF with respect to improving the outcome of IVF is not supported by robust randomized controlled trials (RCTs). The surgical removal of endometriomas may improve access to ovaries and reduce the incidence of post-oocyte-retrieval infection, although the surgical treatment itself may damage/decrease the ovarian reserve. Compared with women with no surgical treatment, women who had their endometrioma surgically treated before IVF/ICSI had a similar LBR (OR 0.90; 95% CI [0.63, 1.28], 5 studies, 655 women, I2 = 32%), a similar CPR (OR 0.97; 95% CI [0.78, 1.20], 11 studies, 1,512 women, I2 = 0%) and a similar mean number of oocytes retrieved (SMD −0.17; 95% CI [−0.38, 0.05], 9 studies, 810 cycles, I2 = 63%) [2]. In general, clinicians will consider surgical management of endometriomas

How to Manage Endometrioma in the IVF Patient

Risks of surgical treatment of endometrioma prior to ART

Risks of intact/untreated endometrioma during ART

Surgical risks – bleeding, pain, infection, visceral injury

Accelerated progression of the disease

Follicular fluid contamination

Impact to ovarian reserve Pregnancy related complications

Premature ovarian failure

Incomplete surgery and disease recurrence

Infection to the endometrioma

Increase requirement of GnRH, cost and side effects Undiagnosed occult malignancy

Surgeon’s competency and learning curve

Chemical peritonitis Risk of cycle cancellation

Potential delay of ART Challenging oocyte retrieval

Figure 10.1 Summary of the risk of surgical treatment of endometrioma prior to ART and risk of intact endometrioma during ART

greater than 3 cm if they were symptomatic or for the improvement of access to the ovaries.

Discussion on Stimulation Protocols and Treatment-Adverse Effects

Individualization of Care

If a GnRH agonist is used prior to the IVF/ICSI cycle, many will continue the treatment using a long-agonist protocol. However, more recent prospective studies have found similar pregnancy rates in women with endometriosis who had undergone GnRH antagonist versus GnRH agonist IVF/ICSI treatment cycles [5, 6]. No studies have examined the results of different protocols specifically on women with and without endometrioma. The ovarian stimulation regime used on women with endometriosis undergoing IVF/ICSI must take into account the center’s individual expertise and the patient’s preference. While GnRH agonist cycles are advantageous in terms of cycle planning, GnRH antagonist cycles are regarded as more patient friendly. When compared with women without endometrioma, women with intact endometrioma have a lower mean number of oocytes retrieved and are nearly three times as likely to have a cancelled cycle compared with those without the disease.

The optimal management of the woman with endometrioma therefore often presents a clinical conundrum. Surgery carries not insignificant risks (Figure 10.1), particularly in women with endometrioma with coexisting severe deep infiltrating endometriosis, and thus individualization of care is crucial; optimal management depends on key fertility determinants such as age, ovarian reserve, symptoms of pain related to the disease and other comorbidities such as previous surgery (which may further increase the risk of surgical complications). It may be that in some, the optimal route is for IVF to be attempted before the consideration of more extensive surgery, while for the more symptomatic patient, or in those with large endometrioma, surgical management may take precedence. Transvaginal localization of the ovaries to assess their accessibility for monitoring follicular growth and oocyte retrieval via transvaginal ultrasound scan can also be useful prior to embarking on surgery or IVF.

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On the Day of Oocyte Retrieval It is best practice not to attempt to aspirate the content of an endometrioma before or during oocyte retrieval. This is because the content of the endometrioma is often viscous, and is likely to obstruct the tiny lumen of the oocyte retrieval needle. During oocyte collection, it is important to avoid puncturing the endometrioma if at all possible. If the endometrioma is inadvertently punctured by the oocyte collection needle, flushing the needle prior to the aspiration of the next follicle may reduce the risk of needle blockage and the subsequent loss of suction pressure due to obstruction by the content of the endometrioma. The administration of prophylactic antibiotics before the procedure is also an acceptable practice and clinicians need to be vigilant regarding the possibilities of late-onset peritonitis [7]. Due to the adhesive and inflammatory nature of endometriosis, anatomical distortion within the pelvis is highly likely, particularly when the disease is severe. This may mean that the pelvic structures such as the ureters and bowel, which are normally mobile, may be adhered to the vaginal vault. Hence, the theoretical risk of complications during oocyte retrieval is higher. It is imperative that an experienced operator is available to perform the oocyte retrieval for patients with severe endometriosis/endometrioma.

Impact of Endometrioma on Oocyte and Embryo Development The quality of oocytes collected from women with endometrioma is also thought to be adversely affected in women with endometriosis in terms of delay of maturation, suboptimal fertilization and embryo development. Hamdan et al. [8] found that the fluid from the follicles of patients with endometriosis inhibits egg maturation by generating Reactive Oxygen Species (ROS), resulting in DNA damage. This damage can delay oocyte maturation, and hence influence fertilization.

Effect of Endometrioma on Pregnancy Endometriosis is associated with a range of pregnancy complications, namely spontaneous hemoperitoneum in pregnancy, obstetric hemorrhage and preterm birth, independent of the ART. These complications can occur during the antenatal or postnatal period and were only recently acknowledged as a potential major contributor to significant maternal and perinatal mortality and

morbidity. The mechanisms around how endometriosis results in many of these complications are still largely unknown, but multidisciplinary input is recommended into care of these patients preconception antenatally and postnatally.

Conclusion Undergoing IVF treatment is a stressful time for our patients. Patients facing the additional burden of endometrioma require not only good-quality, evidence-based care, but also psychological support for their concerns. Clinicians should be holistic in their approach. There is not one dogmatic recommendation as to what intervention women with endometrioma should or should not have prior to IVF/ICSI, but based on current evidence, consideration should be given to individualize the care of these patients.

Suggested Standard Operating Protocol (SOP) Standard Operating Protocol (SOP) for management of women with endometriosis/endometrioma

Guidelines, Reading Material ESHRE endometriosis guideline 2015 [1], Royal College of Obstetrics and Gynaecology (RCOG) green top guidelines and NICE Guidelines 2013 [4].

Further Reading M. Hamdan et al. The impact of endometrioma on IVF/ICSI outcomes: A systematic review and meta-analysis. Hum. Reprod. Update 2015. 21(6):809–25. M. Hamdan et al. Influence of endometriosis on assisted reproductive technology outcomes: A systematic review and meta-analysis. Obstet. Gynecol. 2015. 125(1):79–88. F. Raffi, M. Metwally and S. Amer. The impact of excision of ovarian endometrioma on ovarian reserve: A systematic review and meta-analysis. J. Clin. Endocrinol. Metab. 2012. 97(9):3146–54. A. M. Sanchez et al. The distinguishing cellular and molecular features of the endometriotic ovarian cyst: From pathophysiology to the potential endometrioma-mediated damage to the ovary. Hum. Reprod. Update 2014. 20(2): 217–30. E. Somigliana et al. Risks of conservative management in women with ovarian endometriomas undergoing IVF. Hum. Reprod. Update 2015. 21(4):486–99. S. K. Sunkara et al. Association between the number of eggs and live birth in IVF treatment: An analysis of 400 135 treatment cycles. Hum. Reprod. 2011. 26(7):1768–74.

How to Manage Endometrioma in the IVF Patient

1. G. A. Dunselman, et al. ESHRE guideline: Management of Women with Endometriosis. Hum. Reprod. 2014. 29 (3): p. 400–12.

5. R. Pabuccu, G. Onalan and C. Kaya. GnRH agonist and antagonist protocols for stage I-II endometriosis and endometrioma in in vitro fertilization/intracytoplasmic sperm injection cycles. Fertility & Sterility 2007. 88(4): 832–9.

2. C. Yap, S. Furness and C. Farquhar. Pre and post operative medical therapy for endometriosis surgery. Cochrane Database Syst. Rev. 2004(3): CD003678.

6. J. Rodriguez-Purata et al. Endometriosis and IVF: Are agonists really better? Analysis of 1180 cycles with the propensity score matching. Gynecol. Endocrinol. 2013. 29(9):859–62.

3. H. N. Sallam et al. Long-term pituitary down-regulation before in vitro fertilization (IVF) for women with endometriosis. Cochrane Database Syst. Rev. 2006(1): CD004635.

7. L. Benaglia et al. IVF outcome in women with accidental contamination of follicular fluid with endometrioma content. Eur. J. Obstet. Gynecol. Reprod. Biol. 2014. 181: 130–4.

4. National Institute of Clinical Excellence. Fertility: Assessment and treatment for people with fertility problems. NICE Clinical Guideline 2013. 156:29.

8. M. Hamdan et al. The sensitivity of the DNA damage checkpoint prevents oocyte maturation in endometriosis. Sci. Rep. 2016. 6:36994.

References

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11

How to Manage IVF Patients with Intrauterine Adhesions

Introduction

Hysterosalpingogram (HSG)

Intrauterine adhesions (IUAs) or intrauterine synechiae are abnormal fibrous connections joining tissue surfaces of the uterine cavity. IUAs are scars that result from trauma to the uterus, most commonly associated with any instrumentation of the uterine cavity or infection. When IUAs occur with menstrual abnormalities, infertility or recurrent miscarriages, they may be referred to as “Asherman’s syndrome.” The true incidence of IUAs or Asherman’s syndrome is difficult to ascertain as women with mild IUAs may be asymptomatic. The majority of severe IUAs is related to uterine curettage for pregnancy complications, and there is an increased prevalence of IUAs in women who undergo curettage after a pregnancy loss at a later gestational age when compared with earlier gestation. Moreover, the risk of IUAs is greatest in patients with repeated curettages or in those who undergo uterine curettage two to four weeks after delivery when compared to the early postpartum period [1]. Rarely, IUAs can be caused by infection, especially endometrial tuberculosis, which may result in complete obliteration of the uterine cavity in most of the cases.

In women who had HSG performed as part of the workup for infertility, the HSG film should be reviewed. Apart from assessing the patency of the tubes, HSG may provide information concerning the internal configuration of the uterine cavity and the presence of any uterine anomaly or IUAs, which may present as sharp, irregularly shaped filling defects in variable locations in the uterine cavity. Occasionally, the cavity may simply appear narrowed. In severe cases of IUAs, partial or complete obliteration of the uterine cavity with or without obstruction of the tubes may be evident. Soares et al. reported a sensitivity of 75 percent in detection of IUAs and a positive predictive value of 50 percent [2]. The main drawback of HSG is that air bubbles, mucus, debris and endometrial polyps may also produce filling defects, resulting in relatively high false positive results.

Presenting Features

Transvaginal ultrasound scan (TVS) is a routine investigation prior to IVF treatment. In women suspected to have IUAs, several ultrasonographic (USG) features alert to the diagnosis of the condition: (1) thin or irregular endometrial echo at one or more sites, often due to a combination of adhesions and underlying fibrosis; (2) dense irregular echoes within the endometrial cavity may indicate fibrosis and calcification; (3) one or more echolucent cystic areas interrupting the endometrium may be found, which signifies the trapping of menstrual blood within adhesion bands. The best time to perform TVS evaluation of the endometrium is during the luteal phase of the cycle. TVS also allows visualization of the uterine cavity when complete obstruction of the cervix renders HSG or hysteroscopy impossible. The reported

Jacqueline Chung Pui Wah and Tin-Chiu Li

In women planning to go ahead with in vitro fertilization (IVF) treatment, the following features should alert to the possibility of presence of IUAs: (1) women with scanty or reduced menstrual flow, especially if it started after surgical evacuation of uterine cavity after miscarriage or delivery; or (2) women with recurrent miscarriage or recurrent implantation failure; or (3) ultrasound examination showing a persistently thin and irregular endometrium, or fluid collection within the uterine cavity.

Diagnosis To confirm the diagnosis of IUAs, the following investigations may be considered:

Ultrasound Two-Dimensional Transvaginal Ultrasound Scan (2-D TVS)

How to Manage IVF Patients with Intrauterine Adhesions

sensitivity and specificity of 2-D TVS in detecting IUAs is both around 95 percent [2].

Saline Infusion Sonography (SIS) The use of saline infusion during the ultrasound scan has been shown to improve the diagnostic accuracy. This method is particularly useful in cases when 2-D TVS is inconclusive or uninformative.

Three-Dimensional Transvaginal Ultrasound Scan (3-D TVS) 3-D TVS is being rapidly adopted in the field of reproductive medicine. 3-D TVS permits 3-D rendering and reconstruction of the coronal plane and allows more accurate assessment of the uterine cavity. It can also be used to classify the severity of IUAs. Kim et al. [3] further confirmed the accuracy of 3-D TVS in depicting IUAs and the extent of cavity damage in patients with suspected IUAs using six morphological characteristics of the endometrium on 3-D TVS including: marginal irregularity in the coronal plane, endometrial thinning of < 2 mm, a defect in the endometrial lining, obliteration or undetectable endometrium suggesting extensive adhesion, fibrosis as indicated by hyperechoic lesion without posterior shadowing or calcification. They divided their patients into three groups – Group 1 with one 3-D TVS finding, Group 2 with two findings and Group 3 with three findings or more. The sensitivity of diagnosis in Groups 1, 2 and 3 was 88.23 percent, 97.67 percent and 100 percent, respectively.

Three-Dimensional Saline Infusion Sonography (3-D SIS) When severe IUAs are suspected on clinical or 2-D/ 3-D TVS, it is desirable to proceed with 3-D SIS before embarking on hysteroscopy for a number of reasons. From time to time, in women with severe IUAs, the uterine cavity and endocervical canal may be completely or nearly completely obliterated, in which case hysteroscopy may be rather challenging, even with ultrasound guidance. 3-D SIS may help to delineate prior to hysteroscopy the exact locations of the adhesions, which may help the surgeon to be better prepared in planning the operation, including the type of instrument and anesthetics required. It may well replace the need for a preliminary diagnostic office or outpatient hysteroscopy, which is preferred by some surgeons.

Hysteroscopy Hysteroscopy is the gold standard for the diagnosis and treatment of IUAs. However, it is necessary to

Figure 11.1 Central intrauterine adhesion band upon hysteroscopy

make a choice of hysteroscopy with no-touch techniques in the office setting or in theatres under spinal or general anesthetics. When clinical and USG features are suggestive of mild IUAs, it is possible to perform the hysteroscopy in the office setting and to remove the adhesions using a combination of cold steel (scissors) and balloon dilatation therapy (see later in this chapter). However, if the clinical and USG features are consistent with severe IUAs, it is desirable to arrange the hysteroscopy in operating theatres under spinal or general anesthetics. In recording the adhesions within the uterine cavity, it is recommended to describe not only the location, but also the extent and severity of the adhesions; whether both ostia could be visualized before or after hysteroscopic adhesiolysis should be documented. An assessment should be made of the amount of normal endometrial tissue present at the end of the operation. One should consider the use of either the American Fertility Society (AFS) [4] or European Society for Hysteroscopy (ESH) [5] scoring system in order to provide a baseline with which any subsequent assessment (second-look hysteroscopy) could be compared.

Treatment Hysteroscopic Surgery In the past, blind division using Hegar dilators and curettes were used to treat IUAs. Nowadays, hysteroscopic surgery is the mainstay of treatment. Ideally, hysteroscopic adhesiolysis should be performed

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in removing marginal adhesions, especially those around the ostia, and so may be used in conjunction with conventional hysteroscopic scissor dissection: once the endocervical and central adhesion bands have been removed using hysteroscopic scissors, the Foley balloon technique may be employed to remove the marginal adhesions.

Prevention of Adhesion Reformation

Figure 11.2 Foley catheter with tip cut off for use in the Foley balloon technique

during the proliferative phase of the menstrual cycle. It is important during preoperative counseling to explain to the patient that second-look hysteroscopy is often required, especially in those with dense fibrotic adhesions or obliteration of the cavity. In performing hysteroscopic adhesiolysis it is always wise to use the ostia as a landmark to guide how far and how lateral one should go. Fibrotic bands or patches should be removed by sharp dissection or electrosurgery using a resectoscope with loop electrodes. When using electrosurgery, the minimal effective energy should be used to avoid damage to the endometrial tissue. In cases where the cavity is completely obliterated, USG guidance is recommended. Fluoroscopic guidance has been suggested, but this is nowadays rarely used as it exposes the patient to ionizing radiation.

Foley Balloon Therapy Saravelos and Li reported a novel ultrasound-based interventional technique for treating mild cases of IUAs [6]. The balloon of a 12 Fr Foley catheter with the tip cut off was introduced into the uterine cavity and inflated with 5 mls of normal saline or water, which can be used to stretch open adhesions bands. The advantage of the Foley balloon technique is that it can be performed as an outpatient procedure without the need for anesthesia. However, care must be taken not to overstretch the balloon or the uterine cavity, as it may cause excessive pain and tear open muscle fibers. It is not necessary to inflate the balloon beyond 5 mls. The Foley balloon technique is particularly useful

The reported rate of reformation of IUAs after hysteroscopic adhesiolysis is high, ranging from 3.1 percent to 23.5 percent and up to 62.5 percent in severe adhesions [7]. Thus, steps should be taken to prevent reformation of adhesion. Various strategies have been reported to prevent recurrence. The majority of these strategies aims to mechanically separate the uterine walls after operation so the dissected surfaces can heal without adhering back together.

Intrauterine Contraceptive Devices (IUCDs) One of the most commonly used methods in the past is the insertion of an intrauterine contraceptive device (IUCD) following hysteroscopic adhesiolysis and kept in situ for two weeks to three months postoperatively. The Lippe’s loop was most widely used because of its larger surface area and inertness. However, it is no longer available in many countries. Since then, copperbearing IUCDs have been available, but there is concern that they may induce excessive inflammatory reaction and instead promote the reformation of adhesions. Moreover, T-shaped IUCDs were not useful as the surface area is too small to effectively prevent IUAs. In case an IUCD is considered, either the loop IUCD or a heart-shaped IUCD should be considered.

Foley Catheter A Foley catheter with balloon inflated may be kept inside the uterine cavity for a week or two as a physical barrier to prevent reformation of adhesions. Orhue et al. [8] compared the use of an IUCD or a Foley catheter balloon. In their study, the IUCD was removed after three consecutive withdrawal vaginal bleedings while the Foley catheter was removed 10 days after hysteroscopic adhesiolysis. Menstrual restoration in the Foley catheter group was 81.4 percent when compared with 62.7 percent in the IUCD group. They concluded that the Foley catheter appeared to be a more effective method than the IUCD.

How to Manage IVF Patients with Intrauterine Adhesions

Cook Intrauterine Balloon A triangular, heart-shaped balloon that conforms to the normal shape of the uterine cavity has recently been developed (Cook Medical Inc., Bloomington, Indiana, USA) for use instead of the Foley balloon. In a retrospective cohort study, Lin et al. [9] compared the efficacy of intrauterine balloon, IUCD and hyaluronic gel in the prevention of adhesion reformation after hysteroscopic adhesiolysis. They found that the Cook intrauterine balloon or IUCD was more effective than hyaluronic acid gel. Later, the same group performed a prospective randomized controlled trial (RCT) [10] to compare the efficacy of Cook’s IU balloon and a heart-shaped IUCD in the prevention of adhesion reformation after hysteroscopic adhesiolysis. Both devices were removed seven days after the operation. Both devices appeared to have similar efficacy based on adhesion score reduction during second-look hysteroscopy one to two months after the initial hysteroscopic adhesiolysis. We would recommend the insertion of an intrauterine device for at least a week after hysteroscopic adhesiolysis of severe IUAs. Our current preference is the Cook intrauterine balloon, followed by Foley balloon if the former is not available; we seldom use the IUCD nowadays.

Hyaluronic Acid and Other Anti-Adhesion Barriers Hyalobarrier® (Fidiaa Advanced Biopolymers SRL, Padova, Italy) is a highly transparent viscous gel formed by highly purified, auto cross-linked polymers (ACP) of hyaluronic acid. Hyaluronic acid gel was found to be effective in preventing IUAs’ reformation after hysteroscopic adhesiolysis or de novo formation of adhesion after evacuation of the uterus. However, Lin et al. [10] suggested that the hyaluronic acid gel was less effective than the intrauterine balloon or heart-shaped coil intrauterine coil in preventing adhesion reformation. It is, however, unclear if the use of hyaluronic acid gel would confer additional benefit when used in conjunction with the balloon or coil. Seprafilm® is another mechanical bioresorbable adhesion barrier that is used by some surgeons to prevent postoperative adhesions. Unlike the Hyalobarrier® gel, it is a bioresorbable membrane of chemically modified hyaluronic acid and carboxymethylcellulose. The membrane may be rolled into

a thin cylinder and pushed into the endometrial cavity and endocervical canal; it then turns into a hydrophilic gel around 24 hours after placement and can provide a protective coating around the traumatized tissues up to seven days during reepithelization. The introduction of the membrane into the uterine cavity is more tedious than the application of gel, which explains why the latter is more often preferred. Overall, there is yet insufficient evidence to support the routine use of hyaluronic acid gel or Seprafilm® after hysteroscopic adhesiolysis.

Amnion Graft Both fresh and freeze-dried amnions have been used with some success to prevent adhesion reformation after intrauterine adhesiolysis. After the operation, a Foley catheter with amnion graft attached on its surface was introduced into the uterine cavity. The Foley balloon was then distended with 3.5–5 mls of normal saline. The end of the Foley catheter was then cut off below a knot near the balloon end and the balloon was removed two weeks later with antibiotics prophylaxis given. So far, the number of cases in studies utilizing amnion graft remains small and so further studies are required to verify its safety and efficacy before it can be recommended for routine clinical use.

Antibiotics Although infection and inflammation may be a primary cause for IUAs, current evidence does not recommend the routine use of antibiotics in patients undergoing hysteroscopic surgeries. However, antibiotics prophylaxis may be considered if an intrauterine device such as the balloon or coil is inserted for a period of time after the operation to avoid secondary infection.

Estrogen Therapy and Regeneration of Endometrium Apart from the restoration of normal uterine cavity and prevention of adhesion reformation, there is also a need to promote regeneration of the endometrium to resume normal function. This depends on successful re-epithelization of basal endometrium. Currently, postoperative estrogen therapy is considered beneficial, but there is no consensus about the dose, regimen (continuous or cyclical, estradiol alone or combined estradiol and progesterone) or duration (4–12 weeks) of the hormonal treatment. A commonly used

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protocol is estradiol valerate 4 mg per day for four weeks followed by progestogen (e.g., medroxyprogesterone acetate at a dose of 10 mg/day) in the last two weeks of estrogen treatment.

without adequate secretory transformation is very poor; in such a case, the patient must be carefully counseled regarding the poor prognosis.

Embryo Transfer Postoperative Assessment Second-Look Hysteroscopy It is advisable to perform second-look hysteroscopy a few weeks after hysteroscopic adhesiolysis, especially in women with a desire to conceive. Adhesions are easier to be separated when they are still fresh and flimsy. Given time, the reformed adhesions will gradually become fibrotic and more difficult to remove. Second-look hysteroscopy is particularly important in women with severe IUAs as the likelihood of recurrence of adhesion is rather high.

Assessing Endometrial Function It is necessary to check not only if adhesions have reformed after adhesiolysis but also if endometrial function has returned to normal prior to embarking on IVF treatment. It is important to point out that an adhesion-free uterine cavity does not necessary imply that endometrial function and receptivity are normal. There are two methods to assess endometrial function after IUA adhesiolysis. Ultrasound examination of the endometrium in the mid-cycle and the mid-luteal phase is necessary to provide important information on endometrial thickness and regularity, as well as any unusual features such as collection of fluid. Additionally, 3-D TVS may be used to measure endometrial volume and Doppler study may be considered to examine sub-endometrial blood flow. The inclusion of SIS to the 2-D or 3-D examination (discussed earlier in this chapter) may provide an additional opportunity to check if the cavity is indeed adhesion free. If endometrial thickness is suboptimal, high-dose estrogen therapy such as estradiol valerate 8–10 mg per day should be considered and endometrial thickness monitored again to determine if there is any improvement. If the endometrium remains thin despite hormone therapy, a biopsy should be obtained on day LH+7 or equivalent for histological dating to assess if there is adequate secretory transformation. A thin endometrium is not a contraindication for embryo transfer provided there is adequate secretory transformation. However, the chance of successful implantation in women with a thin endometrium

Whenever possible, only one embryo should be transferred at a time in women who underwent treatment for IUAs because of the increased risk of miscarriage and various obstetric complications in this group of women (see later in this chapter). The overall reproductive outcome correlates well with the severity of IUAs, with higher pregnancy and live birth rates in the mild IUA group compared with lower pregnancy and live birth rates in the severe group.

Managing Future Pregnancy It is well recognized that pregnancy following successful treatment of Asherman’s is associated with increased risk of spontaneous miscarriage, preterm birth, placenta previa, ectopic pregnancy, severe early-onset intrauterine growth restriction and spontaneous uterine rupture during pregnancy [1, 7]. Therefore, it is necessary that women who conceived after treatment for Asherman’s syndrome receive specialist obstetric care with regular, close monitoring throughout the pregnancy.

Suggested Standard Operating Protocol (SOP) Management of women with intrauterine adhesions including clear definitions and diagnostic criteria

Suggested Audits Clinical outcomes with and without surgical interventions of women with intrauterine adhesions

References 1. A. B. Hooker, M. Lemmers, A. L. Thurkow, M. W. Heymans, B. C. Opmeer, H. A. Brölmann, et al. Systematic review and meta-analysis of intrauterine adhesions after miscarriage: Prevalence, risk factors and long-term reproductive outcome. Hum. Reprod. Update 2014;20:262–78. 2. S. R. Soares, M. M. Barbosa dos Reis and A. F. Camargos. Diagnostic accuracy of sonohysterography, transvaginal sonography, and hysterosalpingography in patients with uterine cavity diseases. Fertil. Steril. 2000;73: 406–11. 3. M. J. Kim, Y. Lee, C. Lee, S. Chun, A. Kim and H. Y. Kim, et al. Accuracy of three dimensional

How to Manage IVF Patients with Intrauterine Adhesions

ultrasound and treatment outcomes of intrauterine adhesion in infertile women. Taiwan J. Obstet. Gynecol. 2015;54: 737–41. 4. The American Fertility Society classifications of adnexal adhesions, distal tubal occlusion, tubal occlusion secondary to tubal ligation, tubal pregnancies, mullerian anomalies and intrauterine adhesions. Fertil. Steril. 1988; 49:944–55. 5. K. Wamsteker. European Society for Hysteroscopy (ESH) classification of IUA. 1989. (Class III) 6. S. H. Saravelos and T. C. Li. Ultrasound guided treatment of intrauterine adhesions in the outpatient setting. Ultrasound Obstet. Gynecol. 2016 Jul 15. 7. D. Yu, Y. M. Wong, Y. Cheong, E. Xia and T. C. Li. Asherman syndrome–one century later. Fertil. Steril. 2008;89:759–79.

8. A. A. Orhue, M. E. Aziken and J. O. Igbefoh. A comparison of two adjunctive treatments for intrauterine adhesions following lysis. Int. J. Gynaecol. Obstet. 2003;82:49–56. 9. X. Lin, M. Wei, T. C. Li, Q. Huang, D. Huang and F. Zhou, et al. A comparison of intrauterine balloon, intrauterine contraceptive device and hyaluronic acid gel in the prevention of adhesion reformation following hysteroscopic surgery for Asherman syndrome: A cohort study. Eur. J. Obstet. Gynecol. Reprod. Biol. 2013;170:512–16. 10. X. N. Lin, F. Zhou, M. L. Wei, Y. Yang, Y. Li and T. C. Li, et al. Randomized, controlled trial comparing the efficacy of intrauterine balloon and intrauterine contraceptive device in the prevention of adhesion reformation after hysteroscopic adhesiolysis. Fertil. Steril. 2015;104:235–40.

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What Is the Role of Complementary and Alternative Medicine during IVF? A Review of the Evidence Trevor Wing and Andrew Flower

For the purposes of this chapter, we define complementary and alternative medicine (CAM) as a wide array of health care practices that exist outside of the current biomedical model used to understand, diagnose and treat ill health. As we are discussing evidence-based CAM approaches that may help with in vitro fertilization (IVF), the available research restricts our focus more precisely to the potential role of herbal medicines and acupuncture for this purpose. It is outside of the remit of this chapter to consider the possible role of psychological interventions in assisting IVF. Acupuncture is a practice originating in East Asia several thousand years ago that involves the insertion of fine needles into clearly located acu-points on the surface of the body. Traditional thinking describes this as a process to restore the healthy and balanced circulation of “vital energy” or “Qi” around the body through a network of interconnected acupuncture channels. Herbal medicine involves the use of plants (Oriental systems of herbal medicine also include minerals and animal products) to promote healing and alleviate ill health. Most of the research into herbs to support IVF investigates a form of herbalism called Chinese herbal medicine (CHM), which also developed in East Asian countries and has a recorded history of use of more than 2,000 years. In this chapter we consider the potential role that these approaches may have during IVF and investigate whether there is any robust in vitro (lab-based) and in vivo (clinical) evidence to support any claims for these interventions. We also discuss how to integrate CAM interventions during IVF treatment.

In Vitro Research: Biologically Plausible Mechanisms Assisted reproductive technology (ART) aims to improve female (and male) ability to conceive and

for the female to become pregnant. There are several types of ART, the most common being IVF and intrauterine insemination (IUI). More recent and advanced treatments still are largely based on these two fundamental treatment approaches. We therefore focus on biologically plausible mechanisms for CHM and acupuncture in improving the success rates for IUI and IVF. By “success rates,” we are specifically referring to achieving pregnancies confirmed with beta human chorionic gonadotropin (BhCG) urine pregnancy tests. IUI utilizes either a natural (non-medicated) treatment cycle or stimulated treatment cycle, in which case either ovulation induction pharmaceutical medications (PM) (clomiphene) or mild gonadotrophin medications are administered to increase the probability of ovulation. Acupuncture administered in the follicular phase of an IUI cycle is aimed at improving ovarian blood supply and therefore follicular estradiol secretion, improving the probability and strength of ovulation, and enhancing the endometrial quality, leading to an endometrium that is more receptive to embryo implantation. CHM given for the three months before IUI also probably improves “pipeline” folliculogensis, leading to an increase in follicle numbers in the IUI treatment cycle and therefore an increase in estradiol with its beneficial effects on ovulation and endometrial receptivity. IVF in its many flavors always aims to mature one or more follicles to the size where the oocytes contained within the follicles can be removed under ultrasound guided follicular fluid aspiration for fertilization in vitro and later returning morula or blastocyst embryos to the uterine cavity. In the case of natural cycle, IVF acupuncture and CHM treatments aim to support both the follicular phase and the luteal phase in much the same way as discussed earlier for IUI. There are no ovarian stimulation medications used and therefore no interaction effects to be taken

Complementary and Alternative Medicine during IVF

into account. Ideally again treatment should start three months before the IVF cycle to maximize the positive effect on folliculogenesis and then within the natural IVF cycle to improve ovarian blood supply, improve estradiol levels and improve endometrial quality, resulting in improved receptivity to embryo implantation. In the case of stimulated (short or long protocols), IVF acupuncture and or CHM is still beneficial in the three months before the stimulated IVF cycle to improve folliculogenesis before medication begins. Because the intended physiological effects of acupuncture and CHM and gonadotrophin medications are similar in the follicular phase of the treatment cycle, it is not advisable to use acupuncture and CHM treatments for these purposes during the IVF cycle as the interaction with stimulation gonadotrophins has not been researched to date and the outcome is unpredictable and unknown. Medicated IVF cycles can, however, be supported by focusing the acupuncture and CHM on endometrial quality and enhancing receptivity to embryo implantation when embryos are returned to the uterus.

In Vivo Research: Clinical Trials on Acupuncture and CHM Acupuncture Following the publication in 2002 of a clinical trial [1] that appeared to show that acupuncture administered during IVF resulted in significant improvements in clinical pregnancy rate and live births, there has been an active interest in researching the potential contribution of acupuncture to a successful IVF outcome. Unfortunately, subsequent research in 40 randomized controlled trials (RCTs) and at least 10 different systematic reviews has produced inconclusive and inconsistent results. Major challenges in researching acupuncture have led to this confusion. Unlike conventional medical treatment, traditional acupuncture is usually individualized according to the unique presentation of each patient. This means that there is a wide variety in treatment regimes, both in terms of which points are selected and when and how often treatment is administered, and what level of training is required to deliver this intervention. This diversity in practice means that it can be problematic trying to aggregate the results of trials to enable a more powerful metaanalysis of all the available data. However, while

standardizing the acupuncture intervention might be good from a research perspective, it would mean a significant deviation from traditional notions of best practice. Within the framework of evidence-based medicine, clinical trials comparing active treatment with inert “placebo” controls are considered the gold standard for rigorous evidence. However, it is difficult to provide an inert control for acupuncture. The use of “sham” Streitberger needles, which are designed to look as if they are being inserted, can still exert pressure on acu-points that may have a therapeutic effect, and the needling of non-acupuncture points is hardly an inactive control. In addition to these methodological challenges, many of the acupuncture trials are small in size and may be poorly designed. This means that they may be subject to bias and their findings are unreliable. Because of these factors, it is possible to consider all the evidence from acupuncture trials and to come to quite different conclusions. A systematic review by Cheong et al. [2] considered 14 trials involving 2,670 women undergoing IVF and concluded there was no evidence of benefit of acupuncture during assisted conception. Two years later a different review by Zheng et al. [3] involving 24 clinical trials and 5,807 women found that, once studies excluding the use of (the controversial) Streitberger needle were excluded, acupuncture did in fact increase both clinical pregnancy and live birth rates. These results were challenged by a re-analysis of the data a year later by Meldrum et al. [4] that excluded poorer-quality trials and concluded that there was now no evidence of benefit. These negative findings were confirmed by a Cochrane review published in 2013 [5], although both Cheong and Meldrum did find that acupuncture had a statistically significant improvement on pregnancy rates and live births in studies that did not use ‘sham’ acupuncture as a control. This suggests that these “sham” controls are indeed not inert and that acupuncture may have a useful effect in a “real-world” context. The most recent systematic review at the time of writing [6] considers the results of 21 RCTs involving 5,428 participants. This analysis tries to evaluate the impact of acupuncture delivered at three different time points: (1) immediately before and after embryo transfer (ET), (2) acupuncture administered at 30 minutes post ET, and (3) acupuncture administered before ET in the follicular stimulation phase as well as

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before and after ET. While there were no statistically significant benefits for acupuncture over controls for the first style of intervention, the second and third approaches do show significant improvements in pregnancy rates. However, the trials involved were few in number and small in size (only two trials involving 442 participants and two trials involving 284 participants, respectively) in each of these subgroups, so this can only really be considered as preliminary data that need further investigation. What can we say, then, to women considering acupuncture treatment to support them during IVF? There is little doubt that the initial promise of Paulus’s early trial has had to be muted and qualified. The combined research findings are equivocal and ultimately rather unhelpful. Acupuncture can help produce relaxation and may provide a supportive environment for women undergoing the stressful process of IVF. It may be these associated or “contextual” effects, rather than any “specific” effect of acupuncture, that produces positive findings in studies comparing acupuncture with a non-“sham” control. From a patient perspective, mechanisms are less important than outcome, and acupuncture does appear to confer some advantage in conception and live birth rates, even though it may not work in the way acupuncturists maintain it does. Given that acupuncture also appears to be a safe intervention [7] and shows no increase in risk for miscarriage [5], we maintain that it is valid to recommend it to women undergoing IVF, but with the proviso that more research is required to understand the mechanisms involved and to properly quantify the extent of the benefits that can be expected from this intervention.

Chinese Herbal Medicine Given the importance of succession within Chinese culture, the tradition of CHM has always placed great emphasis on treatments to improve fertility. Clinically the supportive evidence for CHM is, like acupuncture, unsatisfactory. The most up-to-date summary appeared in 2013 as a systematic review of 20 RCTs involving 1,721 women who had undergone IVF in conjunction with CHM. This paper reported a significant improvement in pregnancy rates as a result of the combined CHM and PM interventions [8]. In addition, the use of CHM and PM did not lead to any increase in rates of miscarriage or ovarian hyperstimulation syndrome. However, the methodological quality of the included trials was poor and

subject to a number of biases, and there was little or no reporting on live birth rates, so these data must be interpreted with a degree of caution.

Whole System Traditional Chinese Medicine In clinical practice, Chinese medicine is usually administered as a complex intervention that combines its main therapeutic modalities of acupuncture and CHM with associated interventions such as lifestyle advice, dietary recommendations, breathing and relaxation exercise, and massage. This “package” of care is known as whole system traditional Chinese medicine (WS-TCM). It makes sense to evaluate the impact of WS-TCM on assisted reproduction, and more of this kind of pragmatic research is starting to appear. A retrospective cohort study of 1,231 women attending a private IVF clinic in Oregon in the United States [9] found that in non-donor treatments, WSTCM was associated with greater odds of live birth than IVF alone (adjusted odds ratio [AOR] 2.09; 95% confidence interval [CI] 1.36 to 3.21), or embryo transfer with acupuncture only (AOR 1.62; 95% CI 1.04 to 2.52). Similar but slightly reduced benefits were apparent in a smaller sample of 162 women undergoing donor cycles. The use of WS-TCM was not associated with any increased risk of miscarriage, ectopic pregnancies or multiple pregnancies. This study has obvious limitations. It is a retrospective analysis without randomization to balance any confounding influences – although adjustments were made for known factors such as age, drug dosage and previous IVF cycles. Women also chose the treatments they undertook, which could create a selection bias, and finally there was a disparity in embryo quality between the groups. In fact, women in the WS-TCM group were diagnosed with a diminished ovarian reserve compared to women in the other two groups, so the effect of WS-TCM may have been underestimated. Once again there is the obvious need for more rigorous prospective RCTs to substantiate these preliminary findings. However, until such work is completed, there does not appear to be any reason to dissuade women interested in combining IVF with WS-TCM from pursuing this path.

CAM Interventions during IVF We now take a more precise look at how these CAM interventions can influence IVF at various phases of

Complementary and Alternative Medicine during IVF

the treatment cycle, with particular reference to how acupuncture and CHM are commonly used in clinical practice. Conventional medicine (CM) IVF treatments consist of several stages, with each stage modifying the patient’s natural menstrual cycle. The three main elements of CM IVF treatment and CAM supporting treatment are: 1) Ovarian follicle stimulation to grow and mature as many antral follicles as possible to a size that allows each follicle to be aspirated, resulting in oocyte collection. This stimulation phase is usually between 10 and 14 days. During this phase, acupuncture and or CHM can be used to reduce anxiety and improve blood flow within the pelvis to promote endometrial quality. Care should be taken not to use any CHM medicinals which would directly act on the ovaries, leading to unpredictable ovarian response to stimulation medication. 2) Follicle ripening/ovulation prevention. Once the stimulation phase is completed, oocytes need to be detached from the follicle wall membrane. Granulosa cells referred to as “ripening or triggering” at the same time as natural ovulation must be prevented, and this is achieved by administering a luteinizing hormone-releasing hormone antagonist. This allows around 36 hours in which oocytes can be collected without fear of loss due to natural ovulation. During this phase, there is no specific acupuncture or CHM physiology treatment indicated; however, continuing to treat to reduce anxiety, stress and adverse effects from medication is helpful. 3) Post-oocyte-collection progesterone supplementation to prevent endometrial shedding and pregnancy loss. Between three and five days (usually) post oocyte collection, the fertilized embryo/s will be returned to the uterine cavity and hopefully a receptive endometrium will allow implantation and the initiation of endogenous hCG to prevent the onset of menses and support the early trophoblast. During this phase, the contextual effects of acupuncture may help to improve embryo implantation rates. CHM formulas that support and improve ovarian vascularity and endometrial receptivity can also be employed to help improve the chances of implantation further. Should the cycle

result in pregnancy, usually discovered around 10 days post ET with a home pregnancy test, then acupuncture and CHM treatment can be modified to minimize miscarriage probability. This is particularly helpful in a patient with a history of repeated early pregnancy loss in IVF cycles.

Conclusion We have seen that there are biologically plausible explanations for how acupuncture and CHM may improve the success rates of IVF. We have also presented and tried to make sense of contradictory research about acupuncture and the limited investigations of CHM and WS-TCM. We have also described the possible role of these CAM interventions during the ART cycles. Given that these interventions appear to be safe and may contribute to the success of IUI and IVF, there may be a place for them within ART treatment. While the evidence is not sufficiently robust to routinely recommend the use of CAM, there is no reason to dissuade women with an interest in these supporting treatments from opting to include herbal medicines or acupuncture as part of their fertility care program. Further research is required to provide real clarity on the potential role of these CAM treatments in supporting ART. In the meantime, we recommend that those who are interested in these CAM options ensure they receive treatment from properly qualified, experienced and professionally regulated practitioners who have further training in reproductive gynecology. In the United Kingdom, three main professional registers can be contacted to find a suitably qualified practitioner: RCHM BAcC ATCM

References 1. W. E. Paulus, M. Zhang, E. Strehler, I. El-Danasouri and K. Sterzik. Influence of acupuncture on the pregnancy rate in patients who undergo assisted reproduction therapy. Fertil. Steril. 2002 Apr;77(4):721–4. 2. Y. Cheong, L. G. Nardo, T. Rutherford and W. Ledger. Acupuncture and herbal medicine in in vitro fertilisation: A review of the evidence for clinical

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practice. Hum. Fertil. (Camb.) 2010 Mar;13(1):3–12. doi: 10.3109/14647270903438830 3. C. H. Zheng, M. M. Zhang, G. Y. Huang and W. Wang. The role of acupuncture in assisted reproductive technology. Evidence-based complementary and alternative medicine. eCAM. 2012;2012:543924. doi: 10.1155/2012/543924 4. D. R. Meldrum, A. R. Fisher, S. F. Butts, H. I. Su and M. D. Sammel. Acupuncture – help, harm, or placebo? Fertility and Sterility 2013; 99(7):, 1821–4. 5. Y. C. Cheong, S. Dix, E. Hung Yu Ng, W. L. Ledger and C. Farquhar. Acupuncture and assisted reproductive technology. Cochrane Database Syst. Rev. 2013 Jul 26;7: CD006920. doi: 10.1002/14651858.CD006920.pub3 6. C. Shen, M. Wu, D. Shu, X. Zhao and Y. Gao. The role of acupuncture in in vitro fertilization: A systematic review and meta-analysis. Gynecol. Obstet. Invest. 2015;79 (1):1–12. doi: 10.1159/000362231. Epub 2014 May 16

7. A. Nandi, A. Shah, A. Gudi and R. Homburg. Acupuncture in IVF: A review of current literature. J. Obstet. Gynaecol. 2014 Oct;34(7):555–61. doi: 10.3109/01443615.2014.919997. Epub 2014 Jun 9 8. H. Cao, M. Han, E. H. Y. Ng, X. Wu, A. Flower, G. Lewith, et al. Can Chinese herbal medicine improve outcomes of in vitro fertilization? A systematic review and meta-analysis of randomized controlled trials. PLoS ONE 2013;8:12: e81650. https://doi.org/10.1371/ journal.pone.0081650 9. L. E. Hullender Rubin, M. S. Opsahl, K. Wiemer, S. D. Mist and A. B. Caughey. Impact of whole systems traditional Chinese medicine on in vitro fertilization outcomes. Reproductive Biomedicine Online 2015;30 (6):602–12. doi:10.1016/j.rbmo.2015.02.005. 10. Zheng, C. H., et al. Effects of acupuncture on pregnancy rates in women undergoing in vitro fertilization: A systematic review and meta-analysis. Fertility and Sterility 97(3): 599–611.

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How to Manage the Patient with Hydrosalpinges before an IVF Cycle Arri Coomarasamy and Justin Chu

Background/Context Approximately one-quarter of female infertility is caused by tubal disease [1]. Previous abdominal surgery and endometriosis can cause tubal infertility; however, the majority of these patients have tubal disease secondary to ascending infection (most commonly Chlamydia trachomatis) acquired from sexual transmission. This can lead to the development of pelvic inflammatory disease that, when untreated, causes chronic infection of the fallopian tubes. The anatomical structure of the fallopian tubes can become lost if untreated with loss of ciliary function. This causes dilation of the tube, which is known as hydrosalpinx. The prevalence of hydrosalpinx in women with tubal infertility is approximately 30 percent [2]. Hydrosalpinx is a term used to describe a spectrum of tubal pathology caused by distal fallopian tube occlusion. The tubal occlusion causes fluid to accumulate inside the tube. This can be diagnosed by ultrasonography or identified at diagnostic laparoscopy.

How Does the “Problem” Relate to IVF and Vice Versa? Before in vitro fertilization (IVF) became commonplace, reproductive surgeons would repair hydrosalpinx by restoring tubal anatomy. Women would then be asked to attempt natural pregnancy. To provide a further treatment option, IVF was introduced to help patients with tubal infertility (including those with hydrosalpinx). As the availability of IVF treatment has become more widespread, substantial evidence has emerged to suggest that proceeding with IVF treatment with an untreated hydrosalpinx yielded significantly worse results compared with women who did not have hydrosalpinx. Those women with hydrosalpinges visible on ultrasonography were found to have particularly poor outcomes [3]. The cause of this worsened treatment prognosis in those with hydrosalpinx is not well understood.

However, several theories have been postulated. It has been suggested that the hydrosalpingeal fluid could be embryotoxic or impair the process of embryo implantation and development [4]. There are also theories that the hydrosalpinx fluid may also leak from the fallopian tube and wash out any embryo that is replaced inside the endometrial cavity [5]. In women who have hydrosalpinges visible on pelvic ultrasonography, the treatment of hydrosalpinx by removing or occluding the fallopian tube was shown to increase the chances of clinical pregnancy and live birth by several research groups 15 years ago. Observational and interventional data have demonstrated that removing the affected tube or keeping hydrosalpingeal fluid from entering the endometrial cavity approximately doubled the chances of clinical pregnancy compared to women who had not had this treatment. A Cochrane review published in 2010 by Johnson et al. [6] collated all primary data from studies investigating the treatment of hydrosalpinges prior to IVF and concluded that surgical treatment (salpingectomy or tubal occlusion) should be offered to all women diagnosed with hydrosalpinx before an IVF cycle. In light of these findings, it has now become common clinical practice to perform salpingectomy or tubal occlusion on women with hydrosalpinx before commencing IVF treatment. In some women, hydrosalpinx may not be diagnosed prior to the commencement of the IVF treatment cycle; instead, it may be identified later, during the ovarian stimulation phase. In this challenging scenario, the effectiveness of transvaginal ultrasound guided drainage (performed at the time of oocyte retrieval) has been investigated. Unfortunately, primary studies investigating this treatment strategy have shown that transvaginal drainage of hydrosalpingeal fluid offers comparable clinical pregnancy rates with women in whom hydrosalpinx is untreated [7]. This is likely due to the high recurrence rate of hydrosalpinx when this technique is used. Furthermore, it has been found that transvaginal

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drainage of hydrosalpingeal fluid performed at the time of oocyte retrieval yields worse reproductive outcomes than when hydrosalpinx is treated surgically (by salpingectomy or tubal occlusion) before IVF treatment is started. Some reproductive physicians believe that performing salpingectomy or tubal occlusion on all patients with hydrosalpinx and infertility is a radical strategy. In a woman with bilateral hydrosalpinges, performing these procedures would leave the patient sterile and would not allow for further attempts at natural conception. These women would be left reliant on further IVF treatment cycles in order to achieve pregnancy, which carries risks associated with assisted reproductive treatments (such as ovarian hyperstimulation syndrome), as well as significant financial costs. Therefore, a role remains in restoring anatomy by performing tubal conserving surgery such as salpingostomy, which is done to drain the hydrosalpingeal fluid. Women can then attempt natural conception for a time interval before undergoing more definitive surgery. The rates of natural conception achieved after salpingostomy have been found to be comparable to the clinical pregnancy rates achieved by IVF treatment [8]. In the United Kingdom, the overall live birth rate per embryo transfer recorded by the Human Fertility and Embryology Authority was 36 percent in 2014. This is comparable to the natural conception rates achieved by salpingostomy (27 percent), although this is associated with an ectopic pregnancy rate of 10 percent [8].

Strategies/Solutions Women under investigation for infertility who are found to have hydrosalpinx need careful and thorough counseling. As mentioned earlier, a number of strategies are available to treat the hydrosalpinx before IVF treatment is commenced. However, the current evidence suggests that some form of tubal surgery should be performed prior to beginning IVF treatment to optimize the chances of pregnancy. The correct strategy to adopt will depend on the wishes of the patient as well as the surgical capabilities of the medical team delivering her care. It is important to ensure that the patient understands the consequences of each of the treatment strategies and that the surgeon is aware of her wishes. Before surgery, the potential findings and associated surgical options should be fully explored with the patient. Correct

and comprehensive counseling enables the surgeon to avoid performing irreversible surgery or to avoid repeat surgery due to overly conservative surgery. In women with reduced ovarian reserve or those with more severe forms of hydrosalpinx, the most appropriate strategy would be to perform sterilizing surgery followed by IVF treatment. However, in younger women and those with milder forms of hydrosalpinx, it may be more appropriate to offer tubal conserving surgery as tubal function may be restored and there is more time for the patient to attempt natural conception. Research suggests that women should attempt natural conception for at least 24 months before proceeding with more definitive surgery such as salpingectomy or tubal occlusion followed by IVF treatment [8]. Where sterilizing surgery is to be performed, the surgeon will need to decide whether salpingectomy is required or whether tubal occlusion would be a better method of ensuring that hydrosalpingeal fluid is kept separate from the endometrial cavity. This may be decided intraoperatively. Most clinicians would favor tubal occlusion when anatomy is distorted by adhesions or when there is significant risk of damaging the ovary, making salpingectomy more difficult to perform. The advantage of salpingectomy is to improve access to the ovaries for oocyte retrieval and reduce the risk of infection during the procedure. Where access to the pelvis is anticipated to be complicated (e.g., due to a history of extensive pelvic or abdominal surgery or in the case of frozen pelvis) hysteroscopic tubal occlusion can be considered. The use of ESSURE devices to occlude the fallopian tubes has been found to yield similar reproductive outcomes when compared to salpingectomy or laparoscopic tubal occlusion treatment of hydrosalpinx [9]. However, note that there is the need to perform pelvic ultrasonography three months after the insertion of ESSURE devices to ensure correct placement. Whichever strategy is used, effective counseling and accurate documentation is vital as patients must be made aware of the risks and consequences. In women who are to undergo sterilizing surgery for bilateral hydrosalpinges, it is important to emphasize that they will be reliant on IVF treatment in the future, which may require self-funding. Furthermore, patients should be counseled that there may a reduction in ovarian blood flow and therefore reserve if salpingectomy is performed.

The Patient with Hydrosalpinges before an IVF Cycle

Women who are considering tube-conserving surgery should be made aware that there is an increased risk of ectopic pregnancy if pregnancy is achieved and that they may still need more definitive surgery later on should the hydrosalpinx recur.

Related Research Areas/Websites Although it has become clear that hydrosalpinx reduces the chance of clinical pregnancy achievable by IVF treatment, a number of uncertainties still face a clinician when considering the management strategy. First, it remains unclear which of the treatment strategies mentioned earlier should be offered to patients. More research is required to aid the clinician in deciding whether to perform sterilizing surgery (salpingectomy or tubal occlusion) or tubeconserving surgery. Although the pragmatic approach would be to suggest tube-conserving surgery to those women who are younger and those who have milder forms of hydrosalpinx, the exact subgroup of patients that this strategy should be offered to is not yet clear. A trial comparing the two strategies may improve patient selection. However, due to the technical difficulties with performing salpingostomy, this trial would be difficult to power and to conduct. Furthermore, success rates from tubal surgery would be specific to the surgeon and the surgical techniques performed. Further research is also needed to investigate the best technique to perform salpingostomy. For example, some tubal surgeons advocate the formation of a sutured cuff to prevent the closure of salpingostomy, whereas others perform a simple linear salpingostomy. There is also a lack of available evidence to guide clinicians toward performing salpingectomy or tubal occlusion if sterilizing surgery is the treatment strategy chosen. As mentioned earlier, there is some evidence that performing salpingectomy can reduce the blood supply to the ovary, inhibiting the ovarian response to gonadotrophins during stimulation [10]. However, salpingectomy is also thought to improve access to the ovaries during transvaginal oocyte retrieval and to reduce the risk of complications of hydrosalpinx (such as torsion) in later life. Further research would aid clinicians to perform the best surgery to enable the best outcomes for patients.

Conclusion In many ways, managing a patient with hydrosalpinx identified prior to starting IVF treatment is simple and complex simultaneously. Although it is clear and well accepted that some form of surgical treatment should be performed to the hydrosalpinx, the exact nature of this surgery is not well understood. In many reproductive medicine and surgery centers, the treatment options have become limited due to clinicians becoming de-skilled in tubal surgery. Importantly, this has meant that patients with hydrosalpinx may not be offered a full range of treatment options, with sterilizing surgery and subsequent IVF treatment becoming the most popular strategy to employ in patients with hydrosalpinx. In some countries, where government funding for reproductive treatment is more substantial, having repeated cycles of IVF treatment may be a possibility. However, in other countries, where infertile couples with hydrosalpinx may only receive one cycle of treatment, tube-conserving surgery for hydrosalpinx may have a more important role. These couples have the chance to attempt natural conception. Only when an adequate period of time has elapsed should more definitive tubal surgery be offered followed by IVF treatment.

Suggested Standard Operating Protocol (SOP) Management of women with tubal disease Suggested patient information sheet: Tubal disease and fertility

Suggested Audits Several audits can be conducted to ensure that women with hydrosalpinx identified before the start of IVF treatment are correctly managed. In particular, it would be useful to ensure that all women are offered some form of tubal surgery before starting IVF treatment. Additionally, an audit could be performed to ensure that women are appropriately counseled with all treatment options offered, and that the consequences of their hydrosalpinx treatment are well explained.

References 1. N. Johnson, S. van Voorst, M. C. Sowter and A. Strandell Mol. tubal surgery before IVF. Hum. Reprod. Update 2011;17.

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2. A. Strandell. Treatment of hydrosalpinx in the patient undergoing assisted reproduction. Curr. Opin. Obs. Gynecol. 2007;19:360–5. doi:10.1097/GCO.0b013e3282 1642b9\n00001703-200708000–00013 [pii] 3. A. Strandell. The influence of hydrosalpinx on IVF and embryo transfer: A review. Hum. Reprod. Update 2000;6: 387–95. 4. M. Kassabji, J. A. Sims, L. Butler et al. Reduced pregnancy outcome in patients with unilateral or bilateral hydrosalpinx after in vitro fertilization. Eur. J. Obstet. Gynecol. Reprod. Biol. 1994;56:129–32. 5. R. F. Savaris and L. C. Giudice. The influence of hydrosalpinx on markers of endometrial receptivity. Semin. Reprod. Med. 2007;25:476–82. 6. N. Johnson, S. van Voorst, M. C. Sowter et al. Surgical treatment for tubal disease in women due to undergo in vitro fertilisation. Cochrane Database Syst. Rev. 2010; CD002125. doi:10.1002/14651858.CD002125.pub3

7. N. Hammadieh, A. Coomarasamy, B. Ola et al. Ultrasound-guided hydrosalpinx aspiration during oocyte collection improves pregnancy outcome in IVF: A randomized controlled trial. Hum. Reprod. 2008;23: 1113–17. doi:10.1093/humrep/den071 8. J. Chu, H. M. Harb, I. D. Gallos et al. Salpingostomy in the treatment of hydrosalpinx: A systematic review and meta-analysis. Hum. Reprod. 2015;30:i448. doi:10.1093/humrep/dev135 9. P. Arora, R. S. Arora and D. Cahill. Essure((R)) for management of hydrosalpinx prior to in vitro fertilisation: A systematic review and pooled analysis. BJOG 2014;121:527–36. doi:10.1111/14710528.12533 10. M. Fan and L. Ma. Effect of salpingectomy on ovarian response to hyperstimulation during in vitro fertilization: A meta-analysis. Fertil. Steril. 2016. doi:10.1016/j.fertnstert.2016.03.053

Chapter

14

Tubal Assessment in the IVF Patient Ka Ying Bonnie Ng and Ying Cheong

Background It is estimated that one in six couples requires investigation and treatment of subfertility, and tubal factors account for about 14 percent of causes of infertility in the women, with a much higher prevalence in India due to higher rates of unrecognized pelvic inflammatory disease and tuberculosis. Tubal damage can occur with any external or internal injury and potentially hinders the normal transport of gametes. Examples include pelvic infection, endometriosis and previous abdominal surgery. The Fallopian tube is a dynamic and complex structure, and hence, it is important to understand that demonstration of tubal patency does not always correspond to tubal function. Coordinated neuromuscular activity, cilial action and endocrine secretions are some of the factors required for successful tubal function. Assessment of tubal patency should be done alongside semen analysis and investigations to assess ovulation. An “ideal” or “gold standard” test for tubal patency would be sensitive (i.e., all true positives would be identified by a positive test result and a negative test result would rule out disease in all those without the disease) and specific (i.e., the test result would only be positive in women with the disease). Many tests are available in the assessment of tubal disease and this reflects the fact that there is no one “ideal” test; none yields perfect accuracy and predictive values. They are also subject to operator expertise and potential intraoperative or technical complications.

When Does the Patient Require Tubal Assessment Prior to Assisted Conception? Very often, by the time the patient arrives at the fertility center, they will already have had some form of assessment performed on their Fallopian tubes. If there were abnormalities, these may have been

investigated and treated. But the following scenarios may occur whereby the fertility specialist has to decide if further tubal assessment testing is required before starting the patient on assisted conception treatment.

The Patient with an Untreated, Uninvestigated Abnormal Tubal Assessment Test It is possible that a previous tubal assessment test had highlighted an abnormality at the time when the patient arrives in the fertility clinic. It is crucial, and the onus is on the treating physician, to request for the necessary video/films/reports to clarify the nature of these abnormalities and the treatment strategy instituted prior to further fertility treatment. The following scenario and suggested solutions are tabulated in Table 14.1.

The Patient Who Had a Normal Tubal Assessment Before, but Since Developed Risk Factors for Tubal Blockage These risk factors include the following: complex abdominal surgery, pelvic inflammatory disease, complex appendicitis requiring major surgical intervention and sexually transmitted disease. This patient is now at high risk of having developed a tubal blockage. Depending on the interim event, an investigative laparoscopy in view of treatment, or a further tubal assessment test is warranted to exclude hydrosalpinx.

The Patient Who Has a Uterine Abnormality Diagnosed at Tubal Assessment The methods of tubal assessments also facilitate the diagnosis of uterine abnormalities, including that of intrauterine polyps, fibroids and adhesions. The management of these conditions are dealt with in Chapters 3 and 11, respectively.

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Table 14.1 Suggested solutions and salient counseling points for the patient with an untreated, uninvestigated abnormal tubal assessment test prior to their fertility treatment (IUI or IVF). IUI: Intrauterine insemination; IVF: in vitro fertilization

Tubal assessment abnormality observed

Solution

Salient counseling points

Hydrosalpinx (uni- or bilateral)

Need to advise tubal surgery versus removal (refer to Chapter 13 on the management of hydrosalpinges)

Patient needs to understand that current evidence suggests that if a hydrosalpinx is not removed prior to IVF, the pregnancy success rate may be halved. Patient needs to understand that if bilateral salpingectomy was performed, the only chance of conception will be via IVF. The success of IVF per cycle will need to be explained and understood by the patient. If tubal surgery was discussed, the risk of ectopic pregnancy needs to be highlighted.

Possible proximal blockage

Advice for laparoscopy + dye test with consideration to proceed to hysteroscopic cannulation of proximal blockage; recommend radiological cannulation of the Fallopian tube if this is available

IUI/DI patient: A significant proportion of these will be as a result of tubal spasms. But a discussion prior to surgery of tubal cannulation will allow the surgeon to proceed without relisting the patient for another procedure. IVF/intracytoplasmic sperm injection (ICSI) patient: it is justifiable to proceed to IVF/ ICSI if patients do not wish to further investigate their tubal condition and/or have other fertility factors, e.g., male factor.

Uterine adhesions

Advice for hysteroscopic resection of adhesions under ultrasound guidance +/−Laparoscopy + dye

Hysteroscopic resection of intrauterine adhesion can be a complex operation (see Chapter 11 on Asherman’s syndrome) and referral to a tertiary center for the surgical management of this condition is necessary if local expertise is not available.

Uterine polyp

Advice for hysteroscopic removal of uterine polyp

Avoid removing polyp blindly with polyp forceps as this usually does not remove the polyp completely.

The Patient Who Never Had Any Tubal Assessment

they may choose to proceed straight to assisted conception treatment.

The following section describes the pros and cons of the methods currently available to assess the Fallopian tubes in this group of patients. Tubal assessment should be considered in women wishing to undergo assisted conception. A risk assessment needs to be made jointly between the clinician and the patient, and the outcome of this may be that the patient chooses to undertake tubal assessment, an investigative treatment such as laparoscopy or alternatively,

Methods of Tubal Assessment Hysterosalpingography (HSG) When Is It Performed? HSG is the radiographic evaluation of the uterus and Fallopian tubes; its main use is in assessing the whole tube, the condition of the tubal lumen and the site of

Tubal Assessment in the IVF Patient

block. It is useful in diagnosing endometrial and Mullerian abnormalities, including congenital anomalies, leiomyomas, synechiae, polyps, tubal occlusion, salpingitis isthmica nodosum (SIN) (also known as diverticulosis of the Fallopian tube), hydrosalpinx and peritubal adhesions. HSG is widely used for tubal evaluation in women presenting with subfertility and is a fairly accurate and easy way of identifying tubal damage. Both proximal and distal tubal blockage can be recognized in HSG. Proximal tubal blockage, typically caused by SIN, has a characteristic honeycomb appearance. Multiple small diverticular collections of contrast protrude from the lumen into the wall of the isthmic portion of the Fallopian tube. White flecks of contrast material often persist at the site of suspected blockage even in the post-drainage X-ray. In tuberculous salpingitis, there may be a similar calcification appearance; however, calcification of the uterus, ovaries and lymph nodes can also be present. The etiology of SIN is unknown, but it is probably a post-infectious reaction. Distally obstructed tubes might have small, clubshaped ends on HSG, representing the presence of hydrosalpinges. However, in most cases, distal tubal obstruction is detected by proximal filling with dye, but no spillage of dye into the peritoneum. A meta-analysis of three studies gave pooled estimates of HSG sensitivity as 0.65 (95% CI 0.50–0.78) and specificity as 0.83 (95% CI 0.77–0.88) [1]. Because of HSG’s low sensitivity, it is of limited use for detecting tubal obstruction, but its high specificity makes it useful for ruling out tubal obstruction. HSG may be used as a screening test for couples with no history of pelvic infection and if the test is abnormal, laparoscopic assessment should be performed.

from the fimbrial end, is observed and captured in the form of X-rays. Clear intraperitoneal spill must be demonstrated. If spill is not demonstrated, rotating the patient from side to side or gentle abdominal pressure may increase the likelihood of intraperitoneal dispersion. It has been suggested that the use of intravenous smooth muscle relaxants such as terbutaline and prostaglandins may help to relieve spasm.

How Is It Performed?

What Are the Risks?

HSG is usually performed in the early follicular phase; this avoids the possibility of pregnancy and facilitates maximum uterine visibility as the endometrium is thin in the proliferative phase. HSG is performed with the patient in the lithotomy position on the fluoroscopy table and aseptic precautions apply. After cleaning the vagina and cervix, an HSG cannula is introduced and stabilized in the cervix. A very small amount of radio-opaque contrast solution, warmed to body temperature, is introduced through the cannula into the uterus. The filling of the solution in the uterine cavity and passage through the Fallopian tubes, including its spill

The risks associated with WSCM HSG include potential reaction to the iodine containing radiopaque contrast. The most common complication is cramping at the time of contrast injection. Premedication, minimizing cervical trauma and a consideration of the emotional state of the patient may increase the tolerance of the procedure. Pelvic infection is a contraindication for HSG, and pelvic infection is a complication affecting 1–3 percent of women having HSG; prophylactic antibiotics should be given to those with risk factors for pelvic infection. OSCM may be associated with less pain, probably because there is less irritation of the peritoneum. Despite

Tubal Flushing at HSG The potential therapeutic effect of tubal flushing at HSG has been under speculation for more than 50 years. At HSG, a water-soluble contrast media (WSCM) or oil-soluble contrast media (OSCM) is used. A Cochrane review that collected evidence from 13 randomized controlled trials [2] showed that women having OSCM tubal flushing had a higher rate of pregnancy and live birth compared to women with no intervention; evidence suggests that for subfertile women, the chance of an ongoing pregnancy will be increased from 17 percent without intervention to between 29 percent and 55 percent with intervention. A more recent multicenter randomized controlled trial further confirmed the 10 percent improved live birth rate of OSCM over that of WSCM [3]. WSCM allows better imaging of the tubal mucosal folds and ampullary rugae (internal tube architecture) compared to OSCM. OSCM has a high viscosity, which leads to slow filling of the tubes; often, this requires a late film after 24 hours. However, sometimes, this “late” film can offer additional information, such as adhesions after slow peritoneal spillage. It is slower to resorb and remains in the pelvic cavity for longer.

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Figure 14.1 Tubal assessment in the IVF patient

positive effects on pregnancy rates, extravasation of OSCM into the pelvic cavity has potentially serious adverse effects such as lipogranuloma formation, which occurs if there is accumulation of OSCM within a blocked tube leading to a chronic inflammatory reaction or anaphylaxis if the OSCM enters blood vessels or lymphatics.

Laparoscopy and Dye When Is It Performed? Historically, laparoscopy and dye was the first-line screening evaluation for women with subfertility and is now regarded as the gold standard for assessing tubal patency. However, HSG or hysterosalpingocontrast-sonography (HyCoSy), when available, are less invasive procedures. At present, it is still considered the most accurate diagnostic test available for assessing tubal-related infertility. It enables the

operator to directly visualize the abdominal cavity and pelvic structures, including the external appearance of the Fallopian tubes and the fimbrial ends. In addition, pelvic pathologies such as endometriosis, which may contribute to infertility, may be diagnosed and treated.

How Is It Performed? At laparoscopy, diluted methylene blue is instilled into the uterine cavity and the Fallopian tubes via a cannula inserted into the cervix (traditionally a Spackman). Sometimes dye does not pass through an apparently normal tube due to the preferential flow of blue dye to the opposite tube. To overcome this problem, application of gentle pressure using a laparoscopic forceps over the tube with the preferential flow of dye allows for the redirection of blue dye to the other tube, and avoids the misdiagnosis of “blockage” in an apparently normal Fallopian tube (Figure 14.1).

Tubal Assessment in the IVF Patient

It is also important to ensure a good seal at the level of the Spackman cannula and the cervix to prevent backflow of dye into the vagina; if there is insufficient dye passing up the uterine cavity into the Fallopian tubes, patency cannot be shown. In this scenario, reapplication of the instruments to secure a better seal or the use of two teneculae, one over the anterior and the other over the posterior lip of the cervix, may ensure sufficient seal. However, in the event that the latter is not possible, the insertion of a Foley catheter into the uterine cavity, with the pulling back of the inflated balloon onto the internal cervical os, will ensure a better seal. Methylene blue can then be injected through the catheter without the problem of excessive backflow of dye into the vagina.

What Are the Risks? Some serious though rare complications are associated with laparoscopy, including injury to the bladder, bowel and blood vessels. The risk of a laparotomy as a result of a severe complication is 1.4 to 3.1 per 1,000 cases. This risk is increased in patients with previous abdominal or pelvic surgery, previous pelvic inflammatory disease or obesity. A prospective study reported a complication rate of 5.7 per 1,000 cases, with the most commonly observed being hemorrhage from the epigastric vessels and injury to the bowel [4].

Hysterosalpingo-Contrast-Sonography (HyCoSy) When Is It Performed? The National Institute for Health and Care Excellence (NICE) recommends HyCoSy as a primary investigation for tubal assessment for low-risk women without a history suggestive of tubal damage [5]. It is not a widely available investigation, although it is an effective alternative to HSG when the expertise is available. Women with comorbidities such as inflammatory disease, previous ectopic pregnancies or endometriosis should be offered laparoscopy and dye in order for tubal and other pelvic structures to be assessed at the same time. However, HyCoSy has high diagnostic accuracy in the assessment of the uterine cavity and there is a good statistical concordance with HSG and laparoscopy with dye [6].

How Is It Performed? HyCoSy assesses tubal patency and the uterine cavity by transvaginal ultrasound and the concomitant

instillation of an echogenic contrast medium into the uterine cavity via a catheter inserted through the cervical os. An echogenic contrast medium is required as there is no tissue–fluid interface in normal tubes. Fallopian tubes may only be visualized when there is a hydrosalpinx or free fluid in the pelvis outlining the tubes, or fluid is introduced in the tubal lumen or the pelvis. The cheap and cost-effective option is the use of saline mixed with air, which produces a contrast fluid due to the presence of air bubbles. However, these air bubbles stand for a short period, making tubal assessment practically more difficult. Ex-Foam can be used as an alternative to saline solution and remains stable for several minutes. The foam is created by diluting gel (containing glycerol and hydroxyethyl cellulose) purified in water; little air bubbles are formed by pushing the gel through small openings in syringes or tubes. HyCoSy is usually done in the midproliferative phase (days 6–10 of a 28-day cycle) of the menstrual cycle, after menstruation stops but before ovulation, to reduce the risk of disrupting an early pregnancy.

What Are the Risks? This examination has been shown to be safe and well tolerated with shorter examination times and fewer side effects compared to HSG. It avoids the risk of radiation, while providing the same information as HSG. The procedure has no major complications; the most common complications include nausea and vomiting and abdominal pain. The risk of pelvic infection and associated peritonitis is about 1 percent. Failure to perform or complete the procedure is documented in approximately 7 percent of cases [7].

Fertiloscopy or Transvaginal Hydrolaparoscopy When Is It Performed? Fertiloscopy is relatively new. One prospective multicenter study (the FLY study) compared the two endoscopic techniques of laparoscopy and fertiloscopy in routine evaluation of the pelvis in infertile women; they showed that both have similar sensitivity and negative predictive values, and concluded that fertiloscopy may be considered an alternative to diagnostic laparoscopy in the routine assessment of women

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without clinical or ultrasound evidence of pelvic disease [8].

How Is It Performed? Fertiloscopy is done under either general or local anesthetic and involves introducing 100–200 mls of saline via a Veress needle into the posterior fornix of the vagina and through the pouch of Douglas. After this, a mini laparoscope (less than 3 mm) is introduced via a trocar. This allows visualization of the posterior pelvis, including the posterior aspect of the uterus, ovaries and the Fallopian tubes. The tubal patency test can also be performed using methylene blue. If tubal pathology is found, falloposcopy can be performed at the same time. It is also possible to perform small interventions such as adhesiolysis, ovarian drilling and coagulation of endometriosis spots. Fertiloscopy is minimally invasive, substantially harmless and well accepted by patients. It is easily performed, even in obese patients. It permits a very “physiological approach” as it does not require mobilization of the tubes and ovaries for the examination. The procedure is quick, lasting 10–15 minutes, and is easy to learn. The development of surgical capabilities may in the future reduce the number of laparoscopies for conditions such as mild endometriosis.

What Are the Risks? There is a concern that fertiloscopy is associated with rectal and uterine injuries (although studies seem to show that this is not the case [9]). It is contraindicated in cases where there is evident pelvic pathology, requiring laparoscopy; these include pelvic masses occupying the pouch of Douglas (including posterior myomas, ovarian cysts and women with fixed uterine retroflexion). One of the limitations of fertiloscopy is the inability to visualize the anterior aspect of the uterus and uteri-vesical fold. There is low operative capacity and currently, fertiloscopy is only considered as a diagnostic procedure.

Falloposcopy or Salpingoscopy When Is It Performed? Falloposcopy is mainly used as a research tool and is a highly specialized procedure that allows direct vision of the inside of the Fallopian tube. There are suggestions that falloposcopy may be a more discriminatory test of tubal pathology, as one study has

shown that women with normal Fallopian tubes at falloposcopy have higher rates of spontaneous pregnancy (27.6 percent) compared to those with mild or severe endotubal lesions (11.5 percent to 0 percent) [10].

How Is It Performed? Falloposcopy involves cannulation of the Fallopian tubes via the cervix and uterus with ultrasound or hysteroscopic guidance. There are two main types of falloposcope: coaxial and linear everting catheter. In the coaxial technique, the uterotubal ostium is identified and a guidewire is introduced into the Fallopian tube until the fimbrial end or an obstruction is reached. A fiber-optic endoscope, usually no more than 5 mm in width, then replaces the guidewire. There is constant irrigation and distension of the tubes as the fiber-optic endoscopy is slowly withdrawn; this allows visualization of the Fallopian tube mucosa in a retrograde fashion. The linear everting catheter technique is a different kind of falloposcopy that does not require a guidewire.

What Are the Risks? Substantial training and investment are required for this highly technical procedure. It is used as a research tool, as there are still significant technical difficulties, such as cannulation difficulties and problems with visualization. Complications include pinhole tubal perforations and tubal wall dissection.

Chlamydia Testing Chlamydia antibody testing (CAT) is a low-risk screening modality, with minimal inconvenience to the patient, and should be considered as an initial infertility investigation, prior to laparoscopy. It can be performed at any time during the menstrual cycle. While a negative CAT can be reassuring, a positive test warrants further assessment using invasive diagnostic procedures such as laparoscopy. For CATnegative patients with subfertility, they should have further investigation with, e.g., HyCoSy if available. CAT, however, does have limitations; it is unable to identify women with noninfectious cases of tubal factor infertility (e.g., endometriosis, previous pelvis surgeries), antibody titers may decline over time, and false positives are possible. Chlamydia immunoglobulin G antibodies are thought to persist for many years and so have been assessed as markers of previous C. Trachomatis

Tubal Assessment in the IVF Patient

infection. CAT has been shown to be a noninvasive and cost-effective method of screening for potential past Chlamydia infection and for tubal factor subfertility. A meta-analysis of 23 studies showed that the discriminative capacity of the Chlamydia antibody detected using enzyme-linked immunosorbent assay (ELISA), micro-immunofluorescence or immunofluorescence in the assessment of tubal pathology was comparable to that of HSG in the diagnosis of tubal occlusion [11]. In addition, there was a positive correlation between the level of antibody titer and the degree of tubal damage. Chlamydia screening has a favorable costeffectiveness; the risk of pelvic inflammatory disease (PID) after lower genital tract chlamydia is in the range up to 30 percent. The risks of developing tubal infertility after PID are 10–20 percent. For detection of current infection of chlamydia, samples can be taken from the cervix or urethra for culture. An alternative for urethral/cervical cultures is nucleic acid amplification tests (NAATs) that can be performed on urinary samples or self-administered vulvo-vaginal swabs. The three types of commercially available NAATs are polymerase chain reaction (PCR), transcription-mediated amplification and strand displacement amplification.

Summary For many years, it has been presumed that tubal assessment essentially entails the assessment of tubal patency. Various tests and techniques have therefore been developed to focus on assessing tubal patency. A summary of these tests is shown in Table 14.2. It must, however, be appreciated that a patent tube does not equate to a functional tube and that “routine” tubal testing is not always necessary. The clinician must, in planning for the investigations of a subfertile couple, be able to individualize their care. A change of paradigm in tubal assessment will be when diagnostic tools are available to facilitate the assessment of tubal damage, patency and, last but not least, tubal function.

Suggested Standard Operating Procedure (SOP) • •

Performing HyCoSy (HSG is usually performed in radiology, and so will not normally be covered under the assisted conception center protocol regulation)

Suggested Audit •

Waiting time for tubal assessment

Table 14.2 Table comparing the various tubal assessment tests. −: No, +: Yes

Gold standard – diagnostic and therapeutic

HSG

HyCoSy

Lap and dye

Falloposcopy

Salpingoscopy

CAT

-

-

+

-

-

-

Widely available

+

-

+

-

-

-

Assess tubal patency

+

+

+

+

+

-

Assess tubal function

-

-

-

-

-

-

Learning curve

+

++

+

+++

+++

+

Cost

++

++

+++

++

++

+

Requires general anesthetics

-

-

+

+/-

+/-

-

Outpatient procedure

+

+

_

+/-

+/-

+

Blood test

-

-

-

-

-

+

Ultrasound scan performed at the same time

-

+

-

-

-

-

Research only

-

-

-

+

+

+/-

Ascending infection risk

+

+

+

+

+

-

Procedure-associated discomfort

++

++

+

++

++

-

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•

Clinical outcomes of tubal assessment (rate of adverse events, failed cannulation of the cervix)

Suggested Patient Information Sheet •

Tubal assessment

References 1. P. Swart, B. W. Mol, F. van der Veen, M. van Beurden, W. K. Redekop, et al. The accuracy of hysterosalpingography in the diagnosis of tubal pathology: A meta-analysis. Fertil. Steril. 1995;64:486–91. 2. L. Mohiyiddeen, A. Hardiman, C. Fitzgerald, E. Hughes, B. W. Mol, et al. Tubal flushing for subfertility. Cochrane Database Syst. Rev. 2015; 5:CD003718. 3. K. Dreyer, J. van Rijswijk, V. Mijatovic, M. Goddijn, H. R. Verhoeve, et al. Oil-based or water-based contrast for hysterosalpingography in infertile women. N. Engl. J. Med. 2017; 376:2043–52. 4. F. W. Jansen, K. Kapiteyn, T. Trimbos-Kemper, J. Hermans and J. B. Trimbos. Complications of laparoscopy: A prospective multicentre observational study. Br. J. Obstet. Gynaecol. 1997; 104:595–600. 5. Royal College of Obstetricians and Gynaecologists (RCOG) (2013) Nice clinical guideline. Fertility: Assessment and treatment for people with fertility problems.

6. A. B. Dijkman, B. W. Mol, F. van der Veen, P. M. Bossuyt and H. V. Hogerzeil. Can hysterosalpingocontrast-sonography replace hysterosalpingography in the assessment of tubal subfertility? Eur. J. Radiol. 200035:44–8. 7. C. D. de Kroon, G. H. de Bock, S. W. Dieben and F. W. Jansen. Saline contrast hysterosonography in abnormal uterine bleeding: A systematic review and meta-analysis. BJOG, 2003;110:938–47. 8. A. Watrelot, M. Nisolle, H. Chelli, C. Hocke, C. Rongieres, et al. Is laparoscopy still the gold standard in infertility assessment? A comparison of fertiloscopy versus laparoscopy in infertility. Results of an international multicentre prospective trial: The “FLY” (Fertiloscopy-LaparoscopY) study. Hum. Reprod. 200318: 834–9. 9. S. Gordts, A. Watrelot, R. Campo and I. Brosens. Risk and outcome of bowel injury during transvaginal pelvic endoscopy. Fertil. Steril. 2001;76:1238–41. 10. H. Dechaud, J. P. Daures and B. Hedon. Prospective evaluation of falloposcopy. Hum. Reprod. 1998; 13: 1815–18. 11. B. W. Mol, B. Dijkman, P. Wertheim, J. Lijmer, F. van der Veen, et al. The accuracy of serum chlamydial antibodies in the diagnosis of tubal pathology: A meta-analysis. Fertil. Steril. 1997; 67: 1031–7.

Chapter

15

Which Culture Media to Use Necati Findikli and Mustafa Bahceci

Human Embryo Culture Media: A Short Background and Current Status As it is seen in the most branches of science and medicine, initial work on embryo culture media was performed on cells, reproductive cells and tissues of laboratory and farm animals before translation into human reproduction. Since Ringer defined the first cell culture medium in 1882, subsequent refinements and improvements have progressively been made in the embryo culture media concept. Ironically, the first successful in vitro fertilization (IVF) embryos and babies were produced by using simple somatic cell culture media (Ham’s F-10). For many years, IVF clinics produced their own culture media by mixing and dissolving different cell culture components. A certain need for standardization as well as batch-to-batch consistency have finally led to the production of specific commercial media for human embryos [1]. Subsequent improvements as well as a considerably high demand from IVF treatment providers created a boom in the number of producers as well as changing the nature of the commercial embryo culture products, leading to the production of synthetic embryo culture media systems, including combinations of more than 80 different inorganic/ organic components with the aim of creating a human embryo culture environment that can be as competent as possible or mimics in vivo conditions the best [2]. Conceptual and practical development in this area have contributed to the so-called single-step and sequential culture media systems. Single-step media is also known as the “let the embryo choose” approach and resembles an “open buffet” for an embryo during preimplantation. Due to recent advances in timelapse embryo incubation, this approach, heavily based on a statistical model and computer-assisted algorithm to define the optimal concentration of each media component, has now gained popularity. The latter approach, which is based on a “back to nature” philosophy, states that, in order to have the most optimal embryo development in culture, one

should mimic the natural reproductive environment. It is therefore mainly based on a “two-step culture” composed of different culture environments that try to imitate the oviduct and the uterus.

Which One Is Better? Single Step or Sequential? The initial development of human embryo culture media was born out of academia and research [3]. However, it was quickly commercialized in the late 1980s due to an increasing demand for the widespread use of assisted reproductive technologies (ART) worldwide. The demand in ART has driven the increase in consistency and overall quality in the laboratory. However, the availability of a number of brands with no specific information on the content of the media has created significant confusion and despair among users. Recent studies, in which each ingredient of several commercial embryo culture media was analyzed, indicated a large variation, in terms of concentration, purity and batch-to-batch consistency of inorganic/organic culture components, as well as supplements among different products [4, 5]. The guidance as to which product is best or superior is often not based on human studies and is based on extrapolated animal studies. From a practical point of view, most laboratories usually adopt a single media or brand preference, and unless unexpected laboratory culture failures arise, most utilize a standard consistent protocol. Currently, embryo culture media comparison studies are often sponsored by commercial companies and hence have their associated issues relating to conflict of interest. The prospective comparative approach is the most common, utilizing a patient or sibling oocyte-based approach rather than per-patient comparisons, and so such studies are significantly confounded. In a recent systematic review and meta-analysis, Sfontouris and colleagues have documented that, although more than 500 studies have been designed as randomized controlled trials, only a handful have

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satisfied the eligibility criteria of a robust systematic review [6]. Furthermore, a majority of these studies that were initially included had methodological problems (randomization and concealment issues, absence of a proper power calculation and small sample sizes, high heterogeneity in the patient population, etc.). Based on the current data, drawing a firm conclusion about the superiority of one commercial media over another is not possible [7]. On the other hand, it is generally accepted in the embryology world that all embryo culture media performs more or less similarly, in the hands of qualified embryologists working within the laboratory with good laboratory practices. Until more informative research is conducted, the choice of culture media is often based on personal or laboratory experience for the given clinic. In a clinic that focuses on blastocyst-stage transfer, it may not be pragmatic to increase the number of steps within the embryo culture protocol, as failure to maintain the optimal culture condition may detrimentally impact the desired clinical outcome. The buffering capacity of different culture media may be different due to the variable composition of ingredients, and it takes laboratory personnel time and experience to familiarize with these differences, which are specific to their laboratory environment. A further pragmatic consideration is that, very often, laboratories may have their “preferred” partner from a logistical, pricing and distribution perspective, and hence the choice of “which embryo culture to choose” is unlikely to hinge solely on scientific evidence or merits, but more so on commercial grounds. The lack of availability of in-house quality control and quality assurance systems sometimes favors the selection of a product with an in-built tracking system compared to those without.

pH and temperature stability that are known to have a profound impact on embryo viability [8]. What is yet controversial is which is the best incubator to use as technology advances; there is now a range of choices from classical box-type incubators through multichamber bench-top incubators to all-in-one timelapse embryo culture incubators. Very often, IVF laboratories have more than one type or model of incubator, and it is key that embryologists are mindful of their similarities and differences. For example, incubators with time-lapse monitoring lack real-time pH monitoring, whereas box-type or bench-top incubators, in which a real-time pH monitoring can be implemented, do not have time-lapse morphokinetic assessment options. Clearly, manufacturers upgrade and modify their systems continuously; if these technologies are adopted, it is important that the upgrades are monitored carefully to ensure compliance with local protocols and policies.

Important Criteria to Consider while Choosing the Embryo Culture Media

Embryo Manipulation and Culture Approaches

Physical and Technical Infrastructure Very often, embryologists resist change from what they have been taught and are used to, even when they have left where they originally trained. It is key to appreciate that new laboratory environment brings new optimization challenges, and it is not sufficient to simply replicate the methods of culture based on a fixed formula. This includes optimizing variables such as air quality,

Culture Environment There is evidence that a more physiological environment for human embryos may result in beneficial clinical outcomes. A low-oxygen environment has been shown to be beneficial in the context of embryo culture in terms of a better clinical outcome [4]. However, such an assumption may be too simplistic. A recent elegant study suggests that the effects of culture media on embryo development were both protein and oxygen dependent, and that these interactions are complex. It is therefore fair to say that the culture environment has to be individualized to the laboratory setup and the experience of the personnel involved. It is also not inconceivable that the incubator and laboratory environment can in future be further optimized with the advent of technology to better the outcomes of the embryo culture.

Ideally, the developing embryo should be left in vitro culture with minimal disturbance until the embryo transfer. In contrast to this scenario, the developing embryo in vitro today is arguably more manipulated than ever before; apart from the observation of morphological development at regular intervals, the advent of technology to potentially improve implantation means that embryos can be subjected to procedural interventions such as assisted hatching,

Which Culture Media to Use

biopsy for genetic diagnosis or screening and cryopreservation prior to embryo transfer. While the traditional practice of IVF limited the amount of fluctuation of this environment until the cleavage stage before embryos are returned to the uterus, day 5 to day 6 extended culture exposes the developing embryo to a far longer period of variation in its nonnatural environment. Many culture media provide a standard baseline set of “ingredients” to minimize sudden fluctuation of the culture media environment when the media is being changed. The use of time-lapse incubators favors a single-step embryo culture media without the need for a media change on day 3; this technology has been shown to improve embryo quality, although it has not yet been proven to improve live birth rates [10]. The successful use of single-step embryo culture media needs to go hand in hand with a high-specification laboratory with a good air-handling system in situ. The impact of the various culture media, sequential or single step, on the developmental programming of the embryo is still not well studied, but is a growing area of research [11].

Future Prospects Significant effort has been made to ensure that the culture media, which the embryo grows in, is as physiological, natural and consistent as possible. However, the environment within the reproductive tract and the uterine cavity is not yet fully known. Hence, no matter how one varies the form of culture media use, the exact composition of the reproductive tract cannot yet be completely mimicked in vitro. In the future, technology such as those utilizing microfluidics or other sensing technologies may help scientists provide the in vitro embryo with a more natural environment. This consideration is crucial as the long-term outcome for IVF-born babies is still unknown. While current studies have reassured us in terms of the outcomes of children conceived under ART in terms of the perinatal outcomes such as congenital abnormalities, continued emphasis must be given to research examining the developmental differences, at birth and into adult life, between the embryo implanted from in vitro technology and those from natural conception.

Suggested Standard Operating Protocol (SOP) 1.

SOP on steps to introduce a new culture media system

2.

Risk assessments of changing culture media system

Suggested Audits 3.

Key performance indicators (clinical and laboratory) on embryo development and pregnancy rates after culture media change

References 1. P. Quinn, J. F. Kerin and G. M. Warnes. Improved pregnancy rate in human in vitro fertilization with the use of a medium based on the composition of human tubal fluid. Fertil. Steril. 1985. 44(4):493–8. 2. E. Chronopoulou and J. C. Harper. IVF culture media: Past, present and future. Hum. Reprod. Update 2015. 21 (1):39–55. 3. H. Landecker. It is what it eats: Chemically defined media and the history of surrounds. Stud. Hist. Philos. Biol. Biomed. Sci. 2016. 57:148–60. 4. D. E. Morbeck et al. Composition of commercial media used for human embryo culture. Fertil. Steril. 2014. 102 (3):759–66 e9. 5. D. E. Morbeck et al. Composition of protein supplements used for human embryo culture. J. Assist. Reprod. Genet. 2014. 31(12):1703–11. 6. I. A. Sfontouris et al. Blastocyst culture using single versus sequential media in clinical IVF: A systematic review and meta-analysis of randomized controlled trials. J. Assist. Reprod. Genet. 2016. 7. M. M. Youssef et al. Culture media for human pre-implantation embryos in assisted reproductive technology cycles. Cochrane Database Syst. Rev. 2015 (11):CD007876. 8. J. E. Swain. Optimal human embryo culture. Semin. Reprod. Med. 2015. 33(2):103–17. 9. K. H. Hong et al. Examining the temperature of embryo culture in in vitro fertilization: A randomized controlled trial comparing traditional core temperature (37 degrees C) to a more physiologic, cooler temperature (36 degrees C). Fertil. Steril. 2014. 102(3):767–73. 10. N. Costa-Borges et al. Blastocyst development in single medium with or without renewal on day 3: A prospective cohort study on sibling donor oocytes in a time-lapse incubator. Fertil. Steril. 2016. 105(3): 707–13. 11. S. H. Kleijkers et al. Influence of embryo culture medium (G5 and HTF) on pregnancy and perinatal outcome after IVF: A multicenter RCT. Hum. Reprod. 2016.

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Chapter

16

How to Set Up and Manage Key Performance Indicators What Are Red-Alerts and What to Do When They Occur? Rachel Cutting and Caitriona Meaney

How to Set Up and Manage Key Performance Indicators (KPI) Control charts are used in many different applications and are basically designed to measure the outcome of a process using an indicator that has a range of 0 to 100 (i.e., percentage values). David and Sharon Mortimer [1] described how baseline data should first be gained, for example, monthly averages of the clinic’s own patient population and data for the preceding six months for the indicator. The mean and standard deviation (SD) of these values are calculated and then used to determine the control mean plus two values, the warning limit (+/- 2SD) and the control limit (red-alert) (+/- 3SD) (Figure 16.1). Most professional bodies such as the European Society of Human Reproduction and Embryology (ESHRE) also recommend reference to published national and internal data when defining critical

performance levels. Above all, it is essential that data collected to generate KPIs are true and correct. They must be checked as with any other dataset for accuracy, especially if generated from an electronic patient database. A process is said to be “in control” if values remain close to the mean. Action is only required if indicator values begin to fall inside the warning or control limits. Such scenarios include when indicator values are continuously above the mean, or crossing the warning level in an adverse direction; this may be a sign that a problem is beginning to manifest itself and subsequent action to confirm this should be undertaken. In other cases where the indicator crosses the warning level three times in a row, this is more likely to be nonrandom event and further investigation should be taken to confirm if a problem exists. When the indicator goes to red-alert, then immediate action must be taken; this is discussed later on in this chapter.

100

Figure 16.1 Control chart (modified from Mortimer and Mortimer [1])

Indicator (0–100)

90 80

Upper control limit

70

Upper warning limit

60 50 Control mean

40 30 20

Lower warning limit

10

Lower control limit

0 Jan Feb Mar April May Jun July Aug Sep Oct Nov Dec

How to Set Up and Manage Key Performance Indicators

Conversely, when an indicator is continuously above the mean or crosses the warning or control limits in the beneficial direction, changes to a process that may have affected the indicator should be reviewed and built on in order to maintain the improvement seen. Control data should be regularly recalculated from recent data to account for changes in clinical and laboratory performances. As performances improve and become more standardized, the variability observed in data should be less frequent. Thresholds can also be based on how specific protocols or new techniques impact outcomes and the nature of the indicator [2, 3]; however, it is important to ensure that variability in performance levels does not adversely affect the measurement of improvement. In vitro fertilization (IVF) programs are highly regulated in many countries and the requirement for a quality management system (QMS) is mandatory in some countries such as the United Kingdom and Australia. As part of a QMS there should be a set of comprehensive KPIs that are objective, relevant to the laboratory and measure a broad range of specific events or aspects of treatment [2]. Integration of an international QMS such as ISO 9001:2015 can help clinics to manage KPIs and comply with regulatory requirements. An internal audit team whose task it is to perform internal audits prior to ISO examination ensures that the system is implemented and developed appropriately in order to meet with the standards’ requirements. Auditing a different team, e.g. a nurse auditing embryology processes, enables auditors to gain in-depth knowledge of the team and its processes, which may later contribute to refining and improving the management system. Professional bodies such as the Association of Embryologists have in their Good Practice Guidelines in Clinical Embryology [4] stated that all IVF centers should have a number of KPIs to allow evaluation and continuous quality improvements that should be performed at specified time intervals. The paper suggested a number of KPIs, including benchmarks (Table 16.1). ESHRE, in its revised guidelines for good practice in IVF laboratories [5], recommended an integrated quality management program to encompass and integrate processes and procedures. The guidelines state results should be evaluated on a regular basis, indicators should be objective and relevant, and adequate thresholds determined. ESHRE recommended the following indicators to be regularly reviewed, analyzed and discussed:

• • • • • • • • •

Number/rates of errors and adverse events Rates of normally fertilized oocytes Cleavage rates Rates of embryos of good quality Proportion of patients with failed fertilization Ongoing clinical pregnancy rates (fresh/frozen/ thawed transfers) Multiple pregnancy rates Implantation rates Rates of survival of zygotes and embryos after thawing

Frequency of analysis can depend on the size of the center and number of cycles undertaken. Smaller centers may not have sufficient numbers for weekly analysis and may prefer to examine monthly trends. However, large centers will generate enough data on a weekly basis, which means the KPIs are an extremely valuable tool to monitor performance. Centers may also use “gold standard” patients for their KPIs if the patient demographics are variable and to prevent bias. The definition of a gold standard patient may be defined individually by a center but may, for example, be a patient under the age of 37 years in her first cycle of treatment. However, it is essential not to focus too narrowly as alerts may occur in specific age groups if related to clinical management. As mentioned earlier, it is important not to just focus on laboratory activities in regards to accessing clinic performance. Other clinical or protocol-driven practices can directly affect KPIs such as operator and embryologist at embryo transfer. Audits such as these should be routinely carried out to ensure that all staff are competent in these potentially pregnancy ratelimiting procedures. Total quality improvement (TQI) is a relatively new approach to quality management in assisted reproduction clinics and aims to encompass all of the clinic’s activities, ensuring optimal performance not just through pregnancy rates but also in patient satisfaction and financial performance [2, 6]. Monthly patient satisfaction surveys are a useful tool in auditing if patients are happy with the service provided, if there are any areas that can be improved and if they would use the service again. Questionnaires can include statements describing a characteristic of the service received such as those centered around facilities, staff, communication, provision of information, etc. Responses can be measured using a five-point Likert scale (“Strongly disagree,” “Disagree,” “Neither

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Table 16.1 ACE laboratory key performance indicators (modified from Hughes and ACE [4]) These are suggested benchmarks. Centers should calculate their own benchmarks and trigger points based on their patient population and data.

OVERALL KPI

Calculation (x 100 for %)*

Suggested benchmarks

1

IVF fertilization rate

2PN + 3PN/No. inseminated

> 65%

2

IVF abnormal fertilization rate

≥ 3PN/No. inseminated

< 5%

3

IVF 1PN rate

1PN/No. inseminated

< 5%

4

ICSI fertilization rate

2PN/No. injected

> 65%

5

ICSI 1PN rate

1PN/No. injected

< 5%

6

ICSI damage rate

No. degenerate/No. injected

< 10%

7

Failed fertilization rate

No. of cases with 0 fertilized/No. inseminated

< 5%

8

Low fertilization rate

No. of cases with < 30% 2PN of Met II

< 10%

9

Cleavage rate

No. cleaved/No. 2PN

> 90%

10

IVF oocyte maturity

No. fertilized + unfertilized Met II/No. oocytes collected

> 80%

11

ICSI oocyte maturity

No. of Met II/No. of oocytes collected (at time of injection)

> 80%

12

Utilization rate

No. transferred or frozen/No. 2PN

> 50%

13

Blastocyst formation rate

No. 2PN with progression to D5/D6/Total no. of 2PN

> 50%

14

Frozen embryo survival rate

No. survived/No. thawed

> 70%

15

Follicle yield

No. of oocyte collected/No. follicles punctured

> 80%

Based on patients under the age of 40 who had at least three oocytes collected or more

FRESH CYCLES Pregnancy rates (%)

Suggested benchmarks D2/3

Suggested benchmarks D5/6

Positive hCG per oocyte retrieval (OR)

40%

45%

Positive hCG per embryo transfer (ET)

45%

50%

Clinical preg. (FH on scan at 7 weeks)/OR

35%

40%

Clinical preg. (FH on scan at 7 weeks)/ET

40%

45%

Multiple birth rate/ET

< 10%

< 5%

Pregnancy rates (%)

Suggested benchmarks D2/3

Suggested benchmarks D5/6

Positive hCG per thaw cycle

35%

40%

Positive hCG per thaw cycle + ET

40%

45%

Clinical preg. (FH on scan at 7 weeks)/thaw

30%

35%

Clinical preg. (FH on scan at 7 weeks)/ET

35%

40%

Multiple birth rate/ET

< 10%

< 5%

FROZEN CYCLES

agree nor disagree,” “Agree,” “Strongly Agree”). Examples of statements may include: • “The clinic reception area provides a welcoming environment.”

• •

“The staff always introduced themselves and treated me with courtesy.” “During the nurse consultation I understood all the information given to me.”

How to Set Up and Manage Key Performance Indicators

Performance targets Registration and detail input: 65% ICSI fertilisation rate >65%

Mismatch error rate: 0%

IVF/ICSI insemination?

Mixing of patient sperm and partner eggs

Incorrect type of insemination performed

DISCARD

Incorrect sperm and eggs mixed*

Figure 16.2 Sperm production and preparation process flowchart. * is opportunities for error involving witnessing procedures.

•

“The written information I was given was clear and easy to understand.”

Open-ended questions, i.e. those that do not set specific options for response, should be included as they allow patients to clarify their responses and encourage feedback that may be beneficial in improving or developing the service. Examples of these may include: • “How could written information be improved?” • “What did you feel was done well in your treatment and what could have been improved on?” Evaluation of surveys should be carried out regularly and relayed at clinic meetings to provoke discussions and promote an inclusive and positive workplace.

Yearly audits should be carried out on the survey data and indicators set for the following year. A good approach to beginning a TQI program is to map out all processes in each sector and determine performance targets for each process. Flowcharts are a good tool for process mapping, using a standard symbol set that represents different actions or steps in the process. On completion of a process flowchart it is then possible to identify where problems may occur and areas where improvements can be made. Risk analysis and failure mode and effects analysis (FMEA) can be used to quantify these potential problems [7]. On review of the example flowchart in Figure 16.2, mapping sperm production and preparation

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Initial patient contact

Virol./ Biochem

Referral

Rachel Cutting and Caitriona Meaney

Embryol.

IUI

FER

IVF/ICSI Initial medical consult

Clinical Care Admin.

Pregnancy Testing Notes to daily MDT meeting

Follow up Nurse Consultation

Financial Co-ordination

To counsellor

Monitoring Open appt: patient phones reception Day 1 bleed

Reception check details + registration

Financ. Services Counsel.

Treatment

Further testing

Billing

Available throughout and after process

Figure 16.3 IVF Workflow chart

processes, it is evident that many of the opportunities for error here involve witnessing procedures. Due to the irreversible and dramatic consequences of mismatch error, a performance target of 0 percent means that this procedure needs to function at 100 percent. Extra manual witnessing or introduction of an electronic witnessing system could help to maintain this performance target [7], and provide extra reassurance for patients in this critical area [8]. Determining a workflow plan that visualizes how these processes flow between different sectors gives a fuller picture of how the clinic functions as a whole and how the various processes integrate (Figure 16.3).

What Are Red-Alerts and What to Do When They Occur? If a KPI falls into alert, systematic checking of the implemented quality control checks and audits should commence to try to identify a cause. The troubleshooting process should start with the more obvious checks such as patient demographic data and unforeseen events; for example, if the fertilization KPI is low, it may be influenced by a failed fertilization case, so it is important to work out whether the unexpected KPI is due to a global

issue or an isolated problem. It is also useful to determine if the KPI has been constantly low, in warning limits, or in acceptable limits prior to this trigger. A decrease in any KPI should not be assumed to be solely laboratory based as clinical, technical, procedural and environmental factors can all affect an IVF program. A multidisciplinary, supportive and holistic team approach will ensure staff feel inclusive and prevent a blame culture. Systematic checking should then commence, looking at all areas as both clinical and laboratory aspects can affect KPIs. Mortimer and Mortimer [1] described the troubleshooting process as a structured, scientific process during which every detail should be examined (Figure 16.4). This process can be used for the variables that may affect a KPI, as described in what follows.

Processes Any new procedure or process should be fully validated before being introduced into a program, but even when validation is complete, subtle changes or deviations from a standard operating procedure (SOP) can affect KPIs. All processes should be

How to Set Up and Manage Key Performance Indicators

‘There seems to be a problem...’

Collect data

REINVESTIGATE

Analyse data

Define problems Collect data

Design experiment(s)

Prioritise problem(s) Resolve problems(s): • Identify cause(s) • Analyse relationships • Define/design solution(s) • Project outcomes • Implement solution(s) NOT OK

Monitor outcome

END

Figure 16.4 Troubleshooting process flowchart (modified from Mortimer and Mortimer [1])

reviewed, including drugs involved in stimulation regimens. Embryo transfer is a critical limiting point of an IVF cycle. Meldrum et al., as early as 1987, highlighted that a meticulous embryo transfer technique is essential for a successful program; reviewing operator pregnancy rates is important to ensure consistency in a team and will highlight operator variability [9]. It is important to recognize that the number of embryos transferred may affect the clinical pregnancy rate, so implantation rate is a more accurate measure of lab performance.

General Clinic and Laboratory Air quality has been documented as an important factor in determining the success of an IVF unit as many papers have highlighted the negative effect of poor air quality on outcome [10]. Disruption to positive air flow systems or problems with HEPA filters may affect air quality. This situation can be avoided by continuously measuring air flow and particle counts via an external monitoring system. Legislations such

as the European Union Tissues and Cells Directive (2004, revised 2008) recommend high standards of air quality for viable and nonviable particles in the assisted reproduction technology (ART) laboratory; a background environment of EU GMP Grade D and a working area environment of EU GMP Grade A [11]. Particle and microbial testing should be carried out at least quarterly and/or where a problem is noted, e.g., a KPI is unsatisfactory or in the red-alert. Portable particle counters with continuous measurements (required for Grade A testing), and contact surface plates are recommended methods of testing. As part of a troubleshooting exercise the levels of volatile organic compounds in the lab can be measured. Volatile organic compounds (VOCs) can accumulate in a lab from off gassing from plastic ware, incubators, cleaning agents, monitors, microscopes and furniture. Changes in VOC levels may occur due to building work being carried out in the vicinity, e.g., fumes from organic solvents in paints, cleaning solutions, noxious substances (glues), so the wider hospital/clinic area should be examined [13]. If baseline VOC levels haven’t been already determined, then monitoring should continue. Some commercial VOC meters are capable of detecting contamination at 0.1 parts per million (ppm) levels (0.1 ppm is below an established threshold of embryo toxicity). Published data are limited, but information from the commercial sector from their own validation exercises suggests that the threshold level of toxicity of Acrolein for mouse embryos has been established as 0.58 ppm when dissolved in media. Field experience suggests the 0.5 ppm indicates potential indoor air quality (IAQ) contaminants in laboratories. International consensus provides guideline limits for IAQ in the range from 0.10 to 0.65 ppm depending on location. If high levels are detected, air filtration units with activated carbon filters can be used. Other variables to examine are the air temperature supplied to theatres and laboratory, e.g., has the temperature changed (cooler air may affect egg quality). If a change in temperature does occur, fertilization and subsequent embryonic development can be negatively affected [14, 15].

Laboratory Equipment Rigorous monitoring of critical equipment, whether it is continuous or daily, should be routine in any laboratory and will pick up underperformance; however, in the event of an alert, further checks to

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determine whether the equipment is functioning at desired levels should be undertaken. Checks such as temperature of heated stages, incubator temperature, oxygen and carbon dioxide levels, temperatures of test tube blocks and temperature of fridges containing medium should be carried out. pH measurements should be taken to check if the incubator is providing the correct environment to maintain the appropriate pH of the culture media. As part of these checks questions should be asked to determine if the equipment used to monitor the equipment variable has been recently calibrated and is accurately measuring the variable. A useful process to undertake if suboptimal equipment is suspected is to conduct an equipment review protocol (ERP). This validation process is designed to formally check whether the equipment is installed correctly, whether it is in satisfactory condition, whether the service documentation is up to date and whether calibration has been undertaken. The document also details the performance history and the equipment is assessed formally as a pass, pass with comments or a fail.

Laboratory Consumables/Culture Medium A robust system should be in place for recording batch numbers, date of entry and expiration dates of all consumables. In the event of a red-alert, consumable batch numbers should be checked to determine if any new lot numbers have been introduced. In the United Kingdom, the Human Fertilisation and Embryology Authority (HFEA) states only products which are CE marked for IVF must be used, which does allow some safety aspects to be addressed. Consumables do come with certificate of analysis and often descriptive quality control checks, for example endotoxin checks and mouse bioassay results. Consumables should be stored and used as per manufacturer’s instructions. Use of media, for example for purposes for which it is not intended, and/or addition of exogenous substances, for example protein sources without recommendation, may result in adverse outcomes. In-house testing can be conducted if a specific product is suspected of causing an issue, for example sperm survival assay may detect a potential toxin [16]. In the event of an alert, batch numbers should be checked for the date of their introduction into the laboratory to determine whether any change coincides with a new batch. If an item is suspected of potential toxicity, in-house testing should follow as well as contacting suppliers for them to undertake further investigation with other users.

Staff Depending on the country there are differing training programs for both clinical and scientific staff, with embryological training being the most variable. Robust training will drive consistency, but it is essential to audit performance. Annual performance checks of individuals can include intracytoplasmic sperm injection (ICSI) and embryo transfer. ICSI performance checks should include fertilization rate, damage rate, cleavage rate, utilization rate and clinical pregnancy rate as a minimum. Individual parameters for the embryos transfer audit can include clinical pregnancy rate, implantation rate, ectopic rate and early miscarriage rate. Ensure that the demographics of transfer patient do not give false results in these cases. If a red-alert occurs, these audits should be brought forward and performance checked. Both experienced and new practitioners can be checked through direct observation of procedural skills (DOPs) or SOP competency reviews. This involves a member of the staff using the defined SOP to shadow an individual undertaking a task to check if they are compliant against the SOP. A checklist approach can be used, allowing constructive monitoring and evaluation. This may pick up minor changes in practice that could affect a KPI. Another important variable to consider is embryo grading. Morphological scoring of embryos has shown considerable interand intra-observer variability, which can significantly impact clinical success rates [17]. In order to audit embryo grading, laboratory staff should participate in both internal and external quality assurance (EQA) programs for embryo grading. Commercial schemes such as EQASRM and UK NEQAS offer morphological assessment of embryos where individuals can assess their skills against other professionals in the field. With the introduction of time-lapse and associated algorithms into ART as an embryo-selection tool, assessing dynamic and static morphologic parameters is now also required. Internal QA schemes are recommended to evaluate inter- and intra-rater reliabilities by intra-class correlation coefficient (ICCs) [17].

Conclusion A robust quality control program will help to minimize internal laboratory factors affecting embryo quality and success rates. However, downward trends can still inexplicably occur, but frequent monitoring

How to Set Up and Manage Key Performance Indicators

ensures issues can be quickly highlighted. The integral KPIs and audits should be accepted by the multidisciplinary team as a method to provide a translucent approach to quality with the aim to analyze trends and highlight issues quickly so corrective and preventative actions can be put into place. Staff should be supported and trained constructively if a decrease in performance is seen, and if red-alerts do occur, a systematic approach without a blame culture should be systematically followed. IVF is multifactorial, and despite thorough troubleshooting and investigation, a cause may never be found. However, having a robust quality management process can only be an advantage and will help both prevent and identify problems with the ultimate aim of a highly consistent program.

Suggested Standard Operating Protocol (SOP) Clinic’s approach to total quality improvement program and key performance indices “Alerts” management process Quality control protocol

References 1. D. Mortimer and S. Mortimer. Quality and Risk Management in the IVF Laboratory. Cambridge: Cambridge University Press 2005. 2. J. E. Mayer, E. L. Jones, D. Dowling-Lacey, F. Nehchiri, S. J. Muasher, W. E. Gibbons and S. C. Oehninger. Total quality improvement in the IVF laboratory: Choosing indicators of quality. Reprod. Biomed. Online 2003;7(6): 695–9. 3. W. R. Boone, H. L. Higdon III and J. E. Johnson. Quality management issues in the assisted reproduction laboratory. J. Reprod. Stem Cell Biotechnol. 2010 1 (1):30–107. 4. C. Hughes. Association of clinical embryologists – guidelines on good practice in clinical embryology laboratories 2012. Hum. Fertil. (Camb.) 2012;15:174–89. 5. M. Magli, E. Van den Abbeel, K. Lundin, D. Royere, J. Van der Elat and L. Gianaroli for Committee of the Special Interest Group on Embryology. Revised guidelines for good practice in IVF laboratories. Hum. Reprod. 2008;23(6):1253–62. 6. J. I. Olofsson, M. R. Banker and L. P. Sjoblom. Quality management systems for your in vitro fertilization clinic’s laboratory: Why bother? J. Hum. Reprod. Sci. 2013;6(1):3–8. 7. L. Rienzi, F. Bariani, M. Dalla Zorza, S. Romano, C. Scarica, R. Maggiulli, A. Nanni Costa and

F. M. Ubaldi. Failure mode and effects analysis of witnessing protocols for ensuring traceability during IVF. Reprod. Biomed Online 2015;31(4):516–22. 8. M. Forte, F. Faustini, R. Maggiulli, C. Scarica, S. Romano, C. Ottolini, A. Farcomeni, A. Palagiano, A. Capalbo, F. M. Ubaldi and L. Rienzi. Electronic witness system in IVF – patients perspective. J. Assist. Reprod. Genet. 2016; in press. 9. D. R. Meldrum, R. Chectkowski, K. A. Steingold, D. de Ziegler, M. I. Cedars and M. Hamilton. Evolution of a highly successful in vitro fertilisation embryo transfer program. Fert. Steril. 1987;48:86–93. 10. R. Heitmann, M. Hill, A. James, T. Schimmel, J. Segars, J. Csokmay, J. Cohen and M. Payson. Live births achieved via IVF are increased by improvements in air quality and laboratory environment. Reprod. Biomed. Online. 2015; 31(3):364–71. 11. European Union. EU Good Manufacturing Practice. Medicinal Products for Human and Veterinary Use. Annex 1 Manufacture of Sterile Medicinal Products (corrected version), 91/356/EEC. European Union. 2008. 12. J. Hall, A. Gilligan, T. Schimmell, M. Cecchi and J. Cohen. The origin, effects and control of air pollution in laboratories used for human embryo culture. Hum. Reprod. 1998;13: 146–55. 13. P. A. Almeida and V. N. Bolton. The effect of temperature fluctuations on the cytoskeletal organisation and chromosomal constitution of the human oocyte. Zygote 1995;3(4):357–65. 14. W. H. Wang, L. Meng, R. J. Hackett, R. Odenbourg and D. L. Keefe. Limited recovery of meiotic spindles in living human oocytes after cooling-rewarming observed using polarized light microscopy. Human Reprod. 2001;16(11):2374–8. 15. J. D. Critchlow, P. L. Matson, M. C. Newman, G. Horne, S. A. Troup and B. A. Lieberman. Quality control in an in-vitro fertilization laboratory: Use of human sperm survival studies. Hum. Reprod. 1989 Jul;4(5):545–9. 16. G. Paternot, A. M. Wetzels, F. Thonon, A. Vansteenbrugge, D. Willemen, J. Devroe, S. Debrock, T. M. D’Hooghe and C. Spiessens. Intraand interobserver analysis in the morphological assessment of early stage embryos during an IVF procedure: A multicentre study. Reprod. Biol. and Endocrinol. 2011;9:1–5. 17. L. Sundvall, H. J. Ingerslev, U. B. Knudsen and K. Kirkegaard. Inter- and intra-observer variability of time-lapse annotations. Human Reprod. 2016; 28(12): 3215–21.

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How to Manage the Patient with Thin Endometrium Tse Yeun Tan and Heng Hao Tan

Background Successful embryo implantation during the window of implantation is a complex process, with 31–64 percent contributed by uterine receptivity [1]. Endometrial dating and synchrony assessment via endometrial biopsy are limited by invasiveness, and transvaginal ultrasound (TVUS) is the most commonly used modality for endometrial assessment now. Sonographic evaluation of endometrial thickness (EMT) and pattern are often used as surrogate markers of endometrial receptivity. EMT is defined as the minimal distance between the echogenic interface of myometrium and endometrium measured through the uterine central longitudinal axis and increases with estradiol levels during folliculogenesis. The endometrium pattern appears as a single thin line after menstruation, which changes to a tripleline appearance with the onset of ovulation (see Figure 17.1). Sonographic measurement of anatomical parameters (endometrial volume) and physiological parameters (uterine and endometrial blood flow) have also been evaluated, but none has emerged as a superior prognostic indicator of reproductive outcome.

Definition and Incidence of Thin Endometrium There is no consensus on the definition of thin endometrium, although a pre-ovulatory EMT of ≤ 7 mm is commonly used. A 2014 systemic review and metaanalysis on EMT and in vitro fertilization (IVF) pregnancy rates reported a prevalence of 2.4 percent when a cut-off of ≤ 7 mm measured on day of ovulation trigger was used [2]. However, the reported prevalence of thin endometrium in the literature varies with patient age and EMT cut-off used, as well as day of endometrial measurement.

Clinical Significance of EMT and Endometrial Pattern Clinical Significance of EMT A thin endometrium at the time of transfer is disconcerting for patients. It is also widely accepted among clinicians that a thin endometrium during IVF cycles is associated with reduced endometrial receptivity and negative effects on pregnancy rates. Interestingly, not all studies have confirmed this association, which may be attributed to the use of different protocols, sonographic techniques and differing patient demographics across studies. Most studies have focused on either the EMT or endometrial appearance as possible markers of uterine receptivity. The same 2014 meta-analysis concluded that the EMT has virtually absent capacity to predict pregnancy versus non-pregnancy. However, it did show that there was a lower probability of clinical pregnancy with a thinner endometrium. The probability of conceiving with EMT ≤ 7 mm was significantly lower compared to cases with EMT > 7 mm [23.3 percent versus 48.1 percent, OR 0.42 (95% CI 0.27–0.670) P = 0.0003]. The positive (PPV) and negative predictive values (NPV) for clinical pregnancy were 77 percent and 48 percent, respectively. The meta-analysis also demonstrated a trend toward lower ongoing pregnancy and live birth rates (LBR) [OR 0.38 (95% CI 0.09–1.54), P = 0.18) in patients with thin endometrium. A meta-analysis of miscarriage rates (MR) was not performed, but other studies have reported higher MR in patients with thin endometrium [3]. In this meta-analysis, the cut-off value defining thin endometrium was taken as EMT as ≤ 7 mm due to this threshold value being used in most of the included studies. However, the authors cautioned against using one specific EMT cut-off value to discriminate between thin and normal EMT as the

How to Manage the Patient with Thin Endometrium

Figure 17.1 Ultrasound image of a triple layer endometrium

EMT Triple line

minimum EMT required for implantation remains undefined and pregnancies have still been reported at very low measurements. In addition, analyses for other EMT cut-offs (6–12 mm) also showed that the probability of clinical pregnancy was significantly improved with increased EMT up to a cut-off value of ≤ 10 mm [2]. It is evident from these results that although EMT may be a factor for the probability of conceiving after IVF, it has low predictive capacity for the occurrence of pregnancy. Due to the limited ability of EMT to act as a discriminatory tool for pregnancy prediction, we recommend that the decision for IVF cycle cancellation, freezing all embryos or withholding of IVF treatment should not be based solely on the basis of a thin endometrium alone.

Clinical Significance of Endometrial Pattern A triple-line appearance of the endometrium is commonly viewed as being reflective of endometrial receptivity as compared to a homogenous, hyperechoic endometrial pattern without a central echogenic line. A review analyzing the correlation between endometrial pattern and assisted reproduction technology (ART) pregnancy rates found that a multilayered pattern (triple line), which was more frequently associated with conception, exhibited high sensitivity (95 percent) and NPV (85.7 percent) but low specificity (13.7 percent) and PPV (33.1 percent) in the prediction of

clinical pregnancy post embryo transfer (ET) [1]. Specificity and PPV for conception remained low even when endometrial pattern was used in conjunction with other sonographic parameters (EMT or uterine blood flow resistance) as prediction criteria. These results suggest that the main utility of endometrial pattern classification lies in the high NPV (85.7 percent) of a non-multilayer (i.e., non-triple-line) pattern, rather than the presence of a triple-line appearance, which may indeed be more frequently associated with conception, but still exhibited low PPV and specificity. Regardless, it is important to note that the absence of a triple-line pattern does not preclude the occurrence of pregnancy [1]. There is clear evidence that uterine receptivity is only one of the multiple contributory factors to successful implantation. To better elucidate the effects of uterine receptivity, studies of oocyte donation and preimplantation genetic screening (PGS) cycles help to reduce embryo variability and minimize the impact of the embryo on implantation rates. When outcomes of oocyte donation studies were evaluated, it was found that endometrial patterns exhibited higher PPV (47.8 percent) and specificity (42.8 percent) for conception as compared to autologous IVF cycles [1]. There was also no significant difference in clinical pregnancy rates (CPR) for the group with lower EMT measurements ≤ 6 mm compared to those with higher EMT > 6 mm [3]. When euploid embryos were transferred after PGS,

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the presence of a mid-late secretory endometrial pattern on day of ovulation trigger rather than EMT was also found to be associated with reduced implantation rates [4]. Contrary to the evidence presented earlier, these results suggest that when the confounding effect of the embryo was minimized, endometrial pattern may in fact be better correlated with endometrial receptivity than EMT.

Causes of Thin Endometrium Iatrogenic •

•

•

Intrauterine adhesions (IUAs) are often associated with a thin endometrium. Obstetric curettage carries the highest risk of IUA formation, but trauma to the non-pregnant uterus during gynecological surgery (e.g., myomectomy) is also a risk factor [5]. Certain medications such as clomiphene citrate have a suppressor effect on the endometrium. Long-term combined oral contraceptive pill (COCP) use can infrequently give rise to chronically thin endometrium, even after drug cessation [6], but there is no evidence that its use in IVF causes endometrial thinning. Radiation-induced uterine damage is generally considered irreversible and its effects include uterine vasculature damage, reduced volume and diminished EMT [7].

Other Causes •

•

There is a paucity of data on the association between endometrial infection and thin endometrium, although genital tuberculosis causing endometritis and atrophic endometrium has been reported. Congenital conditions such as Turner’s syndrome can also be associated with thin endometrium due to uterine hypoplasia and hypo-vascularization.

Idiopathic •

Patient factors govern intrinsic endometrial properties and the individual’s potential for endometrial growth regardless of the IVF stimulation protocol used [8].

The etiology of thin endometrium affects endometrial receptivity. In Asherman’s syndrome, functional endometrium may be deficient despite surgical

correction of endometrial cavity. Thus, inflammatory conditions that lead to endometrial destruction and fibrosis have poorer reproductive outcomes than those with thin endometrium resulting from idiopathic causes.

Managing Thin Endometrium in IVF When encountering a patient with thin EMT during IVF, management options include: • Proceeding with ET regardless; • Institution of adjuvant treatment to increase EMT; • Embryo freezing and postponing fresh embryo transfer (cycle segmentation); and • Cycle cancellation as a last resort. As IVF clinicians, we know that successful implantation does not depend on endometrial receptivity alone. Other prognostic factors such as maternal age, oocyte and embryo quality, as well as serum progesterone on the day of ovulation trigger, can influence implantation rates. Based on the current evidence, we do not recommend making clinical decisions based on thin endometrium alone. Instead, we advocate personalizing management options after taking into consideration other prognostic factors, as well as the center’s individual experience with embryo freezing in such instances. If the decision is to proceed with fresh ET despite thin EMT, patients should be counseled about the lower probability of pregnancy but reinforce that the chance of pregnancy is not entirely negated. A survey of medical professionals found that 30 percent would postpone a fresh ET when EMT was ≤ 6 mm [3]. For clinicians considering this option, it is noteworthy that EMT is known to be significantly lower in natural cycles as compared to during controlled ovarian hyperstimulation (COH), and therefore the clinical implication is that an increased EMT is less likely to be achieved at subsequent frozen embryo transfer (FET) [1]. However, despite the higher probability of thicker endometria arising from COH cycles, the adverse effects from supra physiological hormonal levels are known to reduce uterine receptivity and impair reproductive outcomes. This is demonstrated by the higher implantation rates seen during hormone replacement treatment (HRT) FET compared to COH even at low EMT measurements of 4–5 mm (27.2 percent versus 10.6 percent, p = 0.0325) [9]. Thus, it appears the negative

How to Manage the Patient with Thin Endometrium

impact of stimulation may outweigh the benefits that a thicker endometrium may confer. With regards to the FET regime of choice for patients with thin endometrium, there appears to be no FET protocol that can be recommended over others. Many clinicians would utilize HRT rather than relying on the natural cycle of patients who have a history of thin endometrium during their fresh IVF cycle. However, a systemic review and meta-analysis to evaluate the most effective method of endometrial preparation prior to FET found no significant differences in EMT or pregnancy rates between HRT and natural FET cycles [10]. For patients with a thin endometrium during natural or HRT-FET, stimulated FET has been attempted to promote endometrial growth. However, this may not be beneficial as a study comparing HRT and stimulated FET cycles found no difference in EMT and reproductive outcomes [11]. If cycle segmentation is the decided course of action, we also recommend a review for correctable risk factors before embarking upon FET. A cavity assessment should also be performed if not done previously as IUAs have been diagnosed in up to 16 percent of women undergoing hysteroscopy before their first IVF cycles [12]. Hysterosalphingography (HSG) and two-dimensional (2-D) TVUS are the two commonly used modalities for uterine abnormality detection. However, 22 percent of patients embarking upon IVF were found to have uterine anomalies on hysteroscopy that were previously not detected on HSG or 2-D TVUS [12]. Despite the diagnostic accuracy of hysteroscopy, it still carries more risks than HSG or TVUS and as a result, newer but less invasive methods such as 3-D sonohysterography have been used increasingly for uterine cavity assessment. Regardless, hysteroscopy still represents the gold standard for IUA diagnosis, which also allows for concurrent therapeutic adhesiolysis [5]. To date, there has been much research generated on treatment options to increase EMT or improve reproductive outcomes in patients with thin endometrium. These are briefly discussed in what follows, but there is currently a paucity of evidence from good quality studies to validate the efficacy of these treatment options, or to establish the superiority of any one therapy [7].

Estrogen Parenteral, vagina and transdermal routes have benefits of avoiding the first liver pass effect and have been used in conjunction with oral estrogen to build up

thin endometrium. However, there is no clearly superior route of estrogen administration in patients with thin endometrium. Strategies utilizing exogenous estrogen to increase EMT include high-dose oral estrogen or extended treatment duration during FET, as well as estrogen supplementation before or during COH cycles [7].

Human Chorionic Gonadotropin (HCG) Addition of daily HCG (150 IU) from day 8/9 of HRT-FET cycles has reportedly improved EMT, but there may be evidence of detrimental effect beyond a drug threshold [7].

GnRH agonist A randomized controlled trial (RCT) has reported higher EMT, implantation and pregnancy rates from the use of luteal phase support with GnRH agonist (Triptorelin 0.1 mg) given on day of oocyte retrieval, ET day and three days later, as compared to conventional luteal support [7], although more studies of this nature are required to confirm this finding.

Therapies to Improve Endometrial Perfusion Data from HRT-FET and intrauterine insemination (IUI) cycles show that aspirin does not increase EMT, although higher implantation and pregnancy rates were reported. This suggests that the improved reproductive outcomes were not related to increase in EMT, but rather to other end mechanisms such as improved uterine flow. However, recent studies on the use of aspirin for thin endometrium have not demonstrated improved uterine blood flow parameters [7]. Non-randomized studies on the use of vaginal Sildenafil (25 mg QDS) during COH or FET cycles have shown promising results in terms of improved uterine artery blood flow, increased EMT and pregnancy rates, but two RCTs have reported conflicting results of its efficacy [7]. Agents like vitamin E and L-Arginine have been used to improve uterine artery blood flow and endometrial growth. Pentoxifylline (800 mg) in combination with vitamin E (1,000 IU) taken for periods of six to eight months may be beneficial to patients with refractory endometrium resistant to conventional treatment arising from premature ovarian failure or radiotherapy [7].

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Granulocyte Colony-Stimulating Factor (GcSF) Non-randomized studies of intrauterine granulocyte colony-stimulating factor (300 mcg) instillation during COH and FET cycles have demonstrated improvement in EMT, implantation and pregnancy rates but it appears to be non-beneficial in unselected IVF populations with normal EMT [7].

Intrauterine Platelet-Rich Plasma (PRP) Last, the benefit of autologous intrauterine PRP for promoting endometrial growth has been documented in a recent case report [7].

Conclusion Given the absence of high-quality evidence supporting the efficacy of current therapeutic options for patients with thin endometrium, there is an urgent need for more mechanistic and therapeutic research to be undertaken in areas of regenerative medicine and endometrial receptivity [7].

Suggested Standard Operating Protocol (SOP) 1)

Clinical regime for the management of women with thin endometrium

References

4. J. A. Gingold, J. A. Lee, J. Rodriguez-Purata, M. C. Whitehouse, B. Sandler, L. Grunfeld, T. Mukherjee and A. B. Copperman. Endometrial pattern, but not endometrial thickness, affects implantation rates in euploid embryo transfers. Fertility and Sterility 2015;104(3): 620–8. 5. P. H. Kodaman and A. Arici. Intra-uterine adhesions and fertility outcome: How to optimize success? Curr. Opin. Obstet. Gynecol. 2007 Jun;19(3):207–14. 6. R. F. Casper. “It’s time to pay attention to the endometrium.” Fertility and Sterility 2011;96(3): 519–21. 7. J. A. Garcia-Velasco, B. Acevedo, C. Alvarez, M. Alvarez, J. Bellver, J. Fontes, J. Landeras, D. Manau, F. Martinez, E. Muñoz and A. Robles. Strategies to manage refractory endometrium: State of the art in 2016. Reprod. Biomed. Online 2016;32 (5):474–89. 8. M. Scioscia, G. Lamanna, F. Lorusso, G. Serrati, L. E. Selvaggi and R. Depalo. Characterization of endometrial growth in proliferative and early luteal phase in IVF cycles. Reprod. Biomed. Online 2009;18: 73–8. 9. J. H. Check, R. Cohen, C. Wilson, D. Corley and C. Dietterich. Very thin endometria in the late proliferative phase is more associated with poor pregnancy rates following controlled ovarian hyperstimulation than graduated estradiol regimens used for frozen embryo transfer. Fertility and Sterility 2014;101(2):e21.

1. S. Friedler, J. G. Schenker, A. Herman and A. Lewin. The role of ultrasonography in the evaluation of endometrial receptivity following assisted reproductive treatments: a critical review. Human Reproduction Update 1996;2(4):323–35.

10. E. R. Groenewoud, A. E. Cantineau, B. J. Kollen, N. S. Macklon and B. J. Cohlen. What is the optimal means of preparing the endometrium in frozen–thawed embryo transfer cycles? A systematic review and meta-analysis. Human Reproduction Update 2013;dmt030.

2. A. Kasius, J. G. Smit, H. L. Torrance, M. J. Eijkemans, B. W. Mol, B. C. Opmeer and F. J. Broekmans. Endometrial thickness and pregnancy rates after IVF: A systematic review and meta-analysis. Human Reproduction Update 2014;20(4):530–41.

11. K. P. Wright, J. Guibert, S. Weitzen, C. Davy, P. Fauque and F. Olivennes. Artificial versus stimulated cycles for endometrial preparation prior to frozen–thawed embryo transfer. Reprod. Biomed. Online 2006;13(3): 321–5.

3. L. Dain, D. Bider, J. Levron, V. Zinchenko, S. Westler and M. Dirnfeld. Thin endometrium in donor oocyte recipients: Enigma or obstacle for implantation? Fertility and Sterility 2013;100(5):1289–95.

12. D. Galliano, J. Bellver, C. Díaz-García, C. Simón and A. Pellicer. ART and uterine pathology: how relevant is the maternal side for implantation? Human Reproduction Update 2014;dmu047.

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How to Manage the Patient with Fluid in the Endometrium Prior to Embryo Transfer Christian M. Becker and Monica Mittal

Epidemiology The incidence of intrauterine fluid being present at the time of embryo transfer (ET) following ovarian stimulation in a fresh treatment cycle has been estimated to range between 3 percent and 8 percent [4, 5]. It is reported to be highest in women with tubal factor infertility [4, 5], and greater still in the presence of a hydrosalpinx [5]. The overall clinical pregnancy rate has been shown to be significantly less in women with a certain amount of uterine fluid present at the time of ET [4, 5].

Etiology A number of etiological factors have been postulated (Table 18.1).

Clinical Implication The main determinant of the impact of endometrial cavity fluid on assisted reproduction technology (ART) cycle outcomes is the time of development of the fluid. Fluid detected before or at the initiation of

Table 18.1 Factors implicated in the accumulation of fluid within the uterine cavity during assisted reproductive treatment cycles [4, 6]

Factors Tubal infertility with or without the presence of a hydrosalpinx Endometriosis pelvic inflammatory disease polycystic ovarian syndrome Subclinical uterine infections Uterine pathology, e.g. endometrial polyps, submucosal fibroids Cervical canal obstruction physiological fluid of the genital tract Unexplained subfertility Controlled ovarian stimulation

ovarian stimulation has been shown to have a negative impact in contrast to transient fluid accumulation seen during ovarian stimulation and at the time of oocyte retrieval, but that then disappears by the time of ET. The depth of fluid accumulation at the time of ET is also thought to be a significant determining factor of cycle outcomes, with measurements in the order of 3 mm or more in the anterior posterior diameter in the sagittal plane during transvaginal ultrasound scans having a greater detrimental impact on implantation rates [4, 6] compared with lower volumes. Furthermore, the fluid is more likely to diminish if less than 3 mm in diameter [4]. However, the volume of fluid accumulation is not considered significant if transient [7].

Pathophysiology The exact mechanism of action of the endometrial cavity fluid on implantation is unknown. Postulated theories include: embryotoxicity of the accumulated fluid; disruption of embryo implantation; mechanical loss of embryos through the presence of excessive fluid; or, altered gene expression [8] (integrin αvβ3 [9]) and disruption of the cytokine and glycoprotein cascades [10] affecting endometrial receptivity [8]. The development and role of physiologically derived endometrial cavity fluid is less clear. A small study has postulated the potential benefit of this fluid, facilitating implantation through growth factors contained within the fluid [6]. Furthermore, transient intrauterine fluid accumulation that disappears by the ET day, not thought to be secondary to a hydrosalpinx, polycystic ovarian syndrome or an identifiable pelvic pathology, is not thought to negatively impact the outcome of the cycle [7]. Uterine fluid associated with the presence of a hydrosalpinx is thought to be embryotoxic in nature with the inflammatory media interfering with the development of the embryo and impacting the receptivity of the endometrium prior to and during

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(a)

(b)

(c)

Figure 18.1 a. Transvaginal ultrasound assessment of the endometrial thickness in the longitudinal plane, measured between the myometrial and endometrial interface b. Transvaginal ultrasound assessment of the endometrial cavity fluid in the sagittal plan (anteriorposterior diameter) c. Transvaginal 3-D ultrasound assessment of the endometrial cavity fluid

implantation [5, 6, 10]. This theory is supported by higher implantation rates being documented following fluid aspiration or tubal occlusion in the presence of a hydrosalpinx [5, 6, 8]. Animal models and studies utilizing human embryos, however, have differed in their explanation of why the fluid is detrimental to pregnancy outcomes. Specifically, no embryotoxic molecule has been isolated to account for the assumed toxic nature of the fluid so far [9], with several studies documenting negative bacterial cultures from the hydrosalpinx fluid [11]. It has therefore been suggested that the most likely causative factor is the reduction in essential substrates required for embryo development in hydrosalpingeal fluid [9]. Murray et al. [12] demonstrated that the inhibitory nature of the hydrosalpingeal fluid on embryo development can be counteracted to a degree by the addition of energy factors such as lactate. Other authors have described a potential mechanical cause, namely the reversal of the direction of intrauterine peristalsis (opposition of the cervical to fundal peristalsis) in the presence of a hydrosalpinx, impairing implantation [10]. Animal studies have also shown the negative effect of fluid on embryo development, through growth retardation, an increased miscarriage rate and pregnancy loss, when normal saline is administered into the uterine cavity of mice [8]. The extent of these effects appears to be directly correlated to the volume of fluid applied, mirroring the clinical observation [4, 6], with pregnancy rates in human subjects being even lower when the hydrosalpinx can be visualized on ultrasound scan [13]. This theory is further supported by animal and human studies looking at the incubation of embryos in varied concentrations of

hydrosalpinx fluid, with embryo development being negatively correlated with increasing concentrations of hydrosalpinx fluid [14, 15]. Particularly in human studies, blastocyst stage development was shown to be significantly inhibited in the presence of 100 percent hydrosalpinx fluid compared with 50 percent hydrosalpinx fluid. This is further compounded by no statistical difference being documented in the blastocyst stage development in 50 percent hydrosalpinx fluid and controls [15]. Authors have further speculated that the presence of intraluminal fluid may dilute the factors important in implantation [8]. The presence of normal saline in mice models resulted in a change in gene expression of uterine receptivity genes, in particular the expression of integrin αvß3, which has been shown to be a pivotal marker of the window of implantation [16]. This marker has been shown to be downregulated in the presence of hydrosalpinges [17]. This is not supported by studies conducted on murine embryo models that were replaced into the horn of the mice with human hydrosalpinx fluid, whereby no difference in implantation was demonstrated [14]. In contrast, other studies have suggested that the fluid from hydrosalpinges contains growth-promoting factors with the increased production of trophouteronectin and progesterone being demonstrated [9]. Experiments looking at hydrosalpinx fluid and granulosa cells collected at the time of oocyte retrieval from human patients undergoing in vitro fertilization (IVF) treatment have shown increased steriodogenesis with normal embryogenesis from oocytes containing three pronuclei [11]. Another study looked at the effect of hydrosalpinx fluid on human cytotrophoblast viability. It showed increased concentrations

Fluid in the Endometrium Prior to Embryo Transfer

of tropho-uteronectin and β-human chorionic gonadotrophin, molecules critical in the process of implantation [18]. The authors therefore suggested alternative hypotheses for the reduced pregnancy rates demonstrated in the presence of hydrosalpinx fluid, such as mechanical interference [11, 18]. Furthermore, it has been suggested that fluid within the endometrial cavity can negatively impact the endometrial thickness, with a thinner endometrium being documented in the presence of uterine fluid accumulation present on the day of ET compared to patients without fluid accumulation being documented [5].

Management The development and significance of endometrial cavity fluid prior to ET is debatable, with little consensus on its appropriate management. However, the timing and origin of development of the fluid is considered crucial in counseling women appropriately on its potential impact on implantation and cycle outcome. A detailed pelvic anatomy scan prior to the commencement of ART enables parameters known to affect the outcome of the cycle to be addressed at the planning stage. Surgical management of hydrosalpinges through tubal occlusion or removal prior to the commencement of IVF has been widely documented to significantly improve pregnancy and live birth rates through ART [19]. A number of different methodologies have been described for the management of a hydrosalpinx: transvaginal aspiration; hysteroscopic proximal intratubal occlusion (Essure®); salpingostomy; laparoscopic tubal ligation; or, salpingectomy [20]. A Cochrane review compared the different techniques employed over the years for the management of a hydrosalpinx [21]. The primary outcome measure was live birth rate. Secondary outcome measures included ongoing pregnancy rate, ectopic pregnancy rate and the surgical complication rate. Laparoscopic salpingectomy compared with no surgical intervention for the management of a hydrosalpinx has been shown to significantly increase the odds of an ongoing pregnancy rate (three trials; 395 women; OR 2.20, 95% CI 1.26 to 3.82). In contrast, laparoscopic occlusion compared with no intervention did not increase the odds of an ongoing pregnancy (one trial; 65 women; OR 7.21, 95% CI 0.87 to 59.57);

however, the trial was significantly underpowered. The clinical pregnancy rate was shown to be significantly increased (two trials; 209 women; OR 4.66, 95% CI 2.17 to 10.01). Furthermore, laparoscopic salpingectomy versus laparoscopic occlusion did not show one to be superior over the other for ongoing pregnancy rate (one trial; 100 women; OR 1.65, 95% CI 0.74 to 3.71). Tubal aspiration was not shown to significantly improve the clinical pregnancy rate (one trial; 64 women; OR 1.97, 95% CI 0.62 to 6.29). In addition, no difference was reported in the odds ratio of an ectopic pregnancy rate or complication rate with the different interventions, but the studies remained underpowered [21]. A randomized controlled trial comparing the effectiveness of hysteroscopic tubal occlusion using Essure® with laparoscopic salpingectomy, in the management of hydrosalpinges prior to ART, has shown the superiority of the latter in terms of ongoing pregnancy rates beyond 10 weeks’ gestation (absolute difference 24.8 percent per embryo transferred; p = 0.004) [20]. The study further showed statistical significance in favor of a salpingectomy per embryo transferred for secondary outcome measures including implantation rate (absolute difference 23.7 percent) and live birth rate (absolute difference 20.2 percent). No ectopic pregnancy was documented during this study. The miscarriage rate was higher for patients undergoing Essure® (absolute difference 1.2 percent per embryo transferred), but this did not reach statistical significance. In addition, the antimullerian hormone level (AMH) was shown to increase following a salpingectomy but decrease following insertion of an Essure®, to a significant degree. A minimal effect was seen on the other ovarian reserve parameters. The overall significance of this finding was considered debatable due to the small sample sizes and large confidence intervals. Reported complications included wound infection from laparoscopic salpingectomy and pelvic inflammatory disease from intratubal occlusion. Furthermore, the tip of the device was found to be protruding into the uterine cavity at a post-hysteroscopy check-in in some cases [20]. However, if a laparoscopic salpingectomy is not feasible as it carries with it anesthetic and surgical risks, then any surgical intervention capable of disrupting the communication of the hydrosalpinges with the uterine cavity should theoretically improve

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Endometrial Fluid Management Flow Chart

Selection of patients for IVF

Detailed Pelvic Assessment i.e. Imaging Modalities: USS, MRI, Hysteroscopy, HyCoSy, HSG

Reversible Pelvic Pathology i.e. Endometrial Polyp, Submucosal Fibroid, Hydrosalpinx

Yes

No

Surgical Intervention i.e. Polypectomy, Transcervical Resection of Fibroid Salpingectomy

Yes

No

Arrange surgical intervention

IVF cycle commencement

Continued Endometrial Surveillance through transvaginal ultrasound

Yes

Tubal Factor Infertility

Cryopreserve all embryos

Endometrial Cavity Fluid

No

Non-Tubal Factor Infertility

Fluid aspiration

Embryo Transfer

Figure 18.2 Systematic approach to the management of endometrial cavity fluid detected prior to the ET

Fluid in the Endometrium Prior to Embryo Transfer

treatment outcomes [20]. Note that the use of Essure® for the management of hydrosalpinges in ART is currently off label [20]. If used for this purpose, deeper placement of the implant can be considered to avoid protuberance into the endometrial cavity [22]. Two randomized clinical studies, the inSIGHT and TROPHY trials, looked at the benefit of hysteroscopic morphological assessment of the uterine cavity prior to IVF treatment in individuals undergoing their first cycle [23] or those with recurrent implantation failure [24], respectively. The studies concluded that the prevalence of uterine cavity abnormalities within the population is approximately 10 percent, with hysteroscopic procedures not being found to increase pregnancy or live birth rates [23, 24]. In view of this, it has been summarized that pathology not identified by routine transvaginal ultrasonography may be of no clinical significance [25]. Furthermore, no data exist regarding the potential benefits of performing hysteroscopic exploration of the uterine cavity in the presence of uterine fluid on its own. Consideration should be given to assessment of the endometrial cavity during controlled ovarian stimulation through transvaginal ultrasonography to rule out pathology, enabling continuous review of any changes in the endometrium and uterine cavity, and a dynamic process to be individualized. Embryo transfer is considered to be able to proceed without consequence if the presence of the uterine cavity fluid is transient and disappears by the ET date [26]. In contrast, the ET procedure should be abandoned in the presence of endometrial cavity fluid development in cases of tubal factor infertility, secondary to the negative impact of the fluid on pregnancy outcomes. In these cases, cycle cancellation with embryo cryopreservation is considered more appropriate [26]. In some instances, successful cycle outcomes have been documented following aspiration of the fluid immediately prior to ET [27], without negatively impacting implantation [28]. This fluid can, however, re-accumulate [5], if implantation is performed too early. Follow-up diagnostic hysteroscopy, laparoscopy and/or tubal assessment may be warranted to further evaluate tubal functionality and the endometrium should the fluid within the uterine cavity be persistent.

Furthermore, progesterone is widely utilized in ART cycles, providing luteal phase support. It is commonly started soon after oocyte retrieval. Progesterone has been shown to regulate the fluid volume within the uterine cavity by increasing the resorption of sodium and subsequently the endometrial fluid [29]. This may explain the cases in which endometrial cavity fluid is seen at the time of oocyte retrieval but disappears by the ET date.

Summary Intrauterine cavity fluid present at the time of ET can present a diagnostic and management dilemma in the care of patients undergoing ART cycles. The pathophysiology of the fluid is largely unknown with a number of hypotheses being speculated. Careful counseling of these patients is therefore needed, taking account of their past history and previous cycle outcomes. It is largely accepted that the presence of large volumes of endometrial cavity fluid at the time of ET should lead to the cryopreservation of the embryos, especially whereby the success rates associated with frozen embryo transfers is no longer statistically significantly different from a fresh ET. Larger randomized controlled studies are needed before definitive conclusions can be drawn. Figure 18.2 aims to provide a systematic approach to the management of endometrial cavity fluid detected prior to the ET procedure.

Suggested Standard Operating Protocol (SOP) SOP for Management of Women with Fluid in the Endometrium

References 1. W. B. Schoolcraft, E. S. Surrey and D. K. Gardner. Embryo transfer: Techniques and variables affecting success. Fertility and Sterility 2001;76(5):863–70. 2. T. C. Papageorgiou, R. M. Hearn-Stokes, M. P. Leondires, B. T. Miller, P. Chakraborty, D. Cruess and J. Segars. Training of providers in embryo transfer: What is the minimum number of transfers required for proficiency? Human Reproduction 2001;16(7):1415–19. 3. A M. Abou-Setta, H. G. Al-Inany, R. T. Mansour, G. I. Serour and M. A. Aboulghar. Soft versus firm embryo transfer catheters for assisted reproduction: A

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systematic review and meta-analysis. Human Reproduction 2005;20(11):3114–21. 4. R. H. He, H. J. Gao, Y. Q. Li and X. M. Zhu. The associated factors to endometrial cavity fluid and the relevant impact on the IVF-ET outcome. Reproductive Biology and Endocrinology 2010;8(46):1–6. 5. L. W. Chien, H. K. Au, J. Xiao and C. R. Tzeng. Fluid accumulation within the uterine cavity reduces pregnancy rates in women undergoing IVF. Human Reproduction 2002;17(2):351–6. 6. R. K. K. Lee, S. L. Yu, Y. F. Chih, Y. C. Tsai, M. H. Lin, Y. M. Hwu, W. Y. Huang, and J. T. Su. Effect of endometrial cavity fluid on clinical pregnancy rate in tubal embryo transfer (TET). Journal of Assisted Reproduction and Genetics 2006;23(5):229–34. 7. M. Polat, F. K. Boynukalin, I. Yarali, B. D. Erdoğan, G. Bozdağ and H. Yaralı. Transient intrauterine fluid accumulation not due to hydrosalpinx or any identifiable pelvic pathology is not detrimental to IVF outcome. Archives of Gynecology and Obstetrics 2014;290(3):569–73. 8. S. Lu, H. Peng, H. Zhang, L. Zhang, Q. Cao, R. Li, Y. Zhang, L. Yan, E. Duan and J. Qiao. Excessive intrauterine fluid causes aberrant implantation and pregnancy outcome in mice. PLoS One 2013;8(10):1–8. 9. A. Strandell. The influence of hydrosalpinx on IVF and embryo transfer: A review. Human Reproduction Update 2000;6(4):387–95. 10. A. Strandell and A. Lindhard. Why does hydrosalpinx reduce fertility? The importance of hydrosalpinx fluid. Human Reproduction 2002;17(5):1141–5. 11. I. Granot, N. Dekel, I. Segal, S. Fieldust, Z. Shoham and A. Barash. Is hydrosalpinx fluid cytotoxic? Human Reproduction 1998;13(6):1620–4. 12. C. A. Murray, H. J. Clarke, T. Tulandi and S. L. Tan. Inhibitory effect of human hydrosalpingeal fluid on mouse preimplantation embryonic development is significantly reduced by the addition of lactate. Human Reproduction 1997;12(11):2504–7. 13. W. de Wit, C. J. Gowrising, D. J. Kuik, J. W. Lens and R. Schats. Only hydrosalpinges visible on ultrasound are associated with reduced implantation and pregnancy rates after in-vitro fertilization. Human Reproduction 1998;13(6):1696–1701. 14. V. J. Rawe, J. Liu, S. Shaffer, M. G. Compton, J. E. Garcia and E. Katz. Effect of human hydrosalpinx fluid on murine embryo development and implantation. Fertility and Sterility 1997;68(4):668–70. 15. A. Strandell, A. Sjogren, U. Bentin-Ley, J. Thorburn, L. Hamberger and M. Brannstrom. Hydrosalpinx fluid does not adversely affect the normal development of human embryos and implantation in vitro. Human Reproduction 1998; 13(10):2921–5.

16. B. A. Lessey, A. J. Castelbaum, C. A. Buck, Y. Lei, C. W. Yowell and J. Sun. Further characterization of endometrial integrins during the menstrual cycle and in pregnancy. Fertility and Sterility 1994;62(3):497–506. 17. W. R. Meyer, A. J. Castelbaum, S. Somkuti, A. W. Sagoskin, M. Doyle, J. E. Harris and B. A. Lessey. Hydrosalpinges adversely affect markers of endometrial receptivity. Human Reproduction 1997;12 (7):1393–8. 18. S. W. Sawin, J. R. Loret de Mola, F. MonzonBordonaba, C. L. Wang and R. F. Feinberg. Hydrosalpinx fluid enhances human trophoblast viability and function in vitro: Implications for embryonic implantation in assisted reproduction. Fertility and Sterility 1997;68:65–71. 19. N. Johnson, S. van Voorst, M. C. Sowter, A. Strandell and B. W. J. Mol. Surgical treatment for tubal disease in women due to undergo in vitro fertilisation. Cochrane Database of Systematic Reviews 2010;1–69. 20. K. Dreyer, M. C. I. Lier, M. H. Emanuel, J. W. R., Twisk, B. W. J. Mol, R. Schats, P. G. A. Hompes and V. Mijatovic. Hysteroscopic proximal tubal occlusion versus laparoscopic salpingectomy as a treatment for hydrosalpinges prior to IVF or ICSI: An RCT. Human Reproduction 2016;0(0):1–12. 21. N. Johnson, S. van Voorst, M. C. Sowter, A. Strandell and B. W. J. Mol. Surgical treatment for tubal disease in women due to undergo in vitro fertilisation (Review). Cochrane Database of Systematic Reviews 2010;Issue 1:1–42. 22. E. S. Sills, D. J. Walsh, C. A. Jones and S. H. Wood. Endometrial fluid associated with Essure implants placed before in vitro fertilization: Considerations for patient counseling and surgical management. Clinical and Experimental Reproductive Medicine 2015;42 (3):126–9. 23. J. G. Smit, J. C. Kasius, M. J. C. Eijkemans, C. A. M. Koks, R. van Golde, A. W. Nap, G. J. Scheffer, P. A. P. Manger, A. Hoek, B. C. Schoot, A. M. van Heusden, W. K. H. Kuchenbecker, D. A. M. Perquin, K. Fleischer, E. M. Kaaijk, A. Sluijmer, J. Friederich, R. H. M. Dykgraaf, M. van Hooff, L. A. Louwe, J. Kwee, C. H. de Koning, I. C. A. H. Janssen, F. Mol, B. W. J. Mol, F. J. M. Broekmans and H. L. Torrance. Hysteroscopy before in-vitro fertilisation (inSIGHT): A multicentre, randomised controlled trial. Lancet 2016;1–8. 24. T. El-Toukhy, R. Campo, Y. Khalaf, C. Tabanelli, L. Gianaroli, S. S. Gordts, S. Gordts, G. Mestdagh, T. Mardesic, J. Voboril, G. L. Marchino, C. Benedetto, T. Al-Shawaf, L. Sabatini, P. T. Seed, M. Gergolet, G. Grimbizis, H. Harb and A. Coomarasamy. Hysteroscopy in recurrent in-vitro fertilisation failure (TROPHY): A multicentre, randomised controlled trial. Lancet 2016;1–8.

Fluid in the Endometrium Prior to Embryo Transfer

25. A. Pellicer and D. Galliano. Comment. Hysteroscopy before IVF: Can it improve outcomes? Lancet 2016. 26. M. A. Akman, H. F. Erden and M. Bahceci. Endometrial fluid visualized through ultrasonography during ovarian stimulation in IVF cycles impairs the outcome in tubal factor, but not PCOS, patients. Human Reproduction 2005;20 (4):906–9. 27. A. N. Griffiths, S. R. Watermeyer and L. D. Klentzeris. SHORT COMMUNICATION. Fluid within the endometrial cavity in an IVF cycle – a novel approach

to its management. Journal of Assisted Reproduction and Genetics 2002;19(6):298–301. 28. M. H. van der Gaast, K. Beier-Hellwig, B. C. Fauser, H. M. Beier and N. S. Macklon. Endometrial secretion aspiration prior to embryo transfer does not reduce implantation rates. Reproductive BioMedicine Online 2003;7(1):105–9. 29. K. Bhusane, S. Bhutada, U. Chaudhari, L. Savardekar, R. Katkam and G. Sachdeva. Secrets of endometrial receptivity: Some are hidden in uterine secretome. American Journal of Reproductive Immunology 2016;75 (3):226–36.

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Practical Tips on Personalizing Luteal Phase Support Srividya Seshadri and Sesh Kamal Sunkara

Background The luteal phase in assisted reproductive technology (ART) cycles is not sufficient, although the underlying mechanism is unclear [1]. In normal cycles, the luteinizing hormone (LH) surge results in ovulation and the LH induces the granulosa cells in the corpus luteum to produce progesterone, which in turn induces the vascularization and the thickening of the endometrium to facilitate implantation. After implantation, the placenta produces human chorionic gonadotropin (hCG), which stimulates the corpus luteum to produce estradiol and progesterone, which maintains early pregnancy. In a natural pregnancy, LH is produced continuously after the ovulation surge and only decreases when hCG from the growing trophoblast takes over. Conversely, the ovarian stimulation used in ART cycles results in very high steroid levels; this inhibits the pituitary secretion of LH which is thought to shorten the luteal phase, a situation known as premature luteolysis [2, 3]. The other mechanisms thought to result in an inadequate luteal phase function in ART cycles include a supraphysiologic estradiol level, decreased LH level, inhibition of the corpus luteum and asynchronization of estradiol and progesterone [4]. Due to these reasons, adequate luteal support to enhance pregnancy rates is required. The luteal support reported in the medical literature includes different types of progesterone, hCG, estrogen and gonadotrophin-releasing hormone (GnRH) agonists. The addition of estradiol to progesterone luteal support is currently debated and the final situation in luteal phase support needs further studies. The use of GnRH agonist in luteal support has been recommended in more recent studies [2]. Table 19.1 outlines the different hormones used as luteal support and the supporting grade of evidence.

Progesterone Preparations and Routes of Administration and Duration of Luteal Support Progesterone preparations can be divided into two groups: natural progesterone and synthetic preparations. Synthetic derivatives or progestins are 1) 17-hydroxyprogesterone derivatives and 2) 19nortestosterone derivatives. Various formulations of progesterone are now available, including oral, vaginal and intramuscular (IM) progesterone. Levine and Watson [5] concluded that the vaginal administration of progesterone results in a greater bioavailability with less relative variability than oral progesterone. There is no consensus on the standard dose of progesterone in luteal phase support. Studies have been conducted using IM injections (12.5–100 mg/day), various vaginal preparations such as creams, pessaries, sustained release gel and vaginal rings, vaginal applications of oral formulations and oral preparations, including micronized progesterone (600–1200 mg/day) and dydrogesterone (20–30 mg/ day). Table 19.2 summarizes the evidence comparing the different routes of progesterone administration and the effect on clinical outcomes. The ideal luteal support enhances live birth rates and reduces risk of ovarian hyperstimulation (OHSS). Progesterone during the luteal phase is associated with higher rates of live birth or ongoing pregnancy than placebo [2]. The duration for the luteal support with progesterone is unclear for ART cycles. Four studies showed a clear benefit in a statistically significant clinical pregnancy rate (CPR) when the progesterone is taken until 12 weeks’ gestation. However, three studies have shown no clear benefit in CPR when the progesterone support is stopped at the time of the positive pregnancy test versus placebo. The start time of progesterone post oocyte retrieval

With progesterone

GnRH agonists

IntranasalIntramuscularSubcutaneous

OralTransdermalVaginal

With progesterone

Estrogen

LBR statistically significant in progesterone and GNRH agonist group

No difference in LBR in progesterone and oral estrogen supplementation compared to progesterone administration only

No difference in LBR

1) LBR higher in progesterone group 2) No difference in LBR 3) No difference in LBR 4) LBR statistically significant in progesterone and GnRH agonist group 5) Statistical difference in LBR in the vaginal/rectal group 6) No difference in LBR

OralVaginalRectalIntramuscular (IM)Subcutaneous

IntramuscularSubcutaneous

1) Progesterone versus placebo 2) Progesterone versus HCG 3) Progesterone versus progesterone and estrogen 4) Progesterone versus progesterone and GNRH agonist 5) IM versus vaginal/rectal 6) Vaginal versus rectal progesterone

Progesterone only

Outcomes

Routes of administration

HCG

Comparisons

Luteal support

Table 19.1 The different hormones used as luteal support and the supporting grade of evidence

Grade ARandomized controlled trials

Grade ARandomized controlled trials

Grade ARandomized controlled trials

Grade A: Randomized controlled trials

Grade of evidence

Srividya Seshadri and Sesh Kamal Sunkara

Table 19.2 The evidence comparing the different routes of progesterone administration and the effect on clinical outcomes

Comparison of the different routes of progesterone administration

Effect on outcome

Study

Oral versus vaginal progesterone

Similar clinical and ongoing pregnancy rates in both the groups

Friedler S, Raziel A, Schachter M, Strassburger D, Bukovsky I, Ron-El R. Luteal support with micronized progesterone following in-vitro fertilization using a down-regulation protocol with gonadotrophin-releasing hormone agonist: A comparative study between vaginal and oral administration. Hum Reprod. 1999;14:1944–8.

Oral dydrogesterone versus vaginal micronized progesterone

Similar rates of pregnancy in both groups

Chakravarty BN, Shirazee HH, Dam P, Goswami SK, Chatterjee R, Ghosh S. Oral dydrogesterone versus intravaginal micronised progesterone as luteal phase support in assisted reproductive technology (ART) cycles: Results of a randomised study. J Steroid Biochem Mol Biol. 2005;97:416–20.

Oral micronized progesterone versus IM progesterone

No significant difference in pregnancy rate between both groups

Licciardi FL, Kwiatkowski A, Noyes NL, et al. Oral versus intramuscular progesterone for in vitro fertilization: A prospective randomized study. Fertil. Steril. 1999;71: 614–18

Vaginal versus IM progesterone

No significant difference in pregnancy rate between both groups

Van der Linden M, Buckingham K, Farquhar C, Kremer JAM, Metwally M. Luteal phase support for assisted reproduction cycles. Cochrane Database of Systematic Reviews 2011, Issue 10. Art. No.: CD009154.

Vaginal pessaries (Ellios™) versus capsules (Uterogestan™)

Pregnancy rate comparable in both the groups

Germond M, Capelli P, Bruno G, Vesnaver S, Senn A, Rouge N, Biollaz J, Comparison of the efficacy and safety of two formulations of micronized progesterone (Ellios™ and Utrogestan™) used as luteal phase support after in vitro fertilization. Fertil. Steril. 2002;77:313–15.

Vaginal progesterone suppositories (Cyclogest) versus vaginal progesterone tablets (Endometrin)

No clinical outcomes reported

Yu Ng EH, Chan CCW, Tang OS, Ho PC, A randomized comparison of side effects and patient convenience between Cyclogest suppositories and Endometrin tablets used for luteal phase support in IVF treatment. Eur. J. Obstet. Gynecol. Reprod. Biol. 2007;131:182–8.

Crinone versus Utrogest capsules vaginally

Pregnancy rate comparable in both the groups

Ludwig M, Schwartz P, Babahan B, Katalinic A, Weiss JM, Felberbaum R, Al-Hasani S, Diedrich K. Luteal phase support using either Crinone 8% or Utrogest: Results of a prospective, randomized study. Eur. J. Obstet. Gynecol. Reprod. Biol. 200;103:48–52.

Subcutaneous progesterone (Lubion) versus vaginal progesterone (Endometrin)

Pregnancy rate comparable in both the groups

Lockwood G, Griesinger G, Cometti B. Subcutaneous progesterone versus vaginal progesterone gel for luteal phase support in in vitro fertilization: A noninferiority randomized controlled study. Fertil. Steril. 2014;101(1): 112–19.

Practical Problems in Assisted Reproduction

Practical Tips on Personalizing Luteal Phase Support

• Vaginal route of administration crinone 8 percent (90 mg) twice a day OR cyclogest 400 mg twice a day • Continue progesterone up to either until the pregnancy test or six to eight weeks of pregnancy (dependent on clinician experience and patient history)

is unclear; however, some studies have suggested a potential benefit in delaying vaginal progesterone start time to two days post oocyte retrieval [6].

Estrogen Preparations and Routes of Administration A combination of transdermal estrogen and progesterone was found to increase the pregnancy rates compared to a group that received progesterone only; however, findings should be regarded with caution because of the inconsistency observed between studies and the small quantity of data provided [2].

2)

a. Crinone (90 mg/day) and E2 (4 mg/day) + 1,500 IU hCG 35 hours after trigger b. 1,500 IU hCG on the day of oocyte retrieval + 1,500 IU hCG four days after oocyte retrieval c. Recombinant LH (rLH): six alternate doses of 300 IU rLH, starting on the day of OPU and repeated every other day in addition to 600 mg daily of progesterone, administered vaginally d. Estrogen 4 mg/day + 50 mg IM progesterone per day

hCG as Luteal Support There is no difference in the clinical pregnancy rates in the group that received HCG supplementation versus a placebo. The risk of OHSS increased in the HCG group compared to the placebo group [2].

GnRH Agonist

3)

The hypothesis for the role of GnRH agonist as a luteal support is that it extends LH production throughout the luteal phase, thus preventing the occurrence of premature luteolysis and consequently improving pregnancy rates [2]. There is evidence that adding GnRH agonist during the luteal phase improves the likelihood of ongoing pregnancy [7]. This could include a single-dose triptorelin of 0.1 mg given six days after oocyte retrieval or alternatively the nasal spray 100 mcgm three times a day [2]. However, more robust evidence is still required.

Protocols 1)

Agonist cycle with hCG trigger (Figure 19.1): • The day after oocyte retrieval, progesterone is commenced

Antagonist cycle with GnRH agonist trigger (Figure 19.2):

FET (natural cycle): Crinone/cyclogest daily until pregnancy test or six to eight weeks of pregnancy

4)

FET (medicated cycle) (Figure 19.3): Estrogen 4 mg/day + vaginal/rectal progesterone until 10 weeks of pregnancy

Frozen embryo transfer and donor oocyte cycles are replacement cycles, with supplementation typically lasting until a gestational age of 10 weeks [8]. Table 19.1 summarizes the different luteal support regimens [2]. Table 19.2 summarizes the evidence for the different progesterone regimens. Patients need to be counselled that progesterone during the luteal phase is associated with higher rates of live birth or ongoing pregnancy than placebo. The addition of GnRH agonist to progesterone

Agonist long regimen

Gonadotrophin stimulation

Progesterone administration: Oral /vaginal /subcutaneous

GnRH agonist

Menstruation

Mid-luteal phase

Menstruation

hCG

Oocyte Embryo retreival transfer

Pregnancy test

Figure 19.1 Luteal support in a long agonist cycle

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Antagonist regimen 1500 hCG 35 hrs after trigger + estrogen and progesterone OR Recombinant LH 300 IU on alternated days +600 mg progesterone

GnRH antagonist

OR Estrogen and Progesterone

Gonadotrophin stimulation

Lead follicle ≥ 14 mm/ Day 6 of stimulation Day 2/3 of cycle

Menstruation

Agonist trigger

Oocyte retrevial

Embryo transfer

Pregnancy test

Figure 19.2 Luteal support in an antagonist cycle with GnRH trigger

Estrogen administration GnRH agonist

Menstruation

Mid-luteal phase

Menstruation

Progesterone administration

Embryo transfer

appears to improve outcomes. The hCG may increase the risk of OHSS. Moreover hCG, with or without progesterone, is associated with higher rates of OHSS than progesterone alone. Cases of severe OHSS are reported to the regulatory authority, i.e. the Human Fertilisation and Embryology Authority (HFEA) in the United Kingdom. An adequate luteal support regimen is mandatory to maintain clinical pregnancy and live birth rates. A luteal phase deficiency of progesterone is one of the reasons for implantation failure and miscarriages [9].

Research Areas Randomized trials establishing the duration of luteal phase progesterone supplementation Randomized trials establishing the timing of progesterone supplementation post oocyte retrieval Randomized trials establishing the dose and duration of GnRH agonists as luteal support

Summary In summary, progesterone is the ideal support luteal regimen for IVF cycles. The route of administration

Figure 19.3 Luteal support in a medicated frozen embryo transfer cycle

Till 8–10 weeks pregnancy

and the duration would depend on patient choice, patient history and clinician experience.

Suggested Standard Operating Protocol (SOP) Figure 19.1 summarizes the luteal support in IVF cycles triggered with HCG. Figure 19.2 summarizes the luteal support in IVF cycles triggered with an agonist trigger. Figure 19.3 summarizes luteal support in frozen embryo transfer cycles.

Suggested Audits The OHSS rate with the different luteal phase regimens Miscarriage rate with the different luteal phase regimens Perinatal outcomes and congenital malformation rate with the different luteal phase regimens

References 1. R. G. Edwards, P. C. Steptoe and J. M. Purdy. Establishing full-term human pregnancies using cleaving embryos grown in vitro. British

Practical Tips on Personalizing Luteal Phase Support

Journal of Obstetrics and Gynaecology 1980;87(9): 737–56. 2. M. van der Linden, K. Buckingham, C. Farquhar, J. A. Kremer and M. Metwally. Luteal phase support for assisted reproduction cycles. Cochrane Database Syst. Rev. 2015;(7):CD009154. doi:10.1002/14651858. CD009154.pub3 3. H. M. Fatemi. The luteal phase after 3 decades of IVF: What do we know? Reprod. Biomed. Online 2009;19 Suppl. 4:4331. 4. R. Pabuccu and M. E. Akar. Luteal phase support in assisted reproductive technology. Curr. Opin. Obstet. Gynecol. 2005;17:277–81. 5. H. Levine and N. Watson. Comparison of the pharmacokinetics of Crinone 8% administered vaginally versus Prometrium administered orally in postmenopausal women. Fertil. Steril. 2000;73: 516–21.

6. M. T. Connell, J. M. Szatkowski, N. Terry, A. H. DeCherney, A. M. Propst and M. J. Hill. Timing luteal support in assisted reproductive technology: A systematic review. Fertil. Steril. 2015;103(4):939–46. e3. 7. W. P. Martins, R. A. Ferriani, P. A. Navarro and C. O. Nastri. (2016), GnRH agonist during luteal phase in women undergoing assisted reproductive techniques: A systematic review and meta-analysis of randomized controlled trials. Ultrasound Obstet. Gynecol. 47:144–51.doi:10.1002/uog.14874 8. C. B. Lesser. Progesterone supplementation in assisted reproductive technology: making regimens friendly. Supplement to SRM. 2011:S258. 9. L. G. Nardo and H. N. Sallam. Progesterone supplementation to prevent recurrent miscarriage and to reduce implantation failure in assisted reproduction cycles. Reprod. Biomed. Online 2006;13:47–57.

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How to Avoid Ovarian Hyperstimulation Syndrome Best Management Strategies Nikolaos Tsampras and Raj Mathur

Prediction of OHSS

of multiple follicles (> 20) and high number of oocytes retrieved (> 25) are positively related with OHSS. Steward et al., in an analysis of 256,381 cycles, demonstrated that retrieval of > 15 oocytes increases the risk of OHSS significantly, without improving live birth rate [3]. High estradiol levels are also associated with an increased risk of early OHSS, but the predictive value of estradiol levels is poor. Despite the widespread use of patients’ characteristics and stimulation parameters to trigger preventative measures, a significant proportion of patients who go on to develop OHSS are not recognized as being at increased risk of OHSS before or during treatment. A particular problem is the inability to accurately predict late OHSS, which is related to pregnancy and only poorly predictable from ovarian response parameters.

Demographics

Preventative Measures

Introduction Ovarian hyperstimulation syndrome (OHSS) is an uncommon, but potentially serious, iatrogenic complication of controlled ovarian stimulation (COS) during assisted reproductive technology (ART). Moderate-to-severe OHSS occurs in approximately 1–5 percent of cycles, although the true incidence is unknown, due to a lack of consensus in definition and difficulties in ascertainment. Advance in understanding the pathogenesis of the syndrome has led to various preventive strategies. Identification of women at high risk and appropriate intervention can reduce the incidence of OHSS without compromising the treatment outcome.

Pretreatment evaluation of the patient allows the clinician to assess the risk of OHSS and optimize the COS protocol. Evidence suggests young age, polycystic ovaries and a previous history of OHSS as risk factors for OHSS [1].

Ovarian Reserve Markers Ovarian reserve markers have been widely used to quantify the risk for OHSS. High anti-Mullerian hormone (AMH) levels and high antral follicle count (AFC) are related to increased risk of OHSS. However, despite the wide application of AMH- and AFC-based “individualized” protocols, clear cut-off values of these markers have not been prospectively validated [2].

The pathophysiology of OHSS is characterized by enlarged ovaries and an increase in capillary permeability, leading to leakage of fluid from the vascular compartment, with third-space fluid accumulation and hypovolemia. Endocrine and immunological mediators and vasoactive cytokines have been implicated in the pathogenesis of OHSS, such as vascular endothelial growth factor-A (VEGF-A), interleukin 1, 2 and 6 (IL1, IL2, IL6) and tumor necrosis factor (TNF) alpha (Figure 20.1). An understanding of the pathophysiology allows the clinician to understand the rationale behind the various preventative strategies available [4].

Controlled Ovarian Stimulation: Protocols and Regime

Ovarian Response Parameters

Gonadotropin-Releasing Hormone (GnRH) Antagonist

Ovarian response parameters may aid the identification of patients at high risk during COS. Development

Protocols using GnRH antagonist are associated with a reduced incidence of OHSS. A Cochrane review

How to Avoid Ovarian Hyperstimulation Syndrome

LH

HCG Cancellation, cryopreservation

Figure 20.1 Preventative measures in relation to OHSS pathophysiology

Continue GnRHa after cancellation

Macrophages and granulosa cells Coasting

Metformin

VEGF Aspirin

Microthrombi

Cabergoline, Dopamine

Inc. vascular permeability

Angiotensin antagonists Secondary mediators

OHSS

included 73 randomized control trials (RCTs) with 12,212 participants. The authors concluded that GnRH antagonist protocols are associated with a substantial reduction in OHSS without reducing the likelihood of achieving live birth, when compared with long-course GnRH agonist protocols [5].

Choice of Gonadotropin: Starting Dose and Preparation Although it is widely considered that OHSS incidence may vary with the starting gonadotropin dose, there is little evidence to show what the optimal dose of folliclestimulation hormone (FSH) for potentially high responders should be. Pragmatically, doses ranging from 100 to 150 iu daily are commonly used. An individualized approach, taking into consideration previous stimulation history and risk factors for excessive response, is reasonable. The choice of gonadotropin does not seem to affect the incidence of OHSS. A Cochrane review, including 42 trials with a total of 9,606 couples did not find evidence of a difference in the incidence of OHSS rate between patients prescribed different gonadotropin preparations [6].

Choice of Trigger Human chorionic gonadotropin (hCG) is critical to the development of OHSS in cycles with prior exposure to FSH. Cycle cancellation before final oocyte maturation triggering can virtually eliminate the risk of OHSS. Understandably, cycle cancellation has

a significant emotional and financial impact, and may not be acceptable except in very high-risk scenarios. The use of recombinant hCG is neither safer nor more effective than urinary hCG, according to a recently updated Cochrane review [8]. In GnRH antagonist cycles, final follicular maturation can be achieved by administration of a GnRH agonist trigger, inducing an endogenous luteal hormone (LH) surge. As LH has a shorter half-life and produces less sustained luteotrophic stimulation than hCG, this strategy is associated with a lower risk for OHSS. A meta-analysis including 17 RCTs and 1,847 patients concluded that GnRH agonist trigger instead of hCG in fresh autologous GnRH antagonist IVF/ ICSI treatment cycles significantly reduces the risk of OHSS, but to the detriment of the live birth rate [8]. The difference in pregnancy rate was not seen in donor-recipient cycles, suggesting luteal phase inadequacy when agonist trigger was utilized [9]. Dual triggering with co-administration of GnRH agonist (commonly Buserelin 2 mg) and hCG (up to 5000 iu) may improve the reproductive outcome, but the risk of OHSS in this clinical scenario remains to be established. Other measures have been suggested to improve the luteal phase and pregnancy rate in cycles where GnRH agonist trigger has been used, including intensive steroid therapy with intramuscular progesterone and transdermal estradiol, luteal phase recombinant LH injections and daily luteal intranasal GnRH

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agonist [2]. There is no definitive evidence that any of these luteal phase measures improves the live birth rate with GnRH trigger to that obtained by hCG trigger. Where fresh embryo transfer is not a consideration, e.g. oocyte donation or fertility preservation, GnRH agonist trigger provides the safest option for follicular maturation. Despite the effectiveness of the endogenous LH surge produced by GnRH agonist administration, triggering by recombinant LH has not shown to reduce the risk of OHSS.

Coasting Coasting refers to the practice of withholding gonadotropins while continuing pituitary suppression, in cycles with over-response, allowing the response to settle to “safe” levels before a trigger is administered. Retrospective studies suggest that coasting is associated with a reduction in the risk of developing OHSS, but the criteria for when to initiate or stop coasting are not widely agreed. Most clinicians would not recommend coasting until the leading follicles have reached a mean diameter of around 15 mm. Estradiol levels between 2,500 pg/ml and 6,000 pg/ml have been used as indicators of the need for coasting. Evidence from a large retrospective study indicates that delaying the hCG trigger until estradiol drops to 3,000 pg/ml is associated with a low risk of OHSS [2]. In GnRH antagonist cycles the option of GnRH agonist trigger means that the role of coasting may be limited, or alternatively coasting may only need to be for a short duration as the trigger may be safely administered at a higher estradiol level than in cycles with hCG trigger.

69 studies and 16,327 cycles found a significantly lower risk of developing OHSS with the use of progesterone compared with hCG, with a comparable pregnancy outcome [10].

Pharmacologic Interventions Several pharmacologic interventions have been used attempting to reduce the incidence of OHSS in highrisk patients. In decreasing order of the quality of evidence supporting each medication, these are:

Metformin Studies on cultured vascular smooth muscle cells revealed that insulin stimulates VEGF protein expression and secretion. Based on this theory, metformin (500 mg three times a day or 850 mg twice a day) has been investigated as an adjuvant for ART in women with polycystic ovarian syndrome (PCOS), in an attempt to reduce the risk of OHSS. Evidence from meta-analysis shows evidence of benefit.

Dopamine Agonists Dopamine agonists, such as cabergoline, bromocriptine and quinagolide, have been proposed as a preventative measure for OHSS, based on dopamine’s antagonistic action to the vascular permeability-enhancing effect of VEGF through the dopamine receptor type 2. Cabergoline has been the most widely studied, in the dose of 0.5 mg daily for eight days starting on the day of trigger. Evidence suggests that cabergoline does not affect neo-vascularization, enabling implantation to occur. Cabergoline appears to prevent early, but not late, OHSS.

Freeze-All

Hydroxy-ethyl starch (HES)

Elective cryopreservation of all embryos (“freeze-all”) avoids exposure to endogenous hCG of pregnancy and should thereby eliminate the possibility of late OHSS. Despite its theoretical value and widespread use in practice, cryopreservation has not been adequately studied and robust evidence of its effectiveness in preventing OHSS is lacking. However, taking in consideration advances in cryopreservation techniques and the existing moderate evidence, freeze-all should be considered as a key intervention in high-risk patients [2].

Plasma volume expanders, as HES (6 percent HES 500–1,000 ml), have been used to prevent OHSS. Intravenous infusion on the day of oocyte retrieval or/and the day of embryo transfer has been investigated. A small randomized trial supports the value of this intervention.

Luteal Phase Support Increased hCG exposure is associated with an increased risk of OHSS. A meta-analysis including

Aspirin Low-dose aspirin (100 mg, once a day) has been used based on the theory that high VEGF levels may cause increased platelet activation and lead to release of substances, such as histamine, serotonin, platelet-derived growth factor or lysophosphatidic acid, that can further potentiate the physiologic cascade of OHSS.

How to Avoid Ovarian Hyperstimulation Syndrome

Intravenous (IV) Calcium Intravenous calcium infusion (10 mL of 10 percent calcium gluconate in 200 mL normal saline) on the day of oocyte retrieval and days 1, 2 and 3 after oocyte retrieval has been investigated as a preventative measure for OHSS. Increased calcium is hypothesized to inhibit cAMP-stimulated renin secretion, which decreases angiotensin II synthesis and its subsequent effect on VEGF production.

IV Albumin Administration of intravenous albumin (10–50 gr) around the time of oocyte retrieval has been also proposed as a measure to prevent OHSS. The underlying theory suggests that albumin may bind to vaso-active mediators, such as VEGF. Several randomized trials have been conducted, with varying results. On balance, it appears unlikely that albumin has a major preventative effect on the risk of OHSS.

GnRH agonist trigger and elective cryopreservation can significantly reduce the risk of OHSS, and is applicable to a large number of at-risk women. In women with PCOS, the use of metformin reduces the risk of OHSS. Evidence from non-randomized studies supports coasting in cycles with overresponse. Cabergoline may reduce the risk of developing early, but not late OHSS. Aspirin, IV calcium, metformin and HES may have a role that requires further research in high-risk patients. However, no available method guarantees complete avoidance of OHSS, and there is no consensus on criteria for applying various preventative strategies. Further research is needed in order to predict the risk of OHSS and optimize ART as a safe and effective treatment.

Suggested Standard Operating Protocol (SOP) Management of OHSS

Miscellaneous Treatments Luteal GnRH antagonist administration, letrozole, methylprednisolone, intramuscular progesterone and ketoconazole have been proposed as preventative measures. So far there are insufficient data to evaluate the use of these measures and further research is needed.

Suggested Audit Incidence and outcomes of OHSS

Recommended Patient Information Sheet OHSS

Comparison of Pharmacologic Interventions Guo et al. published a comprehensive systematic review and network meta-analysis in January 2016, evaluating the pharmacologic interventions to prevent OHSS [11]. Eleven preventive strategies were included. The incidence of OHSS was the primary outcome and the pregnancy rate was the secondary outcome. Thirtyone RCTs were identified, including 7,181 patients. Five pharmacologic interventions were superior to placebo in decreasing OHSS incidence: aspirin, IV calcium, cabergoline, metformin and IV HES, without affecting the success of the treatment. The rank probability demonstrated aspirin and IV calcium to be the most effective. Additionally albumin was found to decrease the pregnancy rate, without preventing OHSS. [1]

Conclusion Ovarian hyperstimulation syndrome is a significant iatrogenic complication in women undergoing ART. Prediction of women at high risk is crucial to optimize the treatment and apply preventative measures. However, the identification of these patients remains challenging. The use of GnRH antagonist protocols,

References 1. B. Luke, et al. Factors associated with ovarian hyperstimulation syndrome (OHSS) and its effect on assisted reproductive technology (ART) treatment and outcome. Fertil. Steril. 2010;94(4):1399–1404. 2. R. S. Mathur and B. K. Tan. British Fertility Society Policy and Practice Committee: Prevention of ovarian hyperstimulation syndrome. Hum. Fertil. (Camb.) 2014;17(4):257–68. 3. R. G. Steward, et al. Oocyte number as a predictor for ovarian hyperstimulation syndrome and live birth: An analysis of 256,381 in vitro fertilization cycles. Fertil. Steril. 2014;101(4):967–73. 4. B. K. Tan and R. Mathur. Management of ovarian hyperstimulation syndrome guidelines. Produced on behalf of the BFS Policy and Practice Committee. Hum. Fertil. (Camb.) 2013;16(3):160–1. 5. H. G. Al-Inany, et al., Gonadotrophin-releasing hormone antagonists for assisted reproductive technology. Cochrane Database Syst. Rev. 2016;4: CD001750. 6. M. van Wely, et al., Recombinant versus urinary gonadotrophin for ovarian stimulation in assisted

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reproductive technology cycles. Cochrane Database Syst. Rev. 2011;(2):CD005354.

9. L. Meyer, et al. Risk factors for a suboptimal response to gonadotropin-releasing hormone agonist trigger during in vitro fertilization cycles. Fertil. Steril. 2015;104(3):637–42.

7. M. A. Youssef, A. M. Abou-Setta and W. S. Lam, Recombinant versus urinary human chorionic gonadotrophin for final oocyte maturation triggering in IVF and ICSI cycles. Cochrane Database Syst. Rev. 2016;4:CD003719.

10. M. van der Linden, et al. Luteal phase support for assisted reproduction cycles. Cochrane Database Syst. Rev. 2011;(10):CD009154.

8. M. A. Youssef, et al., Gonadotropin-releasing hormone agonist versus HCG for oocyte triggering in antagonist-assisted reproductive technology. Cochrane Database Syst. Rev. 2014;10:CD008046.

11. J. L. Guo, et al., Pharmacologic interventions in preventing ovarian hyperstimulation syndrome: A systematic review and network meta-analysis. Sci. Rep. 2016;6:19093.

Chapter

21

Anesthetic Choices in IVF Practice Singaraselvan Nagarajan and Eileen Lew

Introduction Women undergoing procedures in in vitro fertilization (IVF) practice, such as oocyte retrieval and embryo transfer, often require analgesia and sedation or anesthesia for optimal surgical conditions and comfort. Although most commonly performed by the transvaginal approach under ultrasound guidance, oocyte retrieval can also be performed transabdominally and by laparoscopy. The ideal anesthetic technique should: • Be short acting and easily titratable • Be associated with good recovery profile and minimal postoperative side effects • Provide adequate analgesia • Provide immobilization for excellent surgical conditions • Not adversely affect success of pregnancy IVF procedures are usually performed in an ambulatory setting at a fertility center that is freestanding or part of a hospital complex. The location and set-up of these centers must be given careful consideration to ensure they fulfill standards and requirements pertaining to facilities, equipment and personnel needs in IVF procedures performed under sedation or anesthesia. In particular, there should be easy access to inpatient beds for those who develop serious perioperative complications.

Preoperative Considerations Prior to the planned IVF procedure, every patient should have a thorough pre-anesthesia evaluation. This usually involves history-taking, targeted physical examination and review of past anesthetic records. If there had been previous oocyte retrieval, the type and dose of anesthetic or sedative medications administered and occurrence of any perioperative complications should be noted. Risks and benefits of anesthetic options and pain management strategies should be discussed with the patient. All patients

should be advised to follow standard preoperative fasting guidelines.

Cause of Subfertility The underlying cause of subfertility may influence anesthetic management (Table 21.1). Patients with high body mass index (BMI) have an increased risk of obstructive sleep apnea (OSA), gastroesophageal reflux disease (GERD), diabetes mellitus and hypertension. Those with OSA may exhibit increased opioid sensitivity and frequent desaturations in the intra- and postoperative period. Hence, patients with high BMI may not be suitable for day case IVF procedures performed under general anesthesia. A non-particulate antacid should be administered on the morning for those with aspiration risk. Underlying anemia, pelvic infections and thyroid disorders should be treated before initiating IVF cycles. Women with severe endometriosis may suffer chronic pelvic pain and have higher analgesic requirements.

Table 21.1 Anesthetic considerations in patients with known causes of subfertility

Causes of subfertility

Anesthetic considerations

Polycystic ovarian syndrome

high body mass index, insulin resistance, abdominal bloating

Pelvic inflammatory disease

acute or acute-on-chronic pelvic infections

Endocrine disorders

abnormal thyroid function

Endometriosis

abdominal bloating, anemia, chronic pelvic pain

Fibroids

anemia

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Preexisting Illness As women seeking fertility treatment may be in their late 30s or 40s, they may present with age-related medical conditions, such as hypertension and diabetes. Patients with poorly controlled medical conditions should be co-managed with a physician, prior to initiation of IVF cycles. Antihypertensive medications should be continued on the morning of procedure. Women with Type II diabetes should omit their oral hypoglycemic agents upon fasting while those with Type 1 diabetes should follow standard perioperative insulin regimes designed for ambulatory surgery.

Ovarian Hyperstimulation Syndrome Controlled ovarian stimulation, which results in more eggs and hence a higher chance of pregnancy, is carried out before oocyte retrieval. Ovarian hyperstimulation syndrome (OHSS) occurs due to an exaggerated hormonal response to ovulation induction. Patients may present with clinical features of mild-tomoderate OHSS, including: nausea and vomiting, abdominal bloatedness, pain and diarrhea. In severe OHSS, acute abdominal pain from follicular rupture and hemorrhage, ascites, pleural effusion and oliguria can occur.

Anxiety Subfertility is a source of anxiety for women. Furthermore, uncertainty about the outcome of the IVF procedure and previous failed IVF cycles compound their anxiety. The level of preoperative anxiety can influence the amount of sedation required and intraoperative patient movement [1]. Preoperative assessment should aim to allay any anxiety the patient may have. A clear explanation of what to expect and involving the patient in the decision-making process about the choice of anesthetic technique can help to reduce anxiety. Anxiolytics may be prescribed if deemed necessary.

Anesthetic Choices Impact of Anesthetic Drugs on Clinical Pregnancy Rates Laparoscopic oocyte retrieval, with the associated carbon dioxide pneumoperitoneum, has been noted to significantly reduce follicular fluid pH and oocyte fertilization rates. Although anesthetic agents have

been shown to accumulate in follicular fluid, there is no clear evidence that current clinical doses adversely affect oocyte fertilization and embryonic development. A Cochrane review of 21 randomized control trials in 2013 showed no significant difference in pregnancy rates with respect to anesthetic techniques employed for assisted reproductive procedures [2]. These techniques include: conscious sedation and analgesia, in conjunction with acupuncture or paracervical block; general anesthesia and spinal anesthesia. Trials also compared patient-controlled versus physician-administered conscious sedation. In spite of study limitations due to poor reporting of methods, small sample sizes and heterogeneity of protocols, this Cochrane review suggests that the concurrent use of more than one method of sedation and analgesia resulted in better pain control than one modality alone.

Anesthetic Management Anesthetic options include • Sedation with monitored anesthesia care (MAC) • General anesthesia (GA) • Regional/local anesthesia Although various combinations of anesthetic and sedation techniques have been described, transvaginal oocyte retrieval is most commonly performed under sedation, with deepening of sedation depth to approach that of GA at critical time points to prevent patient movement.

Sedation with MAC Under MAC, sometimes referred as “conscious sedation” or “twilight anesthesia,” medications are given intravenously to induce a state of depression, with deeper planes of sedation achieved at critical time points to prevent movement. The challenge lies in the ability to anticipate these episodes of increased surgical stimulation so that medications can be titrated accordingly, while maintaining spontaneous ventilation. Commonly, an intravenous anesthetic agent (e.g., propofol or thiopentone) or a benzodiazepine (e.g., midazolam) are used in combination with a short-acting opioid (e.g., fentanyl, alfentanil or remifentanil) to achieve sedation and analgesia. Some centers have also reported using ketamine

Anesthetic Choices in IVF Practice

Table 21.2 Advantages and disadvantages of different anesthetic techniques used in IVF procedures

Advantages

Disadvantages

Sedation with Monitored Anesthesia Care

General Anesthesia

Regional Anesthesia

Easy administration

Airway maintained by anesthetist

Patient maintains own airway

Patient maintains own airway

No involuntary limb movements

No involuntary limb movements

Faster recovery time

No discomfort during procedure

No or minimal discomfort during procedure

Involuntary limb movements

Postoperative nausea vomiting

Delayed ambulation

Discomfort during procedure

Prolonged recovery time

Urinary retention

analgesia as part of the propofol/midazolam sedation technique. Propofol, an intravenous anesthetic agent known for its rapid onset-and-offset, is particularly useful. It also has anti-emetic properties, and this reduces the risk of postoperative nausea and vomiting, compared with thiopentone. Propofol can be given as [1] intermittent boluses titrated to required depth, with the addition of a short-acting opioid like fentanyl or alfentanil [2], a target-controlled infusion (effect-site concentration 1.5–2.0 mcg/mL) with remifentanil (effect-site concentration 1.5–2.5 ng/mL). Patient-controlled sedation and analgesia techniques, using propofol and short-acting opioids, have been employed successfully at some centers. A nonanesthetist-led sedation service for IVF procedures has also been described and found to be safe, with a respiratory adverse incident rate of 0.5/1000 and unplanned direct anesthetic assistance rate of 3.5/1,000 cases [3].

General Anesthesia (GA) General anesthesia may be considered in • Very anxious patients • Cases where a long procedure time is anticipated, e.g. large number of oocytes to be retrieved Furthermore, GA provides excellent surgical conditions for the IVF procedure as there would be no involuntary lower limb movements that may increase the likelihood of injury to pelvic structures. A spontaneous breathing technique with laryngeal mask airway or mask anesthesia is preferred. Both volatile and total intravenous anesthesia (TIVA) techniques can be employed, with the addition of an opioid for analgesia. In patients with active GERD, securing the

airway via endotracheal intubation is necessary, aided by the use of muscle relaxants. Use of volatile anesthetic agents poses a risk of postoperative nausea and vomiting and may delay recovery and home-readiness.

Regional/Local Anesthesia Spinal anesthesia may be considered in patients with • Morbid obesity • Moderate-to-severe obstructive sleep apnea • Gastroesophageal reflux disease • Inadequate fasting time The sensory and motor block under spinal anesthesia provides excellent surgical conditions. Disadvantages of spinal anesthesia include potential for prolonged motor block, urinary retention and postdural puncture headache. Use of short-acting local anesthetic agents such as 1 percent chloroprocaine or 0.5 percent prilocaine may allow earlier home-readiness while maintaining a low incidence of transient neurological syndrome, compared with lignocaine. Epidural anesthesia is not routinely used. Paracervical block can provide local anesthesia around the vaginal cervix and fornicular areas, thus minimizing pain during needle insertion. However, it cannot provide adequate analgesia for the ovarian puncture and supplementary sedation and analgesia are usually required.

Other Techniques The use of acupuncture techniques, alone or in combination with sedation and analgesia, has been reported. The efficacy of acupuncture in facilitating assisted reproductive techniques has been under investigation for several years, with conflicting results.

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A Cochrane review [4] conducted in 2013 concluded that there is no benefit or harm in using acupuncture in patients undergoing oocyte retrieval or embryo transfer in terms of improving the live birth rate or clinical pregnancy rate.

Postoperative Care Common Complications in Recovery and Their Management Pain Abdominal pain, if present, can usually be treated with simple analgesics, e.g. paracetamol or nonsteroidal anti-inflammatory drugs. Severe pain may necessitate use of opioids. Pain may be more significant if the procedure is prolonged or multiple eggs (more than six) are retrieved.

Nausea and Vomiting Use of volatile anesthetics, opioids and manipulation of ovaries (both pharmacologically before procedure and surgically during egg retrieval) all contribute to nausea and vomiting. Intraoperative prophylaxis with combination antiemetic agents (e.g., ondansetron, cyclizine and metoclopramide) should be considered. Further rescue therapy may be needed in the recovery.

Fever Raised temperature is commonly seen in patients presenting for IVF procedures. This may be the result of ovarian stimulation. The fever usually settles with the use of paracetamol. If the temperature is persistently high post-procedure, the patient may need admission for further investigation and management.

Urinary Retention One of the recognized postoperative complications of spinal anesthesia is urinary retention. If present, it is advisable to keep the patient under observation. If urinary retention does not resolve, short-term catheterization is warranted.

Ovarian Hyperstimulation Syndrome If abdominal pain, shortness of breath, oliguria and severe nausea and vomiting occur, ovarian hyperstimulation syndrome (OHSS) should be suspected. Mild cases may be managed with judicious hydration and symptomatic treatment. In severe OHSS,

hospitalization for further investigations and management may be necessary after oocyte retrieval.

Hypotension Hypotension (defined as < 20 percent of pre-procedure systolic blood pressure) is common after general and regional anesthesia, contributed by mild dehydration from fasting overnight. Hypotension can be avoided by the infusion of 500–1,000 mL of crystalloid solution during and immediately after the procedure. If hypotension persists after fluid rehydration, other causes of hypotension (particularly bleeding and hypoglycemia) should be excluded and treated accordingly.

Home-Readiness Patients may be discharged if they fulfill the following criteria: • Stable vital signs • Minimal pain, nausea and/or vomiting • Able to pass urine • Able to ambulate without assistance • Oriented to time, place and person • Presence of a responsible escort • Instructed on emergency contact

Conclusion Advances in surgical and anesthetic techniques, coupled with patient-centric safety initiatives, have all contributed toward making IVF procedures safer and more efficient. Close doctor–patient communication to identify patients’ needs and address their concerns remains the cornerstone in ensuring a pleasant and satisfying patient experience. IVF centers should be well prepared to deal with the unexpected complications that may arise as a result of the procedure. Policies and protocols should be in place and regular audits should be conducted to assess for compliance to guidelines. Some areas of audit should include unplanned hospital admissions after procedure, cancellations on the day of procedure and incidence of postanesthetic complications.

References 1 T. M. Osborn and N. A. Sandler. The effects of preoperative anxiety on intravenous sedation. Anesth. Prog. 2004;51(2):46–51.

Anesthetic Choices in IVF Practice

2. I. Kwan, S. Bhattacharya, F. Knox and A. McNeil. Pain relief for women undergoing oocyte retrieval for assisted reproduction. Cochrane Database of Systematic Reviews 2013;Issue 1. Art. No.: CD004829. doi: 10.1002/ 14651858.CD004829.pub3 3. J. A. Edwards, J. Kinsella, A. Shaw, S. Evans and K. J. Anderson. Sedation for oocyte retrieval using target controlled infusion of propofol and incremental alfentanil

delivered by non-anaesthetists. Anaesthesia 2010;65(5): 453–61. doi: 10.1111/j.1365–2044.2010.06264.x 4. Y. C. Cheong, S. Dix, E. Hung Yu Ng, W. L. Ledger and C. Farquhar. Acupuncture and assisted reproductive technology. Cochrane Database of Systematic Reviews 2013;Issue 7. Art. No.: CD006920. doi: 10.1002/14651858. CD006920.pub3

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22

Tips and Tricks of Transvaginal Oocyte Retrieval Durga G. Rao, Sujatha Vellanki and Ying Cheong

Introduction Transvaginal ultrasound-directed oocyte retrieval (TVOR) is the most widely used method of oocyte retrieval for in vitro fertilization (IVF). This technique was first developed by Pierre Dellenbach and colleagues in Strasbourg, France, and reported in 1984. Steptoe and Edwards used laparoscopy to recover oocytes when IVF was introduced, and laparoscopy was the major method of oocyte recovery until TVOR. The use of ultra-sonographic monitoring in the early 1980s has considerably simplified and improved follicle identification and oocyte recovery. In addition, remarkable refinements were eventually made on ultrasound technology, aspiration equipment and anesthesia procedures that contributed to efficacy, safety and comfort of oocyte retrieval. Complication rates of TVOR are low, with most studies citing a < 5 percent complication rate [1]. The most common complications include severe abdominal pain (3 percent), vaginal bleeding (3–8 percent), mild pelvic infections (~1 percent) and pelvic abscesses (0–0.6 percent) [2, 3]. Injury to pelvic viscera including the bowel, ureter and bladder is far less common. In this chapter, we would like to discuss, first, the importance of identifying patients and conditions that may cause difficulty at oocyte retrieval, and, then, the various solutions to these problems to make the procedure effective and safe.

Preliminary Steps When a woman is being evaluated for fertility treatment, her medical history is the starting point for identifying risk factors for TVOR. This is probably the most important step in avoiding complications, that is, identifying high-risk patients and taking adequate measures to ameliorate the risks. A history of: a. Dysmenorrhea: need to rule out endometriosis and adenomyosis and adhesions of the ovaries causing altered position of the ovaries. Also

b.

endometriotic cysts in the ovaries and their sizes can impact the retrieval rates and increase the complication of infection post procedure. Pelvic inflammatory disease (PID): investigating and treating pelvic infection is very important before TVOR and should be a routine step before scheduling the start of IVF/intracytoplasmic sperm injection (ICSI). A high vaginal swab to detect chlamydia and bacterial infection is mandatory. Pelvic infection complicating TVOR is approximately 0.3–1.5 percent [4]. There have been case reports of even tuberculosis reactivation post TVOR [5], and Actinomyces tubo-ovarian abscess can be identified on rare occasions.

Even endometriotic cysts can be a risk for postTVOR sequelae with the formation of tubo-ovarian masses; therefore utmost care must taken when attempting retrieval in women with severe endometriosis and especially in the presence of endometriotic cysts. Follicles should be aspirated without piercing the cysts, but if follicles are inaccessible, then pre-IVF laparoscopy and cystectomy and adhesiolysis could reduce the complications post TVOR, and should be considered and discussed with the woman. Traditional practice has placed emphasis on surgery only if ovaries are inaccessible or prohibit the easy aspiration of follicles, with concerns of surgery placed on possible reduction in ovarian reserve. c. Previous complex abdominal-pelvic surgeries: examples of these include vaginal reconstruction surgeries, adhesiogenic procedures such as myomectomy, those with severe endometriosis, surgery for inflammatory bowel disease such as Crohn’s disease, which would require accurate baseline evaluation for ease of retrieval and accessibility of the ovaries. In cases where ovarian repositioning has been performed in women who underwent radiotherapy for pelvic malignancies, either transabdominal ultrasound

Tips and Tricks of Transvaginal Oocyte Retrieval

d.

e.

f.

guided retrieval or returning the ovaries to their normal anatomical position in a repeat laparoscopy to enable TVOR should be considered. Abnormal situation of kidney: presence of pelvic kidney should be diagnosed before the procedure to avoid complications, especially in women with Mullerian abnormalities. Women with raised BMI: the ovaries may not be easily visualized in women with significant obesity, and can lead to a higher risk of complications as well as a lower egg yield. Weight loss appears to improve the visualization of the ovaries as a result of reduced visceral fat. Chronic constipation: in women with loaded rectum, use of laxatives for bowel clearance from the day prior to oocytes retrieval may be helpful. During the process of TVOR Always verify whether the trigger injection was taken and the time and date it was taken to ensure that the TVOR is done at the appropriate time, so that “empty follicle syndrome” due to faulty administration and timing of ovulation trigger injections is avoided. Types of pain relief used for transvaginal oocyte retrieval include local, conscious sedation and epidural, spinal and general anesthesia (see Chapter 21). The primary goal is to provide safe and effective analgesia facilitating optimum surgical conditions and speedy postoperative recovery. There is also concern about potential effects of any drugs used on reproductive outcome. A transvaginal probe with a transducer frequency of 5–7 MHz gives a sufficient penetration depth and enough resolution for accurate scanning of the lower pelvis and is easy to handle during follicular puncture. A sterile probe cover cleansed in saline can be used to cover the probe. The patient should be advised to empty her bladder before the procedure. Lithotomy position is generally recommended. Alternatively when it is not possible due to any muscle contractures or knee joint problems, lateral or Sims position can be attempted. The vagina is cleansed two to three times with normal saline to remove the mucus and discharge. Antiseptic solutions and povidone iodine are toxic to gametes, and are therefore not recommended. Different needles are available commercially for oocyte retrieval. If the inner diameter of the needle is 0.8–1 mm, the oocyte cumulus complex appears to be unaffected, provided that the aspiration pressure is < 120 mmHg. A 17- to 18-gauge needle (outer diameter) thin walled with an inner diameter of 20 gauge

is ideal for oocyte retrieval. The needle is made for single use and thereby guarantees sterility and is nontoxic to the oocyte. There is some evidence of a trend for lower pain scores with transvaginal collections performed with a 19-gauge needle (used in in vitro maturation [IVM] retrievals) when compared with a 16- or 17-gauge needle (used in IVF retrievals), although not statistically significant [6]. The needle guide should be properly fixed to the probe, so that there is no inadvertent damage due to the aspiration needle.

Procedure for aspiration: The ultrasound probe should be positioned intravaginally to facilitate direct needle access to the ovarian follicles without the risk of puncturing other pelvic visceral or blood vessels. If required, ultrasound Doppler can be used to verify the course of smaller blood vessels. The needle should puncture the follicle at right angles at the point of the largest diameter.

Movement of Aspiration Needle The speed of the needle insertion should be controlled in several ways suggested in what follows (Figure 22.1). 1) The best way to undertake retrieval is for the operator to grasp the needle with their fingertips (like a pencil) and advance it using an overhand motion. Advancing the needle using the small muscles of the hand and fingers with a wrist motion allows the operator to have full control of the needle’s movement and speed. 2) The alternative and more common aspiration approach is to advance the needle with an underhand motion using the muscles of the wrist and forearm. Whichever method is adopted, further stabilization of the arm, to avoid muscle strain, can be achieved by resting the elbow on the knee. The question as to whether small, controlled rotation of the aspiration needle, while aspirating, is advantageous is still controversial. Dahl et al. [7] showed that rotating the needle in a follicle during aspiration increased the number of oocytes obtained. Some surgeons believe that this needle motion causes scraping or curetting of the wall of the follicle. The possible benefits of gentle needle rotation are 1) decreasing the likelihood of the needle lumen becoming prematurely blocked by a collapsing follicle wall or debris; 2) preventing the needle tip from unintentionally exiting

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within the palm of the assistant’s hand when the tubes are transported into the laboratory. It is useful to have the oocyte collection procedure performed in a theatre that is adjacent and has direct access to the laboratory. Antibiotic prophylaxis is recommended prior to oocyte retrieval in patients with a history of endometriosis, PID, ruptured appendicitis or multiple prior pelvic surgeries [9].

on of d moti n a h r Ove eedle tion n aspira

Difficult Oocyte Collection Scenarios Underhand motion of aspiration needle Figure 22.1 Over and underhand motion for egg collection

the collapsing follicular wall; 3) enhancing visualization of the needle tip; and 4) providing a “tunneling” motion that will facilitate the complete entry of the needle tip through the flexible follicular wall. Applying vacuum suction prior to the needle entering the follicle will aspirate follicular fluid that may escape with the oocyte as the wall is being punctured [8].

Follicular Flushing Follicular flushing can be achieved with a double lumen aspiration needle, as creating turbulence within the follicle by flushing is potentially another approach to freeing the oocytes. The usage of a wider double lumen needle may increase the complications of bleeding and postoperative pain and also the duration of the procedure, and hence is to be used with caution especially in women with fewer than four oocytes.

Complications Can Be Avoided by Proper Visualization of the Needle Tip throughout the Procedure The needle is connected to polyethylene tube (10 cc), which is in turn connected to the foot pump; hence good hand, foot and eye coordination is required throughout the oocyte aspiration procedure.

Temperature Control Temperature control is the most important of all the steps as delay in delivering of oocytes to the laboratory can result in detrimental cooling effect to the oocytes. Use test tube warmers or have the test tubes held

Ovary Located too Far Above the Vaginal Fornices Intramyometrial (Terumo & Kato technique) and intraabdominal approaches can be tried. The intramyometrial approach is associated with an increased risk of bleeding and overt manipulation of the needle should be avoided, to avoid breaking the needle in situ. A tenaculum placed on the posterior lip of the cervix and applying downward traction may bring the ovary nearer to the vagina. Partially filling the bladder may also alter the position of the ovaries in an advantageous way. The laparoscopic surgical mobilization of ovaries may be considered prior to commencing IVF. Antiadhesion barriers may be used to reduce the chance of postoperative adhesion reformation.

Extremely Mobile Ovaries Occasionally, when there are very few stimulated follicles and a mobile ovary, the ovary may move away from the needle, making aspiration of the follicles within the ovary a challenge. Adequate anesthesia can reduce breathing movements and may make the ovary accessible for needle insertion. External abdominal manual pressure by an assistant is often all that is required to stabilize the ovary for aspiration.

Follicle/Blood Vessel When in doubt, use the Doppler to identify the blood vessel. The ultrasound probe should also be rotated through 90°, where a vessel will look tubular, while the follicle will still appear round. If a puncture to the pelvic vessels is suspected, the needle should be immediately removed gently and the patient monitored closely. In general, bleeding from a small puncture in the

Tips and Tricks of Transvaginal Oocyte Retrieval

pelvic vessels will cease, but if bleeding was observed and the patient shows signs of acute blood loss, and free fluid can be seen accumulating within the pelvis via ultrasound, one may have to resort to laparoscopy/ laparotomy. A vascular surgeon should be called, in addition to the emergency support team within the hospital. Early recourse to laparoscopy or laparotomy in the event of uncontrolled bleeding reduces the morbidity and the chances of oophorectomy.

Endometriosis/Hydrosalpinx One should avoid aspirating the endometriotic cyst or a hydrosalpinx. If these structures were aspirated inadvertently, the oocyte retrieval needle will require repeated flushing to avoid infection and blockage. Consider broad-spectrum antibiotics to avoid infection or tubo-ovarian mass.

Large Uterus (Fibroids) The pelvic cavity has a limited capacity to accommodate two enlarged ovaries with follicles as well as a large fibroid uterus. If fibroids are large and immobile, the mobile ovaries may be displaced above the uterus, making transvaginal retrieval difficult. Therefore, it is important to consider the accessibility of the ovaries prior to the ovarian stimulation. If necessary, a transabdominal approach is planned, with adequate anesthesia. Alternatively, a myomectomy should be considered beforehand.

Empty Follicle Syndrome Empty follicle syndrome occurs when no oocytes are recovered from the follicular fluid. This may be because the oocyte failed to mature due to the failure to “trigger ovulation.” The situation can be clarified by checking, in the case of an agonist trigger, the patient’s serum luteinizing hormone (LH) and progesterone levels, and in the case of HCG trigger, HCG levels. A three times rise in LH from the basal level is indicative of adequate LH surge, when agonist trigger is given. If in spite of an adequate LH surge or HCG levels, no oocytes are retrieved, the condition can then be attributed to a genuine empty follicle syndrome.

Conclusion Transvaginal oocyte retrieval is now the standard method for oocyte retrieval. Transvaginal follicular

aspiration is usually preferred due to its shorter operation time and less invasive nature, but the transabdominal ultrasound (TAUS)-guided follicular aspiration can be an alternative in women with ovaries inaccessible by TVUS. TAUS can be considered in patients with raised BMIs or history of pelvic surgeries, or where the ovaries are not easily visualized. TVOR is a “simple” procedure, in the hands of the skilled and trained specialist, with a relatively low risk of complications. As a general principle, raised index of suspicion, early identification of adverse events and prompt response and decision-making, if required for laparoscopy or laparotomy, will limit unwanted sequelae.

Suggested Standard Operating Protocol (SOP) Patient information sheet on what happens during oocyte collection

References 1. T. Catanzarite, L. A. Bernardi, E. Confino and K. Kenton. Ureteral trauma during transvaginal ultrasound-guided oocyte retrieval: A case report. Female Pelvic Med. Reconstr. Surg. 2015;21:e44–5. [PubMed] 2. A. K. Ludwig, M. Glawatz, G. Griesinger, K. Diedrich and M. Ludwig. Perioperative and postoperative complications of transvaginal ultrasoundguided oocyte retrieval: Prospective study of > 1000 oocyte retrievals. Hum. Reprod. 2006;21:3235–40. [PubMed] 3. S. J. Bennett, J. J. Waterstone, W. C. Cheng and J. Parsons. Complications of transvaginal ultrasound-directed follicle aspiration: A review of 2670 consecutive procedures. J. Assist. Reprod. Genet. 1993;10:72–7. 4. C. Aragona, M. A. Mohamed, M. S. Espinola, A. Linari, F. Pecorini, G. Micara and M. Sbracia. Clinical complications after transvaginal oocyte retrieval in 7,098 IVF cycles. Fertil. Steril. 2011;95:293–4. 5. H. I. Annamraju, R. Ganapathy and B. Webb. Pelvic tuberculosis reactivated by in vitro fertilization egg collection? Fertil. Steril. 2008;90 (5):2003.e1–3. doi: 10.1016/j.fertnstert.2008.02.147. Epub 2008 May 27. 6. A. Seyhan, B. Ata, W. Y. Son, et al. Comparison of complication rates and pain scores after transvaginal ultrasound-guided oocyte pickup procedures for in vitro maturation and in vitro fertilization cycles. Fertil. Steril. 2014; 101:705–9.

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7. S. K. Dahl, S. Cannon, M. Aubuchon, D. B. Williams, J. C. Robins and M. A. J. Thomas. Follicle curetting at the time of oocyte retrieval increases the oocyte yield. Assist. Reprod. Genet. 2009; 26(6):335–9. 8. R. Horne, C. J. Bishop, G. Reeves, C. Wood and G. T. Kovacs. Aspiration of oocytes for in-vitro fertilization. Hum. Reprod. Update. 1996; 2(1):77–85. 9. N. Pereira, A. P. Hutchinson, J. P. Lekovich, E. Hobeika and R. T. Elias. Antibiotic prophylaxis

for gynecologic procedures prior to and during the utilization of assisted reproductive technologies: A systematic review. J. Pathog. 2016;4698314. doi: 10.1155/2016/4698314. Epub 2016 Mar 7. 10. K. Sharpe, A. J. Karovitch, P. Claman and T. N. Suh. Transvaginal oocyte retrieval for in vitro fertilization complicated by ovarian abscess during pregnancy. Fertil. Steril. 2006; 86 (1):219.e11–3.

Chapter

23

What to Do When the Electric Oocyte Aspiration Pump Stops Working Ying Cheong

Introduction

The Setup

Transvaginal oocyte recovery is a routine process undertaken in any IVF center. One crucial aspect of the entire oocyte collection process in modern practice now involves the use of an electric suction pump, of which there are numerous makes and models. The electric aspiration pump, when activated by the foot pedal and hence “hands free,” will provide a rapid suction response at the needle tip, and hold constant vacuum settings accurately for long periods of time compared to manual aspiration. While the aspiration pump reliably functions well most of the time, experienced clinicians will have faced challenges in follicular aspiration due to failure of the suction system. Thus, it is crucial that every clinician skilled in reproductive medicine knows what to do in this scenario to enable successful oocyte retrieval.

Figure 23.1 diagrammatically illustrates the system for oocyte collection, with Figure 23.2 demonstrating the system in practice. The principle of the aspiration pump is straightforward, with fluid within the follicle being aspirated by negative pressure suction (generally between 120 mmHg and 170 mmHg) into a test tube, which is warmed by the test tube warmer. A disposable vacuum (ideally with a filter) tube is also attached to the test tube, and connects to the suction machine. The negative pressure generated by the suction machine relays a negative pressure within the test tube, which, in turn, facilitates the aspiration of the follicular fluid and oocyte via the aspiration line into the test tube. The test tube is filled to around three-quarters full and then handed to the embryologist for examination (to confirm presence/absence of oocyte[s]), with an

Suction machine Aspiration line Disposable vacuum like +/– hydrophobic filter

Syringe (if double lumen)

Test tube Direction of aspiration Direction of flushing Direction of suction

Single or double lumen needle

Test tube heater

Figure 23.1 Usual setup for oocyte aspiration

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Oocyte retrieval pre-pack sterile set Aspiration needle Syringe for double lumen needle flush

Suction filter attached to aspiration pump

Bung over test tube

Suction tubing from test tube to aspiration pump

Figure 23.2 Photograph of actual oocyte aspiration setup at Complete Fertility Centre Southampton

empty test tube replacing the tube that has been removed. The function of a new aspiration pump requires validation prior to use, and must be serviced regularly as per its manufacturer’s instructions. It is essential to have a spare pump available and ready for use. The manufacturer’s manual should be retained for future reference. Prior to the start of every oocyte retrieval, a small amount of flush should be aspirated as a test run to ensure the aspiration setup and system is functioning normally before commencing the oocyte collection. The suction pressure is normally set within the range of 120 mmHg to 170 mmHg. A lower range of suction is preferred. Indeed, aspiration pressures of 90 mmHg to 120 mmHg have been associated with good oocyte yield and minimal damage. Despite adjusting the pressure on the suction machine being far from an exact science, higher pressures have been associated with decreased quality of oocyte retrieved, and potential loss of ovarian granulosa cells that may contribute to luteal phase insufficiency. The pressure of 180 mmHg was historically used when oocyte retrieval was performed laparoscopically and morphologically abnormal oocytes were noted with higher frequency than current retrieval methods.

Issues Relating to Machine/System Setup 1)

Error in the way the tubes are connected – if the flow of the aspiration and vacuum tubes are incorrectly connected, there will be no suction, and no flow.

Solution: Check connection with manual or standard operating procedure (SOP) and reconnect the system. 2) Damage to the test tube – if there is a crack in the test tube, the suction will not be sufficient. 3)

Solution: Change to a new, undamaged test tube. The seal of the test tube is insufficient.

If the aspiration flow rate is slow or at drip rate, it is always useful to check that the seal over the test tube is complete. An incomplete seal will not facilitate adequate suction pressure to build up within the test tube. Solution: Tighten the seal by placing gentle pressure over the bung that sits on top of the test tube, but beware not to apply too much force over the bung, as this may create tiny cracks on the lip of the test tube, aggravating the problem. 4) Follicular fluid overflow – fluid has aspirated into the suction machine. Solution: This may have damaged the aspiration machine, and the suction machine should be changed. Contact the manufacturer for advice regarding the need for cleaning and repair. 5) Suction pedal damaged – if the aspiration works with manual suction but not an electrical pump, then the foot pedal may be damaged. Solution: Change the foot pedal and contact the manufacturer for repair or replacement. Always be careful to place the suction pedal cables away from heavy trolleys and/or foot traffic for safety reasons, and also to avoid unnecessary damage to the cables.

When the Electric Oocyte Aspiration Pump Stops Working

Issues Relating to Technique of Oocyte Aspiration 6)

The tip of the aspiration needle is not in the follicle.

Solution: Try repositioning the needle or ask for senior help. 7) The aspiration needle is blocked by blood or endometriotic aspirate. Single lumen aspiration

Solution: Disconnect the aspiration system and flush the needle, and if this is still under high pressure, then change the aspiration needle.

Other Possible Issues 8)

Socket or fuse issue with power supply

Solution: Always check the power source. Change power socket/source. 9) Internal machine malfunction Solution: If all the foregoing steps have been checked and the machine is still not working, then contact the manufacturer and switch to the spare suction machine.

What if There Is No Spare Aspiration Machine? In the event that no spare aspiration machine is available, the operator can switch to manual aspiration. This method essentially uses a larger syringe (e.g., 50 mls) to create the suction that the suction machine would have generated within the test tube (this is the same setup as Figure 23.1 except swap the suction machine to a suction syringe and manually aspirate). While this method works, it is not ideal as there will be no means of monitoring the pressure applied, or to be able to generate a consistent pressure. Some centers may have a single lumen aspiration line, which can connect directly to a syringe. If that is the case, the aspirated fluid will be retrieved directly from the syringe, once filled with follicular fluid and oocyte(s) (Figure 23.3).

Post-Aspiration Machine Failure Actions It is important that machine failure is clearly documented, reported on an institution’s reporting system

Figure 23.3 Single lumen aspiration setup in case no spare suction machine is available

and reviewed for learning and actions to mitigate future risk. Aspiration machine failure is also a clinical risk, with the patient potentially exposed to a longer procedural time and longer anesthetic exposure. The patient may experience increased postoperative pain and symptoms as a result. More importantly, if the issue was not resolved, no oocytes may be retrieved in the small window of time that is available after ovulation trigger. Hence, while it is important that the clinical risk is reported and assessed and actions are taken to prevent recurrence, the patient concerned will require a full explanation and discussion as per the Duty of Candour (Health & Social Care Act, Regulation 20).

Suggested Standard Operating Protocol (SOP) 1) 2)

SOP for oocyte retrieval SOP for actions to take if oocyte aspiration machine fails

Suggested Audits 1)

Issues related to equipment failure during oocyte retrieval

Reference 1. A. Kumaran, P. K. Narayan, P. J. Pai, A. Ramachandran, B. Mathews and S. K. Adiga. Oocyte retrieval at 140 mmHg negative aspiration pressure: A promising alternative to flushing and aspiration in assisted reproduction in women with low ovarian reserve. J. Hum. Reprod. Sci. 2015, Apr.–Jun.;8(2):98–102.

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How to Manage the Patient Who Has No Egg Retrieval Nikoletta Panagiotopoulou and Abha Maheshwari

Poor Response to Controlled Ovarian Stimulation and Limited Follicular Recruitment

IUI, with live birth rates for either option of approximately 5 percent [3].

It is estimated that 5–18 percent of all assisted reproductive technology (ART) cycles are complicated by poor response. Poor response can be expected, such as for women with poor ovarian reserve or previous poor response to controlled ovarian stimulation (COS), or can be unexpected. Possible explanations for unexpected poor response include inappropriate use of gonadotropin injections, decreased number of follicle-stimulating hormone (FSH) receptors in granulosa cells or the presence of FSH receptor-binding inhibitors in follicular fluid. In the face of poor ovarian response to COS, the chance of failure to retrieve oocytes after follicular aspiration is high. Moreover, the majority of poor responders continue to respond poorly in subsequent COS cycles. Therefore, these cycles are associated with very low cumulative pregnancy rates [1]. Management options for poor responders include: cancellation, conversion to intrauterine insemination (IUI) or continuation with the ART cycle and oocyte retrieval (Figure 24.1).

In bifollicular recruitment, available evidence suggests a better chance of a live birth with ART than with IUI, especially in patients younger than 40 years of age [4]. Moreover, cycle cancellation and initiation of a new cycle did not provide better outcomes than continuation with the ART cycle [5]. In addition to evidence on ART effectiveness, economic evaluation studies based on probabilistic decision analysis suggest that continuation with ART is more cost-effective than conversion to IUI [6]. However, the current literature on management options for poor responders is limited to retrospective cohort studies or case series and, therefore, caution is recommended when making recommendation to patients.

No Follicular Recruitment In cases of no follicular recruitment, management options are limited. The dose or the duration of gonadotropins can be increased in order to promote follicular recruitment and follicular development, respectively, even though evidence suggests that an FSH dose greater than 300 IU is of minimal benefit [2]. Cycle cancellation, though, may be inevitable for those who fail to respond to either approach.

Monofollicular Recruitment In cases of monofollicular recruitment, available evidence suggests that there is no significant difference between continuation with ART and conversion to

Bifollicular Recruitment

Subsequent Cycles There is no evidence that any particular protocol is more effective for poor responders [7]. Further treatments should be started after careful counseling with regards to treatment options and guarded prognosis.

Adequate Follicular Recruitment and Development The incidence of failure to retrieve oocytes after follicular aspiration in women with adequate response to COS is low and estimated to occur in 0.05–3.5 percent of women undergoing oocyte retrieval [8]. The commonest cause for this failure is inappropriate or insufficient administration of human chorionic gonadotrophin (hCG), even though causes may also include premature ovulation, technical problems associated with oocyte retrieval, empty follicle

How to Manage the Patient Who Has No Egg Retrieval

Poor response to COS

Expected

Unexpected

Poor ovarian reserve

Good ovarian reserve

Check gonadotropin injection technique

Increase dose & duration of gonadotropin treatment

No follicular recruitment

Monofollicular recruitment

Bifollicular recruitment

Cancel cycle

Offer continuation with ART or conversion to IUI after careful counselling

Offer continuation with ART (optimal option) or conversion to IUI after careful counselling

Figure 24.1 How to manage the patient with poor response to controlled ovarian stimulation resulting in limited follicular recruitment

syndrome, impaired folliculogenesis, suboptimal absorption of hCG and, less frequently, genetic conditions. Identifying the underlying cause of failure to obtain oocytes at the time of retrieval should be the basis for formulating a care plan for these patients (Figure 24.2). To this end, routine fertility investigations, such as pelvic ultrasound scan and hormone assessment, could be helpful. Depending on the drug used for ovulation triggering, serum hCG or luteinizing hormone (LH)/progesterone levels at the time of oocyte retrieval can help distinguish genuine empty follicle syndrome from other causes in women with an adequate number of mature follicles. Indeed, in the latter cases, insufficient circulating hCG or LH/progesterone levels would be observed even though no cut-off levels have been agreed; hCG concentrations from 5 to 161 IU/L have been proposed to suggest adequate ovulation triggering with hCG, but no LH or progesterone cut-offs have been proposed for ovulation triggering with a gonadotropin-releasing hormone (GnRH) agonist. Presence of free fluid in the pelvis on ultrasound scan with sufficient circulating hCG levels can indicate premature ovulation.

Reduced Human Chorionic Gonadotrophin or Luteinizing Hormone/Progesterone Bioavailability In hCG-related cases, various management strategies have been proposed. If hCG was administered at inappropriate time points, delaying the oocyte retrieval of the second ovary or carrying out oocyte retrieval later have been proposed [9]. Alternatively, a second, rescue course of hCG with repeat oocyte retrieval of intact follicles 35–37 hours later has been proposed, even though data to support its effectiveness are scarce and only based on case reports or case series. Inconsistency of reported outcomes [10] and the inherent limitations of available research studies’ design should be taken into consideration when proposing the latter approach to patients. Patient education and information provision is key in avoiding hCG-related causes of failure to retrieve oocytes. Yet medication errors are not always unavoidable. Some fertility clinics have employed office hCG administration, selection of premixed hCG preparations or assessing serum hCG levels prior to oocyte retrieval in order to further reduce the chance of hCG-related failures.

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No eggs retrieved with adequate response to COS Check timing of trigger injection ≥36hrs from trigger

+60°, (4) UCA > –45°

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and a full bladder allows the clinician to identify the uterus and cervix easily using transabdominal ultrasound. As yet there is no strong evidence that filling the bladder does reduce ET difficulty, but it undoubtedly allows the practitioner to track the ET catheter more clearly.

Ultrasound Guidance Until the introduction of ultrasound, the main embryo transfer method was the “clinical-touch” technique. The clinician would feed the catheter through the cervix blindly and use proprioceptive feedback to decide whether the catheter tip was in the uterus appropriately. Ultrasound offers the clear advantage of seeing the catheter along its course through the cervical canal and into the uterus. Furthermore, ultrasound ensures that the practitioner, and the patient, is confident that the catheter tip is in the uterine cavity and yet is not too close to the fundus. Clinicians were initially slow to accept ultrasoundguided ET, but as the volume and quality of studies grew, so did the evidence that clinical pregnancy rates improved with ultrasound guidance [5]. It is now a fairly well-established concept and it is the authors’ opinion that ultrasound guidance should be the default approach to embryo transfer.

ET Catheter Selection The ET catheter has evolved over the years, especially with the advent of coaxial and echogenic catheters that permit easier visualization with ultrasound. The fragile embryo is loaded into the tip of the catheter, protecting it on its passage into the uterus, and so choosing which catheter to use might influence outcome. Furthermore, the way in which the catheter moves through the cervix could be important in terms of distortion and biomechanical stress applied to the tip of the ET catheter. The two different types of available catheters have been the soft and firm catheter systems. The advantage of firmer, malleable catheters is that they can be molded to suit the cervical canal shape, overcoming resistance more easily. While the soft catheter produces marginally higher clinical pregnancy rates, it is also more likely to fail to negotiate the cervix in difficult procedures [6]. Clinical practice around the world varies considerably based on the clinician and clinic’s preference and this makes it

nearly impossible to undertake detailed research in this area.

Personal Choice It is the authors’ personal preference to insert a reasonably stiff outer sheath as far as the internal cervical os under ultrasound guidance. If an empty (dummy) internal catheter is inserted, it should only go as far as the tip of the external sheath, i.e. just into the uterine cavity, so minimizing any disturbance to the endometrium. The outer sheath remains in the cervical canal and only then is a fresh inner catheter loaded with the embryo(s). This approach ensures that the passage of the loaded catheter will be straightforward. It also means that any impediment to the passage of the catheter is recognized and addressed before the embryos are removed from their incubator.

Predicting a Difficult Embryo Transfer It is a good maxim that if you can make a procedure as simple and straightforward as possible, it is easier to resolve difficult situations when they are encountered. This means understanding the potential pitfalls and being prepared for them. In terms of embryo transfer, this means using a catheter with which the practitioner is familiar, having a reasonably full bladder to maximize ultrasound visualization and straighten the utero-cervical angle (with an anteverted uterus), introducing an unloaded inner catheter (if that is unit practice) and using ultrasound to follow the path of the catheter in to the uterine cavity.

Past Embryo Transfers Attention to previous embryo transfers and any mock transfers undertaken prior to a cycle will help inform the clinician of the risk of a difficult transfer.

Cervical Dilatation The value of cervical dilatation is contentious as the longevity of the effect is questionable. An RCT found that women who had had two previous difficult transfers achieved higher pregnancy rates if cervical dilatation was performed one to three months prior to the planned ET [7]. Whether this was due to making the procedure easier or the potentially beneficial effects on implantation of endometrial trauma is open to debate, and while cervical dilatation is safe, it is not without financial cost.

Management of a Difficult Embryo Transfer

The Mock Embryo Transfer The purpose of the mock transfer is to simulate as closely as possible the conditions of the real ET procedure, but the key question is whether the mock ET is a good way to predict a difficult transfer. What we can say is that patients who had a mock transfer had fewer difficult transfers and improved clinical pregnancy rates [8]. The timing of the mock ET has always been contentious. With ovarian stimulation, not only does the uterine position change, particularly in cases of retroversion, but also the measured utero-cervical canal length changes in almost 60 percent of cases [9]. The mock ET has been shown to increase the chance of fluid migration with increased uterine contractility, but when performed close to the ET day itself, there was no detrimental effect on implantation and clinical pregnancy rates [10]. It seems counterintuitive to stimulate the uterus twice if it may be detrimental. Importantly this potential drawback can be mitigated by performing the “mock” transfer at the time of the embryo transfer, ensuring the mock soft inner catheter does not advance beyond the tip of the external catheter (i.e., not beyond the internal cervical os).

Use of a Stylet A soft catheter will naturally follow the line of least resistance, but if negotiation of the utero-cervical canal is difficult, a malleable stylet might be used. This allows shaping of the catheter and more force to be applied to overcome resistance, but it runs the risk of causing pain (and hence myometrial contractions) and in extreme circumstances, creating a false passage. Elective use of the malleable stylet in the first instance where a difficult transfer is expected is preferable to having to resort to this measure after the soft catheter fails to negotiate the cervix. In a retrospective case-control study, Tiboni et al. [11] found a statistical reduction in implantation rates and live birth rates in women where a stylet was required. A stylet may also be helpful in cases when a pinhole external cervical os is identified as it will widen the external os. Once this has been achieved, it is usually straightforward thereafter to pass a transfer catheter up the utero-cervical canal without the need for a stylet.

Use of a Tenaculum If the utero-cervical angle is very acute, it could make passage of the catheter difficult. However, application

of a tenaculum to apply traction on the cervix, in order to straighten out the UCA, will inevitably cause myometrial contractions. Intuitively this cannot be beneficial, but there are no RCTs to confirm or refute this. However, application of the tenaculum can be painful and on these grounds alone it should be avoided if possible.

Embryo Transfer under Sedation To our knowledge there are no studies on the benefit or otherwise of sedation at the time of embryo transfer. However, in women who cannot tolerate speculum examination, sedation does allow the practitioner to take his/her time with optimal positioning of the speculum and ET catheter. It follows that if a previous transfer has proved difficult, sedation before embarking on the transfer would be prudent, especially if additional maneuvers may prove necessary.

Transmyometrial Embryo Transfer If a malleable stylet cannot be threaded through the utero-cervical canal and a tenaculum cannot straighten the canal sufficiently to allow the passage of the catheter, then transmyometrial embryo transfer (TMET) using a Towako needle (or similar device) is an option. The needle is introduced through a vaginal fornix under ultrasound guidance in much the same way as a transvaginal egg retrieval but the needle passes through the myometrium and the tip is positioned in the endometrium. The inner needle is withdrawn and the loaded embryo catheter passed through the lumen of the external needle. It is a straightforward procedure, but passing a needle through the myometrium will inevitably cause myometrial contractions, which intuitively can only be detrimental. In more than 10 years of practice, the author (MG) has only ever had to resort to this on three occasions. Khairy et al. [12] reported their experience of 46 TMETs over 10 years and found no statistical difference in live birth rates compared to persevering with a transcervical transfer. With such small numbers, clearly a meaningful RCT is virtually impossible but it is the authors’ opinion that TMET should be a procedure of last resort.

Avoiding a Difficult Embryo Transfer The key to avoiding a difficult embryo transfer is meticulous patient preparation and attention to the specific contributory factors discussed in this chapter.

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Top Tips • • • • • • • • • • •

Every effort should be made to avoid “difficult” transfers and all critical factors carefully considered. Take heed of the difficulty of past ETs. If a difficult ET is envisaged, consider prior sedation to minimize the distress to the patient. Optimize patient position, cervical preparation and bladder volume before ET to ensure patient comfort. Ultrasound guidance will result in easier transfers with improved outcomes. Soft catheters should be used when feasible. Mock transfers allow better preparation for difficult transfers. Excessive cervical mucus should be removed to potentially decrease bacterial contamination and mucus plugging of the catheter. Avoid stylets and application of a tenaculum if possible. A transmyometrial ET is a procedure of last resort. The procedure should be performed in a minimum amount of time.

Suggested Standard Operating Protocol (SOP) Clinical SOP for embryo transfer includes clinical protocol and equipment list

Suggested Audits Incidence of difficult embryo transfer – stratified by clinicians, embryologist and patient groups Patient information sheet Embryo transfer

References 1. R. Matorras, R. Mendoza, A. Expósito and F. J. Rodriguez-Escudero. Influence of the time interval between embryo catheter loading and discharging on the success of IVF. Hum. Reprod. 2004;19(9):2027–30. 2. D. R. Listijono et al. An analysis of the impact of embryo transfer difficulty on live birth rates, using

a standardised grading system. Hum. Fertil. (Camb.) 2013;16(3): 211–14. 3. L. Craciunas, N. Tsampras and C. Fitzgerald. Cervical mucus removal before embryo transfer in women undergoing in vitro fertilization/intracytoplasmic sperm injection: A systematic review and meta-analysis of randomized controlled trials. Fertil. Steril. 2014;101(5):1302–7. 4. H. N. Sallam et al. Ultrasound measurement of the uterocervical angle before embryo transfer: a prospective controlled study. Hum. Reprod. 2002;17 (7):1767–72. 5. J. Brown, K. Buckingham, W. Buckett and A. M. AbouSetta. Ultrasound versus ‘clinical touch’ for catheter guidance during embryo transfer in women. Cochrane Database of Systematic Reviews 2016;Issue 3. Art. No.: CD006107. doi: 10.1002/14651858.CD006107.pub4 6. W. M. Buckett. A review and meta-analysis of prospective trials comparing different catheters used for embryo transfer. Fertil. Steril. 2006;85(3):728–34. 7. N. Prapas, Y. Prapas, Y. Panagiotidis, S. Prapa, P. Vanderzwalmen and G. Makedos. Cervical dilatation has a positive impact on the outcome of IVF in randomly assigned cases having two previous difficult embryo transfers. Hum. Reprod. 2004;19(8): 1791–5. 8. R. Mansour, M. Aboulghar and G. Serour. Dummy embryo transfer: A technique that minimizes the problems of embryo transfer and improves the pregnancy rate in human in vitro fertilization. Fertil. Steril. 1990;54(4):678–81. 9. K. L. Miller and J. L. Frattarelli. The pre-cycle blind mock embryo transfer is an inaccurate predictor of anticipated embryo transfer depth. J. Assist. Reprod. Genet. 2007;24(2–3):77–82. 10. K. O. Katariya et al. Does the timing of mock embryo transfer affect in vitro fertilization implantation and pregnancy rates? Fertil. Steril. 2007;88(5):1462–4. 11. G. M. Tiboni, E. C. Colangelo, E. Leonzio and E. Gabriele. Assisted reproduction outcomes after embryo transfers requiring a malleable stylet. J. Assist. Reprod. Genet. 2012;29(7):585–8. 12. M. Khairy, H. Shah and M. Rajkhowa. Transmyometrial versus very difficult transcervical embryo transfer: efficacy and safety. Reprod. Biomed. Online 2016;32(5):513–17.

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How to Manage the Patient Who Fails to Produce a Semen Sample Alpha K. Gebeh and Mostafa Metwally

The Subject of the “Problem”

Normal Erectile Function and Ejaculation

Introduction

Erection and ejaculation are initiated by a combination of physical and psychological stimuli and controlled mainly by parasympathetic stimuli.

Many interventions in assisted conception are focused mainly on the female patient and the critical role of the man is usually when a semen sample is required. The success of such interventions is time dependent and therefore it is crucial that the man can produce a sample within a specific timeframe. In the vast majority of cases, this is not a problem, but it is not uncommon for some men to suffer temporary erectile dysfunction or ejaculatory failure during treatment with potential financial loss and psychological stress for the couple. Moreover, prolonged time intervals between oocyte collection and insemination or microinjection, especially when this exceeds six hours, are associated with adverse in vitro fertilization (IVF)/intracytoplasmic sperm injection (ICSI) outcomes [1]. Knowledge of this can place the male partner under considerable pressure to produce a sample within a specific timeframe, which can further compound the problem.

Table 27.1 Risk factors for erectile and ejaculatory dysfunction

Classification Psychogenic

Causes/Associations Anxiety Stress Mental health problems Psychological trauma Relationship problems Family/social pressures Depression

Endocrine disorders

Hypogonadism Hyperprolactinemia Diabetes mellitus

Vascular

Peripheral arterial disease Hypertension

Etiology and Risk Factors The prevalence of inability to produce a semen sample in men who have previously had no history of sexual dysfunction is approximately 1 in 200 and therefore many units will expect to see at least three to five cases per year. Although there may be an organic cause to the problem, in many cases the overriding etiology is related to stress or psychological factors that adversely impact the man’s ability to maintain erection or ejaculate. A summary of risk factors is given in Table 27.1 [2]. Where an underlying problem is identified, appropriate referral for relevant specialist input should be considered. For instance, where underlying psychosexual factors are thought to be contributory, counseling from a psychosexual counselor may provide some benefit. In practice, temporary erectile dysfunction or anejaculation during treatment is usually unexpected and can occur even in those without preexisting risk factors.

Veno-occlusive disease Traumatic injury Drug induced

Antiandrogens Antihypertensives Antidepressants Heroin Marijuana

Medical disorders

Renal insufficiency Hepatic insufficiency Sleep apnea Dyslipidemia

Neurological

Neuropathy Spinal cord injury Multiple sclerosis Stroke

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Table 27.1 (cont.)

Classification

Causes/Associations

Genitourinary

Peyronie’s disease Cavernous fibrosis Benign prostatic hyperplasia Lower urinary tract symptoms

Iatrogenic

Post radiation

When activated, nerves that enter the corpora cavernosa release nitric oxide and acetylcholine, causing smooth muscle relaxation and influx of blood flow to tissues via cGMP and cAMP dependent pathways. The activation of sympathetic tone following ejaculation leads to smooth muscle relaxation, which results in the penis returning to a flaccid state. Phosphodiesterase-5 in penile tissue contributes to maintaining flaccidity by breaking down cGMP. The ejaculatory reflex is controlled by sympathetic nerves in the spinal cord and receives somatic input from nerves supplying the gland penis. Higher centers also play a role in ejaculation, though the exact mechanisms are not fully understood. Given the complex interplay between physical and psychological stimuli in erection and ejaculation, it is not surprising that men undergoing stressful fertility treatments may not be able to produce a semen sample.

Strategies/Solutions and Management Options The management of unexpected cases where the male partner is having difficulty producing a semen sample on the day of oocyte retrieval will depend on the facilities available within the unit, expertise of medical staff, time required to inseminate the oocytes and patient’s choice. However, as insemination or ICSI will need to be performed within six hours of oocyte recovery [1], the most practical options would include conservative measures, oocyte cryopreservation, surgical sperm retrieval (SSR) or cycle cancellation.

Pretreatment Risk Assessment and Sperm Cryopreservation Men who have a history of temporary erectile dysfunction or who are deemed at risk should ideally

have been identified at the initial infertility work-up. A focused sexual history should be taken prior to assisted conception treatment and where the risk is judged to be significant for example, in those with prior history of erectile dysfunction, they should be offered pretreatment cryopreservation of sperm.

Conservative Measures It is important to provide adequate support for the man and his female partner to minimize the stress associated with failure to produce semen as further stress risks compounding the problem. Sensitive inquiries or history of potential contributory factors may help direct treatment strategies. Audiovisual aids and female partner support: Initial basic measures that can be employed include allowing the man more time to produce a semen sample with or without the aid of audiovisual support or to ask the female partner for assistance in helping him produce a semen sample within the unit. Semen sample production at home: In men deemed at high risk for having difficulty producing a sample in the unit on the day of oocyte collection, the man could be allowed home to produce a semen sample within his own familiar environment or possibly using his own sourced audiovisual support as this may reduce the stress associated with timed sample production within an IVF unit. In men who find it difficult to produce a semen sample by masturbation, a nonspermicidal condom at home could be useful. Such condoms need to be validated as some brands of condoms sold as non-spermicidal are in fact toxic to sperm [3]. A risk with this approach of sample production at home is that specimens may be interfered with or may be clinically ineffective by the time they are received at the center. In the United Kingdom, although centers may use sperm produced by a man at home, the treating center should follow protocols to confirm the man’s identity, that the sample was produced by him and within one to two hours of receipt in the center and that it was not interfered with. The couple should consent to the use of such gametes produced at home and that this information is documented in the patient records. Timed intercourse: With regards to intrauterine insemination (IUI) treatment, one Cochrane review concludes that there was no difference in cumulative live birth rates, multiple pregnancy rates and other

The Patient Who Fails to Produce a Semen Sample

adverse effects for couples with unexplained subfertility undergoing IUI when compared with timed intercourse [4]. An option therefore is for the couple to have timed intercourse at home if the man finds the assisted reproduction technology (ART) environment stressful and contributory to his inability to produce a semen sample. This option may not be advisable in IVF as the superovulation induced with gonadotrophins risks multiple pregnancies.

Drug Treatment Sildenafil inhibits cGMP-specific phosphodiesterase5, leading to accumulation of cGMP in the corpus cavernosum and accumulation of blood in the penis, thus maintaining erection following sexual arousal. It also appears to reduce the refractory period following ejaculation. It is given as a 50 mg tablet and the man can attempt to produce a sample within 30 to 60 minutes of injection. Onset of action may be delayed if taken with food. Common side effects include nausea, vomiting, flushing, headaches and visual disturbance. Contraindications include unstable angina, recent stroke and myocardial infarction, those already receiving nitrates, renal and liver disease and uncontrolled hypertension. Sexual stimulation is still necessary for Sildenafil to produce its desired effects. The female partner could therefore play a role with sexual stimulation if this appears to be a challenge for the man, or audiovisual support may be useful. Other phosphodiesterase-5 inhibitors include Avanafil, Tadalafil and Vardenafil.

Mechanical Vibrators and Electroejaculation Penile vibratory stimulation (PVS) is a recognized treatment for anejaculation in men with spinal cord injury, but it can be also used in men to facilitate semen production for IVF/ICSI. The benefits are, however, less pronounced than in spinal cord injury patients. In a study by Saleh et al. [5], only 20 percent of men without a prior history of sexual dysfunction were successful in producing a sample using PVS. Generally, the device is placed on the glands penis and set at specified amplitude and frequency, which activates the ejaculatory reflex, and an antegrade ejaculation is initiated. It is vital that the urethra is milked manually to retrieve as many sperm cells as possible. When successful, PVS induces an ejaculation within a few minutes of stimulation. Subsequent attempts are made following a brief interruption of

stimulation if the initial attempt fails. Relative contraindications for PVS are severe inflammation of the penile skin, untreated hypertension or cardiac disease, as well as a patient’s inability to comprehend instructions about the procedure. Other methods include prostatic massage and rectal probe electroejaculation, though these are more invasive approaches. However, assisted conception centers often lack the familiarity, training, equipment and experience regarding PVS and electroejaculation methods and generally resort early on to surgical sperm recovery methods when faced with the problem of anejaculation [6, 7].

Urinary Sperm Retrieval in Retrograde Ejaculation Retrograde ejaculation is not uncommon in men with organic conditions such as diabetic neuropathy, multiple sclerosis or previous prostrate surgery, or with pharmacologic treatment of lower urinary tract symptoms with alpha-receptor blockers. Where possible, the medication should be discontinued following input from the appropriate specialist if the benefit of resolved retrograde ejaculation outweighs the benefit the medication provides. In cases of neuropathy-induced, iatrogenic or idiopathic retrograde ejaculation, sympathomimetic drugs have had modest success. These approaches are generally not feasible in the acute setting when a semen sample is required for ongoing IVF/ICSI/IUI treatment. Where this is the case, management aims at increasing the number of motile and viable sperm in the ejaculate by attempting to decrease urinary acidity, which is toxic to sperm. This can be accomplished by emptying the bladder and instilling a sperm wash medium or a sterile culture medium for gametes into the bladder prior to attempting sample production. The bladder is emptied after ejaculation and the urine obtained is centrifuged and the pellet examined for sperm. The recovered sperm can be used for treatment or cryopreserved.

Surgical Sperm Recovery (SSR) Sperm can be surgically obtained by epididymal sperm aspiration using percutaneous or microsurgical methods, or by extracting sperm from testicular tissue and used for ICSI. The choice of method will usually depend on the available experience and familiarity of the surgeon with each procedure. However, there is insufficient evidence to recommend any specific sperm retrieval technique over the other in men undergoing ICSI. It is therefore recommended to use

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the least invasive and simplest technique available [10]. Spermatic cord block can be feasible for percutaneous epidymal sperm aspiration (PESA) and testicular sperm aspriation (TESA), which makes it suitable for emergency situations but for open procedures such as scrotal explorations and multifocal testicular sperm extraction (TESE), a general anesthetic would be required, which may limit their use in the acute setting. SSR can be complicated by pain, swelling, infection, hydrocele and hematoma formation. Hematoma formation is more common with percutaneous than open techniques.

Oocyte Cryopreservation Oocyte cryopreservation is a technique that was relatively ineffective at first due to the problem of crystal formation and cell injury, but more recently modifications to it have led to better success rates. Pregnancy rates are at least similar with both vitrification and slow freezing techniques, but vitrification offers a simple, rapid and inexpensive method with acceptable survival rates after thawing. However, oocyte survival rates following thaw may not be as high as those for embryos [8]. Pregnancy rates per cycle of frozen embryo replacement (FER) have improved over the years and are comparable to rates from fresh transfers [9]. As such, oocyte cryopreservation, especially where a large number of oocytes was retrieved, is a viable option and avoids cycle cancellation and discarding of oocytes. The cryopreserved oocytes could then be used in a subsequent treatment cycle using either donor or partner sperm. Where donor sperm is used, appropriate counseling is vital to discuss the implications of using donor gametes.

Cycle Cancellation Although there are numerous useful options, their implementation would depend on the couple’s choice, availability of resources, training and expertise and medical conditions. There are situations where there is no option but to cancel the cycle and discard oocytes. This comes with adverse financial and emotional consequences to patients, but is nonetheless an option where all else fails or where other options are not feasible.

Conclusion Many management options exist for temporary erectile and ejaculatory dysfunction. These need to be tailored to the patient’s individual circumstances and choice, the expertise and resources available, the

time interval when sperm is required in order not to compromise IVF/ICSI success rates and the type of treatment the patient is undergoing. It most cases, the conservative measures described earlier would be successful and only occasionally do more invasive approaches become necessary.

References 1. M. Jacobs, A. M. Stolwijk and A. M. M. Wetzels. The effect of insemination/injection time on the results of IVF and ICSI. Hum. Reprod. 2001;16: 1708–13. 2. A. W. Pastuszak. Current diagnosis and management of erectile dysfunction. Curr. Sex. Health. Rep. 2014;6: 164–76. 3. M. J. Tomlinson, A. Naeem, J. F. Hopkisson and B. Campbell. Brief communication: Assessment and validation of nonspermicidal condoms as specimen collection sheaths for semen analysis and assisted conception. Human Fertility 2012;15: 140–3. 4. S. M. Veltman-Verhulst, E. Hughes, R. O. Ayeleke and B. Cohlen. Intra-uterine insemination for unexplained subfertility. Cochrane Db. Syst. Rev. 2016. 5. R. A. Saleh, G. M. Ranga, R. Raina, D. R. Nelson and A. Agarwal. Sexual dysfunction in men undergoing infertility evaluation: a cohort observational study. Fertil. Steril. 2003;79:909–12. 6. A. Kafetsoulis, N. L. Brackett, E. Ibrahim, G. R. Attia and C. M. Lynne. Current trends in the treatment of infertility in men with spinal cord injury. Fertil. Steril. 2006;86:781–9. 7. M. Fode, S. Krogh-Jespersen, N. L. Brackett, D. A. Ohl, C. M. Lynne and J. Sonksen. Male sexual dysfunction and infertility associated with neurological disorders. Asian J. Androl. 2012;14: 61–8. 8. D. Glujovsky, B. Riestra, C. Sueldo, G. Fiszbajn, S. Repping and F. Nodar et al. Vitrification versus slow freezing for women undergoing oocyte cryopreservation. Cochrane Db. Syst. Rev. 2014. 9. D. Adamson, C. Calhaz-Jorge, J. A. C. Alcala, C. De Geyter, J. de Mouzon and T. D’Hooghe et al. Assisted reproductive technology in Europe, 2011: results generated from European registers by ESHRE. Preliminary results. Hum. Reprod. 2014;29: 54–5. 10. A. van Peperstraten, M. L. Proctor, N. P. Johnson and G. Philipson. Techniques for surgical retrieval of sperm prior to intra-cytoplasmic sperm injection (ICSI) for azoospermia. Cochrane Db. Syst. Rev. 2008.

Chapter

28

Prevention and Management of Incubator Failure Charissa Watchorn and Julia Paget

Gas Supply Failure The correct supply of gas to the internal incubator chamber is imperative to enable it to function effectively by providing an appropriate CO2 concentration and hence pH to support gamete and embryo development. Oocytes and embryos have limited ability to regulate their internal pH [2], so severe problems with the gas supply to the incubator may result in detrimental effects as observed by failure of fertilization, cleavage or further development. However, any minor discrepancies with the supply of gas such as leakages can often be difficult to detect and may go unnoticed for long periods of time, yet manifest in reduced outcomes of in vitro fertilization (IVF) as identified by key performance indicator (KPI) data.

Prevention Upon initial installation of a laboratory gas system, all elements of the supply from gas cylinders through to the incubator, including associated regulators, manifolds and pipework, should be thoroughly checked. A secure storage area for a number of spare cylinders, which is restricted to authorized personnel and allows protection from extreme weather conditions and potential damage, is required to ensure not only the supply but also the quality of gas provided. Subsequently, any deliveries of new cylinders should be monitored and, prior to changing empty cylinders, it is essential that the correct gas for the incubator is reconnected. Autochangeover units are highly advisable while switching the supply from an emptied cylinder to a full one during use, enabling an uninterrupted gas supply to the incubator. Alarm systems are available that can be connected to cylinders and incubators to alert staff when a tank is empty or if the gas supply fails. Some laboratories prefer an incubator environment of reduced oxygen to improve clinical outcomes [3] while others function only with carbon dioxide (CO2). Using both systems concurrently allows for a back-up if one system fails. In the low oxygen system, nitrogen

may be used to control oxygen concentration through cylinders, generators or utilizing vapor from liquid nitrogen storage tanks. This additional equipment requires careful monitoring as it provides another source for potential failure, subsequently leading to incorrect culture conditions from excess or insufficient nitrogen. This also has safety implications if leakage occurs into the laboratory. The regular inspection of the entire gas system, including levels and pressures, the changeover unit and any filters, should be incorporated into routine laboratory checks, with records kept alongside daily logs of incubator CO2 or pH. For benchtop incubators, a common source of problems with the gas supply is blockage of the tubing leading from the water bottle. This can be due to an accumulation of water in the tube or a kink, or a failure in the filter attachment. Therefore it is prudent to apply physical checks daily as part of routine laboratory set-up and shutdown procedures. Filters that aim to improve the quality of gas supplied to the incubator should be changed at regular intervals and care taken when reattaching them to ensure proper connections. The use of media containing phenol red either for culture or as a control dish within an incubator can also provide a visual indication of pH. This is helpful following any cleaning or maintenance procedures prior to returning incubators to clinical use, but should not be solely used as a method of detecting deviations.

Identification The media in a dish containing phenol red will appear yellow in acidic conditions, noting a high level of CO2 and pink in alkaline conditions indicative of a low CO2 environment. Although a useful visual indicator, once noted, the culture environment will have already been affected and subsequently any tissues cultured within it. However, with the introduction of clear media into many laboratories, to reduce toxicity, this visual aid would be absent. Some incubators display

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the percentage gas levels on their exterior, which if recorded daily can pick up discrepancies in the supply. However, this should not be accepted as true readings and independent regular recordings using a CO2 analyzer or pH meter are recommended. These devices must be carefully calibrated and threshold levels set in order to provide an accurate method of monitoring the culture environment. Although direct measurement of pH within a culture dish provides the best and most appropriate indication of culture conditions, this is often difficult to measure. CO2 measuring is more routinely employed instead, which requires the pH to be calculated with laboratory altitude factored in. The specific type and composition of media used should also be considered, as manufacturers can vary in their recommendations of optimal pH when using their media. Continual monitoring of gas, which can be attached to an automatic telephone dialer system to alert staff, is widely available for conventional larger box-type incubators. However, this is more difficult to install for incubator designs with small chambers, which poses an increased risk that staff should be aware of [1].

Resolution CO2 and pH within the incubator environment may fluctuate slightly over time, and sometimes simple adjustments in incubator set points are sufficient to correct this. If the incubator display or a CO2 or pH measurement appears to be outside of acceptable thresholds, then immediate action should be taken to attempt to locate the source of the problem and resolve it. Any gametes or embryos in culture should first be carefully relocated to another suitable incubator. The attached pipework or tubing, along with any filters, should be examined to note any obvious cracks or defects and to ensure the connections are tight. A commercial detection spray or soapy water may be sprayed at connections from cylinder to incubator to detect leakages in the pipework by observing for bubbling. The cylinders should be checked to ensure that the appropriate gas is connected, a sufficient level remains and that the pressures are correct. Confirm that any auto-changeover system is functioning correctly. Where no problem with the inlet of gas to the incubator can be noted, the fault may be with the internal gas sensors, which could require replacement. In this instance, the incubator supplier or servicing company should be contacted to assess the equipment and repair it.

Power Supply Failure When electrical power to an incubator fails, the subsequent loss of gametes and embryos, mostly due to a fall in temperature but also CO2, is inevitable unless it is resolved within a limited timeframe. The negative impacts of reduced temperatures are well documented, such as its effects on meiotic spindle stability, cleavage and possibly embryo metabolism [4]. Therefore the prompt resolution of an incubator power failure is imperative to avoid irreversible damage.

Prevention Given the severe consequences of a power failure, it is essential that a back-up power supply is available to incubators through a battery and/or generator to minimize risk. If the building offers an uninterrupted power supply, then incubators should be top priority for the circuit. An independent alarm system consisting of continuous temperature monitoring should be installed and attached to an automatic telephone dialer alerting a member of staff to respond outside of working hours. During routine cleaning, care should be taken to ensure that incubator power leads are not dislodged, plugs detached from sockets or switches inadvertently turned off. Hardwiring of incubators within the laboratory eliminates some of these sources of accidental loss of electrical power; however, it also reduces the ease of moving equipment, which can itself be a problem. All incubators should be validated prior to commissioning for use as part of the quality management system. This includes the temperature mapping of dishes within different locations and determining a critical timeframe in the event of power failure for the recovery of gametes and embryos, in association with an emergency call-out alarm system. A back-up incubator is an essential piece of kit if a power failure is confined to an isolated incubator or section of the laboratory.

Identification Power failure is usually easily identifiable as most incubators have LED displays to indicate that it is switched on and connected to an electrical source. If these lights are not visible, the power switches on both the incubator and socket should initially be checked along with any attached power leads. An alarm system with continual independent

Prevention and Management of Incubator Failure

monitoring can detect any loss of power outside of working hours, either by attaching this directly to the input supply or as a result of the internal temperature falling below a set point. Any electrical surges or transient losses of power can be identified if observed daily as part of routine laboratory checks; displaying the readings in a graph format is particularly useful.

Resolution Immediately check if the problem is confined to one plug socket or distributed throughout the laboratory. If it is a localized failure, then plug the incubator into a live socket, which may be outside the laboratory, by means of an extension cable if necessary. If not, then check any relevant circuit breakers or fuse boxes, and if no immediate solution can be found to return power, then an external power supply is necessary. Some desktop incubators are equipped with an internal back-up battery that may provide power for a couple of hours, but a generator is preferable. It is imperative that any failed incubator is not opened, even to check on the embryos cultured, until an alternative functioning incubator is available to move dishes to. Large boxtype incubators can retain heat better than desktop incubators due to the larger internal chamber volume [5].

Equipment Malfunction Another source of incubator failure is malfunction of the equipment itself. Any deviations from the optimal culture conditions of temperature or pH can cause damage to gametes and embryos.

Prevention Upon initial installation of incubators into the laboratory it is essential that correct installation, operational and performance qualification procedures are performed as part of the clinic’s quality management system. Incubator calibration and validation is important prior to commissioning the new equipment for clinical use and also following any further servicing episodes. Service contracts should be set up through the manufacturer, distributor or a company specializing in incubators with appropriate third-party agreements put in place. Alongside this, maintenance and cleaning schedules, taking into account manufacturer

instructions, are necessary in order to ensure that the incubator remains in good working order. A system for monitoring when routine incubator maintenance and servicing are due is essential so that the correct timeframes are adhered to. Smaller benchtop/topload incubators provide faster recovery of environmental variables following door opening, although there is no consensus on their advantage based upon clinical outcomes [5]. They are, however, more sensitive to fluctuations in external environment so their location within the laboratory is important. Placing incubators away from sources of air flow and heat minimizes the risk of the internal environment being affected. Performing daily physical checks of the incubators as part of routine set-up procedures can identify any potential problems at an early stage. Monitoring of temperature and CO2 or pH must be incorporated into the laboratory and relevant set points established with acceptable levels above or below which action should be taken to rectify the deviation. It is important that all measuring devices are properly calibrated and fit for purpose, as the detection of a malfunction could be attributed to either the equipment or the device used to measure it. Staff training is essential with standard operating procedures (SOPs) established for equipment use. All laboratory staff should be knowledgeable in the use and function of the incubator, including an awareness of basic troubleshooting, where manuals can be located, how to identify if the incubator is not functioning correctly and subsequently what to do and who to contact in an emergency situation. Management of incubator use is also an important aspect of its correct functioning, the patient volume and workflow must be considered in order to keep the frequency of door openings to a minimum. In the event of identifying an incubator failure, having another functional incubator available in which to transfer any dishes or tubes is essential. If this is impossible, then contingency plans that may involve other departments or local IVF laboratories should be considered.

Identification Methods for identifying equipment malfunction include those covered in the previous sections.

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Independent pH/CO2 and temperature monitoring along with an alarm system is a crucial resource for the identification of deviations within the incubator environment. As part of a clinic’s quality management system, audits, KPIs and monitoring charts are important measures for the ongoing observation of an incubator’s performance. These are essential to detect reduced laboratory or clinical outcomes that would then require investigation, including but not limited to the assessment of equipment function.

Resolution Upon identification of a potential problem with the functioning of an incubator the gametes and embryos cultured inside should immediately be moved to an alternative incubator. Relevant SOPs and instruction manuals should be consulted to

resolve any common problems, with the necessary toolkits and spare consumables located if needed. If a fault cannot be easily resolved by basic troubleshooting, then the servicing company should be contacted for advice and to attend to assess and repair any potential fault. If no equipment fault is identified but a noticeable fall in KPIs beyond acceptable levels has been identified, then it is prudent to assess the workload of the incubator in order to better manage its usage and reduce door openings. Temperature mapping of various positions within an incubator is useful to identify any differences that may have an effect on embryo development and outcomes of treatment [4]. Checking of the incubator cleaning schedule is also recommended with additional cleaning and decontamination procedures performed to eliminate infection as a source of reduced function.

PROBLEM WITH INCUBATOR IDENTIFIED

Move embryos/ gametes to back up incubator if possible; if not possible, avoid opening door

GAS SUPPLY PROBLEM

ELECTRICAL PROBLEM

MALFUNCTION PROBLEM

Check main cylinders are full and connected

Check if another plug socket can be used and move to new socket if possible (remove embryos from incubator prior to movement)

Refer to Standard Operating Procedure for use of equipment and/or work through instruction manual troubleshooting guide

Check manifold is open to correct tank and pressure set points are correct

Check all lines are sealed use soap spray

Gas sensors inside incubator may be faulty and should be checked by supplier

Check circuit breaker and fuse box

Use back up generator/ battery as alternative if possible, while problem resolved

Call power supply company

Figure 28.1 A flowchart to troubleshoot incubator failure

Contact service company for advice or repair

Prevention and Management of Incubator Failure

Summary

References

It is important for each IVF laboratory to identify the hazards associated with the use of its own incubators and compile risk assessments with contingency plans for the event of a failure. The implementation of the outlined preventative measures can reduce but not eliminate the risk of an incubator failing, while methods for identifying deviations in function such as installing continuous monitoring systems with an emergency call-out alarm allow for quick detection. Once a problem has been identified with an incubator, it is essential that this is immediately resolved (Figure 28.1). Appropriate incubator SOPs must be in place and staff should be competent in knowing how to deal with any potential problems to avoid harm to patients’ gametes and embryos cultured within this vital piece of laboratory equipment.

1. Human Fertilisation and Embryology Authority. HFEA Clinic Focus article (January 2015): Incidents Case Study – A Cautionary Tale on the Use of Benchtop Incubators. Available at www.hfea.gov.uk/9457.html#it em4, Accessed September 10, 2016.

Suggested Standard Operating Protocol (SOP) SOP for incubator management

2. J. E. Swain. Optimizing the culture environment in the IVF laboratory: Impact of pH and buffer capacity on gamete and embryo quality. Reprod. BioMed. Online 2010;21:6–16. 3. S. Bontekoe, E. Mantikou, M. van Wely, S. Seshadri, S. Repping, and S. Masenbroek. Low oxygen concentrations for embryo culture in assisted reproductive technologies (Review). Cochrane Database of Systematic Reviews 2012;7. 4. G. Anifandis. Temperature variations inside commercial IVF incubators J. Assist. Reprod. Genet. 2013;30:1587–8. 5. J. E. Swain. Decisions for the IVF laboratory: Comparative analysis of embryo culture incubators. Reprod. BioMed. Online 2014;28:535–47.

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Troubleshooting Poor Survival after Oocyte and Embryo Cryopreservation Victoria Ryder and Dawn Yell

Introduction A high-quality cryopreservation program is a critical component of the fertility clinic’s service provision. A successful cryopreservation program will ensure that oncology and social fertility preservation patients are provided with the very best chance to have a family, using their own gametes or embryos, in the future. For patients undergoing fertility treatment now, the ability to consistently achieve excellent survival of cryopreserved embryos allows a clinic to confidently implement a single embryo transfer policy. Furthermore, frozen embryo transfer cycles are less of a physical and financial burden for patients than fresh in vitro fertilization (IVF) cycles. With the recent advancements in blastocyst culture and vitrification, there is now evidence to suggest that birth outcomes (live birth rate, birth weight) of frozen embryo transfers are better than for fresh IVF cycles. As a result, many clinics are now considering an elective “freeze-all” policy for some IVF patients. Of course, a clinic must not implement an elective freeze-all policy without sufficient clinical evidence and, more importantly, a high level of justified confidence in its cryopreservation program. For a clinic to achieve and maintain superior cryopreservation results it must quickly identify, investigate and correct any downward trends in survival rates, as part of a robust quality management system (QMS). This chapter covers: • How to investigate a single case of poor survival • How to identify and investigate a downward trend in cryopreserved oocyte/embryo survival rates • Corrective actions • Preventative measures.

Investigating an Isolated Case of Poor Survival Unfortunately, it is not possible to recover an oocyte or embryo that has degenerated during the freezing and

thawing process; however, lessons can be learned and steps can be taken to optimize survival in future cases. Each clinic should set its own criteria for investigating individual clinical cases with poor survival, based on their protocols and results. Survival rates are generally accepted to be poorer for slow freezing methods compared to vitrification, for both oocytes and embryos. Similarly, poorer survival rates are expected for cleavage stage embryos versus blastocysts. As an example, the authors would recommend investigating cases when three or more vitrified blastocysts are warmed with less than 50 percent cell survival. Many factors can influence oocyte and embryo survival at various stages in the freezing and thawing process, and consideration of each individual factor forms the basis of the investigation (Figure 29.1). As a guide, the following actions should form the immediate investigation. The writers assume that a policy to only cryopreserve embryos of good quality is in place.

Patient History Research the patient history, identify if they have had any samples thawed previously, and the survival rate of those samples. Post-thaw survival is related to lipid membrane permeability and flexibility. In theory, some oocytes and embryos may be more susceptible to cryo-damage than others. However, at this juncture, we have no test suitable to assess the quality of oocytes, or indeed their membrane integrity.

Freeze/Thaw Materials Identify whether other samples have been successfully thawed from the same batch of freezing and thawing media and loading device. Check the expiry dates of the used batches. Identify whether any new consumables have been recently introduced.

Operator and Protocol Identify the freeze/thaw technicians, the protocol followed and their experience and competence with that

Poor Survival after Oocyte and Embryo Cryopreservation

Operator experience

Cooling rate

Time of freeze

Cryopreservation media

Biological variability

Freezing/ vitrification

Figure 29.1 The oocyte/ embryo cryopreservation process and factors that may influence survival

Loading device Lab consumables

Human error Room temperature

Slow freezer

Transfer into/ out of storage

Storage

Warming rate

Storage temperature

Operator experience

Timing

Thawing/ warming media

Biological variability

Thawing/ warming

Human error

Room temperature Lab consumables

Temperature

Lab consumables Post thaw culture

pH Temperature Gases

Culture media

protocol. Each practitioner’s competence should be reviewed annually/biennially against the standard operating procedure (SOP). Where new SOPs are introduced, each technician should be trained and supervised by a senior, experienced practitioner until signed off as competent. SOPs should be audited annually to assess suitability. When oocytes and embryos are imported from another center, the relevant SOPs should be obtained and followed, under supervision and with prior training where possible.

Equipment/Environment Temperature monitoring of key equipment in the laboratory should be performed and recorded at least every morning. Identify any possible nonconformities or changes in temperatures or gas levels in incubators, heated stages, lab air and storage vessels at the time of the freeze and thaw. Consider testing for volatile organic

compounds and compare results with regular air quality monitoring. Identify whether any other samples have been successfully thawed from the same storage container. Identify any changes in lab/storage facilities during the time that samples were in storage/frozen/thawed.

Subsequent Culture Conditions If the sample had survived immediately after thaw, but deteriorated later, there may have been a problem with the culture dish, culture media or incubator used post thaw. Identify whether the development of any fresh embryos using the same incubator/consumables batches is satisfactory.

Human Factors Identify the circumstances in the laboratory at the time of the freeze/thaw, consider the workload and any distractions that may have taken place. Encourage

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Table 29.1 Example key performance indicators (KPIs) and benchmarks for cryopreserved oocytes and embryos. This is not an exhaustive list. Adapted from Alpha Scientists [1]

KPI

Calculation

Benchmark

Vitrified oocyte survival rate

# oocytes survived/# thawed

85% (95% for donors < 30 years)

Vitrified oocyte fertilization rate

# oocytes fertilized/# oocytes injected

No more than 10 percentage points lower than that for the comparable population of fresh oocytes at the center

Vitrified oocyte implantation rate

# fetal hearts/# embryos transferred

No more than 10–30% (relative) lower than that for the comparable population of embryos from fresh oocytes at the center

Vitrified blastocyst survival rate

# embryos survived > 75%/# embryos thawed

95%

Vitrified blastocyst implantation rate

# fetal hearts/# embryos transferred

The same as for the comparable population of fresh embryos at the center

an open culture with regards to incident reporting and learning from human error.

Investigating a Downward Trend in Survival Rates

Reporting

Before an in-depth investigation can take place, first perform a preliminary investigation to assess the quality of the data. Identify a timeframe to investigate. This should be the interval between the last on-target and the first off-target KPI result and may cover several months. For the selected time period, investigate the following.

Report the case as an internal nonconformity; record each step of the investigation and the outcome. Where an adverse incident has occurred, report the case to the appropriate external regulatory body within the required timeframe. Remember that at the end of a single investigation, there may not be a clear root cause. A robust QMS with detailed reporting and regular review of nonconformities will help to highlight any potential trends at an early stage.

Identifying a Downward Trend in Survival Rates Centers should perform regular analysis of key performance indicators (KPIs) and nonconformity reports in order to monitor their performance and identify any downward trends in survival rates. KPI benchmarks should be set based upon professional body recommended standards and the clinic’s own patient population and previous year’s performance (Table 29.1). Lower limits of acceptable performance should be set, as well as a policy of immediate investigative action if that lower limit is reached. Performance of individual operators in freezing and thawing should also be examined.

Volume of Data Consider the total number of samples thawed and the number of patients involved over a defined period of time. If either is significantly lower than previous periods, consider re-analysis after a further defined time period/number of cases.

Bias Identify any outliers – individual cases with very poor survival caused by a singular, known event (e.g., imported embryos damaged in transit) – and exclude these from the analysis.

Patient Demographics Investigate the patient demographics, for example, age at the time of freezing/cause of infertility. If patients are older, they may be expected to have a poorer outcome.

Poor Survival after Oocyte and Embryo Cryopreservation

Embryo Demographics Consider the stage at which embryos were frozen and the relevant proportions, and whether this is similar to previous time periods. For blastocyst embryos, one would expect slightly better survival rates for day 5 embryos than day 6. Similarly, blastocysts within the zona may survive at higher rates than fully hatched blastocysts. Also, if the clinic thaws embryos/oocytes cryopreserved using both slow freezing and vitrification methods, consider analyzing these separately. If these preliminary investigations indicate that the data are good quality and unbiased, therefore indicating a true downward trend, consider the following.

Changes in Laboratory Identify any changes in the clinic/laboratory immediately prior to/during the period of interest. This may include, but is not limited to, changes to: • Freezing/thawing protocols • Culture/freeze/thaw media • Laboratory equipment • Premises • Staff.

Changes to Upstream Factors Give consideration to wider changes in clinic policies and procedures, for example, stimulation regime. However, keep in mind that these changes are unlikely

to solely affect post-thaw survival. Most likely there would be a concurrent effect on fresh embryos and oocytes, including fertilization rates, fresh embryo pregnancy rates, embryo quality, blastocyst development rate and embryo utilization rates. Due to the delay caused by storage, any effects on fresh embryos may have been observed much earlier. Examine the original freeze dates for the samples that were thawed within the period of interest.

Common Factors Try to identify any other common factors across all or the majority of cases. Use the basic template for investigating an individual case of poor survival – all the same factors apply (Figure 29.1). If the in-house team can identify potential causes, it may be prudent to consider employing an external consultant for further investigations.

Corrective Actions and Preventative Measures Where causative or contributory factors are identified in an investigation, corrective actions should be taken immediately. These may include: • Retraining/refresher training of staff (team or individuals) • Introduction of new/revision of existing SOPs • Introduction of new procedural checklists • Servicing/replacement of equipment • New/replacement batch consumables

Table 29.2 A summary of the common causes of failure of gametes and/or embryos to survive freezing and thawing, the preventative measures that should be employed to minimize the risks and the associated SOPs required

General causes

Specific causes

Preventative measures

SOP

Environmental factors

Air quality deviations

• Regular monitoring of particles, volatile organic compounds (VOCs) and microbial contamination of laboratory and workstation air

• Laboratory air quality monitoring

Room temperature deviations

• Daily measuring of room air temperature with defined acceptable limits. Action taken to correct temperature if outside those limits

• Daily laboratory equipment monitoring

Storage temperature deviations

• 24-hour monitoring and alarming of storage vessels • Speed when transferring material between storage locations/entering/removing from storage

• Daily laboratory equipment monitoring • Storage vessel failure

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Table 29.2 (cont.)

General causes

Specific causes

Preventative measures

SOP

Culture equipment/ consumables

Incubator gas/pH/ temperature deviations

• Daily monitoring of incubator and heated stage temperature and gas supply/pH in incubators with defined acceptable limits. Alarms and action taken to correct deviations outside those limits • Regular servicing of critical equipment • Equipment validated before use

• Daily laboratory equipment monitoring • Equipment servicing and maintenance • Laboratory equipment validation

Consumables batch problem

• Robust method for batch traceability of all laboratory consumables and media, including in-use dates and expiry dates • Use reputable, industry-recognized suppliers • Recommended to use CE-marked medical devices wherever possible • Mouse embryo assay, sperm survival, bacterial endotoxin tested consumables

• Traceability of gametes, embryos and consumables in the laboratory • Supply and use of laboratory consumables

Procedural error

Deviation from SOP

• Process validation before SOP implemented • Annual procedural audit to confirm suitability of SOP • Annual competency assessment of operators against SOP

• Laboratory protocol validation • Oocyte/embryo freezing/ vitrification • Oocyte/embryo thawing/ warming • Procedural audit • Staff training and competency assessment

Operator skill

Limited experience/new technique

• Rigorous induction and training procedures for new staff and trainees • Annual competency assessment of operators against SOP • External SOPs followed/ culture media used when samples received from other centers

• Staff training and competency assessment • Receiving gametes/ embryos from another center

Sample quality

Poor-quality sample less likely to survive

• Strict freeze criteria • Regular IQC and EQC of embryo morphology grading to reduce interoperator variability

• Morphological assessment of embryos • Selection of embryos for cryopreservation • Quality control in the laboratory

Human factors

Wrong culture media used, e.g., vitrification media used for warming and vice versa

• Operator vigilance • Visual cues, e.g., different-colored packaging for different types of media • Possible witnessing of adding media to dishes/samples

• Oocyte/embryo freezing/ vitrification • Oocyte/embryo thawing/ warming • Witnessing in the laboratory

Poor Survival after Oocyte and Embryo Cryopreservation

Any changes should be immediately communicated with all affected staff and the date of implementation should be recorded in the investigation report. Table 29.2 summarizes the common causes of failure of oocytes and embryos to survive freezing and thawing, the preventative measures that should be employed to minimize the risks and the associated SOPs that should be in place. Once actions have been implemented, performance should be monitored through subsequent KPI analysis. If no improvement is observed within a defined timeframe, reinvestigate and consider external consultation and ceasing oocyte thaw cycles/frozen embryo transfers as applicable.

Conclusion Achieving and maintaining a successful cryopreservation program is intrinsically linked to a clinic’s wider quality management system, of which all staff and

managers should be committed participants. The first steps toward a high-quality program are regular analysis of individual operator and laboratory-wide performance and detailed reporting of individual clinical cases of poor survival. Trends in performance should be identified through minimum quarterly KPI analysis and quickly investigated, with prompt and appropriate corrective actions taken and dedicated follow-up analysis. Unfortunately, while this does not make possible the recovery of already damaged oocytes and embryos, lessons should be learned and steps should be taken to optimize survival in the future.

Reference 1. Alpha consensus meeting on cryopreservation key performance indicators and benchmarks: Proceedings of an expert meeting. Reproductive BioMedicine Online 2012;25:146–67.

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The “Lost” Embryo in the Laboratory Virginia N. Bolton

Introduction The human oocyte is a spherical cell, enclosed in a glycoprotein matrix coating, the zona pellucida, and measuring 0.1 mm in diameter. Following fertilization, the embryo remains the same size throughout the first four days of development, with the cells (blastomeres) within the embryo halving in size at each cleavage division. Only once the blastocyst has formed does the embryo begin to increase in size, with expansion of the blastocoelic cavity, reaching a maximum diameter of around 0.25 mm before hatching from the zona pellucida at around six days after fertilization, when implantation begins. Each of the numerous procedures carried out in the assisted conception laboratory poses technical challenges inherent in the handling and manipulation of microscopic oocytes and embryos, whether moving them from one culture dish to another, from culture dish to embryo transfer catheter, from culture dish to cryopreservation device, or from cryopreservation device to culture dish. Moreover, at multiple stages during procedures, culture dishes containing oocytes or embryos will be moved to and from the incubator and the microscope stage, and for the purposes of cryopreservation, oocytes and embryos will be transported in their cryopreservation devices between containers of liquid nitrogen. Each process involving moving and manipulation of individual oocytes and embryos, or the dishes and devices in which they are cultured or cryopreserved, inevitably holds an element of risk. During all procedures, it is essential that standard operating procedures (SOPs), which will have been prepared with a view to minimizing recognized risks, are followed meticulously. When an incident resulting in loss of an oocyte or embryo occurs, root cause analysis is undertaken, and any contributory factors that are identified are addressed, with revision of the SOPs as necessary. Any loss is serious, but those resulting from process failure, or from failure of

individuals to follow prescribed processes, must be viewed seriously. No SOP can account for human error and lapses in concentration that may lead to avoidable errors and possibly result in catastrophic losses. Many tasks in the laboratory are repetitive, and distraction while performing them may result in errors where, for example, incorrect media or reagents are used, wells of culture dishes are missed during preparation so that aliquots of culture medium and/or oil overlay are overlooked, or even where oocytes are missed during insemination. It is essential that practitioners are aware of and remain vigilant and alert at all times to the risks of attention lapses and involuntary automaticity. Even where all precautions are taken, accidents may occur and, by definition, these are unpredictable and difficult to prevent. This chapter considers errors that may lead to the loss of oocytes or embryos and the frequency with which they may occur, along with measures that may be taken to minimize risk.

Incidents Resulting in Loss of Oocytes and Embryos In the United Kingdom, it is a regulatory requirement that all incidents resulting in the loss of oocytes or embryos are reported to the Human Fertilisation and Embryology Authority (HFEA). During 2015, a total of 73 such incidents was reported (Table 30.1), occurring during a total of more than 72,000 treatment cycles, representing an incident rate of around 1 per 1,000 treatment cycles.

Moving Oocytes and Embryos between Dishes The device used for the collection and movement of oocytes and embryos between dishes is usually a purpose-designed, commercially available, handheld pipetting device to which is attached a disposable

The “Lost” Embryo in the Laboratory

Table 30.1 Summary of incidents resulting in the loss of oocyte(s) or embryo(s) reported to the HFEA during 2015

Nature of incident

No

Process or procedure

Technical difficulty/failure

28

Not recovered from micro pipette (18*); pipette tip not secure (2); damaged during use of micro pipette (4); loss during denudation (2); difficulty with cryopreservation device (1); other (1)

Failure to follow process

26

Discarded in error (6); oocytes not inseminated (2); cryopreservation error (3); exposure to incorrect media/ reagents (5); error in preparation of culture dish (3); oocyte/ embryo not accounted for (4); other process error (3)

Physical accident

18

Dish knocked (4); dish dropped (6); cryopreservation device dropped (3); pipette tip knocked (5)

Culture failure

1

Bacterial contamination

*

Multiple causes (see Table 30.2)

Table 30.2 Incidents where oocyte(s)/embryo(s) were lost when moved between dishes, and suggested preventative measures

Nature of incident

Details (number of incidents)

Technical difficulty associated with use of micropipette

Oocytes/embryos not recovered from micropipette (15)

Depress pipette plunger fully and flush pipette tip fully before aspirating oocyte/embryo Aspirate medium into “dead space” of pipette tip before aspirating oocyte/embryo, providing additional medium for expulsion

Pipette tip not secure (2)

Method of securing tip into pipette varies between models; check tip is securely attached: (a) pull tip gently close to point of attachment, ensuring tip does not detach; (b) flush and aspirate medium into “dead space” (see above) before attempting to aspirate oocyte/embryo

Air bubbles expelled with gamete/embryo (1)

Aspirate medium into “dead space” of pipette tip before aspirating oocyte/embryo, providing additional medium for expulsion and preventing expulsion of air

Damaged (4)

Ensure internal diameter of disposable tip selected is larger than the diameter of the embryo being aspirated

Denudation (2)

When denuding multiple oocyte-cumulus complexes (OCCs) simultaneously, select number to denude according to size of OCC to ensure each oocyte can be observed throughout the denudation process; too many OCCs in one denudation drop may mean oocytes are obscured by density of dispersed cumulus cells

Oocyte (s)/embryo(s) not accounted for (4)

When aspirating and expelling oocytes/embryos into and from pipette between culture drops, count them “in” and “out,” ensuring all are accounted for; check final culture drop/dish for correct total number before returning to incubator; check none remain in emptied drop/dish before discarding

Failure to follow process

Preventative measure(s)

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Table 30.3 Incidents resulting from process failure

Incident Oocyte(s)/embryo(s) discarded due to error

Oocytes not inseminated

Cryopreservation error

Exposure to incorrect media/reagents

Other

Process

Measures to avoid failure

2

Embryo should have been frozen along with sibling embryos, but was overlooked and discarded

2

Two oocytes discarded prior to use

1

Embryo should have been checked for suitability for freezing, but dish was discarded in error

Keep meticulous records to ensure good inter-operator communication; confirm all documentation against contents of all dishes before discarding

1

Uninjected eggs mixed with injected eggs in same dish, and therefore had to remain uninjected

Keep meticulous records to ensure good inter-operator communication; confirm documentation and contents of dishes before adding or removing oocytes or embryos

1

Egg of uncertain provenance in tube of follicular fluid

Confirm and document that all processes are completed at the end of all procedures

2

Total number of eggs collected recorded incorrectly; some oocytes not inseminated

Keep meticulous records to ensure good inter-operator communication

Sperm not introduced into IVF insemination drop at insemination

Always confirm visually that insemination step has been completed

2

Cryopreservation device overlooked and not placed in liquid nitrogen

1

Cryopreservation device overlooked and not placed into freezing machine

Confirm and document that all processes are completed at the end of all procedures; count, confirm and document that the correct number of cryopreservation devices are moved from one phase to the next

2

Culture dish lacking culture medium

1

Culture dish lacking oil overlay

5

Embryos exposed to inappropriate medium/reagent for stage of development/procedure

3

Embryo transfer – embryologist lost concentration; error in loading catheter and embryo lost

plastic or glass pipette tip, the internal diameter of which is appropriate to the procedure to be undertaken. Thus, for denudation (removal of cumulus cells from the oocyte prior to intracytoplasmic sperm injection [ICSI], or following IVF insemination to assess fertilization), a pipette tip is used that is only slightly larger than the diameter of the oocyte (around 145 µm), to facilitate “stripping” all the adhering cumulus cells away from the zona pellucida; for cleavage-stage embryos, a larger diameter tip (around 170 µm) is used; while for handling the

Vigilance, care and concentration are essential at all times

Vigilance, care and concentration are essential at all times

larger expanding or expanded blastocysts, tips that are used will have a diameter of around 290 µm. Reported losses associated with aspiration of oocytes and embryos into such pipette tips, and suggested preventative measures, are listed in Table 30.2. One recognized risk is the “knocking” or flicking” of a pipette tip, into which oocyte(s)/embryo(s) have been collected in order to move them between dishes, against the side of the culture dish. In order to minimize loss from such an accident, it is

The “Lost” Embryo in the Laboratory

Table 30.4 Incidents where oocyte(s)/embryo(s) were lost due to an accident

Nature of accident

Details (number of accidents)

Pipette tip knocked

5

Vigilance, awareness of risk and care; aspirate oocyte(s)/ embryo(s) into micropipette individually when moving between dishes

Dish dropped

6

Vigilance, awareness of risk and care, particularly when wearing gloves

Dish knocked

4

Vigilance, awareness of risk and care when carrying and moving dishes between microscope and incubator; optimize laboratory design to ensure unobstructed routes between workstations and incubators

Cryopreservation device dropped

3

Vigilance, awareness of risk and care when collecting cryopreservation devices from storage tanks; if not securely attached to the holding device, cryopreservation devices may spring free and cryopreserved material warm and perish before the device can be retrieved

Minimization of loss/risk

recommended that oocytes and embryos are aspirated into the pipette tip and moved between dishes individually.

Failure to Follow Process Procedures should be laid out clearly and unambiguously in SOPs in such a way that it is clear what each process entails, and what checks should be made and documented at its completion. Wherever there is a likelihood of catastrophic loss, such as failure to complete insemination of IVF culture drops effectively, checks should be put in place for visual confirmation, by a second witness if appropriate, together with written confirmation that the check has taken place, to avoid such errors. Failures to follow processes that have been reported and that resulted in loss of oocytes and embryos are listed in Table 30.3. These are almost universally the consequences of lapses in concentration, and while practitioners must be alert and guard against this, human error can never be eliminated. Measures to reduce errors include ensuring that when tasks are shared by more than one practitioner, there is clear and efficient communication between individual staff members, with adequate systems of documentation and record-keeping in place so that each practitioner can, at a glance, understand precisely what stage in any given process each set of oocytes and embryos has reached.

Accidents Accidents are, by definition, unforeseen and unpredictable. However, where there is a known risk, as evidenced by precedents (see Table 30.4), practitioners must be made aware of these and care taken to avoid them. Uncluttered workstations and laboratory areas between workstations and incubators will minimize the risk of knocking and dropping dishes; care should be taken in handling dishes when wearing gloves; practitioners must practice and be fully competent and confident in gauging the distance between the ends of pipette tips and the edges of culture dishes.

Conclusion Detailed SOPs, drafted carefully to encompass all confirmation, checking, witnessing and documentation steps, must be adhered to if the risks of incidents resulting in the loss of oocytes or embryos are to be minimized. When tasks are shared between several members of a team, systems must be sufficiently robust to ensure that all practitioners within the laboratory are able to identify easily and accurately the stage within a process that each cohort of oocytes or embryos has reached. The loss of an oocyte or embryo is distressing for all involved, whether directly or indirectly. It is of paramount importance that experiences of such incidents are shared, both

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among staff members within the center where they occur, and among the wider professional community, in order that lessons may be learned, and steps taken to minimize each risk when it is identified. Ideally, such losses will never occur, but in reality, it is the direct experience of dealing with the consequences of such incidents that heightens awareness, and that usually provides the motivation and incentive to ensure such lapses are not repeated.

Suggested Standard Operating Protocol (SOP) SOP for confirmation, checking, witnessing and documentation steps

Acknowledgements Thanks are extended to Erin Barton and Paula Nolan at the HFEA for their help in collating data for the incidents reported to the HFEA during 2015.

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What to Do When There Is a Suspected Infection in the IVF Laboratory Philippa Tucker and Julia Paget

Fortunately, infection within the in vitro fertilization (IVF) laboratory is rare, with studies reporting an infection rate of between 0.35 percent and 0.69 percent [1, 2, 3]. This low rate is predominantly due to effective quality management systems that reduce risk and the use of media containing antibiotics to target common infections. Nonetheless, when extrapolated to the hundreds of thousands of assisted reproduction technology (ART) cycles performed in Europe each year, this equates to thousands of potential incidences of infection. Therefore, remaining vigilant is extremely important, particularly when identification is often too late and the source of infection is ambiguous.

Types of Infection The most common forms of infection within the IVF laboratory are fungal and bacterial (including mycoplasma), and one survey of 32 experienced laboratory directors reported an almost equal split between bacterial and fungal (yeast or mold) infections [4]. However, the incidence of bacterial infection may be affected by the antibiotic used, with another study reporting that prior to using Gentamycin in the culture media, the bacteria Escherichia coli constituted 59 percent of infections identified, while the fungus Candida caused 25 percent of infections, but once Gentamycin was introduced only Candida infection was identified [3]. Although Gentamycin may prevent many bacterial infections, those reported in Table 31.1 have all been identified in IVF culture systems. Based on figures from [2] and [3]: more than 50 percent are relatively common, 25–50 percent are common, 5–25 percent are relatively rare and less than 5 percent are extremely rare.

Sources of Infection Infection within the IVF laboratory can originate from a host of different sources including patients, staff and the laboratory environment (culture media, consumables, equipment, air conditioning systems

and imported gases). However, the most common source, causing approximately a third of culture media infections, is seminal fluid [3, 4]; a further 23 percent of infections are transmitted by staff using improper aseptic technique [4], while the overlay oil may also cause contamination [4]. As seminal fluid is not sterile and easily contaminated, it is unsurprising that semen causes most infections within the laboratory. Studies using culture assays and polymerase chain reaction (PCR) revealed that around 65 percent of semen samples are contaminated with bacteria [2, 5], which can occur from poor personal hygiene involving the hands, penis, nose, hair or even the pot and persist into the embryo culture system [2]. However, the studies found different bacteria to be prevalent within the semen samples, perhaps because PCR is more sensitive than colonization studies and can detect bacteria that cannot be cultured easily. The colonization study found Staphylococcus epidermis (from the skin) to be most common [2] while the PCR study determined gram positive anaerobic cocci to be most prevalent [5]. Mycoplasma has also been identified in approximately 17 percent of semen samples [2] as has Candida trachomatis with detectable DNA levels found in 13 percent of semen samples analyzed, including from patients who tested negative for Chlamydia. The next most common source of infection is staff working within the IVF laboratory; this is particularly true if aseptic techniques are not used. Sources of contamination include the mouth, nose, skin and hair. When aseptic techniques are not employed during culture dish and tube preparation, media can become contaminated. This is problematic if the stock media become infected or if, in rare cases, commercial media are contaminated upon receipt, as these media are used in the preparation of numerous patients’ gametes and the culture of their embryos. Another environmental contaminant is Aspergillus, an airborne fungus that colonizes central heating and

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Table 31.1 Types of microbial infection within ART

Type of microbe

Species

Common sources

Prevalence

Appearance

Bacterial

Escherichia coli

Semen, skin

Relatively common

Rods

Mycoplasma hominis

Semen, follicular fluid

Relatively rare

Cocci

Nonhemolytic streptococci

Semen, follicular fluid

Relatively rare

Cocci (spheres)

Achromobacter anthropi

Opportunistic pathogen found in water

Extremely rare

Rods

Agrobacterium species

Opportunistic pathogen found in plants

Extremely rare

Rods

Citrobacter koseri

Opportunistic pathogen found in human digestive system

Extremely rare

Rods

Klebsiella pneumoniae

Normal flora of skin (staff and patients)

Extremely rare

Rods

Pseudomonas species

Opportunistic pathogen found in hospitals

Extremely rare

Rods

Stenotrophomonas maltophilia

Semen

Extremely rare

Rods

Candida species

Semen, skin, follicular fluid

Common

Round to oval cells

Aspergillus terreus

Paraffin oil, heating and air conditioning systems

Extremely rare

Spore-like appearance

Fungal

air conditioning systems and has contaminated the oil overlay of culture dishes [3]. Once introduced into the laboratory, fungi grow in incubators, water baths, sinks, refrigerators and on surfaces that are not regularly cleaned, and colonize moist environments such as air filters, heat exchangers, humidifiers, water pumps, cooling units, ceiling tiles and even condensation on the windows. Finally, follicular aspirates are an additional source of bacterial and fungal contamination, with more than 99 percent of follicular fluid samples tested containing bacterial contaminants [2, 4, 6]. In 71 percent of cases, the same bacteria were isolated from the patient’s vagina, suggesting the fluid was contaminated when the needle used to aspirate oocytes from the ovarian follicles passed through the vaginal wall. Indeed, even in healthy women the vagina contains several different bacteria, including lactobacilli, Staphylococcus epidermides, Clostridium perfringens and ureaplasma. However, a further 29–35 percent were colonized by bacteria that had not been isolated from the patient [4, 6], suggesting these bacteria may come from the environment.

Identification Infection is initially identified visually, with the media changing color if it contains phenol red or appearing cloudy or turbid under a microscope. Upon closer examination under a high-power microscope, microorganisms may be observed. However, in some cases, there may be no visible signs of infection and the first indication may be an insemination dish containing all immotile sperm, global arrest of embryo development or embryo/oocyte degeneration within a culture dish. Indeed, low levels of bacterial contamination that are not immediately apparent can interfere with embryo development by metabolizing certain components of the media, disrupting the balance of constituents and releasing toxins. Mycoplasma is an example of a bacterial infection that inhibits growth, shifts pH, depletes substrates, induces oxidative stress, secretes harmful metabolites and affects nucleic acid and protein synthesis. Given the effect bacteria can have on the media, it is unsurprising that in the vast majority of infections

Suspected Infection in the IVF Laboratory

embryos appear dark and atretic [4]. Interestingly, bacterial Escherichia coli infections are more likely to be detected on day 1 post oocyte collection and fungal Candida infections on days 2–4 [3]. Furthermore, fungal contamination is more likely to affect multiple dishes, and a contaminated incubator may first be identified when several infected dishes are observed.

Consequences The consequences of an infection can be drastic and far reaching depending upon the source and number of patients affected. Bacterial infection can result in failed or low fertilization, poor or slow embryo growth and even embryonic arrest [3], with the majority of bacterial infections rendering embryos nonviable [4]. This may result in no transfer for the patient and if a transfer does occur, there is often a reduced pregnancy rate [4]. In contrast, Candida infection has been reported to have no impact on embryo development [1, 3] or clinical outcome [1], suggesting this form of infection is not as detrimental to embryonic development.

Resolution If an infection is identified within the lab, it is crucial to determine and contain it immediately (Figure 31.1). This is imperative to reduce the overall harm to the infected patient and prevent the infection from spreading to further patients or contaminating the media or equipment used in embryo culture. Initially the number of patients affected should be determined, as if this is more than one patient, the source of infection is likely to be from equipment, media or the environment rather than an individual patient. A fresh dish should be set up and equilibrated immediately. Once the dish is equilibrated, the affected eggs and embryos should be washed in media containing antibiotics using a fine pipette (to reduce the risk of transferring microorganisms from the contaminated dish) and moved to this new culture dish. While culture media do not contain antifungal components, fungal infection can be removed by repeated washing [1]. An aliquot of the infected media should be saved and sent for microbiology culture and sensitivity (M, C & S) testing along with seminal and follicular fluid or vaginal swabs if these are available. If the infection originated from one of the patients, the patient should be informed and antibiotics administered. Meanwhile, the incubator used for

culture should be cleaned and the patients’ gametes and embryos isolated in a separate incubator. If several patients are affected and equipment is thought to be the source of the infection, then the equipment should be cleaned and decontaminated, according to the manufacturer’s instructions using the inbuilt sterilizing cycle if appropriate. When decontaminating the incubator, ensure that all surfaces, the water pan and fan blades (which are extremely effective at dispersing spores) are cleansed. The cleaning solutions used should be checked to ensure that they do not react with the components of the incubator. OosafeTM (benzyl-akyldimethyl chloride), FertisafeTM (chlorine dioxide, hypochlorites, ozone and free radicals) and Fertisafe PlusTM (silver dehydrate) are all solutions developed specifically for use in IVF laboratories, and OosafeTM has been shown to be safer to embryos than solutions of 70 percent ethanol and 6 percent hydrogen peroxide [7]. Once decontaminated, the incubator should be wiped down with sterile water to remove any residual cleaning solution, which could volatize and contaminate cultures. If media are thought to be the source of infection, the media should be sent for testing the old batch should be removed from use and a new batch opened. If commercial media are used and infection prior to arrival in the lab is suspected, then it would be prudent to contact the supplier.

Prevention Preventing infection within the IVF laboratory is key, with efficient quality management systems being essential to minimize risk. These systems include training staff and assessing their competencies, using and maintaining appropriate equipment, including laminar flow hoods, and undertaking regular cleaning. Quality control should also be performed and regularly reviewed and should incorporate the use of standard operating procedures combined with audits and key performance indicators or quality objectives. Strict rules should be employed within the laboratory to minimize infection. Access should be limited to authorized personnel, who should wear appropriate clothing and footwear, cover their hair with a surgical hat and be bare below the elbow. Face masks may be worn; however, these may be ineffective at preventing infection transmission, but can reduce transmission from the technician to the sterile work surface when using horizontal laminar flow hoods, and they are essential in the absence of flow hoods. On entering and leaving the lab and immediately

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Infection identified

Determine number of patients affected

Resolution for embryo culture

Set up new dishes (with fresh media if contamination is suspected) and equilibrate in non-affected incubators for all affected patients

How to identify source of contamination

1 patient

More than 1 patient

More likely to be from individual patient or partner

More likely to be media or equipment

Send semen follicular fluid or vaginal swabs for M, C + S

Send aliquots of media or swab from affected equipemt for M, C + S

If positive, administer antibiotics. If negative, consider other sources of contamination

Consider opening fresh media battles and/or changing media batch and decontaminating equipment

Clean contaminated incubators

Once dishes have equilibrated, wash embryos through media containing antibiotics with a fine pipette

Transfer to new dishes. Isolate from unaffected patient’s culture dishes and contaminated equipment

Inform appropriate persons including, but not limited to, management, regulatory authorities and patients as appropriate Figure 31.1 Resolution of microbial infection within the IVF laboratory

before clinical procedures, hands and wrists should be washed thoroughly and dried. The laboratory should be regularly cleaned with suitable products that are sufficient to prevent fungal contamination. Embryology should be performed in an air-filtered environment and in Europe the EU Tissue and Cells Directive [8] states that air quality must be a minimum of grade D background and grade

C when manipulating eggs and embryos. This means eggs and embryos should, wherever possible, be handled in laminar flow hoods, preferably class II, which protects both gametes and embryos from the external environment, but also the operator from any potentially contaminated tissue or fluid. Techniques should be performed aseptically, particularly when aliquoting media and preparing dishes

Suspected Infection in the IVF Laboratory

and tubes for culture. Tubes and dishes used for culture should be examined thoroughly for cracks or flaws before use. Batches of media and consumables should be recorded, and when a new batch is opened, this should be noted to ensure media are not open for extended periods. Media should be used only within the manufacturer’s expiry date, and aliquoting the media into single-use tubes upon opening can help prevent contamination. If an infection is observed in the patient prior to IVF, this should be treated first; for example, if leukocytospermia is observed, the semen should be sent for M, C & S and, if required, appropriate antibiotics given to treat the infection prior to IVF. For all patients, detailed instructions on how to produce the sample should be given to ensure they follow strict hygiene protocols and help prevent contamination from other sources. If the infection persists, semen should be collected in an antibiotic-containing medium, and sperm washed and prepared using the swim-up method, which reduces bacterial contamination from 65 percent to 5 percent. Gradient centrifugation can be used, but this technique may not remove all bacteria and cannot completely remove Chlamydia [9].

Conclusion Although there are several sources of potential contamination within the IVF laboratory, fortunately many of the bacterial contaminants found in seminal fluid, from staff and within follicular fluid, rarely cause infection within the culture media. This is due to strict hygiene and cleaning procedures, the use of laminar flow hoods and air conditioning systems, the washing of gametes prior to insemination and the use of antibiotics within culture media. Nonetheless, although infection within the laboratory is unlikely, when it does occur, it can be extremely detrimental, and quick identification, isolation and treatment are required to prevent spread of the infection.

Suggested Standard Operating Protocol (SOP) Manage infection in the lab

References 1. A. Ben-Chetrit, O. Shen, A. Haran, B. Brooks, T. GevaEldar and E. J. Margalioth. Transfer of embryos from yeast-colonized dishes. Fertil. Steril. 1996;66(2): 335–7. 2. E. Cottell, J. McMorrow, B. Lennon, M. Fawsy, M. Cafferkey and R. F. Harrison. Microbial contamination in an in vitro fertilization-embryo transfer system. Fertil. Steril. 1996;66(5):776–80. 3. P. M. M. Kastrop, L. A. M. de Graaf-Miltenburg, D. R. Gutknecht and S. M. Weima. Microbial contamination of embryo cultures in an ART laboratory: Sources and management. Hum. Reprod. 2007;22(8):2243–8. 4. K. O. Polmeroy. Contamination of human IVF cultures by microorganisms: A review. J. Clin. Emb. 2010;13 (4):11–30. 5. A. A. Kiessling, B. M. Desmarais, H. Z. Yin, J. Loverde and R. C. Eyre. Detection and identification of bacterial DNA in semen. Fertil. Steril. 2008;90(5): 1744–56. 6. E. S. Pelzer, J. A. Allan, M. A. Waterhouse, T. Ross, K. W. Beagley and C. L. Knox. Microorganisms within human follicular fluid: Effects of IVF. PloS One. 2013;8 (3):e59062. 7. S. Catt, E. Lingham, W. Lee et al. A randomized trial investigating the effectiveness and safety of three IVF laboratory disinfectants. ESHRE 2013 conference, London. 8. Official Journal of the European Union. Commission Directive 192006/86/EC. Available at http://eur-lex .europa.eu/homepage.html. Accessed September 9, 2016. 9. N. Al-Mously, N. A. Cross, A. Eley and A. A. Pacey. Real-time polymerase chain reaction shows that density centrifugation does not always remove Chlamydia trachomatis from human semen. Fertil. Steril. 2009;92: 1601–15.

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Dealing with Practical Problems Encountered in PGD and PGS Junhao Yan and Zi-Jiang Chen

Definitions of PGD and PGS Preimplantation genetic diagnosis (PGD), introduced in 1989, was developed as an alternative to prenatal diagnosis for couples who are at risk of transmitting inherited diseases to their offspring. Later, in 1995, preimplantation genetic screening (PGS) was also introduced in some laboratories to screen aneuploid embryos for couples with normal karyotype. Nowadays, PGD/ PGS is to identify unaffected and/or euploid embryos in a cohort of embryos produced in an in vitro fertilization/intracytoplasmic sperm injection (IVF/ICSI) program [1].

Indications of PGD/PGS For couples who are carriers of monogenic diseases and do not want to risk termination of an affected pregnancy, PGD is the treatment of choice. PGD can be used for sex-linked diseases (e.g., hemophilia, Duchenne muscular dystrophy), autosomal recessive disorders (e.g., cystic fibrosis, thalassemias) or dominant Mendelian disorders (e.g., myotonic dystrophy, retinoblastoma). In addition, human leukocyte antigen (HLA) matching and chromosomal disorders such as reciprocal translocation, Robertsonian translocation, inversion and deletion may also be considered for PGD [2]. However, PGD is ineffective in diseases where the genetic components are unclear or the pathogenic mutations have not been identified. On the other hand, PGS, or aneuploidy screening, is considered when advanced maternal age (AMA), recurrent miscarriages (defined as three or more consecutive spontaneous miscarriages), repeated implantation failure (defined as three or more transfers of high-quality embryos or placement of four or more embryos without establishing a pregnancy) or severe male factor infertility is present in couples with normal karyotype. PGS has also been used in other indications, such as having a previous chromosomally abnormal pregnancy, to improve the chance of a healthy live birth from IVF/ICSI treatment [3].

While there seems to be no dispute over the benefits of PGD, the use of PGS remains controversial. In the meta-analysis by Mastenbroek et al. [4], nine randomized controlled trials comparing IVF/ICSI treatment with or without PGS were included and yet no evidence showed an increase in live birth rate after PGS. On the contrary, patients in the advanced maternal age group had a reduced live birth rate after PGS. With the advancement in molecular technology, embryo biopsy performed at the blastocyst stage and a cryopreservation policy that allows the transfer of euploid embryos to a synchronized endometrium, more clinical trials are under way to further evaluate the efficacy of PGS.

Biopsy Procedure and Timing Cell samples for PGD/PGS can be obtained from oocytes before and after fertilization or from embryos either at the cleavage or at the blastocyst stage. The biopsy procedure generally involves the breaching of the zona pellucida and then the aspiration of cell(s) of interest. Nowadays, most laboratories are equipped with non-contact laser for zona drilling, although mechanical and chemical means are still employed in some places. While the different timing of biopsy may confer clinical benefits as well as limitations, the procedure itself is invasive and technically demanding. The time of heat exposure during laser ablation of the zona pellucida, the duration of the cell biopsy process and the amount of cell material removed may all compromise the implantation and developmental potential of the embryo.

Polar Body Biopsy Polar body (PB) biopsy has been considered an alternative to blastomere biopsy because PBs are the byproducts from meiosis of the oocyte and thus do not pose similar ethical dilemmas as the genetic analysis of human embryos. Both PBs from a mature oocyte before and after its fertilization have to be obtained for PGD/PGS. As PBs are obtained early in the in vitro culture process, genetic results are often available to

Practical Problems Encountered in PGD and PGS

allow fresh embryo transfer. However, PB biopsy on every oocyte can be time-consuming and the genetic data obtained reflect only the maternal but not the paternal contribution.

Blastomere Biopsy Biopsy at the cleavage stage is still currently widely practiced. Usually on the morning of day 3, one to two blastomeres with visible nucleus are removed from every embryo containing six or more cells. The embryos are then returned to in vitro culture while the biopsied materials are being tested. With a two-day turnaround time for the results, blastocysts with normal results can then be selected for transfer or cryopreservation. The criticism on blastomere biopsy is that the single cell being analyzed may not be representative of the whole genetic information of the embryo due to the phenomenon of mosaicism in embryos at cleavage stage. Besides, even when only one cell is biopsied from an eight-cell embryo, it represents removal of 12.5 percent of the entire embryonic mass and so may be detrimental to the development of the embryo. Indeed, a number of studies have shown reduced blastocyst formation and implantation rates in embryos biopsied at the cleavage stage. Therefore, there is a trend toward trophectoderm biopsy in recent years [5].

Trophectoderm Biopsy In trophectoderm (TE) biopsy, 5–10 cells are usually removed from a blastocyst while leaving its inner cell mass undisturbed. Although mosaicism may still be present in blastocysts, more genetic material can be obtained in TE biopsy as compared to blastomere biopsy, allowing the PGD/PGS analysis to be more robust and reliable. However, IVF centers must have the capability of blastocyst culture and cryopreservation in order to offer TE biopsy. A good blastocyst vitrification program allows more time for the genetic diagnostic process. As some embryos are slower in growth and may not become blastocysts until day 6 of development, most laboratories perform TE biopsy for both day 5 and day 6 blastocysts. Euploid morulae on day 5 could reach blastocyst stage on day 6, with a similar implantation potential to a day 5 euploid blastocysts [6].

Noninvasive Sampling Sampling by noninvasive methods is the ideal option for PGD/ PGS to avoid the need for a direct biopsy of

embryonic cells. By obtaining blastocoele fluid (BF) or spent culture media (SCM), both of which have been shown to contain embryo DNA, whole genome amplification (WGA) could be performed for genetic analyses. However, contamination of the low volume of fluid available for analysis, requiring ICSI to bypass issues around sperm contamination, and the need to completely remove granulosa cells prior to the transfer of the embryo to blastocyst culture medium, are all significant considerations using this technique. Hence these methods require further mechanistic studies and clinical validation prior to their use in daily practice.

Diagnostic Techniques Used in PGD/ PGS Many molecular technologies have been developed and applied to PGD/PGS procedures. In the past, fluorescent in situ hybridization (FISH) was widely used to detect chromosomal abnormalities, while Sanger sequencing after polymerase chain reaction (PCR) was used to detect specific point mutations. The introduction of the whole genome amplification (WGA) method has increased the amount and type of information obtainable from a single or a small number of biopsied cells. The WGA products can then be applied onto different platforms according to various situations, which will be discussed in detail as follows.

Platform for Couples with Genetic Disorders For patients with Mendelian disorders, the selection of a suitable embryo for transfer is mainly based on molecular analysis after PCR amplification. To do this, it needs two informative short tandem repeat (STR) linked markers, flanking each side of the mutation site, to minimize the risk of misdiagnosis caused by allele dropout at any one locus or unexpected contamination. The disadvantage of PCR analysis is that an individual test has to be developed for each couple, which is time-consuming and expensive. Moreover, although WGA, nested PCR, reverse transcription PCR and real-time PCR are all available nowadays for the molecular analysis of embryos in PGD-monogenic disease cases [7], the PCR reaction used in the sample amplification may introduce artificial mutations into the sample, which may affect the accuracy of the diagnosis. To decrease the potential artificial error, the cycles of the PCR amplification have been limited according to the protocol of the

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kit used. Many other factors can also lead to misdiagnosis in PGD, such as contamination, allele drop-out and, as discussed later, mosaicism of the embryo. Given these considerations, quality control measures are crucial to prevent or detect possible contamination, allele drop-outs and ambiguity caused by embryo mosaicism. Genetic results must be carefully interpreted to avoid misdiagnosis.

Platform for Couples with Chromosome Disorders FISH is commonly used in chromosome analysis for embryos with Robertsonian translocation. In addition, chromosome-specific centromeric and telomeric probes can be designed to distinguish balanced embryos from unbalanced ones produced by translocation carriers, although these probes need to be tested in the patients first. FISH results of embryos can be influenced by many factors such as the operator’s skill in cell-spreading, the hybridization efficiency and the probe quality. Since PGD-FISH is mainly carried out on one or two blastomeres at the cleavage stage, the result is not so compelling for the limited biopsied cells. In the past decade, FISH has gradually been replaced by much higher resolution technology such as array comparative genomic hybridization (array CGH) and single nucleotide polymorphism (SNP) arrays. SNP arrays have the added advantage of identifying the parental alleles in the embryo as to exclude uniparental disomy (UPD). However, chromosomal microarray testing cannot distinguish embryos carrying balanced translocation from those with normal karyotype.

Platform for Couples with PGS Indications FISH was initially used in PGS to detect common aneuploidy of chromosomes 13, 16, 18, 21, 22, X and Y. Limited by the number of fluorochrome labels, different sets of fluorescent probes have to be used in multiple rounds of hybridization, and only up to 12 chromosomes can be assessed. Not surprisingly, several RCTs showed discouraging PGS-FISH outcome [8]. Fortunately, several new technologies have been developed to simultaneously analyze 24 chromosomes, including array CGH, SNP arrays and quantitative realtime PCR (qPCR). Chromosomal microarray-based analysis (CMA) such as array CGH and SNP array is the most often used aneuploidy screening method. It has been reported that PGS based on array CGH is associated with better IVF outcome when the same

number of embryos is transferred in both PGS and control groups [9]. Quantitative PCR is another aneuploidy screening method, which avoids the amplification bias of WGA and saves time, and allows blastocyst biopsy with comprehensive chromosome screening (CCS) followed by fresh embryo transfer.

Next-Generation Sequencing Recently, next-generation sequencing (NGS) is available for PGD/PGS after WGA. With known advantages of robust high-throughput and customizable parallel analysis of multiple samples in a single sequencing run, NGS has been applied in PGD to detect specific mutations of nuclear and mitochondrial genome. In PGS, both retrospective and blind validation studies have shown that NGS shares high concordance with array CGH in aneuploidy screening for the same WGA products [10]. In addition, sequencing has been shown to detect lower levels of mosaicism in trophectoderm biopsies than other diagnostic methods. Moreover, NGS can correct the potential bias caused by WGA and is therefore more sensitive in detecting chromosomal structural abnormalities of small fragments. The analysis of the output data relies on the well-known online databases, such as HGMD, Decipher, OMIM, dbSNP and ExAC, as a basis for variant detection and interpretation.

Mosaicism Embryo mosaicism is the presence of two or more cell populations of different genotypes within one embryo. In some specific cases, the CMA result may indicate an embryo being mosaic. This probably occurs when the cells biopsied do not carry the same chromosomal composition, implying that the embryo may contain both euploid and aneuploid cells. The precise incidence and degree of mosaicism in any embryo biopsied is unknown, although it is generally thought to be low. It will be unusual not to have any euploid embryos for embryo transfer. However, the clinical conundrum arises should only mosaic embryos be available. aCGH and NGS may detect mosaicism levels as low as 20 percent [11]. While the actual rate of mosaicism in blastocysts is not well defined, when NGS is performed, preliminary data suggest that 10–30 percent of blastocyst TE biopsies may be diagnosed as mosaic [12]. The clinical significance of chromosomal mosaicism diagnosed by PGS is not well delineated. As for the embryo’s robust self-correction process and the limited biopsy cells, embryos deemed mosaic by PGS

Practical Problems Encountered in PGD and PGS

have the potential to develop into a fetus that is chromosomally normal, chromosomally abnormal or mosaic to a lesser, greater or similar degree to that predicted by the biopsy results [13]. Application of the appropriate risk assessment in this process is a challenge. It has been reported that the transfer of embryos with mosaic 2, 7, 13, 14, 15, 16, 18 and 21 might pose the highest risk for trisomy or monosomy syndrome [14]. For couples who only have mosaic embryos available, genetic counseling is essential. They should be informed of the risk of mosaic embryo and the necessity of prenatal screening after transfer.

Quality Control Standards in PGD/PGS The ESHRE PGD consortium has recommended that PGD/PGS be carried out only in an accredited laboratory while HFEA in the United Kingdom has made this a mandatory requirement. PGD/PGS laboratories should conform to international laboratory standards such as ISO 15189 or its equivalent. Each platform in the laboratory needs to be validated before clinical application. Equipment needs to be on a regular maintenance program. Appropriate control samples and quality control measures need to be specified to ensure the accuracy of the results. Periodic analyses of diagnosis rates, contamination rates and root cause analysis of errors identified through auditing enable laboratories to identify good practice and potential problems. In reporting diagnostic/genetic results, at least two identifying information of the patients should be presented, such as the patients’ name, hospital number or unique patient number. The date of receipt of the biopsy samples and the date of report should be clearly stated. For single gene disorders, the report should have the disorder name, the gene involved and their respective OMIM. The mutation tested should be written using the latest Human Genome Variation Society mutation nomenclature along with the reference sequence used to identify the mutation. For chromosomal disorders, the chromosomal rearrangements should be described using the latest International System for Human Cytogenetic Nomenclature. Interpretative comments to indicate suitable embryos for transfer need to be clearly and correctly presented. An explanation of the results may be provided to describe the minimum criteria used to report a result, including the expected error rates. The report should be signed by the person(s) who carried out the diagnosis and authorized by an appropriately qualified senior staff member.

Strategies and Policies in PGD/PGS Programs For Women with Poor Ovarian Reserve or Poor Responders Some patients may have clear indications for PGD/ PGS but at the same time have a poor ovarian reserve and/or poor response to controlled ovarian stimulation. Consequently, they may have only one to three embryos after an IVF/ICSI treatment cycle. These patients should be carefully counseled regarding the decision whether to proceed with PGD/PGS. If the patient chooses to implant the embryo(s) untested, prenatal genetic testing should be advised to lower the risk of birth defect or genetic abnormality. However, if the patient chooses to proceed with PGD/ PGS, the odds for having a normal embryo among the limited number of embryos available for testing need to be clearly explained. Alternatively, one may suggest banking the embryos over two or more treatment cycles to increase the chance of having one or two embryos with normal genotype.

For Patients with Untested Cryopreserved Embryos Some patients may consider PGD/PGS for their cryopreserved embryos in storage from a past IVF/ICSI treatment cycle. Depending on the number and the developmental stage of these embryos, their thawing/ warming for a one-time testing or in combination with embryos from a new treatment cycle for PGD/ PGS should be feasible. One important factor that requires further patient counseling is the likelihood of having to cryopreserve again the already thawed/ warmed embryos. While healthy live births after repeated cryopreservation have been reported in the literature, the potential detrimental long-term effect is unknown and cannot be excluded.

Should PGD Always Be Coupled with PGS? In most PGD/PGS cases, the biopsied cells have to first undergo WGA. The amount of WGA product is often sufficient in quantity for both PGD and PGS processes. Some argue that patients requiring PGD are usually fertile and do not require aneuploidy screening. However, the chance of aneuploidy in embryos derived from young egg donors can be as high as 60 percent. Therefore, the option of concurrent PGS in women

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seeking PGD should be offered. In this way, one can be sure that the embryo to be replaced following PGD is euploid, and avoid the unfortunate situation of an aneuploidy pregnancy.

When the Result is Inconclusive or Ambiguous PGD/PGS results can be inconclusive or ambiguous when an embryo of poor quality is biopsied, the biopsy contains very few cells or the genetic material is lost during handling and processing, or very rarely, equipment fails or malfunctions. Depending on the nature of the null result, the embryo may be disposed of or subjected to biopsy again. This decision may need input from the different disciplines, including the embryology laboratory and clinical geneticists, to ascertain or negate the necessity of a second biopsy.

Conclusion Above all, PGD is an evolving technique that could end the transmission of genetic diseases or chromosome abnormalities from at-risk couples to offspring before embryo transfer in IVF/ICSI treatment. PGS could also improve the success rate of IVF/ICSI treatment in given patient groups. However, there are important issues relating to the practice of PGS and PGD that are still controversial and require more evidence before being routinely introduced. It is essential that PGD/ PGS laboratories maintain minimum quality control standards. Patients considering PGS or PGD must receive appropriate counseling with regard to the usefulness and limitation of the investigative and diagnostic treatment.

Suggested Standard Operating Protocol (SOP) Clinical-patient pathway for starting PGS/PGD Laboratory SOPs for performing PGS/PGD (this differs if PGS/PGD is performed in-house versus externally) Information sheets Patient information on PGS Patient information on PGD

References 1. S. Palini et al. Pre-implantation genetic diagnosis and screening: Now and the future. Gynecol. Endocrinol. 2015;31(10):755–9. 2. J. C. Harper and S. B. Sengupta. Preimplantation genetic diagnosis: State of the art 2011. Hum. Genet. 2012;131(2):175–86. 3. J. Harper et al. What next for preimplantation genetic screening? Hum. Reprod. 2008;23(3):478–80. 4. S. Mastenbroek et al. Preimplantation genetic screening: A systematic review and meta-analysis of RCTs. Hum. Reprod. Update 2011;17(4):454–66. 5. D. Cimadomo et al. The impact of biopsy on human embryo developmental potential during preimplantation genetic diagnosis. Biomed. Res. Int. 2016;2016:7193075. 6. J. D. Kort et al. Aneuploidy rates and blastocyst formation after biopsy of morulae and early blastocysts on day 5. J. Assist. Reprod. Genet. 2015;32:925–30. 7. Y. M. Zheng et al. Whole genome amplification in preimplantation genetic diagnosis. J. Zhejiang Univ. Sci. B, 2011;12(1):1–11. 8. T. Hardarson et al. Preimplantation genetic screening in women of advanced maternal age caused a decrease in clinical pregnancy rate: A randomized controlled trial. Hum. Reprod. 2008;23 (12):2806–12. 9. E. M. Dahdouh, J. Balayla and J. A. Garcia-Velasco. Impact of blastocyst biopsy and comprehensive chromosome screening technology on preimplantation genetic screening: A systematic review of randomized controlled trials. Reprod. Biomed. Online 2015;30(3):281–9. 10. Z. Yang et al. Randomized comparison of next-generation sequencing and array comparative genomic hybridization for preimplantation genetic screening: A pilot study. BMC Med. Genomics, 2015; 8:30. 11. E. Greco, M. G. Minasi and F. Fiorentino. Healthy babies after intrauterine transfer of mosaic aneuploid blastocysts. N. Engl. J. Med. 2015;373(21):2089–90. 12. S. Munne, J. Grifo and D. Wells. Mosaicism: “Survival of the fittest” versus “no embryo left behind.” Fertil. Steril. 2016; 105(5):1146–9.

Suggested Audits

13. A. G. Besser and E. L. Mounts, Counselling considerations for chromosomal mosaicism detected by preimplantation genetic screening. Reprod. Biomed. Online, 2017;34(4):369–74.

Pregnancy results (clinical pregnancy rates and live birth rates) for PGS/PGS per woman per cycle versus cumulative pregnancy rates

14. N. M. Sachdev et al. Diagnosis and clinical management of embryonic mosaicism. Fertil. Steril. 2017;107(1):6–11.

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Tips and Tricks in Performing 3-D Ultrasound before an IVF Cycle Sotiris H. Saravelos

Background Three-dimensional (3-D) ultrasound (US) is in fact an extension of two-dimensional (2-D) US imaging, in the way that computerized tomography (CT) is an extension of traditional radiography. Three-dimensional US requires the use of a specialized probe, which typically contains a motor that will allow the movement of the piezoelectric crystals aligned at the probe tip, to perform a so-called sweep during a US examination. This “sweep” essentially acquires a sequential series of 2-D US images and collates them into what is called a “3-D volume.” This is then stored into the US machine where it can be further analyzed, similar to how a CT or magnetic resonance imaging (MRI) can be analyzed in the control room following an examination. Threedimensional US is additionally capable of storing 3-D volumes containing power Doppler (PD) flow, which will give information regarding blood flow speed and degree of vascularization of the anatomical structures examined. From the patients’ perspective, it is not typically possible to distinguish between a 3-D and a 2-D US examination. In addition, the time required for the examination and the number of US waves emitted do not vary significantly between 3-D and 2-D US, making it entirely safe for the patients. Interestingly, a common misconception is that only the latest and most expensive US machines will offer 3-D capabilities. In fact, the majority of US machines on the market today offer a 3-D function, and within the context of assisted reproduction, 3-D US machines of even more than 10 years old may often suffice for basic/ routine diagnostic purposes.

Assessment of the Uterus Assessment of the uterus is pivotal in any woman undergoing assisted reproduction technology (ART) for a number of reasons. First, an attempt to ascertain uterine receptivity may help with the planning of the treatment and the approximation of the prognosis.

Second, the diagnosis of specific congenital and acquired uterine anomalies may prompt surgical and/or medical treatment prior to an in vitro fertilization (IVF) cycle of ART, which obviously has important consequences for the women. The value of 3-D US within this context is presented systematically in what follows.

Endometrium and Sub-Endometrium For several years, US has been used to determine the endometrial receptivity before an ART cycle. Although it is known that 2-D US assessment of the endometrial thickness is a predictor of not only the likelihood of pregnancy but also of complications such as pregnancy loss and ectopic pregnancy [1], the value of 3-D US assessment within this context has been less clear. Three-dimensional US has now allowed the estimation of the endometrial volume through software such as VOCAL (Virtual Organ Computer-aided AnaLysis) (GE Medical Systems Kretztechnik GmbH & Co., Austria). VOCAL allows the clinician to manually outline the endometrium as the uterus is rotated along a stable axis over 180° in order to estimate the entire endometrial volume. Originally, this was thought to be very promising and potentially a better marker for uterine receptivity than endometrial thickness alone. However, through the years, it has transpired that endometrial volume assessed with 3-D US is not superior to traditional endometrial thickness measured with 2-D US. In particular, a recent systematic literature review showed that 11 out of 16 studies published in the literature found no relation with clinical outcomes, while a meta-analysis of studies was not possible due to the large heterogeneity of the studies available [2]. When looking at endometrial and sub-endometrial vascularization, the story appears quite similar. Initially, there was some evidence to support the direct or indirect measurement of endometrial blood flow using 2-D US in women undergoing IVF [3]. The emergence of 3D US with PD further increased the hopes of finding a more sensitive marker for endometrial receptivity.

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However, the evidence was once again lacking, with to date 6 out of 10 heterogeneous studies showing no relation with clinical outcomes [2].

Verdict Three-dimensional US assessment of the endometrial volume and endometrial/sub-endometrial vascularization is currently not recommended prior to ART treatment due to the lacking and inconsistent evidence available to support its use.

Congenital Anomalies Congenital uterine anomalies are undoubtedly important to diagnose before proceeding with an IVF cycle, as they are known to be associated with several adverse reproductive outcomes, such as reduced pregnancy rates, increased miscarriage rates, increased preterm delivery rates, increased rate of malposition at delivery, decreased birth weight and increased perinatal mortality rates [4]. The way to diagnose congenital uterine anomalies has been debated for several years since there are many different types of imaging modalities available to use (hysterosalpingography [HSG], hysteroscopy, laparoscopy, MRI, 2-D US, 3-D US, saline infusion sonography [SIS]). However, this debate finally seems to be coming to an end, since the publication of a recent international consensus on the diagnosis of female genital tract anomalies [5]. This

Normal

Unicornuate

consensus included a systematic review on the diagnostic accuracy of all aforementioned methods (with hysteroscopy/laparoscopy acting as the gold standard), which included 38 studies in order to estimate the overall accuracies (here presented with 95% CI): 3-D US 97.6% (94.3–100), SIS 96.5% (93.4–99.5), HSG 86.9% (79.8–94.0) and 2-D US 86.6% (81.3–91.8). Although MRI appears not to have been tested by any study in the literature as a screening tool, it was considered at least as accurate as 3-D US, based on the available studies to date. In view of these findings, it was concluded in this consensus that 3-D US should be the new gold standard method for diagnosis of female genital anomalies, facilitated by endoscopy and MRI in complex cases or those of uncertainty [5]. From a practical point of view, compared with other investigative modalities, 3-D US offers the additional advantages of being noninvasive, time-efficient, cost-effective, readily available, objective, highly reproducible and can be used simply at the patient’s bedside to convey a self-explanatory image of the diagnosis (Figure 33.1). Furthermore, it can be used pre- and postoperatively to objectively assess the depth of the septum and residual fundal wall thickness, and confirm its successful treatment prior to IVF.

Verdict Three-dimensional US assessment for congenital uterine anomalies is nowadays one of the most valuable

T-shape

Septate

Bicorporeal

Figure 33.1 Examples of different uterine anomalies as diagnosed with 3-D US alongside their respective illustrations from the ESHRE-ESGE classification (2013)

Performing 3-D Ultrasound before an IVF Cycle

and evidence-based applications of 3-D US within the context of IVF treatment. A recent international consensus recommends its use above all other investigations for any woman suspected to have a congenital uterine anomaly.

Acquired Anomalies Acquired uterine anomalies, such as polyps, fibroids, adenomyosis and intrauterine adhesions, are important to diagnose prior to ART treatment cycles as they are known to hamper clinical outcomes. Interestingly, there is less data in the literature regarding the use of 3-D US for the diagnosis of acquired uterine anomalies versus congenital uterine anomalies, although most authors would advocate that it is superior to using 2-D US alone. With regards to diagnosing polyps, one study screening 3,850 infertile women found 3-D US to have a sensitivity of 100 percent, a specificity of 99 percent, a positive predictive value (PPV) of 99 percent and a negative predictive value (NPV) of 100 percent in diagnosing endometrial polyps when compared to hysteroscopy [6]. Similarly, 3-D US has been found to be superior in diagnosing fibroids compared with 2-D US [7], with the benefit of providing a clearer visualization of the uterine cavity and a better estimation of the size, shape and appearance of the fibroids. With regards to adenomyosis, in a group of women undergoing hysterectomy rather than IVF, one study found 3-D US marginally more accurate than 2-D US with an overall accuracy for diagnosis of 89 percent versus 83 percent, respectively. In particular, the sensitivity, specificity, PPV and NPV for 3-D US versus 2-D US was 91 percent, 75 percent, 88 percent, 90 percent, and 85 percent, 86 percent, 92 percent and 82 percent, respectively [8]. However, note that there appears to be no 3-D US study concerning adenomyosis, or indeed the junctional zone, to have been performed specifically in a population of women undergoing IVF. With regards to diagnosing intrauterine adhesions, sparse studies in the literature show improved diagnostic accuracy rates with the use of 3-D US. However, most authors in the literature would recommend its use. This is reflected in a recent editorial in the White Journal presenting a diagnostic algorithm, where 3-D US is specifically recommended to map out the location and extent of cavity obliteration and obstruction in women with suspected intrauterine adhesions [9].

It is highly relevant that, when considering the findings of a recent systematic review and metaanalysis that found the accuracy of 2-D SIS to be comparable to the gold standard of hysteroscopy, there is no doubt that the application of 3-D SIS has the potential to become the new gold standard method of choice in the near future [10].

Verdict Preliminary studies suggest that 3-D US, particularly with the application of SIS, has the potential to become the gold standard method of diagnosis for polyps, fibroids and adhesions within the context of ART. Three-dimensional US assessment of adenomyosis can be used in conjunction with traditional 2-D US imaging, but should be reserved predominantly for research purposes at the present time.

Assessment of the Fallopian Tubes Assessment of the Fallopian tubes forms a pivotal part of the work-up for infertility for two main reasons: first, in order to gauge whether the infertility is related to blocked tubes, and, second, to gauge whether there is a hydrosalpinx that could hamper the chances of ART success. For the first, HSG and laparoscopy have been the two commonest forms of investigation, while for the second, 2-D US, HSG and laparoscopy have been the commonest methods of choice. However, developments with 3-D US now mean that a third option is available at the bedside that does not include radiation exposure or invasive surgery.

Patency Patency can nowadays be assessed via hysterosalpingo-contrast sonography (HyCoSy), which refers to the introduction of a sonographic-enhancing positive-contrast fluid into the uterine cavity and Fallopian tubes, in order to visualize the tubes on US and ascertain patency. Two-dimensional HyCoSy has been performed in this context for several years now; however, it does suffer from a number of limitations, which have perhaps hindered its more widespread use. In particular, 2-D HyCoSy requires an experienced operator in order to follow the typically tortuous path of the Fallopian tubes. Furthermore, the echogenic bowel can often distort the image, making it difficult to distinguish the echogenic contrast. However, the newest-generation 3-D US machines allow the visualization of the entire tubal length within a single image by taking a 3-D volume

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acquisition of the pelvis. Furthermore, dedicated software, such as Coded-Contrast-Imaging (CCI) (GE Medical Systems Kretztechnik GmbH & Co., Austria), allows better differentiation between the contrast medium and various artefacts from the adjacent tissue and organs. As a result, studies have shown a sensitivity of 93.5 percent, specificity of 86.3 percent, PPV of 87.8 percent and NPV 92.6 percent and overall diagnostic accuracy of 90.0 percent when comparing 3-D HyCoSy with the gold standard laparoscopy and dye test [11]. Furthermore, it has been shown that there is a significantly shorter learning curve for operators, which makes this examination more amenable for use within an IVF clinic. An example of a 3-D HyCoSy is shown in Figure 33.2.

patency in women with infertility. Newer software such as the 3-D Inversion Mode can be used in conjunction with 2-D US to support the diagnosis of hydrosalpinges, although further research should be carried out to substantiate its use.

Hydrosalpinges

Antral Follicle Count

Although 2-D US can be used to diagnose hydrosalpinges, this can often be challenging as the operator has to reconstruct in their mind the several 2-D planes they are seeing in order to judge whether it is indeed a hydrosalpinx, versus a distorted ovarian or paraovarian cyst. Once again, 3-D US can capture the entire structure in question, within a single volume, therefore allowing its visualization within a single image. Furthermore, a new display modality called the 3-D Inversion Mode can convert anechoic US fluid to solid, with a press of a button, which will show the entire tube as a solid structure without surrounding interference [12]. This can potentially increase accuracy versus 2-D US and once again allow the findings to be conveyed more convincingly to the patients.

The AFC has been shown to be one of the best predictors of ovarian response to ART, and is therefore performed in many centers worldwide. Threedimensional US assessment of the ovary, along with the application of specialized software called Sono Automatic Volume Calculation (Sono AVC), has been studied in the literature in the past few years. This software uses mathematical algorithms to differentiate hypoechoic, fluid-filled areas within an acquired volume (in this case the follicles within the ovary), therefore allowing for the automated estimation of the AFC (GE Medical Systems Kretztechnik GmbH & Co., Austria). It can be performed with a press of a button and within a few seconds, although it is of course open to error, as it can misinterpret physiological fluid spaces for follicles, or fail to detect follicles that are not as hypoechoic as usual (e.g., when the ovary is behind the uterus, or when bowel shadows are present). Nevertheless, a recent study demonstrated that using this 3-D US software, the AFC retrieved was an independent significant predictor of pregnancy following ART treatment [13]. The question, however, remains whether 3-D US assessment of AFC is superior to the manual 2-D US measurement performed by most clinicians to date.

Verdict There is now evidence to support the use of 3-D HyCoSy as the first-line investigation for the assessment of tubal

Assessment of the Ovaries Assessment of the ovaries undoubtedly forms an important part of the ART treatment process. This includes the assessment of ovarian reserve through antral follicle count (AFC) measurements, and also the assessment of the growth of the follicles during ovarian stimulation. The role of 3-D US within this context is analyzed in what follows.

Follicle Tracking Figure 33.2 Three-dimensional HyCoSy with HDlive demonstrating the uterine cavity and both Fallopian tubes in a single image

Other than the AFC, the 3-D US SonoAVC software can assess the number and sizes of follicles during ovarian stimulation. In this case, the software actually provides an automatic estimation of the absolute dimensions of each follicle, both in terms of millimeters

Performing 3-D Ultrasound before an IVF Cycle

in the x-y-z axes, and in terms of actual volume in milliliters. However, a recent randomized controlled trial (RCT) comparing the use of 3-D US and 2-D US in the timing of oocyte maturation and egg collection of standard ART cycles found no significant differences in clinical outcomes between the two groups [14]. Therefore, although potentially quicker and requiring less expertise to perform, 3-D US has not been demonstrated to be superior within this context. An example of an assessment of follicle tracking with 3-D US is shown in Figure 33.3.

Verdict There is currently no evidence to support that 3-D US is superior to 2-D US in the assessment of the ovaries prior or during an IVF cycle. However, there is sufficient evidence to allow its use in the clinical practice setting, if a unit has the particular machines/software available and undergoes appropriate training.

Conclusion In conclusion, 3-D US is useful within the context of IVF, but only for certain types of assessments. It has

not consistently proved to provide a good assessment of uterine receptivity, through either the assessment of the endometrial volume or endometrial/subendometrial vascularization. However, it has proven a highly accurate tool in diagnosing both congenital and acquired uterine pathology. In particular, it is nowadays considered the gold standard for diagnosis in a recently published international consensus. Furthermore, the addition of SIS makes it particularly accurate in diagnosing and mapping submucosal/intramural fibroids, polyps and intrauterine adhesions and may well prove the gold standard within this context also. Furthermore, although not as widespread currently, 3-D HyCoSy ticks all the boxes to become the new first-line method of choice for assessing tubal patency, while other new features of 3-D US, such as inverse rendering, may improve the diagnosis of hydrosalpinges. Finally, 3-D US in conjunction with SonoAVC has been shown to be similar but not superior to traditional 2-D US assessment of the ovarian follicles. Based on the current evidence and possible future developments, 3-D US is likely to become an indispensable tool for any IVF clinic.

Figure 33.3 Three-dimensional US with SonoAVC assessment of a stimulated ovary

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References 1. X. Yuan, S. Saravelos, Q. Wang, Y. Xu and T. C. Z. Li. Endometrial thickness as a predictor of pregnancy outcomes in 10787 fresh IVF–ICSI cycles. Reproductive Biomedicine Online 2016; 33(2):197–205. 2. S. Saravelos, K. Jayaprakasan, K. Ojha and T. Li. Assessment of the uterus with three-dimensional ultrasound in women undergoing ART. Human Reproduction Update 2017;23(2):188–210. 3. E. H. Ng, C. C. Chan, O. S. Tang, W. S. Yeung and P. C. Ho. The role of endometrial blood flow measured by three-dimensional power Doppler ultrasound in the prediction of pregnancy during in vitro fertilization treatment. European Journal of Obstetrics, Gynecology, and Reproductive Biology 2007;135:8–16. 4. C. A. Venetis, S. P. Papadopoulos, R. Campo, S. Gordts, B. C. Tarlatzis and G. F. Grimbizis. Clinical implications of congenital uterine anomalies: A meta-analysis of comparative studies. Reproductive Biomedicine Online 2014;29:665–83. 5. G. F. Grimbizis, A. Di Spiezio Sardo, S. H. Saravelos, S. Gordts, C. Exacoustos, D. van Schoubroeck et al. The Thessaloniki ESHRE/ESGE consensus on diagnosis of female genital anomalies. Human Reproduction 2016;31:2–7. 6. S. Kupesic, A. Kurjak, S. Skenderovic and D. Bjelos. Screening for uterine abnormalities by three-dimensional ultrasound improves perinatal outcome. Journal of Perinatal Medicine 2002;30:9–17. 7. C. Sylvestre, T. J. Child, T. Tulandi and S. L. Tan. A prospective study to evaluate the efficacy of two- and three-dimensional sonohysterography in women with intrauterine lesions. Fertility and Sterility 2003;79: 1222–5.

8. C. Exacoustos, L. Brienza, A. Di Giovanni, B. Szabolcs, M. E. Romanini, E. Zupi et al. Adenomyosis: Three-dimensional sonographic findings of the junctional zone and correlation with histology. Ultrasound in Obstetrics & Gynecology 2011;37: 471–9. 9. T. N. Amin, E. Saridogan and D. Jurkovic. Ultrasound and intrauterine adhesions: A novel structured approach to diagnosis and management. Ultrasound in Obstetrics & Gynecology 2015;46:131–9. 10. S. Seshadri, T. El-Toukhy, A. Douiri, K. Jayaprakasan and Y. Khalaf. Diagnostic accuracy of saline infusion sonography in the evaluation of uterine cavity abnormalities prior to assisted reproductive techniques: A systematic review and meta-analyses. Human Reproduction Update 2015;21:262–74. 11. L. Zhou, X. Zhang, X. Chen, L. Liao, R. Pan, N. Zhou et al. Value of three-dimensional hysterosalpingo-contrast sonography with SonoVue in the assessment of tubal patency. Ultrasound in Obstetrics & Gynecology 2012;40:93–8. 12. I. E. Timor-Tritsch, A. Monteagudo, T. Tsymbal and I. Strok. Three-dimensional inversion rendering: A new sonographic technique and its use in gynecology. Journal of Ultrasound in Medicine 2005;24:681–8. 13. S. Deb, M. Batcha, B. K. Campbell, K. Jayaprakasan, J. S. Clewes, J. F. Hopkisson et al. The predictive value of the automated quantification of the number and size of small antral follicles in women undergoing ART. Human Reproduction 2009;24:2124–32. 14. N. Raine-Fenning, S. Deb, K. Jayaprakasan, J. Clewes, J. Hopkisson and B. Campbell. Timing of oocyte maturation and egg collection during controlled ovarian stimulation: A randomized controlled trial evaluating manual and automated measurements of follicle diameter. Fertility and Sterility 2010;94:184–8.

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How to Draw Up a Management Plan When an IVF Cycle Has Failed Carol Coughlan

Management

Lifestyle Changes

A multidisciplinary approach should be adopted in the management of a couple who have experienced a failed in vitro fertilization (IVF) cycle. It should involve not only an experienced fertility specialist but also a senior embryologist and, where appropriate, a reproductive surgeon or a counselor. Patients need to be reassured that their treatment is under the supervision of an experienced clinician. The couple should be offered ample time for their questions to be addressed and a clear treatment plan agreed. The appointment should include a thorough review of the diagnosis of the underlying cause of infertility, the investigation results, the treatment protocol, the response to ovarian stimulation, the quality of the oocyte and embryos and possible explanation as to why they have not produced a successful pregnancy. The couple should have explained to them that any treatment plan recommended would be discussed and confirmed in a multidisciplinary team meeting and the final decision confirmed in writing. Second, there ought to be an agreed local protocol as to how couples with a failed IVF cycle should be further investigated and managed. The protocol ought to be updated regularly to take into consideration the findings of recent studies. The protocol should contain sufficient details to ensure that patients and staff clearly understand the plan of action, and the rationale behind any decisions made. Appropriate counseling of the couple is of the utmost importance prior to proceeding with further treatment. The couple should be advised as to the likelihood of success in future cycles and cautioned not to pursue further treatment if their prognosis is poor. The service of an independent counselor should be offered at these difficult times. If it is deemed reasonable to pursue further treatment, it is beneficial to instigate appropriate investigations and review previous unsuccessful IVF treatment cycles with a view to modifying or changing the treatment protocol if indicated.

In addition to a review of investigations and treatment, clinicians should discuss and advise as to lifestyle changes that could improve the likelihood of treatment success. Smoking: Women who smoke should be advised to stop. Overall, the literature strongly supports an association between cigarette smoking and infertility. There is evidence that smoking is associated with an increased gonadotrophin requirement for ovarian stimulation, fewer oocytes retrieved, higher numbers of cancelled cycles, lower implantation rates and more cycles with failed fertilization in smokers compared with nonsmokers [1]. Male partners should also be advised to abstain from smoking as there is good evidence that semen parameters are 22 percent poorer in smokers than in nonsmokers [2]. BMI: Couples should be informed of the adverse effect of obesity on fertility treatment outcome and be encouraged to improve their general health. Underweight women (BMI < 19 kg/m2) should be encouraged to gain weight and obese women (BMI ≥ 30 kg/m2) should be advised to lose weight prior to further attempts at IVF treatment. Women should be provided with assistance to lose weight, including psychological support, dietary advice, exercise classes and, where appropriate, weightreducing agents or bariatric surgery. Obesity also has an adverse effect on male fertility and longterm health. Alcohol Consumption: Women who are trying to become pregnant should be advised to reduce alcohol consumption to one or two units once or twice per week and avoid episodes of intoxication to reduce the risk of harming a developing fetus. Men should be informed that excessive alcohol intake is detrimental to semen quality [3].

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Ovarian Stimulation Protocol Ovarian response to gonadotrophin stimulation should be reviewed. If the response is deemed satisfactory, it is not necessary to change the stimulation protocol. In a small proportion of women who are deemed to have suboptimal response to ovarian stimulation, the dose of gonadotrophin may be increased. There is no firm evidence that antagonist protocol is better than agonist protocol, or vice versa [4]. There is some evidence to suggest that “poor responders” to ovarian stimulation and women over the age of 35 years may benefit from the addition of luteinizing hormone (LH) to cycles [5]. In women with endometriosis and adenomyosis, the use of ultra-long protocol involving the administration of gonadotrophin-releasing hormone (GnRH) agonists for a few months prior to IVF or intracytoplasmic sperm injection (ICSI) may increase the pregnancy rate [6]. Currently, standard IVF protocols utilize human chorionic gonadotropin (hCG) as a surrogate for the LH surge. In contrast to hCG, the GnRH agonist induces a surge of both FSH and LH resembling the natural mid-cycle surge. For patients where a significant number of immature oocytes were retrieved in previous cycles, some studies have suggested that the GnRH agonist trigger results in the retrieval of more mature oocytes but does result in luteal phase insufficiency. Studies, although retrospective, suggest an advantage to using a GnRH agonist in combination with hCG as a double trigger in IVF cycles to increase the number of mature oocytes [7, 8].

Sperm DNA Fragmentation There is insufficient evidence to recommend the routine use of sperm DNA integrity tests in both the evaluation and treatment of the infertile couple and for patients undergoing IVF/ICSI [9]. However, sperm DNA fragmentation testing may be useful for a subgroup of patients with repeated IVF failures without an apparent cause in which “unrepairable” sperm DNA damage may be the limiting factor responsible for their fertility problem [10]. When suboptimal quality sperm is considered a contributory cause of IVF failure, supported by an increased amount of sperm DNA fragmentation, several treatment options may be weighed. First, oral antioxidant treatment has been shown to reduce the incidence of sperm DNA fragmentation [11]. Second, based on the observation that sperm DNA damage is lower in the seminiferous tubules as compared to the cauda epididymis and

ejaculated sperm [12], it has been proposed that men with high levels of DNA damage in ejaculated sperm have sperm removed surgically from the testis for ICSI. The use of testicular sperm in couples with repeated IVF failures associated with high sperm DNA fragmentation in semen has been reported to result in a significant increase in pregnancy rates [13] and reduction of miscarriage rates [14], but further studies are required to confirm the benefit.

Improving Embryo Quality and Selection Blastocyst transfer: Several studies have suggested that extending embryo culture to day 5/6 in order to transfer the embryo at the blastocyst stage increases the implantation rate [15]. In women with a history of IVF failures, blastocyst transfer ought to be considered if not performed in previous treatment cycles. Assisted hatching (AH): Hatching of the blastocyst plays an integral role in the implantation process. Failure to hatch (due to intrinsic abnormalities in either the blastocyst or zona pellucida [ZP]) is a possible cause of implantation failure. Assisted hatching involves the artificial thinning or breaching of the ZP and has been proposed as one technique to improve implantation and pregnancy rates following IVF [16]. The American Society of Reproductive Medicine published recommendations regarding assisted hatching and concluded that there is evidence that assisted hatching slightly improves clinical pregnancy rates in poor-prognosis patients, including those with previously failed IVF cycles [16]. Preimplantation genetic screening (PGS): The main purpose of preimplantation genetic screening is to achieve a healthy pregnancy as a result of screening embryos for chromosomal disorders and selecting healthy embryos for transfer into the uterus, thereby selecting embryos with the highest chance of implantation. The current embryo biopsy technique for PGS, day 5 multiple cell trophectoderm biopsy and the current testing method of next-generation sequencing have resulted in significantly improved implantation rates as compared to the initial day 3 single blastomere biopsy and the initial genetic testing method fluorescence in situ hybridization (FISH). The role of preimplantation genetic screening is controversial as recent research has showed that some embryos diagnosed as abnormal by PGS may have the potential to develop into healthy babies [17]. Embryo mosaicism and the trophectoderm sampling

Management Plan When an IVF Cycle Has Failed

technique presently utilized are two reasons for the possible inaccuracy of PGS [17]. However, PGS is suggested as a successful strategy for patients with multiple failed IVF attempts [18], but further studies are required to conclusively confirm whether PGS is beneficial in women with previously failed IVF cycles.

Embryo Transfer Embryo implantation has been found to be dependent on embryo quality, endometrial receptivity and transfer efficiency [19]. In women with previous failed IVF cycles, the details of previous embryo transfers should be reviewed, paying particular attention to any technical difficulties encountered. If there had been difficulty with previous embryo transfers, identified as a procedure taking longer than usual, causing significant pain, requiring a change of catheter, cervical dilatation or use of a tenaculum, it is accepted that the pregnancy rate would be lower [20]. Difficult embryo transfer may be due to cervical stenosis, acute anteversion/retroversion or to acute anteflexion/retroflexion of the uterus. Several techniques may be considered in women with a history of difficult embryo transfer. First, the transfer should be performed under ultrasound guidance [21]. Second, a trial transfer should be discussed and considered. Third, filling the bladder in women with acute anteversion or anteflexion is a simple measure that may sometimes be useful, but will not be helpful in cases of acute retroversion or retroflexion, where an empty bladder is preferable. The application of a tenaculum to the anterior lip of the cervix and applying traction gently downward may help to straighten an acutely flexed uterus but may compromise pregnancy rates by inducing uterine contractions [20]. The use of a rigid catheter may help to negotiate the cervix if difficulty was previously encountered with the use of a soft catheter. Alternative methods to transcervical embryo transfer include transmyometrial and tubal transfer, but they should be reserved for cases that are extremely difficult or impossible. There is insufficient evidence to show that bed rest after transfer improves outcome.

Endometrial Receptivity Embryo implantation is a complex process requiring a receptive endometrium. Supra-physiological hormone levels during controlled ovarian stimulation may result in reduced endometrial receptivity, which may in turn have an adverse effect on implantation and pregnancy

rates in IVF/ICSI cycles. Many studies have suggested a freeze-all policy in which all embryos are electively cryopreserved for transfer in a subsequent frozen-thaw cycle in the hopes of providing a more physiological environment for embryo transfer. A recent prospective cohort study identified a statistically significant improvement in ongoing pregnancy and implantation rates following a freeze-all policy for patients with recurrent implantation failure [22].

Hysteroscopy A recent multicenter randomized controlled trial demonstrated that routine outpatient hysteroscopy does not improve livebirth rates in recurrent implantation failure [23], but where 2-D/3-D ultrasound suggests the presence of endometrial pathology, hysteroscopy is a valuable investigative and treatment modality. It may be performed as an outpatient procedure; small lesions may be removed at the same time, but more significant pathology may need to be dealt with later under general anesthesia.

Intra-Cavity Lesion Submucous fibroid: A meta-analysis showed that submucous fibroids significantly reduced the implantation rate, clinical pregnancy rate and live birth rate and significantly increased the miscarriage rate [24]. A submucous fibroid in women with a previous failed IVF cycle, regardless of the size, should be removed as it has been shown in the meta-analysis that removal of submucous fibroids improves clinical pregnancy rates [24]. Endometrial polyps: Similarly, endometrial polyps in women with recurrent implant failure (RIF) ought to be removed. It has been shown that the removal of endometrial polyps in women undergoing IUI treatment resulted in doubling of the clinical pregnancy rate (CPR) [25]. Uterine septum: In women with previously failed IVF cycles and women with a history of recurrent miscarriage, uterine septae should be removed, regardless of the size. The various techniques used to remove uterine septae have been reviewed [26]. Intrauterine adhesions: It is accepted that intrauterine adhesions would interfere with the implantation process and adversely affect the implantation rate, so if they are present in women with recurrent implantation failure, they should be removed [27].

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Nevertheless, there is as yet no firm literature data to confirm that removal of intrauterine adhesions improves the implantation rate. Furthermore, intrauterine adhesions often recur after surgical removal and there is a high rate of complication (10 percent or more) in cases of severe intrauterine adhesions resulting in partial or complete obliteration of the cavity. The procedure should be carried out by an experienced reproductive surgeon under ultrasound guidance with a view to minimize complications [28].

Myometrial Pathology Intramural fibroid: While women with recurrent implantation failure should have submucous fibroids removed, the possible contribution of intramural fibroids that are not distorting the uterine cavity to implantation failure is far from clear. There is no consensus on whether intramural fibroids in women with RIF should be removed [29]. Many clinicians would recommend removal of intramural fibroids if they are more than 4 cm in diameter. There is a lower threshold to remove an intramural fibroid if it is situated in the anterior lower uterine segment as it may pose problems in delivery of the fetus, especially if caesarean section is required. The pros and cons of myomectomy should be carefully explained in each case. Couples should be aware of the possible complications of myomectomy, including the likelihood of blood transfusion, a small risk (1 percent) of hysterectomy and a relatively high risk of adhesion formation over the uterine scar, as well as a small but serious risk of scar rupture during the ensuing pregnancy. On the other hand, couples should also understand that intramural fibroids may cause not only implantation failure but also a number of other problems, including miscarriage (both first and second trimester), red degeneration, preterm delivery, placental abruption, fetal growth restriction, malpresentation and difficulty with delivery as well as intra- and postpartum hemorrhage. The final decision must be individualized, and the involvement of a reproductive surgeon in the decision-making process is recommended. Adenomyosis: The role adenomyosis plays in reproductive failure is receiving increasing attention [6]. Unlike intramural fibroids, adenomyosis is not as amenable to surgical treatment, but Tremellen and Russell [6] reported on cases of recurrent implantation failure associated with adenomyosis, all

successfully treated with an ultra-long pituitary downregulation protocol.

Hydrosalpinges There is now good evidence that the removal of hydrosalpinges improves the implantation and live birth rates in women undergoing IVF [30]. There are conflicting reports on whether salpingectomy compromises ovarian response to stimulation during subsequent IVF treatment. It is prudent, when carrying out salpingectomy, to diathermize and incise as close to the Fallopian tube as possible and as far away from the ovary as possible, to avoid disruption to the ovarian blood supply. Salpingectomy is not the only surgical treatment option for women with hydrosalpinges contemplating IVF treatment. Salpingostomy may be a possible alternative as it not only “removes” the hydrosalpinges but also produces the possibility of natural conception. It seems logical, therefore, to recommend that women with hydrosalpinges and Fallopian tubes with minimal damage consideration should be given salpingostomy whereas tubes that are severely damaged (especially for those with intraluminal adhesions) ought to be removed (salpingectomy). The drawback of salpingostomy is the possible recurrence of hydrosalpinges, which then necessitates a further procedure to remove the tube, further delaying treatment and incurring extra cost.

Clinical Trials Couples who experience unsuccessful IVF cycles are desperate to seek a treatment that will lead to a successful outcome. Many of them will have searched the Internet looking for a “new” treatment on the horizon. They may not be able to judge for themselves the scientific credibility of such claims they come across on the Internet. They often seek the advice of their specialist to confirm if certain “new” treatments are worth trying. Clinicians should be able to judge if certain treatments have been proven to be of value. If not, treatment should not be initiated, or the treatment should be offered only as part of a clinical trial. In this situation, prior ethics approval for the trial should have been obtained and national research governance guidelines followed. Written consent from each patient should be obtained.

Management Plan When an IVF Cycle Has Failed

Gamete Donation and Surrogacy Couples with recurrent IVF failures need guidance on the appropriateness of proceeding with further IVF attempts. If implantation fails to occur despite repeated treatment attempts or if the prognosis of further IVF treatment is considered poor, alternative treatment options ought to be explored. If the likely source of the problem lies with the embryo, gamete donation should be advised. On the other hand, if the problem lies with the uterus, for example multiple fibroids or Asherman’s syndrome that has failed to respond to surgical treatment, surrogacy should be discussed with the couple.

Conclusion Couples who experience a failed IVF cycle should be offered appropriate investigations to rule out an underlying cause for the cycle failure. The main treatment strategy in couples with IVF failure is to improve the quality of the embryo transferred and the receptivity of the endometrium. 1. Hysteroscopy should be considered prior to a further treatment cycle if ultrasound scan is suggestive of uterine pathology. 2. Appropriate investigations should be carried out to exclude hydrosalpinx as it has been shown to reduce implantation rate, increase miscarriage rate and reduce live birth rate. Removal of hydrosalpinges has been shown to improve the outcome of IVF cycles. 3. Submucosal fibroids have been shown to reduce implantation, pregnancy and live birth rates; the removal of submucosal fibroids improves implantation rates. 4. Endometrial polyps should be removed, although there are no data on the impact of endometrial polyps on women undergoing IVF, it has been shown to improve outcome in women undergoing intrauterine insemination. 5. Uterine septum increases miscarriage rate and its removal improves outcome. 6. The use of ultra-long protocol may improve outcome in women with endometriosis and adenomyosis. 7. Consider removal of intramural fibroids of more than 5 cm. 8. Intrauterine adhesions are a recognized cause of thin endometrium not responding to ovarian

steroid stimulation; if present, intrauterine adhesions should be removed. 9. A multidisciplinary approach should be adopted in the management of recurrent IVF failures. 10. Empirical therapies should, whenever possible, be considered only in the setting of carefully conducted clinical trials.

References 1. H. Klonoff-Cohen, L. Natarajan, R. Marrs and B. Yee. 2001. Effects of female and male smoking on success rates of IVF and gamete intra-fallopian transfer. Hum. Reprod. 2001;16:1382–90. 2. American Society Reproductive Medicine. Smoking and infertility: A committee opinion. Fertil. Steril. 2012; 98 (6):1400–6. 3. National Institute for Clinical Excellence (NICE). 2016. NICE Guidance: Fertility problems: Assessment and treatment (Updated August 2016). 4. H. G. Al-Inany, M. Youssef, M. Aboulghar, F. Broekmans, M. Sterrenburg, J. Smit and A. M. Abou-Setta. Gonadotrophin-releasing hormone antagonists for assisted reproductive technology. Cochrane Database of Systematic Reviews 2011;19, CD008046. 5. M. H. Mochtar, F. van der Veen, M. Ziech. M. van Wely and A. Musters. Recombinant luteinizing hormone (rLH) for controlled ovarian hyperstimulation in assisted reproductive cycles. Cochrane Database of Systematic Reviews 2007;18: CD005070. 6. K. Tremellen and P. Russell. Adenomyosis is a potential cause of recurrent implantation failure during IVF treatment. Aust. N. Z. J. Obstet. Gynaecol. 2011;51, 280–3. 7. D. Griffin, R. Feinn, L. Engmann et al. Dual trigger with gonadotropin – releasing hormone agonist and standard dose human chorionic gonadotropin to improve oocyte maturity rates. Fertil. Steril. 2014;102 (2):405–9. 8. M. Lin, F. S. Wu, R. K. Lee et al. Dual trigger with combination of gonadotropin – releasing hormone agonist and human chorionic gonadotropin significantly improves the live-birth rate for normal responders in GnRH antagonist cycles. Fertil. Steril. 2013;100(5):1296–302. 9. American Society for Reproductive Medicine (ASRM). The clinical utility of sperm DNA integrity testing: A guideline. Fertil. Steril. 2013;99(3):673–7. 10. D. Sakkas and J. G. Alvarez. Sperm DNA fragmentation: Mechanisms of origin, impact on

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reproductive outcome, and analysis. Fertil. Steril. 2010;93,1027–36. 11. A. M. Isidori, C. Pozza, D. Ganfrilli and A. Isidori. Medical treatment to improve sperm quality. Reprod. Biomed. Online 2006;12(6):704–14. 12. E. Greco, F. Scarselli, M. Iacobelli, L. Rienzi, F. Ubaldi, S. Ferrero, G. Franco, N. Anniballo, C. Mendoza and J. Tesarik. Efficient treatment of infertility due to sperm DNA damage by ICSI with testicular spermatozoa. Hum. Reprod. 2005b;20: 226–30. 13. C. K. Bradley, S. J. McArthur, A. J. Gee, K. A. Weiss, U. Schmidt and L. Toogood. Intervention improves assisted conception intracytoplasmic sperm injection outcomes for patients with high levels of sperm DNA fragmentation: A retrospective analysis. Andrology 2016;27 (Epub). 14. A. Borini, N. Tarozzi, D. Bizzaro, M. A. Bonu, L. Fava, C. Flamigni and G. Coticchio. Sperm DNA fragmentation: Paternal effect on early post-implantation embryo development in ART. Hum. Reprod. 2006;21:2876–81. 15. E. G. Papanikolaou, E. M. Kolibianakis, H. Tournaye, C. A. Venetis, H. Fatemi, B. Tarlatzis and P. Devroey. Live birth rates after transfer of equal number of blastocysts or cleavage-stage embryos in IVF. A systematic review and meta-analysis. Hum. Reprod. 2008;23:91–9. 16. American Society for Reproductive Medicine (ASRM). The role of assisted hatching in in vitro fertilization: A guideline. Fertil. Steril. 2014;102(2):348–51. 17. R. Casper, J. Haas, T. B. Hsieh, R. Bassil and C. Mehta. Recent advances in in-vitro fertilization. F1000Res. 2017;6:1616. 18. E. Greco, S. Bono, A. Ruberti, A. M. Labascio, P. Greco, A. Biricik, L. Spizzichino, A. Greco, J. Tesarik, M. G. Minasi and F. Florentino. Comparative genomic hybridization selection of blastocysts for repeated implantation failure treatment: A pilot study. Biomed. Res. Int. 2014. (Epub). 19. R. J. Paulson, M. V. Sauer and R. A. Lobo. Factors affecting embryo implantation after human in vitro fertilization: A hypothesis. Am. J. Obstet. Gynecol. 1990;163 (6 Pt. 1):2020–3.

20. L. Mains and R. J. van Voorhis. Optimizing the technique of embryo transfer. Fertil. Steril. 2010; 94: 785–90. 21. J. Brown, K. Buckingham, A. M. Abou-Setta and W. Buckett. Ultrasound versus “clinical touch” for catheter guidance during embryo transfer in women. Cochrane Database of Systematic Reviews 2010; 20(1): CD006107. 22. Y. Magdi, A. Damen, A. Fathi, A. M. Abdelaziz, M. Youssef, A. Abd-Allah, M. Elawady, M. Ibrahim, and Y. Edris. Revisiting the management of recurrent implantation failure through freeze-all policy. Fertil. Steril. 2017;108(1):72–7. 23. T. El-Toukhy, R. Campo, Y. Khalaf, C. Tabanelli, L. Gianaroli and S. S. Gordts et al. Hysteroscopy in recurrent in-vitro fertilisation failure (TROPHY): A multi-centre, randomised controlled trial. Lancet 2016;387:2614–21. 24. E. A. Pritts, W. H. Parker and D. L. Olive. Fibroids and infertility: An updated systematic review of the evidence. Fertil. Steril. 2016;91:1215–23. 25. J. Bosteels, S. Weyers, P. Puttemans, C. Panayotidis, B. Van Herendael, V. Gomel, B. W. Mol, C. Mathieu and T. D’Hooghe. The effectiveness of hysteroscopy in improving pregnancy rates in subfertile women without other gynaecological symptoms: A systematic review. Hum. Reprod. Update 2010; 16:1–11. 26. H. A. Homer, T. C. Li and I. D. Cooke. The septate uterus: A review of management and reproductive outcome. Fertil. Steril. 2000; 73:1–14. 27. C. M. March. Management of Asherman’s syndrome. Reprod. Biomed. Online 2011; 23, 63–76. 28. D. Yu, Y. M. Wong, Y. Cheong, E. Xia and T. C. Li. Asherman syndrome–one century later. Fertil. Steril. 2008; 89, 759–79. 29. M. Metwally, C. M. Farquhar and T. C. Li. Is another meta-analysis on the effects of intramural fibroids on reproductive outcomes needed? Reprod. Biomed. Online 2011; 23:2–14. 30. A. Strandell, A. Lindhard, U. Waldenstrom, J. Thorburn, P. O. Janson and L. Hamberger. Hydrosalpinx and IVF outcome: A prospective, randomized multicentre trial in Scandinavia on salpingectomy prior to IVF. Hum. Reprod. 1999; 14: 2762–9.

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When Fertilization Fails Tanya Milachich and Atanas Shterev

Introduction

What Is Fertilization Failure?

Fertilization failure (FF) is fortunately a rare event. The current in vitro fertilization (IVF) rate is between 70 and 80 percent. In this chapter, we discuss the various circumstances when fertilization failure may occur and make suggestions as to how to overcome this. Practitioners in the assisted reproductive technology (ART) field need to realize, however, that at this juncture in our field, the mechanistic understanding of FF is yet incomplete. While more research and clinical trials are being planned and are under way, it is important that we are honest, both to ourselves and to our patients, about the limitations of the technology currently available to “fix” FF. Currently, IVF laboratories achieve fertilization rates of 70–80 percent of all mature oocytes [1, 2]. Fertilization is usually assessed 16–18 hours post insemination and if there is a presence of two pronuclei (2PN) (male and female) and two polar bodies (Figure 35.1), the fertilization is judged as successful. Fertilization can also occur abnormally where potentially all or the majority of zygotes have more than two or multiple pronuclei observed 16–18 hours post insemination or later [2].

FF in clinical IVF and intracytoplasmic sperm injection (ICSI) is manifested by the absence of PN, the presence of only one PN (male or female) or the presence of multiple PNs. When fertilization fails, it also ironically sheds some light on potential key factors resulting in the couple’s subfertility, and, in many instances, also gives us insight into possible solutions, in the laboratory but also clinically [2, 3]. FF may be a recurrent phenomenon (up to 30 percent) occurring after conventional IVF or after ICSI [4]. In order to improve the outcome in cases of previous fertilization failure (PFF), several strategies have been proposed [2–4] (Figure 35.2), although one must be aware that many of these options are experimental, and not supported by robust evidence. Fertilization failure can be “total” (total fertilization failure or TFF), where all the oocytes after insemination (16–20 hours) do not show any sign of pronuclei formation after IVF or after ICSI. This can occur in 1–5 percent of all ICSI cases, which in the United Kingdom affects 1,200 couples annually [5]. TFF is often attributed to very severe malefactor infertility in IVF, such as cryptozoospermia, oligoasthenoteratozoospermia (OAT), necrospermia and globozoospermia [3, 4], although TFF in ICSI is more likely related to crucial oocyte activation factors. It is still controversial if the assessment of sperm genetic parameters, e.g., DNA fragmentation is useful in improving pregnancy rates in IVF/ ICSI, but it may be possible to counsel patients if these parameters are known to be poor [3]. In a normal cycle of IVF or ICSI, approximately 70 percent of mature oocytes are expected to fertilize. Most laboratories set a key performance index of 50–60 percent of fertilization of all oocyte yield per cycle. A lower fertilization rate of less than 30 percent after ICSI or 10–40 percent after

Figure 35.1 Fertilized oocyte with 2PN and two polar bodies

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Environmental and laboratory factors (air, %CO2, Temp., HEPA filters, experience, performance)

Evaluation of the maturity and quality of the oocytes and sperm cells

Majority of oocytes in M2

Minority of oocytes in M2

None of the oocytes in M2

Severe male factor (bad morphology, OAT, necrospermia, TESE etc.)

Reassessment of: COH strategy, periconception health (BMI, diet smoking), re-evaluation of medically relevant conditions e.g. endometriosis, chronic pelvic inflammatory Disease, chronic prostatitis, cystitis

Karyotype female and male partner

Possible new genetic testing for maturation arrest-optional

Anti-inflammation and endometriosis therapy, new diet, COH optimization of protocol, physiotherapy etc.

Additional treatment strategies according to results and female age

Total fertilization failure (TFF) in M2 oocytes

Male karyotype, sperm DFI , Kruger morphology, SAT, sperm membrane polarization MSOME/IMSI, MACS, PGS, donor sperm, anti-inflammation therapy

Artificial oocyte activation Oocyte spindle birefringenceevaluation and position

Low fertilization rate (LFR) in M2 oocytes

PGS

Donor oocytes

Abnormal fertilization e.g. majority 3PN zygotes

Perform ICSI only Oocyte spindle birefringenceevaluation and position

Figure 35.2 Possible practical solutions when fertilization fails

conventional IVF can sometimes be observed due to reasons such as the lack of mature oocytes, low overall oocyte numbers, abnormal oocyte and severe male factor [2–4]. Hence, the fertilization rate of any laboratory depends, in part, on the patient characteristics, with poorer-prognosis patients, e.g., older patients and those with poorer ovarian reserve, having poorer fertilization rates than the “gold standard” patients. Interestingly, unexplained infertility is the most frequent indication in total TFF cycles (usually more than 50 percent) and cycles with recurrent TFF (up to 65 percent). Among the patients with successful cycles, the most prevalent indication is male infertility (about 40 percent).

Optimizing the Laboratory Environment The conditions of the environment in the IVF laboratory are crucial for the success of the fertilization process [6]. Hence, there are strict criteria to follow during the in vitro culturing of the human embryos, and the process needs to be subjected to regular audits. Environmental conditions such as purified air, temperature of equipment and relevant surfaces, strict control of the appropriate level of gaseous aeration and laboratory air qualities all contribute to the quality of fertilization and embryo development. Guidelines from European and American societies provide recommendations on the general organization of an IVF

When Fertilization Fails

laboratory, including quality management and laboratory safety [6, 7]. These guidelines are focused on the specific aspects of the procedures performed in IVF laboratories from the identification of patients and reproductive cells, both oocytes and spermatozoa, through consumables and biological material to oocyte and sperm preparation, insemination of oocytes and scoring for fertilization [6] (Figure 35.2). Despite achieving the optimal laboratory conditions, fertilization can still fail. However, optimizing the drug regime and dosage for controlled ovarian hyperstimulation (COH) and the timely administration of human chorionic gonadotropin (hCG) for triggering ovulation is useful for ensuring that an adequate number of mature oocytes is retrieved. This will be important if a review of the previous cycle showed an exceedingly high number of immature oocytes [8].

Multidisciplinary Input In reviewing patients with a history of TFF, it is crucial that the embryology team work together with clinicians to derive an appropriate diagnosis, if any, taking into account the prognostic factors such as diminished ovarian reserve (DOR), primary ovarian insufficiency (POI), premature ovarian failure (POF), advanced maternal age (AMA), endocrine disorders and/or the presence of chronic diseases [8, 9]. If treatable, these medical conditions should be attended to prior to another cycle of IVF treatment.

Optimizing Peri-Conception Factors Peri-conception factors are important for developmental programming [9, 10]. Several lifestyle factors, including body mass index (BMI), diet, alcohol intake, smoking and use of recreational drugs, can influence the outcome of IVF. Certain occupational hazards, such as prolonged exposure to significant levels of air pollution, radiation and heat, also have a detrimental impact on fertility [11] and have been associated with disturbed regulation of oocyte signaling, relocation of organelles and interference with metabolic pathways that are crucial to fertilization, cumulus penetration by the sperm, fusion of sperm with oocyte, egg activation and pronuclei formation. Other chronic inflammatory diseases implicated in TFF include pelvic inflammatory disease (PID), endometriosis, cystitis and prostatitis [12]. Previous studies have shown that fertilization rates were reduced

(< 25 percent) in stage I/II endometriosis and poorer pregnancy outcomes were observed in women with stage III–IV endometriosis compared to those without [9, 12, 13].

Male Factor Emerging evidence suggests that poor semen quality could account, in part, for some cases of TFF. Many instigate investigations, including genetic assessment of the male partner, evaluation of DNA fragmentation of the semen and magnetic activated cell sorting (MACS) on sperm, although none of these tests are evidence based for a better clinical outcome in terms of live birth rate [3, 4]. It has been reported that very high DNA fragmentation (> 50 percent) can result in poorer prognosis [14]. Additional techniques include the sperm aneuploidy test (SAT) and sperm membrane polarization evaluation in men with subfertility with low fertilization rate (< 25 percent), although these are not commonly adopted evidence-based techniques [15]. A new genetic test for maturation arrest for female patients with a TFF history has been described, but again is not commonly used due to its unproven efficacy [16]. Genetic counseling will be necessary if there was an obvious abnormality highlighted by karyotyping, e.g., translocation. In these cases, PGD may be an option. An ultimate solution, in the presence of recurrent TFF, which may not be suitable for everyone, is the use of donor gametes.

Individualization of Care Treatment for couples with TFF needs to be individualized. Some additional treatments before the next IVF cycle relate to the optimization of the patient’s general health [9, 10], improvement of lifestyle in terms of smoking, alcohol consumption and drug abuse, and optimizing or at least being mindful of occupational and environmental hazards [11]. In men with hypogonadotropic hypogonadism, treatment with gonadotrophins can improve the quality of spermatogenesis. Diets rich in vitamins, antioxidants and minerals have been proposed [3, 4, 9, 10, 11]. A practical approach is suggested in Figure 35.2.

Conclusion In conclusion, the mechanistic intricacies involved in normal and abnormal fertilization are still not completely understood. In perfectly normal conditions, in

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the presence of a single polar body, homogeneous cytoplasm, appropriate zona thickness and perivitelline space together with the injection of the “perfect spermatozoa,” TFF is still not completely avoidable and can be distressing. Hence, it is crucial that patients’ expectations are managed and explanations, where available, are provided clearly to assist them through what may otherwise be a difficult fertility treatment journey.

Suggested Standard Operating Protocol (SOP) Clinical SOP – Management of FF in the clinic

Suggested Audits Key performance indicators – fertilization rate including TFF rates

References 1. D. Payne, S. Flaherty, M. Barry and C. Matthews. Preliminary observations on polar body extrusion and pronuclear formation in human oocytes using time-lapse video cinematography. Hum. Reprod. 1997;12:532–41. 2. J. E. Swain and T. B. Pool. ART failure: Oocyte contributions to unsuccessful fertilization. Hum. Reprod. Update, Sep. 2008; 14:431–46. 3. P. Rubino, P. Viganò, A., Luddi and P. Piomboni. The ICSI procedure from past to future: A systematic review of the more controversial aspects. Hum. Reprod. Update, Mar. 2016; 22:194–227. 4. H. E. Chemes and V. Y. Rawe. Sperm pathology: A step beyond descriptive morphology. Origin, characterization and fertility potential of abnormal sperm phenotypes in infertile men. Hum. Reprod. Update, Sep. 2003; 9:405–28. 5. M. Yeste, C. Jones, S. N. Amdani, S. Patel and K. Coward. Oocyte activation deficiency: A role for an oocyte contribution? Hum. Reprod. Update, Jan. 2016; 22:23–47. 6. ESHRE Guideline Group on Good Practice in IVF Labs, M. J. De Los Santos, S. Apter, G. Coticchio, S. Debrock, K. Lundin, C. E. Plancha, F. Prados, L. Rienzi, G. Verheyen, B. Woodward and N. Vermeulen. Hum. Reprod. 2016 Apr.;31(4):685–6. 7. Practice Committees of American Society for Reproductive Medicine and the Society for Assisted Reproductive Technology. Recommendations for development of an emergency plan for in vitro fertilization programs: A committee opinion. Fertil. Steril. 2012 Jul.;98(1):e3–5.

8. S. G. Hillier. IVF endocrinology: The Edwards era. Mol. Hum. Reprod., Dec. 2013; 19:799–808. 9. J. F. Strauss and R. L. Barbieri. Yen & Jaffe’s reproductive endocrinology: Physiology, pathophysiology, and clinical management. In Expert Consult – Online and Print. 7th edn. Philadelphia, PA: Elsevier Saunders; 2014. 10. R. P. M. Steegers-Theunissen, J. Twigt, V. Pestinger and K. D. Sinclair. The periconceptional period, reproduction and long-term health of offspring: The importance of one-carbon metabolism. Hum. Reprod. Update, Nov. 2013; 19:640–55. 11. NICE Clinical Guidelines, No. 156. Fertility: Assessment and Treatment for People with Fertility Problems. National Collaborating Centre for Women’s and Children’s Health (UK). London: Royal College of Obstetricians & Gynaecologists; 2013. 12. H. M. Harb, I. D. Gallos, J. Chu, M. Harb and A. Coomarasamy. The effect of endometriosis on in vitro fertilisation outcome: A systematic review and meta-analysis. BJOG. 2013 Oct.; 120 (11):1308–20. 13. M. E. Coccia, F. Rizzello, G. Mariani, C. Bulletti, A. Palagiano and G. Scarselli. Impact of endometriosis on in vitro fertilization and embryo transfer cycles in young women: A stage-dependent interference. Acta Obstet. Gynecol. Scand. 2011; 90(11):1232–8. 14. M. Esbert, A. Pacheco, F. Vidal, M. Florensa, M. Riqueros, A. Ballesteros, N. Garrido and G. Calderón. Impact of sperm DNA fragmentation on the outcome of IVF with own or donated oocytes. Cited in Scopus 25; Reprod. BioMed. Online. 23(6): 704–10. 15. S. G. Brown, S. J. Publicover, S. A. Mansell, P. V. Lishko, H. L. Williams, M. Ramalingam, S. M. Wilson, C. L. Barratt, K. A. Sutton and S. M. Da Silva. Depolarization of sperm membrane potential is common feature of men with subfertility and is associated with low fertilization rate at IVF. Hum. Reprod. 2016 Apr. 6. pii: dew056. [Epub ahead of print]. 16. R. Feng, Q. Sang, Y. Kuang et al. Mutations in TUBB8 and human oocyte meiotic arrest. The New England Journal of Medicine 2016; 374:223–32. 17. J. Harper, J. Geraedts, P. Borry, M. C. Cornel, W. J. Dondorp, L. Gianaroli, G. Harton, T. Milachich, H. Kääriäinen, I. Liebaers, M. Morris, J. Sequeiros, K. Sermon, F. Shenfield, H. Skirton, S. Soini, C. Spits, A. Veiga, J. R. Vermeesch, S. Viville, G. de Wert and M. Macek Jr. ESHG, ESHRE and EuroGentest2. Current issues in medically assisted reproduction and genetics in Europe: Research, clinical practice, ethics, legal issues and policy. Hum. Reprod. 2014 Aug.;29(8): 1603–9. doi:10.1093/humrep/deu130. Epub 2014 Jul 8.

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What Does Poor Embryo Development Mean, and How Does It Influence Subsequent Cycles? Emma S. Adolfsson

The overall aim of the embryologist is to create, culture and select the best available embryo for transfer and/or cryopreservation, with the ultimate goal of giving the infertile patient a healthy baby. However, sometimes we are faced with the difficult task of delivering bad news either indirectly to the physician or directly to the patient. Sometimes none of the embryos is of good quality, all the embryos have developed poorly and the odds of achieving a pregnancy are very low. What does poor embryo development mean? This chapter aims to explain the background of poor embryo development, and what, if anything, can be done in the subsequent cycles to achieve a better outcome.

Embryo Development as It Should Be In order to discuss poor embryo development in the in vitro fertilization (IVF) cycle, normal development needs to be covered briefly. The mature oocyte and the sperm are haploid, each containing 23 chromosomes. When the sperm fertilizes the oocyte, the diploid zygote is formed. Development of the embryo after fertilization initially depends on a reserve of accumulated maternal mRNA. The first and second division up to the four-cell stage is driven exclusively by material from the oocyte. The unpacking of highly condensed paternal DNA from the sperm takes time, so contribution of paternal RNA on embryo development starts between the second and third embryo division (at the four-to-eight-cell stage). This is the same point in time when the embryo itself starts driving the development through embryonic gene activation and maternal genome degrades. Early human embryo development follows a strict pathway. One cell becomes two cells, each half the size of the first, two become four, four become eight and so on. Cell division continues, a morula of densely packed cells forms, and then a blastocyst evolves, with an inner cell mass and the

throphodectoderm. To select a high-quality embryo, the embryologist traditionally use microscopic assessments at fixed time points during the culture period. Cleavage-stage embryos are scored on the number of cells, the size and symmetry of these cells, the number of visible nuclei and the degree of fragmentation. On day 2, a four-cell embryo with even cell size, low degree of fragmentation and absence of multinucleation is considered ideal, and on day 3 an eight-cell embryo with the same features gives the best chance of pregnancy. At the blastocyst stage, it should be expanded with an inner cell mass consisting of many tight packed cells, and dense trophodectoderm.

What Is Poor Embryo Development? The strict and precise development required to make a normal embryo can go wrong at a number of places. The definition of poor-quality embryos is embryos that are displaying a high degree of fragmentation, uneven blastomere size or multinucleation. For instance, an embryo with fewer than six cells on day 3 of development, and/or more than 50 percent of fragmentation will have significantly reduced chances of achieving pregnancy. Formation of fragments, i.e. anuclear cellular debris formed by externalization cytoplasm parts, is another feature of poor embryo quality. Two distinctly different types of fragmentation have been documented by timelapse analysis in human embryos: definitive fragmentation, characterized as stable persistent fragments, and pseudo-fragmentation, characterized as transient fragments appearing during, or shortly after, cell cleavage. Small amounts of fragmentation do not impair the implantation ability of the embryo, but embryos with strong and persistent fragmentation are less likely to be viable and a higher percentage of fragmentation correlates with aneuploidy [1].

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Multinucleation is another phenomena included in the description and definition of poor-quality embryos. The magnitude of multinucleation on the viability of the embryo depends on at which stage the embryo displays multinucleation. An embryo with multinucleation at the two-cell stage has 20 percent probability of becoming a good-quality blastocyst. If a blastocyst is formed despite this multinucleation, the pregnancy rate remains reduced compared to a mononucleated embryo [2]. Multinucleation at the four-cell stage is correlated with aneuploidy and reduced implantation ability, but at the eight-cell stage, embryos are more likely to be mosaic and the impact of multinucleation is less pronounced. By using time-lapse information, the embryos can be assessed by their cleavage patterns and timing, known as morphokinetics. An embryo with normal morphokinetics develops from a single cell into two cells, then progresses to four, eight, 16 cells and so on, with enough time in between cell cycles to allow for correct DNA duplication and replication. Embryos dividing either too slow or too fast, or in an irregular fashion, may have metabolic and/or chromosomal defects. The first division when the zygote splits into a two-cell embryo is crucial. Embryos that cleave irregularly at this stage are more often aneuploidic with an extremely low implantation rate [3]. Common causes for direct cleavage are fertilization by more than one sperm or failure of extrusion of the second polar body. Another factor correlated with poor implantation ability is reversed cleavage. This too can only be captured using time lapse.

What Are the Causes of Poor Embryo Development? The fate of the fertilized egg depends on three things: the egg, the sperm and the IVF laboratory in which it grows. The quality of the oocyte may be compromised by the type of stimulation used, and the sperm selection process influences the embryo development. Finally, the in vitro culture and the skill of the embryologist may further reduce the potential of the growing embryo.

Oocytes: Cause of Aneuploidy and Influence on Embryo Development The oocyte contributes half of the genetic material, but almost all of the building blocks, the energy and the

shell in which the embryo forms. Therefore, the quality of the egg is essential for the quality of the embryo. The oocyte ages as the woman ages. At the start of the woman’s fertile period, about 80 percent of the oocytes are euploid, and as her fertile period ends, only 10–20 percent of the oocytes are euploid, while the rest display various aneuploid features. The oocytes have rested in meiosis I since the fetal period. Meiosis resumes during ovulation and completes during fertilization to form the haploid gamete. The process to release a highquality oocyte one at a time during a woman’s fertile period is controlled by complex interactions between the oocyte, the surrounding cells and hormones. The oocyte’s potential to turn into a good-quality embryo is largely determined by genetics. But the ability to express that potential depends on a complex interaction between the oocyte and the environment, driven by hormones. The controlled ovarian stimulation used in IVF may therefore reduce that quality [4]. Preimplantation genetic screening (PGS) analysis has shown that the type of stimulation regime results in a different degree of aneuploidy; mild stimulation results in more euploid embryos compared to longtime stimulation with higher doses [5]. Moreover, the choice of triggering method affects the outcome. The triggering of oocytes can be achieved by the administration of human chorionic gonadotropin (hCG) or gonadotropin-releasing hormone agonist (GnRHa), where the latter causes the release of endogenous luteinizing hormone (LH) and folliclestimulation hormone (FSH) and therefore mimics the natural cycle surge. Comparing outcomes between the regimes shows that GnRHa is comparable, or in favor, in regards to the yield (number of oocytes in relation to number of follicles), the percentage of mature oocytes and the number of top-quality embryos generated when fertilized [6]. The link between aneuploidic oocytes and morphology is poor. Therefore, the embryologist cannot deselect against aneuploidic oocytes based on their appearance after oocyte retrieval. The only oocytes to discard prior to fertilization are giant oocytes that are diploid and give rise to triploid embryos [1]. Aneuploid oocytes will give aneuploidic embryos, and the link between aneuploidic embryos and morphology is poor too. Some aneuploidies result in arrested development – defined as absence of cleavage for more than 24 hours. Most often embryos arrest on day 3 of development. This coincides with the time point at which the embryonic gene activation

Poor Embryo Development

takes place. If the embryonic genome is nonfunctional, the development halts and does not proceed past the cleavage stage. Degan Walls and coworkers showed that some trisomies and monosomies are not tolerated past the cleavage stage. But many aneuploidies did not impact the development or quality of the embryo as measured using standard morphological observations. Despite being aneuploidic, the embryo continued to develop into a blastocyst. This highlights that blastocyst culture itself is no guarantee for euploid embryos, and that embryologists cannot deselect aneuploidic embryos based on morphological features or stage of development. However, since the majority of aneuploidies are not seen in later pregnancy, the window for deselection against these embryos seems to be around the time of implantation or shortly thereafter [7]. It appears as if morphokinetics is affected by the chromosomal status of the embryo, i.e. the growth and division of healthy embryos is different from aneuploidic embryos. Several aneuploidy models to identify embryos at risk of being aneuploidic without performing PGS have been proposed. Further research is needed to refine and confirm these models.

Sperms: The Effect of Sperm Integrity and Sperm Selection Methods The sperm contributes half the chromosomal content of the embryo. Damage to the genetic contents of the sperm will show as abnormal chromatin structure, chromosomes with microdeletions, aneuploidies or DNA strand breaks, i.e. sperm DNA fragmentation. In vivo, a selection process occurs that allows only sperm with intact DNA to fertilize an oocyte. In vitro, sperms with damaged DNA may fertilize an oocyte. Since the first two divisions are exclusively maternal derived, the effects will be noticeable first at the end of the second or third cleavage (from the four-to-eightcell stage). The effect will manifest as slower embryo development, poor morphology, or at the extreme as arrested development [8]. Several studies have shown an association between sperm DNA fragmentation, altered sperm nuclear morphology and poor embryo quality. Therefore, the use of intracytoplasmic morphologically selected sperm injection (IMSI) may improve embryo development. Indeed, several studies show that IMSI is associated with improved implantation and clinical pregnancy rates as well as lower abortion rates when compared to ICSI. Selection of the sperm at 6,000

x magnification compared to 200–400 x will aid in selecting a sound sperm for the fertilization [9]. Another selection tool is magnetic activation cell sorting (MACS). MACS detects all stages of apoptosis and is based on loss of membrane integrity. When the sperm is selected for apoptosis, one of the first signs is loss of membrane integrity with subsequent externalization of phosphatidylserine. This substance has a high affinity for Annexin V, and Annexin V bound to magnetic beads can therefore be used to remove affected sperm from normal sperms. MACS efficiently reduces sperm DNA fragmentation levels and separates apoptotic from non-apoptotic sperms. This selection improves sperm quality and functionality, and sperm selection based on MACS increases pregnancy rates [10].

Laboratory: In Vitro Culture and Reprotoxicity All handling and processing of gametes and embryos has detrimental effects on the developing embryos. Reprotoxicity is defined as the negative influence on the physiology and viability of human embryos by the laboratory and the consumables used in the culture of the embryos. The reprotoxicity causes cumulative damage, resulting in reduced viability and possible reduced pregnancy rates and ongoing pregnancy rates. Under the term reprotoxicity almost all things can be gathered: air quality, temperature, culture media, incubators, consumables – specifically plastics – education and experience of the embryologists. Each area needs to be optimized in order to minimize those negative effects. Some processes within the lab are well known to affect embryo development: – Mode of ICSI. The fertilization rate, the degeneration rate and the rate of embryo arrest significantly improve with time; this reflects a learning curve of the ICSI procedure itself. Moreover, selecting a morphological normal sperm may improve outcome as they are less likely to be aneuploidic [11]. – Exposure to low temperatures. Low temperature depolymerizes the meiotic spindle that separates the chromosomes during cell division. Resuming normal temperatures will resume the meiotic spindle, but with higher risk of aneuploidy in the developing embryo. Incubators, oocyte retrieval workstations, ICSI work stations etc. need to be calibrated to maintain correct temperature.

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Hormone stimulation

Reprotoxicity from in vitro culture Accumulated negative effects of suboptimal culture conditions. Toxicity from consumables. Early and long-term effects of culture media.

Oocyte quality

Sperm integrity

Sperm processing

ICSI technique

Sperm selection

Arrested development Sperm RNA impact Nonfunctional embryonic genome.

Figure 36.1 This figure illustrates the main causes of poor embryo development. The gametes themselves have a major impact on the fate of the embryo. Most important is oocyte quality, which can be compromised either due to being genetically abnormal, or due to the effects of supra-physiological hormone levels during controlled ovarian stimulation. Sperm integrity, for example sperm DNA fragmentation, also affects embryo development. Some processing techniques reduce the number of abnormal sperms. Moreover, selecting sperm for ICSI using higher magnification may improve embryo quality. The IVF laboratory and embryologists can negatively affect embryo development, a phenomena described as reprotoxicity. This is due to stress from suboptimal conditions, toxicity from the consumables, culture media and/or additives. Since the initial cell divisions are driven by the maternal genome and maternal building blocks, any effects of damaged sperm RNA or nonfunctional embryonic genome are noticed around the eight-cell stage on day 3 of development, where many embryos arrest.

–

Choice of culture media. Interestingly, culture media affects not only early embryo development but also birth weight. This is in accordance with the Barker hypothesis (Developmental Origins of Health and Disease). Small differences in birth weight may reflect more subtle disturbances that will manifest later in life [12].

Time-lapse incubators can reduce reprotoxicity. They allow for a less disturbed culture with more stable temperature and pH, while the embryologist gains knowledge about the embryos from detailed embryo development analysis.

Can We Know the Cause of Poor Embryo Development? Clues regarding the cause of poor embryo development can be found from laboratory notes. If the oocyte yield is poor (a low number of oocytes retrieved compared to

the number of follicles), or if the oocytes appear dark pitted or atreic, it indicates a stimulation issue. A high number of immature oocytes indicates triggering issues. If embryo development deteriorated prior to the activation of the embryonic genome, i.e. prior to the eight-cell stage, it indicates oocyte quality issues. If embryo development deteriorated or halted at or past the eight-cell stage, the cause might be nonfunctional embryonic genome, or effects of paternal RNA.

Will Poor-Quality Embryos in One Cycle Mean Poor-Quality Embryos in the Subsequent Cycles? Having poor embryo development of all embryos in an IVF cycle, with possibly no embryos available for clinical usage, is devastating news. Is it a one-off or a repetitive finding? As this chapter has highlighted, the causes for poor embryo development (Figure 36.1) might be 1) poor gamete quality as in genetically abnormal oocytes

Poor Embryo Development

or sperms; 2) induced poor-quality oocytes due to suboptimal ovarian stimulation; or 3) induced poor embryo development due to laboratory suboptimal conditions. Repeating exactly the same treatment regimes in the subsequent cycle is likely to lead to the same outcome. By individualizing the controlled hormone stimulation, any negative effects on oocyte quality might be reduced, leading to improved embryo development. Sperm DNA damage analysis and selection of sperm based on MACS or the use of IMSI might aid with selecting euploid sperms. Tight quality control in the laboratory and the elimination of any suboptimal culture conditions will improve the outcome for all patients at the facility. The ultimate goal for the IVF team is a healthy baby for the infertile patient. Only when all things affecting embryo development are considered and optimized can that dream be achieved. Hopefully this chapter has highlighted the contribution of the gametes on embryo development, and the importance of proper individualized stimulation not to compromise on oocyte quality. Once fertilized with a high-quality sperm, the oocyte needs to be cultured in an environment as free of reprotoxicity as possible to achieve its full potential.

References

5. E. Baart et al. Milder ovarian stimulation for in-vitro fertilization reduces aneuploidy in the human preimplantation embryo: A randomized controlled trial. Hum. Reprod. 2007 Apr;22(4):980–8. 6. R. Orvieto. Triggering final follicular maturation: hCG, GnRH-agonist or both, when and to whom? J. Assist. Reprod. Genet. 2017;34(9):1231–2. doi:10.1007/ s10815-017-0982-7. Epub 2017 Jun 27. 7. E. Fragouli, S. Alfarawati, K. Spath, S. Jaroudi, J. Sarasa, M. Enciso and D. Wells. The origin and impact of embryonic aneuploidy. Hum. Genet. 2013; 132(9): 1001–13 doi:10.1007/s00439-013-1309-0. Epub 2013 Apr 26. PMID: 23620267. 8. J. Tesarik. Paternal effects on cell division in the human preimplantation embryo. Reprod. Biomed. Online 2005;10:370–5. 9. G. Lo Monte, F. Murisier, I. Piva, M. Germond and R. Marci. Focus on intracytoplasmic morphologically selected sperm injection (IMSI): A mini-review. Asian J. Androl. 2013 Sept.;15(5):608–15. 10. M. Gil, V. Shar-Shalom, Y. Melendez Sivira, R. Carreras and M. A. Checa. Sperm selection using magnetic activated cell sorting in assisted reproduction. J. Assist. Reprod. Genet. 2013 Apr.;30(4): 479–85. doi:10.1007/s10815-013-9962-8. Epub 2013 Mar 7.

1. L. Rienzi, B. Balaban, T. Ebner and J. Mandelbaum. Atlas of Human Embryology: From oocytes to preimplantation embryos. Hum. Reprod. 2012;27 (suppl. 1).

11. J. Dumoulin, et al. Embryo development and chromosomal anomalies after ICSI: Effect of the injection procedure. Hum. Reprod. 2001;16(2): 306–12.

2. E. Adolfsson and B. Bartholomeuw. Unpublished results.

12. S. H. Kleijkers, E. Mantikou, E. Slappendel, D. Consten, J. van Echten-Arends, A. M. Wetzels, M. van Wely, L. J. Smits, A. P. van Montfoort, S. Repping, J. C. Dumoulin and S. Mastenbroek. Influence of embryo culture medium (G5 and HTF) on pregnancy and perinatal outcome after IVF: A multicenter RCT. Hum. Reprod., 2016;31(10): 2219–30. doi:10.1093/humrep/dew156. Epub 2016 Aug 23.

3. C. Rubio et al. Limited implantation success of direct-cleaved human zygotes: A time-lapse study. Fertil. Steril. 2012 Dec;98(6):1458–63. 4. M. A. Santos, E. W. Kuijk and N. S. Macklon. The impact of ovarian stimulation for IVF on the developing embryo. Reproduction. 2010 Jan;139(1):23–34.

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The ABCs of Quality Management in IVF Oksana Stidston

Introduction The quality management system (QMS) is a method of streamlining and standardizing every process within a fertility clinic, encompassing clinical and laboratory pathways, procedures and processes. This chapter aims to describe the fundamental aspects of a QMS and what helps to make it robust and efficient. The chapter gives a brief overview of: • the rationale for having a QMS in an in vitro fertilization (IVF) clinic • the essential components of an efficient QMS • the quality management cycle and how it helps to control the clinic’s quality processes.

What Is a Quality Management System and Why Have It? It is a legal and regulatory requirement on IVF centers to establish and maintain QMSs that set standards of quality and patient safety.

Personnel

A QMS is a method of defining how an organization should meet the requirements of its customers and the associated stakeholders [1]. The overview of the QMS is shown in Figure 37.1. In addition to being a government requirement, there is a distinct growing recognition that quality management not only ensures improvement of the clinical aspects of the operations of a clinic but also leads to improved financial performance and increased staff satisfaction [2]. The QMS is an approach that puts the emphasis on “prevention rather than cure.” The system strives to facilitate a clinic to gain control, plan ahead, consider risks, have back-up plans and promote longterm continuous improvement of the service. Given the nature of IVF treatment where multiple factors can contribute to the result at many points of the treatment cycle, it is critical that a robust QMS is established to incorporate the workings of several teams: • clinical • nursing • embryology/lab

Role of the quality manager Quality policy

Quality documents Premises and equipment

Quality manual Quality objectives

Third party agreements KPIs Standard operating procedures Audits Validation The Quality Management System

Patient satisfaction Performance monitoring Staff satisfaction and feedback Quality control IQC Document control EQC Record keeping Non-conformities and incidents Accreditation

Figure 37.1 Key components of a QMS in an IVF clinic

The ABCs of Quality Management in IVF

• •

administration/management counseling.

The system should be prepared prior to the start of clinic activity and modified as the workload increases. The QMS should be reassessed at regular intervals and adjusted accordingly. A useful tool for monitoring quality is the classic cycle of Total Quality Management (the Deming cycle), or PDCA (Plan →Do → Check →Act). Following these stages of a quality cycle can be really helpful in maintaining consistency when trying to identify the root cause of a problem and implementing corrective actions (Table 37.1).

The Crucial Elements of the QMS Personnel Your clinic is only as good as your staff. While it is critical to hire suitably qualified and licensed

professionals with superior skills, it is also very important to keep staff enthusiastic and motivated about the quality of the clinic’s performance. Practices vary according to the culture of the clinic and the leadership style, from more hierarchical decision-making processes to that which is pro egalitarianism, with decisions made consensually or “top down.” Whichever the method adopted suitably in the right cultural context, staff should be involved in the major operational decisions and appropriately appreciated. Management must provide staff with suitable working conditions appropriate for the role and workload. Unambiguous job descriptions with clearly delineated responsibilities are conducive to higher efficiency and clearer working relationships between staff. Staff is instrumental in maintaining and improving quality. This part of the clinic has very often been

Table 37.1 Example of a clinic’s quality activity monitoring

Nature of review

Monitoring method

Monitoring frequency

Review of internal errors

• Operations meeting discussion

Biweekly

Patient complaints

• Management team discussion

Weekly or Biweekly

Incidents and nonconformities

• Team discussions • Root cause analysis

Weekly or Biweekly As needed

Patient satisfaction evaluation

• Audit of patient questionnaires • Semi-structured patient interviews

Quarterly

Semen production room feedback

• Patient questionnaires

Quarterly

Personnel feedback, suggestions and complaints

• Online survey • Suggestions box • All-staff meeting

Biannually

Review of standard operating procedures (SOPs)

• Scheduled reviews by relevant staff • Based on new research developments and changes in protocols

Annually (or as scientific evidence base changes)

Internal audits

• Written reports • Electronic quality management systems

Annually

Key performance indicators

• Data analysis and team discussion

Quarterly (or more frequently in case of incidents, outliers or unusual drop in success rates, larger clinics)

Quality management review

• Management meeting with prepared analysis of data: trends, success and underperformance factors.

Annually

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overlooked by clinics, but staff involvement is paramount to the success of the QMS [3]. One of the biggest challenges is to convince people that the quality system is not just a “boring requirement” but an essential part of the clinic’s work and everyone is a participant. Raising adequate awareness, and placing emphasis in staff training in terms of customer service and “hostmanship” qualities, should ensure customer satisfaction and minimize complaints, which will in turn improve staff morale and work satisfaction. The latter should be regularly measured and suggestions collected (Table 37.1).

Quality Management, Policy and Manual and Quality Objectives The quality management could be handled by an individual or a group of individuals whose role is to ensure that the clinic provides a consistent and constantly improved service to the desired level of excellence. The individual or group will be trained and supported to oversee a quality system that effectively controls all the administrative, clinical and laboratory processes. A good quality manager will successfully champion everyone in the clinic to participate in a “quality approach” and be able to use soft skills to support an effective quality culture [4]. Quality policy describes the clinic’s commitment to quality and provides the framework for setting quality objectives, which are the defined goals for the clinic. The goals are measurable and consistent with the quality policy. The quality manual should be a specific and concise overview of the system, including the organizational chart defining reporting relationships.

Equipment and Premises The premises for the clinic should be fit for purpose and provide comfort, privacy and dignity for the patients and a safe working environment for the staff. All consultations should be carried out in a private environment. All equipment must be suitable for laboratory and clinical use, fulfill the existing quality, health and safety standards and be regularly upgraded and maintained. Critical items of equipment, including incubators and frozen gamete and embryo storage facilities, should be validated before use, calibrated

and serviced at regular intervals, monitored and alarmed [5]. Consumables must be appropriately marked, fully traceable and tested, e.g., 24-hour sperm survival, mouse embryo assay, and boxes of consumables should have appropriate storage outside the laboratory. In a good IVF center, the clinical and laboratory quality standards should be high, with emphasis placed on attention to details (Figure 37.2).

Third-Party Agreements Third-party agreements should be in place, where required, for procurement, testing or processing of gametes and embryos or in supplying services where quality and safety of gametes and embryos may be affected (e.g., sperm procurement, transport services etc.).

Clinic Documents The initial establishment of a QMS will involve mapping out in detail all the relevant clinical and laboratory pathways, processes and procedures, concurrently with the identification of key performance indicators (KPIs) [2]. Where relevant, flowcharts and tables can be derived to assist the process (Figure 37.3).

Document Control Standard operating procedures must be reviewed at regular intervals. Regulatory bodies often require annual review of clinical SOPs. Standard operating procedures must also be amended immediately if the process is changed. It is important that only the current versions of the documents are in use. Any old versions should be made inaccessible. The appropriate investment in an electronic QMS that enables paperless or “paper-light” storage of records will reduce the burden of storage of paper records, and facilitate easy access by all the staff members to key laboratory or clinical documents required. The ISO 9001 standards allow flexibility within the organization to specify minimum and maximum retention requirements. National regulations differ, e.g., in the United Kingdom, the Human Fertilisation and Embryology Authority (HFEA) requires 30 years’ storage of all patient records. A center must also keep an incident logbook (spreadsheet). There should also be a log of morbidity associated with assisted conception such as ovarian hyperstimulation syndrome cases

The ABCs of Quality Management in IVF

CE Marking

MEA Tested

Appropriate Storage Conditions

Logs of Batches

Unique Codes for Treatment Cycles

Patients Appropriate Labelling System

Consumables

Donors

Lab Design

Exceptional Air Quality

VOC Control

Temperature

Clean Air Technology

Daily Logs

CO2 Levels

Humidity

Infection Transmission Prevention

Dewar Levels

Cleaning

Figure 37.2 Key elements of lab design (not exhaustive)

SOPs

General

Methods

Denudation

ICSI

Cryopreservation

Oocyte Retrieval

etc...

Labelling

Witnessing

Lab Safety Procedures

etc...

Equipment

Incubators Microscopes

Biosafety Cabinets

etc...

Figure 37.3 Example of laboratory SOPs flowchart (information is not exhaustive)

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and ectopic pregnancies. Emergency procedures should be clearly defined and made available to staff, and out-of-hours instructions provided to patients.

Quality Control A standardized, consistent and well-documented microscopic and morphological assessment of human gametes and embryos is a critical point for quality assurance. Such data should be stored for easy accessibility for KPI purposes. Internal quality control (IQC) is a process that allows for comparison of laboratory results, taking into account within-laboratory and inter-operator variation. External quality control (EQC) provides an independent method of assessing accuracy and overall quality of laboratory performance, in addition to internal quality control. Many national (UK) and international quality control schemes are available (e.g., the National External Quality Assessment Service), and there is usually a cost to participate. While computer-assisted analysis programs may provide an opportunity to enable accurate and reproducible semen analysis results, the system has to be appropriately validated and verified to provide the necessary quality assurance. Where semen analysis, be it computed assisted or by a trained andrologist, is performed with robust quality control in place, there is an increased probability of correlation between quality assurance at semen analysis and pregnancy [6]. Both IQC and EQC are important for standardization of results.

Validation Validation refers to establishing what the “product” (equipment, process) should actually do and checking it to ensure it does precisely that. It is ensuring that the process will consistently produce an outcome or a “product” that meets predefined acceptance criteria. Validation is a requirement from the HFEA in the United Kingdom and the EU Tissue and Cells Directive (EUTCD) in Europe in general, and what is to be validated is very much predefined by the clinic itself. Monitoring clinic performance is important to ensure the system is working. Several aspects of QMS assist with this:

1. 2. 3. 4.

KPIs audits nonconformities and incidents reporting and patient feedback.

Key Performance Indicators (KPIs) The criteria for KPIs are defined using the clinic’s own data or relevant scientific literature. When data are beyond the acceptance criteria, the process is out of control and should be investigated and steps taken to correct the issue. KPIs should be monitored regularly and KPI meetings held to discuss results. The meetings should be scheduled at least quarterly. For larger clinics, it might be practical to analyze KPIs more frequently, monthly or biweekly due to the greater size of their datasets.

Audits Audits help the clinic achieve its objectives by systematically evaluating compliance with processes, SOPs and records. Audits can be: • procedural (e.g., oocyte retrieval) • horizontal (e.g., IVF consents conformity) or • vertical (e.g., IVF 1-patient pathway). Audits can be performed by any member of staff who has received suitable training, but they should not audit their own area of work (e.g., nurses audit lab etc.).

Nonconformities and Incident Reporting A clear process for reporting incidents and nonconformities should be established. Nonconformities should be carefully evaluated to assess for causation and effective plans made for future avoidance of these incidents. Serious or recurring problems require corrective action. A plan should be documented and actioned (e.g., staff offered additional training and support, batches of consumables removed, suppliers changed, procedures reviewed etc.) and an audit performed following implementation.

Patient and Staff Satisfaction and Feedback It has been emphasized in recent years that although pregnancy rates are important, they provide no information about the treatment process itself and opportunities for improvement [7]. High-quality assisted reproduction technology (ART) is inextricably patient-centered [8].

The ABCs of Quality Management in IVF

Table 37.2 Example of problem analysis (increased egg damage rate during ICSI) and management using PDCA quality cycle approach. (Modified from Embryolab TQM workshop discussions, 2016) [4]

Plan

Do

Check

Act

Identify the problem, collect pertinent data and determine the root cause, where possible

Develop and implement a solution; decide upon a measurement to assess its effectiveness

Confirm the results by comparing data before and after a new solution

Document the results, inform staff about process changes. Make recommendation for the next PDCA cycle.

Problem: increased egg damage rate during ICSI Investigate the following potential causes (the list is not exhaustive): Product • patient age/egg quality • blocked ICSI pipettes • size of ICSI pipettes • batches of needles • batches of media, PVP • ICSI rig serviced recently? Process • multiple rounds of ICSI People (doctors; embryologists) • Level of training Procedures • aspiration force • pump pressure in procedures • single lumen needle • draining all follicles at egg collection, even the very small ones

• Discuss • Suggest and agree on the corrective action • Remember to only change one parameter at a time. Otherwise you will not know what made a difference.

• Benchmark against “gold standard patients”; donor eggs and donor sperm • Gather data and see if solution has worked

• Keep the results transparent • Discuss openly, seek solution • Do not apportion the blame • Despite the best efforts, the explanation might never be found • Start fresh PDCA cycle (see Plan)

An invaluable source of information for the clinic’s continuous improvement and ultimately, profitability, is the feedback from the patients themselves. Satisfaction questionnaires assist in identifying any gaps in service and sources of dissatisfaction, and provide insight into consumer behavior. Importantly, there is also evidence that active patient participation helps patients successfully cope with the extreme psychological stress of their treatment journey [9]. Complaints should be dealt with quickly and in a cordial manner. Good and clear communication throughout the treatment will reduce misunderstanding and complaints. The clinic that is responsive and caring significantly enhances the quality of the patient experience.

Accreditation Although there are no fixed accreditation standards for ART centers, ISO 9001–2015 for QMSs, is considered the gold standard fitting for an IVF clinic. The ISO standards offer clinics recognition as an internationally endorsed QM system.

Conclusion An effective QMS provides the framework for monitoring the performance of an IVF clinic (see Table 37.2), sets the targets, picks up the trends and provides the tools for analysis of underlying reasons if things go wrong. A QMS is a way of standardizing the inputs and outputs of the clinic at all times.

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Experience shows that the key word in quality management in a clinic is “trust.” A culture of fear, blame or retribution is lethal to quality efforts. The key reasons for failure to implement the quality system are: • insufficient human or financial resources • lack of mutual respect and appreciation for everyone at the organization • lack of support from the management • inappropriate environment, resistance to change and wrong attitudes [10]. Promoting a safe, supportive, no-blame learning atmosphere, where open exchange of views and information is welcomed, is instrumental to high levels of staff motivation and enthusiasm, effective communication and, as a result, to improved delivery of fertility treatment. Quality is an attitude, a habit, a philosophy; it is never finalized, but is a process that takes time and joint effort to build and maintain.

References 1. International Organization for Standardization. ISO 9001–2015 – How to Use It. Geneva, 2015. 2. J. Olofsson, M. Banker, and P. Sjoblom. Quality management systems for your in vitro fertilization clinic’s laboratory: Why bother? J. Hum. Reprod. Sci. 2013;6(1):3–8. 3. K. Tilleman et al. Belgium: ISO 9001:2000 Certification as a base for total quality management in ART.

In F. Bento, S. Esteves and A. Agarwal (eds.) Quality Management in ART Clinics. Boston, MA: Springer, 217–23. 4. Embryolab Academy TQM Workshop Notes. Belgium: Leuven, 2016. 5. M. C. Magli, E. van den Abbeel, K. Lundin, D. Royere, J. van der Elst, L. Gianaroli et al. Revised guidelines for good practice in IVF laboratories. Hum. Reprod. 2008;23:1253–62. 6. J. Bonde, E. Peter et al. Relation between semen quality and fertility: A population-based study of 430 first-pregnancy planners. Lancet 1998; 352(9135): 1172–7. 7. I. W. van Empel, W. L. Nelen, R. P. Hermens and J. A. Kremer. Coming soon to your clinic: High-quality ART. Hum. Reprod. 2008;23:1242–5. 8. W. L. Nelen, R. P. Hermens, S. M. Mourad, E. C. Haagen, R. P. Grol and J. A. Kremer. Monitoring reproductive health in Europe: What are the best indicators of reproductive health? A need for evidence-based quality indicators of reproductive health care. Hum. Reprod. 2007;22:916–18. 9. S. Gameiro, J. Boivin, L. Peronace and C. M. Verhaak. Why do patients discontinue fertility treatment? A systematic review of reasons and predictors of discontinuation in fertility treatment, Hum. Reprod. Update 2012; 18(6):652–69. 10. S. Mortimer and D. Mortimer. Quality and Risk Management in the IVF Laboratory. Cambridge: Cambridge University Press, 2005.

Chapter

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Human Factors in IVF Practice Rachel J. Broadley

Human Error Is Normal Even the most highly skilled and knowledgeable health care professionals and best-performing teams make errors, without intent to cause harm. Given the rigor of pre-employment checks and competency-based training, it is unlikely you would be doing your job (unless in a supervised/ training capacity) if you did not have the requisite knowledge and ability. Blame directed toward an individual, or yourself, is not ultimately a productive process. Solutions are not drawn from blame and can lead to very damaging consequences for group climate and people’s willingness to share information. Professionals are only too aware how accountable they are for their mistakes [1, 2]. Consider what happens when error occurs. Individual blame, particularly in a public arena, rarely has a productive outcome. Indeed, punitive measures (including public reprimand) generally drive information underground. Any chance of staff trusting in a transparent culture after witnessing that behavior is all but lost. Now, imagine a culture whereby errors and near-misses are openly discussed in staff forums. The individual, supported by senior colleagues if necessary, presents the facts of the case in question. Senior staff openly discussing the same with transparency (and maybe a little humility) is very powerful and is a huge step toward supporting an open culture. This could be where a technical error was made or, with the benefit of hindsight, it was considered a patient could have been offered a different course of treatment that could have led to an improved outcome. All staff then see genuine discussion and learning and understand the value of their contribution. Even the most highly skilled and knowledgeable clinicians make errors. What if you were about to make a mistake? Would you want one of your

colleagues to tell you? If their last experience was you publicly reprimanding them or another colleague, or overhearing gossip about a “stupid” mistake Dr. X had made, how inclined do you think they will be to speak up on your behalf?

Human Performance Is Variable At the 2009 World Championships, Usain Bolt broke his own world record with a 100 m time of 9.58 seconds. This remains Bolt’s personal best (despite his incredible performance at the 2016 Olympic Games) [3]. The point is that we only have one personal best. The very best moment on our very best day. Every other time we are operating below our personal best. We can think about this in terms of having a finite mental capacity. We have a professional task to complete, but any number of other (often interrelated) issues are also occupying some of that capacity and thus affecting our performance relating to that task at any given moment. Physiology Lack of sleep, lack of food/hydration and intercurrent illness are all obvious factors that can impact performance. Psychology There is much more to us than our professional lives, but we have to bring our whole self to work. Personal pressures (home/family/finance) and worry are distracting to task performance. We are probably all aware of how the sudden impact of bad news immediately affects our ability to think about anything else. Life does not stop, but we can be mindful of when we might be vulnerable and take reasonable mitigating actions wherever possible (even a simple conversation with a coworker can help). Environmental The immediate environment, not surprisingly, influences our performance (temperature, humidity,

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lighting, ergonomics of workspace). Unfamiliar new technology (or failing old technology) can also quite clearly impact how well a task is executed. Consider also overall workload. While it is easy to recognize that task and time pressure from very high workload demands will affect performance, our individual ability to control workload in health care is frequently limited. Indeed, when an unanticipated medical emergency occurs, our mental capacity can swiftly become saturated. We are suddenly required to mentally process the unexpected event, weigh the options and immediately execute an intervention: “Don’t just stand there, do something!” It is virtually impossible to think and act at the same time, which is why well-rehearsed scenarios are vital (e.g., life support training). Through this we develop a common vocabulary to suit the incident, clear expectations about roles and responsibilities are set and, because this is “rehearsed,” we enhance our ability to see the wider situation and thus provide ourselves with that capacity to “think.” Without a doubt, these examples are not exhaustive, but, if we accept that error is normal and we are likely working somewhere below our absolute personal best, increasing standardization may seem the obvious answer to mitigate these human fallibilities, although this is not without risk either. Professionals value latitude and, if this ability is too highly constrained, staff can quickly disengage, resulting in systems being ignored, “work-arounds” becoming the norm or even flagrant disregard of the “rules.” Indeed, after a safety incident, it is not uncommon for more checks and steps in a process to be implemented with staff feeling further constrained and limited in autonomy. Overall, however, removal of unwarranted variation in process does improve safety and enables teams to work more cohesively and communicate more efficiently [4]. An aircraft captain could not just simply arrive, fire up the engines and take off without all the requisite checks and measures being complete (thank goodness!). Health care delivery is no different.

Teamwork The whole purpose of working in a well-functioning team is that this is a more effective and powerful unit than simply a group of people carrying out the instructions of the person in the position of authority. Briefing The team brief is not just the purview of the operating theatre. It is here that clear expectations and goals for

the working session are set out. Without an articulated goal, how will the team know if something is happening to pose a threat to a successful outcome? The briefing is crucial to ensuring everyone has the same picture of how the tasks of the day will proceed. Elements to consider for an effective team brief include: - Choose the correct environment, away from interruptions with all team members present; nothing is more devaluing to a team member than considering their input in the brief is not required. - Use professional language. Social conversations are important, but that is not the purpose of this exercise. - Define the goals and expectations. - Define the roles and responsibilities of the team members. - Invite (and expect) participation. - Define what could go wrong and what the contingency actions will be. As we have discussed, our ability to “think” as events are unraveling can be severely compromised. If potential threats have been considered and planned for, the team can swiftly jump into action if necessary. Evidently, not all scenarios can be considered, but do you and all your team know where the nearest cardiac arrest trolley is? Do you have an agency member of staff who does not know how the bleep system works? Do you need a translator in order to consent the third patient on the list? - What other threats may there be to successfully completing the activity (e.g., staff sickness)? - Other operational factors – for instance, if a major incident has been declared in the area that is occupying staff, blood-bank and radiology, where is your back-up if you need assistance? Leadership and Authority Health care, like aviation, involves a hierarchy (usually several different lines) that largely defines accountability for judgments made and reporting relationships. While there must be a final decision maker, the team should expect their opinions to be solicited. Now, given that aforementioned hierarchy, junior team members may feel reticent to speak up, especially if it is not the norm for an authority figure to ask

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others for their opinions and input. If you are leading the brief, start by giving “permission” for your colleagues to speak up, for example: “Have I missed anything?” “Is there anything anybody wishes to ask or didn’t quite understand?” “If something happens that gives you concern, you must tell me.” “If you see ‘x’ happen, please say ‘y’” – e.g., “This is a difficult intubation. If the oxygen saturations reach 95 percent, you must tell me to stop and ventilate the patient,” or “If the blood in the suction reaches 500 ml, you must state this and ensure I acknowledge you.” Our hearing can often be impaired when we are becoming overloaded and we may not “hear” statements of concern if attention is entirely task-focused elsewhere. N.B. avoid the use of leading questions, e.g., “nobody has any questions, do they?”

Consider also how you will respond. If the prevailing environment is one of devaluing an individual’s input or the last person who spoke up had their head bitten off, it is unlikely the next person will want to speak up. But, thinking back to the earlier question, if you were about to make a mistake, would you want somebody to tell you? A positive and professional response toward someone with the foresight to raise a query is imperative. If you are in the decision-making position, you will be aided and supported and your judgments enhanced by the knowledge, experience and insights of others. You will be performing better than you ever could as an individual and have the entire team watching your back – a much more comfortable place to be. At this point in time, it is not about organizational culture or whether “management” support people speaking up (although this is clearly important). At that moment, it is about the relationships in the room where the conversation is being held and that is why the briefing is so important in this regard.

Checks and Checklists It is outside the scope of this chapter to discuss in detail the protocols and processes required in in vitro fertilization (IVF) practice [5]. However, certain general principles should be borne in mind to minimize error. While relevant to all areas, these may be particularly pertinent to laboratory-based tasks.

– Organizational protocols are mandatory and staff must be well-inducted to local procedure and policy. – Ensure the environment is conducive to the task and workflow. Interruptions are a common source of error and distractions must be minimized (limitation of noise/interruptions, e.g., telephone). – Understanding of other colleagues that some processes cannot be interrupted (synonymous with the nursing staff wearing “do not interrupt” tabards on medication rounds – this really does mean “do not interrupt” – even if you are a consultant!). If possible, in dialogue with colleagues, develop “protected” times for critical task completion. – Acknowledge the fatigue that comes with high cognitive intensity and repetitive tasks. While there is some inter-individual variability, the maximum attention span for a complex task is in the region of 20 minutes. Using the principles discussed earlier in relation to performance variability, recognize when you are at risk and not operating toward your personal best and how you can manage those individual pressures. – Are there too many checks? o If you have generated work-arounds to checks, stop and rethink. o Do you assume that someone else will pick up if there is actually an error further downstream? Let’s assume you do not go to work to do a bad job or to put patients in danger, but, if you make an error and are not following the recognized protocol, you are exposing the patients, yourself and the organization to considerable risk. It is very difficult not to apportion blame when procedure is not followed, but it is not unreasonable to question and discuss the protocol itself, gain a deeper understanding and discuss with senior colleagues about whether current procedures should be reviewed. – Are errors repeatedly identified during checks? o Do not rely on double-checks if a more fundamental redesign of the process is required.

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Double-checking is considered standard practice in many scenarios in health care. The problem is the two practitioners involved are rarely operating independently and bias can easily be illustrated. In other words, our brain is incredibly good at seeking information that confirms what we are expecting to see. Indeed, double-checking is frequently an element in highly time-pressured environments and in situations where the error rate is very low, i.e. you are not expecting there to be an error, putting further pressure on our cognitive ability to identify a mistake. Inevitably, when you have one fallible person checking the work of another fallible person, error will, on occasion, occur. Indeed, if you are the junior person performing the check and judge there to be an error, do you assume you are incorrect? However, these procedures are a crucial element in the whole package of approaches to enhancing safety. The most important element of dual checking is independence. If the second check is performed independently (i.e., without cues from the first person), a second person is highly unlikely to make precisely the same mistake as the first (e.g., when calculating a drug dose), an example of endogenous error. Exogenous error (e.g., poor work environment, lack of standard labeling protocols, poor design of packaging) is more difficult to eliminate with dual (even independent) checks. An additional element to consider with dualchecking procedures is the requirement for witnessing and double-checks, by their nature, cause frequent interruptions in the laboratory and clinical environment. Consider how the work in your local environment can be structured to minimize interruptions and what procedure to follow should you be interrupted during a checking sequence. In addition, if there is dependency on automation (e.g., barcoding), know what the manual contingencies are to ensure safety is not compromised in the event of system failure.

– What did we learn? – Are we going to do anything differently in the future? – Is there anything to report or any learning to be shared?

Incident Analysis Errors are past events. Try as we might, we cannot turn back the clock. The only course of action is to mitigate wherever possible the consequences of any error and learn from the incident. A non-punitive system for reporting incidents and near-miss events is to be encouraged, as is dialogue and participation of staff in generating solutions. This can only be achieved with a strong reporting culture and systems approach to investigation, i.e. individual error is not considered the root cause of an incident, rather a reflection of a larger organizational problem [6]. Indeed, analysis of historical incidents for system deficiencies, rather than considering an event in isolation, will allow identification of process weaknesses for targeted intervention. Adverse event and near-miss reporting systems will not overcome underreporting in a system where staff fear blame and disciplinary action in a culture unaccepting of error [7]. A philosophy whereby people are encouraged to expose flaws, even in processes for which they are responsible, is to be encouraged and staff rewarded in the relentless drive for harmfree care.

References 1. A. Green, L. Duthre, H. Young and T. Peters. Stress in surgeons. British Journal of Surgery 1990;77(10): 1154–8. 2. A. Sharma, D. M. Sharp, L. G. Walker, et al. Stress and burnout in colorectal and vascular surgical consultants working in the UK National Health Service. Psychooncology 2008;17(6):570–6.

Debrief/After-Action Review

3. www.biography.com/people/usain-bolt-1920702091

Something disastrous does not need to have happened, nor do you have to be in an operating environment, in order for a debrief to be valuable. Reflect on the goals set out in the brief: – Were the goals achieved? – Did anything happen that we were not expecting? – Were there any “near-miss” events?

4. NHS England: Standardise, educate, harmonise – Commissioning the conditions for safer surgery. Report of the NHS England Never Events Taskforce (February 2014). www.england.nhs.uk/patientsafety/never-events /surgical/ 5. Human Fertilisation & Embryology Authority: Code of Practice (8th edn.). www.hfea.gov.uk 6. Human Factors Analysis & Classification System. www .hfacs.com/hfacs-framework.html

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7. V. F. Nieva and J. Sorra. Safety culture assessment: A tool for improving patient safety in healthcare organizations. Qual. Saf. Health Care 2003;12:ii17–23.

The High Performing Surgical Team: A Guide to Best Practice. London: Royal College of Surgeons, 2013.

NHS England. National Safety Standards for Invasive Procedures (NatSSIPs). www.england.nhs.uk/patientsafety/ never-events/natssips/

S. Gordon, P. Mendenhall and B. B. O’Connor. Beyond the Checklist: What Else Health Care Can Learn from Aviation Teamwork and Safety (The Culture and Politics of Health Care Work). ILR Press, 2012.

National Patient Safety Agency. Five Steps to Safer Surgery. www.nrls.npsa.nhs.uk/

A. Gawande. The Checklist Manifesto: How to Get Things Right. Profile, 2011.

Suggestions for Further Reading

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39

Customer Service and Marketing in IVF Practice Helen Kennard

Introduction Fertility marketing requires a well-choreographed and integrated marketing strategy and communications plan. Steps are required to ensure the fertility clinic stands out from its competitors with clear and unique selling points that its prospective patients recognize and relate to, and also to offer easily accessible and first-class customer service. Marketing is defined by the Chartered Institute of Marketing as “the management process responsible for identifying, anticipating and satisfying customer requirements profitably.”

Where We Are Initially, you will need to analyze the fertility marketplace you’re in from all perspectives, including those of patients, referring doctors, hospitals, charity support organizations such as the Infertility Network UK (INUK), regulatory bodies such as the Human Fertilisation and Embryology Authority (HFEA) and vendors, including pharmaceutical firms. This will involve looking at external factors such as politics, economic stability, social factors, technological advances and legal and environmental influencers, and is known as a PESTLE analysis. According to the HFEA trends and figures report for 2013, the number of women receiving in vitro fertilization (IVF) and donor insemination (DI) treatment continues to grow. You need to fully understand where your fertility clinic currently sits in the fertility service arena. You will also consider your own clinic’s position, and a strength, weakness, opportunities and threats (SWOT) analysis is useful for this. This analyzes your strengths, weaknesses, opportunities and threats. It’s also important to arm yourself with as much information as possible about your competitors and their strategy.

Where We Want to Be Segmentation, Targeting and Positioning You will need to identify market segments or groupings of customer types you would like to pursue. Some common ways to categorize, or segment, people are: age, gender, ethnicity, geographical location, education level, income level, risk factors, pastimes and interests. For example, a segment might be professional women up to the age of 40 years old, who live within 30 miles of the practice and enjoy going on holiday two times or more each year.

Choosing a Target Market A critical decision in marketing is choosing the target market for each of your services. In fertility marketing, you may have a number of service propositions and associated target markets. Understand the characteristics and behaviors of the group. For example, examine what media they use, what they do with their leisure time (magazines, cinema, music, websites), how they communicate with one another, what languages they speak, what their opinions and knowledge are about infertility and what their values are. For example, you may target infertile couples where the female has poor-quality or no eggs at all for your egg donation program. You may also target healthy males aged between 18 and 40 years to become sperm donors for your clinic.

Positioning and Branding Positioning your fertility clinic and its services looks at how you wish your target audience to perceive your company and your product. For example, quality and success may be the most important merits for your customers to align your clinic or one of your services with. Careful thought needs to be placed around patients’ associations with these qualities. For example, a patient may consider that higher prices relate to

Customer Service and Marketing in IVF Practice

a premium service, and so they would expect all features of the clinic, from customer service, to the clinic facilities and ambience, to results, to be first class. One of the key challenges for marketers in this arena is the balance of creating a powerful presence while also maintaining the caring and compassionate aspects that surround infertility and IVF. The brand of a fertility clinic is the customers’ total experience from the clinic name, slogan, and design through to how staff answer the telephone. It’s a company’s personality, customer promise and business values. The elements should cohesively work together to identify and differentiate a fertility clinic and its services from its competitors and maintain a lasting competitive edge. A fertility clinic should clearly deliver the brand message to its customers to confirm credibility and emotionally connect with target prospects. This will increase confidence, motivate buying into your clinic and create loyalty to return and spread the word.

How Are We Going to Get There? Develop a Marketing Plan Marketing mix is the framework for your marketing plans. It facilitates the process of how you are going to develop your service and focuses on how each element will impact your customer.

Service/ product Price

Place

Target Physical

Market Promotion

environment

Process

Figure 39.1 The marketing mix

People

Product/Service You will need to consider the unique selling points (USPs) of your fertility service such as what makes your services better than those of your competitors and how the needs of your customers are being met. It includes the service name and its messaging. Any new services or products should be assessed in relation to patients’ needs and the demand for the service.

Price This involves your pricing strategy. Pricing always helps to shape the perception of your service in a consumer’s eyes. Often, a low price is inferred as an inferior product and a price too high will make the costs outweigh the benefits. Ensure you are aware of your competitors’ pricing as well as the full cost of the service provision.

Place Is your clinic in an easy-to-access location? Is there car parking and, if so, what are the costs? If you are looking at reaching out to a greater geographical area, you may explore outreach clinics to make it easier for your patients to reach you initially. It may be that they access your services at an outreach clinic and travel to your main clinic for their fertility treatment.

Promotion and Marketing Promotion is a very important element of the marketing mix. It involves how you will communicate your fertility unit and its service to your prospective patients. Your promotional strategies will be dependent on your budget, the message you want to convey and your target audience. Digital Marketing Digital marketing encompasses your website, search engine optimization (SEO), blogs, forums, social media and email marketing. There are many benefits of digital marketing: it connects you with your customers on the Internet and the mobile consumer, it can be economical and offers measurable return on investment, it enables precise and personalized marketing to target audiences, it allows real-time customer service, it makes it possible for you to compete with larger clinics or fertility groups, and you’ll find your competitors are using digital media more and more so you need to remain competitive and visible too. Website –– Your website is your shop front. It needs to be attractive, convey your branding, be trustworthy

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and be easy to navigate. You will also need to ensure it is optimized for mobile phones and that visitors can see how to contact you from every page. Continually review the content and keep the site up to date. Once you have a website, you need to develop a well-planned series of steps to make it visible on the first page of a search engine’s results. You can utilize pay-per-click online advertising, whereby you develop an ad for a search engine and use your marketing budget to show up in the search results. Current SEO is linked to excellent content, and this can be achieved by adding value to your website to make it a unique information resource to patients and less of an advertising tool. It’s important to go beyond the basic information site and offer unique content such as free patient education in the form of blogs, newsletters, videos, links to other helpful sites and a search tool. Think about what fertility information and news your prospective patients might be interested in knowing about or are looking for and create interesting and informed content. Blogs –– Blogging is an inexpensive way to enhance your inbound marketing efforts, drive traffic to your website and attract more prospective customers, connect and build trust with your website visitors by providing valuable expert information and present a personal side to your business. Social Media –– People love social media and so should your fertility clinic. It’s advisable to have a presence on Facebook, Twitter, Google+, Pinterest, Tumblr and Instagram and post news and blogs regularly. Newsletters –– Newsletters are a valuable email marketing tool for keeping your fertility clinic in the conscious mind of your prospective client base, and for tracking who is reading, clicking through and deleting what. Content needs to be interesting, concise and resonate with your target customers. Newsletters provide the opportunity to promote your clinic in relation to your prospective patients’ needs or issues such as new services, research, changes in the clinic, staff promotions. Other Media In addition to digital marketing, there are more traditional forms of advertising to make up your marketing mix. These include: • fertility events (open evenings or days for patients or educational meetings for referring medical

• •

•

•

• •

practitioners). Face-to-face opportunities can be invaluable in this service industry. print publications (for adverts, editorials and articles) press releases and other public relations materials (Build links with local and national publications whenever possible. Write professional press releases for newsworthy events, research, new services or service developments that go to the right person at the publication. It’s worth following up with a telephone call.) marketing collateral (brochures, flyers, promotional merchandise, corporate identity materials including business cards, letterhead, logo and envelopes) build case studies and utilize the results of customer surveys (Testimonials are an invaluable marketing resource so that patients can connect with other people’s stories who are similar to theirs.) direct mail pieces and postcards posters, banners and signage (in key locations where prospective patients are likely to take notice).

People and Customer Service The employees of your fertility practice are important in marketing because they are the ones who deliver the service. It’s imperative to hire and train the right people, and this is especially true in the service industry. If you have staff who genuinely believe in your fertility clinic and its services and who feel valued and respected, then it’s highly likely that they will perform the best they can, and this positive atmosphere will be passed on to your customers. They will also be more open to honest feedback about the business as well as willing to input their own observations, thoughts, recommendations and passions toward growing the business. Staff can learn a lot about their patients’ wants and needs and the fertility practice they work at by listening carefully to patients. A fertility practice’s front-line staff are its biggest asset. If you’re searching for something as important as a fertility clinic to give you the chance of making your dream of parenting a reality, you’re not going to hang around if you have a negative first impression. All staff, from the administration office to the laboratory staff, nurses and doctors, need to be empathetic to each patient’s individual circumstances and have a calm and understanding perspective. For example, a patient who is contacting the clinic about donor eggs may feel they are at the end of their tether,

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exhausted and overwhelmed. Patients seek personalized care regarding their unique situation, and staff need to provide this level of care. Patients may be confused with complex and medical technology that they don’t understand. It’s important for staff to talk in terms that the patient knows and communicate complex concepts clearly using understandable terms. Talking through each step or treatment with patients and taking the time to address any questions or concerns will reduce a patient’s stress and worry and keep the whole process as uncomplicated and convenient as possible. It’s important to offer information, advice and instructions at every step of the journey. Fertility patients expect and deserve care and support in all aspects of their liaison with a fertility clinic. A thoughtful approach, a caring attitude and experience will go a long way.

Your customer service is what keeps customers coming back. In today’s world, we expect an instant response to our inquiries, and if these aren’t timely, then prospective patients will go elsewhere. Staff should respond immediately to customer inquiries and their feedback, both positive and negative.

Process Your fertility clinic will need well-tailored systems and processes in place to ensure it runs smoothly and provides an efficient service for patients. These should be reviewed regularly to tweak and enhance as necessary and to minimize costs and maximize profits. Patients will benefit from knowing their fertility journey with your clinic and the processes involved. Payment and paperwork should be as simple as possible. Appointments should be convenient and patients should know how to access the clinic should they require.

Table 39.1 Marketing content and type tailored to patients’ need

Behavior stage

Patients’ thoughts and needs

Marketing/content type

Awareness

A couple have started trying for a baby and haven’t had success. They start to research fertility issues such as “how long should I try for before seeking help?” and “what can I do to improve my chances of natural conception?”

General research around infertility website of regulatory bodies, e.g., HFEA Blogs and articles on infertility Educational content

Consideration

The couple understand they are having fertility issues. They are committed to researching the different treatment options and the clinics that provide these. They may read extensively around the subject trying to make sense of their situation and what they can do to realize their dream.

Engage with advertisements of fertility clinics SEO and organic website ranking Compare clinic websites Understand service provisions more fully Blogs relevant to the patient’s specific fertility problem Contact clinics: open evenings, brochure/ information request Patient may allow connection with clinic, for example social media, email marketing to nurture

Decision

The couple have decided how they will solve their fertility problem. They may choose a fertility clinic because of its first-rate reputation, using the latest techniques, excellent fertility success rates, recommendations, location, price.

Endorsements may help make this final decision First impressions of the clinic are very important Prompt, professional customer service is imperative Smooth processes are important and informed decisions

Ambassador

Success or not, with treatment the patient will endorse the service provision if it has met or exceeded their expectations.

Gain testimonials from ambassadors by gaining and using a variety of media Ask past patients to talk at prospective patient events.

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Physical Evidence This involves the physical environment of your fertility practice, such as the reception area, the design of your clinic, the soft furnishings, scanning services and the facilities available such as a private room for every egg collection and embryo transfer. These things are all tangible elements that your patients will experience and need to be aligned with your brand and your positioning.

Buyer Behavior It’s also interesting to look at your buyer behavior and understand the stages your prospective patients may go through on their way to making a buying decision and beyond.

How Do We Know We Got There? Clarify, Evaluate and Monitor Your Controls Your marketing strategies and goals are theoretical objectives. Your marketing plan is your action plan and it will have constraints, such as budget and resources, that need to be taken into consideration and worked within. No fertility center has an unlimited marketing budget, and sometimes resources are tight. Your marketing mix and activity needs to be within these parameters. It’s important to outline a wish list and then select the most appropriate marketing tools and actions that will provide the best return on investment for your fertility clinic. If staffing is an issue, perhaps during the holiday

season, then marketing activity may well be best channeled once staffing is back to optimum levels. Your marketing plan should contain key performance indicators so that marketing activity can be reviewed and assessed regularly.

References Anon. The Marketing Mix – Marketing Mix Definition of the 4P’s and 7P’s. [Accessed July 2017]. Available at http:// marketingmix.co.uk/. Anon. Marketing theories – PESTEL analysis. Professional Academy. [Accessed July 2017]. Available at www .professionalacademy.com/blogs-and-advice/marketingtheories—pestel-analysis. Anon. SWOT analysis – A guide to the SWOT model, process, advantages and disadvantages. October 15, 2015. [Accessed July 2017]. Available at www.cipd.co.uk/knowl edge/strategy/organisational-development/swot-analysisfactsheet#8010. T. Berry. What is a SWOT analysis? [Accessed July 3, 2017]. Available at https://articles.bplans.com/how-to-performswot-analysis/. D. Court, D. Elzinga, S. Mulder and O. J. Vetvik. The consumer decision journey. McKinsey & Company Marketing & Sales. June 2009. [Accessed July 2017] Available at www.mckinsey.com/business-functions/mar keting-and-sales/our-insights/the-consumer-decisionjourney. Human Fertilisation and Embryology Authority (HFEA). 2013 trends and figures. [Accessed 2017]. Available at www.hfea.gov.uk/media/2081/hfea-fertility-trends2013.pdf.

Chapter

40

The Role of the Research Nurse in an IVF Center Susan Wellstead and Jane Forbes

Definition The research nurse is a registered nurse who has undergone specific research training in order to comply with the International Conference of Harmonization Good Clinical Practice (ICH-GCP) requirements [1] and who has the knowledge and training to follow a study’s protocol. The role and responsibilities of the research nurse are predominately concerned with recruitment of participants to studies within the recommendations from ICH-GCP guidelines. Throughout the participants’ involvement in the study, the research nurse is an integral coordinator of the process that ensures adherence of the relevant study protocol.

National Research Agenda In the past, research in the United Kingdom tended to be sporadic and uncoordinated. In 2006, the formation of the National Institute of Health Research (NIHR) provided specific support in the form of more specially trained research staff along with the infrastructure required to carry out research to a higher standard. The National Health Service (NHS) had a clear vision about the research agenda. “The NHS will do all it can to ensure that patients … are made aware of research that is of particular relevance to them” [2]. In reproductive health, the research activity was mainly seen in the area of obstetric research, closely followed by gynecology and uro-gynecology. Fertility-related research in the United Kingdom took some time to follow this trend, although large multi-centered studies in fertility are now commonplace, in comparison to the past when single-site studies were the predominately funded studies on the national research portfolio. It is envisaged that this increase in fertility-related research activity can help improve the success and long-term outcomes of fertility treatment. Such a development is in keeping with the general ethos of the NHS NIHR to be inclusive so that “everyone could have

the opportunity to benefit from new treatments, interventions and medicine” [3].

Fertility Research Fertility research is multifaceted, and can be clinical or laboratory based. Research nurses tend to be involved with clinical studies, although they will also play a crucial role in assisting in sample collection for more laboratory-based studies. The chapter is presented in two parts: 1. The role and responsibilities of the research nurse 2. Challenges and barriers within the research nurse role in the fertility center.

The Role and Responsibilities of the Research Nurse in the Fertility Center The responsibilities of the research nurse span the duration of the study. These responsibilities are multiple and complimentary to the roles of others involved. From the very inception of an idea to completion of data collection, the research nurse has valuable insights and experience into the successful randomization of a study. The research nurse’s role and responsibilities are depicted in the sections that follow and are summarized in Figure 40.1.

Study Setup The research nurses can make invaluable contributions at the study set-up phase. As protocols are being written and study requirements set out, the in-depth knowledge and experience the research nurse has on the organizational and operational aspects of the research landscape can help drive and shape the project plan appropriately, and facilitate establishing a study that will work alongside the clinical care pathway. Furthermore, an experienced fertility research nurse often possesses a comprehensive knowledge of

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Study completion and disseminati ng results

Study set up Obtaining informed consent

Communica ting with the fertility team

Randomise

Study activities

Data input Data collection

Figure 40.1 Summary of the roles and responsibilities of a research nurse from inception of study to study completion

the research process and the legislations involved as well as having an adequate understanding of the fertility specialty and the patient population. Possessing background knowledge within the fertility research context is useful, particularly in troubleshooting problems, which may arise on a day-to-day basis, but also helps avoid study and data compromise. Regardless of a research nurse’s background experience, there is always a vital role for them within the fertility research arena. Therefore, while a familiarity of the fertility specialty is helpful, it is not an essential requirement of a fertility research nurse as such knowledge and experience can be accumulated with time.

Informed Consent Ethically, the most important part of the research nurses’ responsibilities is obtaining informed consent. As part of fertility research, it is often the case that informed consent is required from the woman and the man undergoing treatment. One of the unique areas of research pertaining to assisted reproductive technology (ART) is that it often may involve the study of gametes or even the embryo. In addition to the legislative issues relating to the use of the latter for research purposes, embryo research requires consent from the male and female in a couple. The use of donor gametes within the research context can add further complexities to the consenting process. Without a thorough understanding

of the ART pathway, and the variations of treatment strategies now increasingly being offered to a variety of family units, for example, that of a single person, samesex couples, heterosexual couples using donor gametes and surrogacy treatment, the consent processes can be easily confused and research protocols breached and the results of studies compromised.

Randomization Randomized controlled trials are now commonplace in fertility research. This often includes the requirement to randomize participants at the site of recruitment. The randomization process can vary from study to study and will require stringent adherence to the protocol. This method of allocating a treatment option for a participant has many implications. In addition to the practical procedure of randomization, the research nurse also has to manage the participant expectations and reactions to the allocated treatment option. This requires the nurse to have a good understanding of the randomization process and the need for randomization in the study, so as to provide a clear explanation of the details of each of the treatment options to the patients. Being sensitive and clear is imperative in retaining the participants’ cooperation and satisfaction during their involvement in the study.

Study Procedures and Activities Once participants have been recruited, the research nurse will need to make the necessary study procedure arrangements in accordance to the study protocol. This can range from collecting demographic data, data regarding fertility, the medical history, venesection, tissue and/or fluid samples. Some examples of biological fluids that may require collection include endometrial fluid, endometrial biopsy, sperm, semen and seminal fluid. It is important that the lead research nurse ensures that the research nurses within the team understand their roles and responsibilities in this process, and that they have received the appropriate training and have the capability and capacity to carry out the process.

Data Collection It is important to ensure that the data collected in any research study are accurate and complete as this has significant implications on the validity of the study conclusion. The research nurse may need to use different sources of information to obtain the correct data. Commonly, this is in the form of hospital databases as

The Role of the Research Nurse in an IVF Center

well as patient notes. Sometimes this information is held in different organizations, and the research nurse may need to open a dialogue with respective organizations to obtain further information required to complete the data collection process. At this juncture, it is important for one to be aware of the rules and regulations of information governance (IG) within any organization, and respect the participants’ confidentiality and data protection where appropriate. Often such details are specified within the study protocol, in terms of the coverage of information sharing; a good communication and frequent reference to the study protocol if required can help avoid breaching patient confidentiality. Occasionally, participants may be approached in a sensitive way to obtain further data, although this is not an ideal strategy, as the burden on the participant should always be kept to a minimum.

Data Input For researchers to analyze data, it needs to be available in an organized way. Case report forms (CRFs) are used to ensure all the information required is clearly set out and easy to populate. This can be in paper form, but more commonly now these are on electronic databases or secure websites (eCRF). The CRFs set out by studies ensure that questions that require answering are not missed, and that all the data required are present. Again, it is the responsibility of the research nurse to ensure these are completed in a timely manner and in full.

Safety Reporting Throughout the participants’ involvement in the study, the safety of each participant is paramount. The ICH GCP guidelines state that “the rights, safety, and wellbeing of the trial subjects are the most important considerations and should prevail over interests of science and society” [4]. Most studies will require the research nurse to collect safety data from participants. The reporting of adverse events requires a specific process, and depending on the nature of the events, events may need to be reported regardless of whether there is a known direct link to the study. The specific categories are listed within the recommendations from ICH GCP, and the mode of reporting should be laid out clearly in every study protocol. Some studies require the participant and/or the clinicians and research staff involved with the participant’s care to remain “blinded” to the treatment. The research

nurse needs to know how this treatment allocation is “unblended” in the event of a patient safety issue. However, these instances are usually rare and “unblinding” is avoided wherever possible.

Summary In addition to the study set-up process, the running of the study, ensuring participant satisfaction and the collection of good-quality data, the roles and responsibilities of the research nurse in the fertility center are primarily concerned with the participants’ safety. The research nurse needs to have excellent organizational skills along with the ability to work flexibly and at times autonomously. When working in the clinical area where clinical workload competes with research needs, these skills are essential. Good communication and the ability to work with a range of health care workers and clinicians help ensure a study succeeds in terms of recruitment to target with a complete dataset.

Challenges and Barriers within the Research Nurse’s Role in the Fertility Center The Impact of Research Recruitment on the Success of IVF Studies Patient recruitment is a significant part of the research nurse’s role [5]. There is a need to reflect upon recruitment. Improving the process of identification and accessing eligible participants are essential to the success of a study. It has been identified that one-third of clinical trials failed to reach recruitment targets. Such difficulties in patient recruitment can inhibit the completion of research and delay results. These failures impact all research stakeholders with the IVF patient at the center (see Figures 40.2 and 40.3).

The Referral Process The need for access to potentially eligible IVF patients is a key theme for recruitment. Collaboration between health care workers and researchers can avoid a barrier between research nurse and patient, also known as “clinical gatekeeping.” The rate of patient referral directly affects recruitment rates, and poor recruitment rates can lead to underpowered, expensive and delayed results [6]. The literature and research around the referral process is scarcely published, and there is little guidance on how to manage it [7], yet it is considered highly

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Figure 40.2 Research stakeholders

Clinic

Chief Investigator

R&D

Patient

Staff

Sponsor

CTU

Disclose all studies available at the institution The clinician decides which study to refer the patient to Employ a randomizing process of selecting a study Institutions prioritize study Figure 40.3 Strategies employed when allocating studies to a patient population

influential. It continues to be common practice within the United Kingdom and other countries for the primary caregiver or clinician to refer a patient to the research team before contact about a study can be made. This is considered a layer of protection for the IVF patient, who could be considered to be in a vulnerable group. One study identified that only 19 percent of eligible patients were informed about trials from their clinician (Bain 2005, cited by [8]). The primary point of research involvement (that initial conversation) is key to good recruitment rates. Research has suggested that 98 percent of patients don’t

read the patient information leaflets but rely solely on the initial verbal communication with the clinician [7]. With patients considered potentially vulnerable, and clinicians holding the key to access, the “gatekeeper principle” occurs. This is a common phenomenon within health care and the research arena where someone feels protective over a patient in their care, and influences what they are exposed to [9]. Researchers are relying on the engagement and compliance of these gatekeepers to the patient population. With the potential and power to withhold access to recruits, a culture of trust is required between the clinical team and researcher for a study to succeed.

The Role of the Research Nurse in an IVF Center

It should be considered that every time a suitable candidate for a study is missed, a failure in the system has occurred. Although there may be many contributing factors to poor recruitment, clinicians missing research opportunities with their patients have been identified as key [10]. Understanding this relationship and overcoming the barriers to it working effectively would appear fundamental to successful recruitment.

Potential Barriers That Influence Clinicians and Their Research Engagement Preconceptions about IVF research and the lack of familiarity about the research process by the clinical team may be obstacles to them identifying themselves as having a role within this field. It would appear that research is far from ingrained into practice culture, but is seen as a bolt on a role that is optional. It has been suggested that promoting staff engagement could be the primary factor for successful recruitment [11]. Within a busy IVF clinic, numerous barriers to research engagement have been identified throughout the enrollment process, from institutional, staffing and patient issues. Nixon et al. [14] highlight the key issues perceived by staff at Brighton and Sussex hospitals, regarding their engagement in research activity. They are listed here in priority order: Lack of time Staffing issues Support Lack of awareness Resources Lack of experience Other Graffy et al. [12] discussed nesting studies within host trials addressing these issues and providing robust strategies and offering alternative evidencebased approaches to referrals and recruitment.

Moving Forward It is important to explore creative and practical methods of engaging front-line clinical practitioners. Harter et al. [13] identifies “Patient Engagement Teams” (PETs) that provide a multidisciplinary approach. This not only looks at the patient, the

research and treatment as a whole collaborative process but also discusses trial adaptations that were made, as study recruitment and referral efficacy were assessed. Evaluating best practice and achievements while implementing the required changes is an ongoing process throughout this study. With multicenter sites working together, a culture of reflective support and collective wisdom could be applied. Where it was identified that clinicians were reluctant to engage with recruitment, effort was made to identify the causal factors and target specific staff issues. Where clinicians lacked familiarity with the study, efforts were made to teach them about the study, including the anticipated positive clinical outcomes and benefits. Where the burdens of clinical work impeded the research process, the PETs offered practical solutions to alleviate them. This study showed results of 20 percent recruitment increase in one site in one year, with an additional increase from 34 percent to 50 percent in two sites in year two and a further 16 percent in year three. This approach addressed real problems with viable solutions, while embedding research firmly within clinical practice.

Is Choice Always Empowering? As options for IVF studies increase, so does the burden of choice for the patient. The management of multiple competing studies in a limited patient population needs to have carefully applied strategies to ensure this burden does not have a negative impact on all research stakeholders, especially the patients themselves. There is little evidence to draw from when deciding how to approach patients about multiple studies, especially in the field of IVF. We do, however, know from surrounding literature that increased choice can be paralyzing and result in disengagement from the research process. In what follows are some of the options employed to allocate patients to studies. These options will require an individual approach from researcher and unit and for them to decide the best “fit” for their situation. There is no right way of approaching this dilemma, but it is vital to consider the impact competing studies have on the potential participant, as well as on the research and clinical teams.

Summary It would appear that there is a dearth of research about the research process, yet we understand that the recruitment process must be managed with skill and delicacy to ensure ongoing patient satisfaction and trial

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success. Nested studies within our current ongoing trials could offer evidence on both how to best manage the recruitment process successfully and how to enhance the patients’ satisfaction with research and their treatment. The fear a patient feels about making the wrong choice about which study, if any, to participate in, with anticipated regret, can lead to anxiety, choice paralysis and research disengagement. The harder the choice and the more that is at stake, the less able they are to make the final decision. Adding research options to patients’ care pathways can lead to higher expectations, and it is essential that the research nurse manages those expectations. Poor treatment outcomes can be devastating. When we have offered choices to the patients through research opportunities, there may be associated dissatisfaction with the choice they made.

Conclusion The research nurse requires specific skills and knowledge to undertake the role. The responsibilities attached to the role involve a detailed knowledge of the research process in order to effectively assist with study design and setup. This input helps to ensure the smooth running of a study along with the recruitment target required to answer the study question. The most important responsibility for the research nurse is advocating for the participant throughout the study. As discussed, the participant can face complex and multiple decisions throughout the IVF treatment, and research opportunities add to this. The research nurse needs to manage this burden and support the participant throughout the process while offering choice and ensuring informed consent.

Suggested Standard Operating Protocol (SOP) 1. 2.

Steps for how to include a patient in research Obtaining feedback from patients after participation in research

References

3. National Institute for Health Research. Building a Research Career Handbook. 2015. www.nihr.ac.uk/ building-a-research-career-handbook 4. ICH GCP. http://ichgcp.net/2-the-principles-of-ichgcp-2 5. L. Rowland and C. Jones. Research midwives: Importance and practicalities. British Journal of Midwifery 2013;21(1):60–4. 6. M. Campbell, C. Snowdon, D. Francis, D. Elbourne, A. McDonald, R. Knight, V. Entwistle, J. Garcia, I. Robertson and A. Grant. Recruitment to randomised trials: Strategies for trial enrolment and participation study. STEPS study. Health Technology Assessment 2007; 11. 7. L. Newington and A. Metcalf. Factors influencing recruitment to research: Qualitative study of the experiences and perceptions of research teams. BMC Medical Research Methodology 2014;14(10). 8. M. Picillo, N. Kou, P. Barone and A. Fasano. Recruitment strategies and patient selection in clinical trials for Parkinson’s disease: Going viral and keeping science and ethics at the highest standards. Parkinsonism and Related Disorders 2015; 21(9): 1041–8. 9. E. Ward, J. Miller, J. Graffy and P. Bower. Contrasting approaches to recruitment in primary care research. Primary Health Care Research and Development 2009;10(4):368–73. 10. L. Afrin, J. Oates and D. Kamen. Improving clinical trial accrual by streamlining the referral process. International Journal of Medical Informatics 2015;84 (1):15–23. 11. B. Fletcher, A. Gheorghe, D. Moore, S. Wilson and S. Damery. Improving the recruitment activity of clinicians in randomised controlled trials: A systematic review. BMJ Open 2012. http://bmjopen.-2011-000496 12. J. Graffy, P. Bower, E. Ward, P. Wallis, B. Delaney, A. Kinmonth, D. Collier and J. Miller. Trials within trials? Researcher, funder and ethical perspectives on the practicality and acceptability of nesting trials of recruitment methods in existing primary care trails. BMC Medical Research Methodology 2010. http://bmc medresmethodol.biomedcentral.com/articles/10.1186/ 1471-2288-10-38

1. International Conference on Harmonization. ICH Tripartite Guideline. 1996. www.ich.org/fileadmin/Pub lic_Web_Site/ICH_Products/Guidlines/Efficacy/E6R1

13. G. Harter, J. Darden, N. McMenemy, T. McElvy and A. Hendrich. Consent and enrollment process: achieving high enrollment rates for obstetric research. Applied Nursing Research 2016;29:101–6.

2. Department of Health. Handbook to the NHS Constitution. 2009. www.gov.uk/government/publica tions/the-nhs-constitution-for-england

14. E. Nixon, S. Young, V. Sellick and K. Wright. An innovative approach to facilitating nursing research. Br. J. Nurs. 2013;22(3):160–2, 164–7.

Chapter

41

Financial Models in an IVF Practice Kiran Gedela

What Is a Business Plan and Why Is It Required? A business plan presents the simulation of the likely overall performance of the company. It reflects the company strategy, and creating the plan forces the business owners to think deeply and clearly about the environment in which a company operates, and to help lay down the road map and requirements to achieve success. The business plan helps the business owner and his team in multiple ways, including: 1) Clarify direction: a. b. c. 2)

c.

Factoring future growth and direction; Incorporating market trends, seasonality and new innovations; Understanding the competitors better and factoring in competitive response to your participation in the market.

Help the business owner answer key decisions: a.

b. c. 4)

6)

7)

Develop new business alliances and partnerships based on the business plan. Attract team members: business plans can be designed as a sales tool to attract partners, secure supplier accounts and attract executive-level employees into the new venture. Plan for operations and business expenses: a. b.

c.

d. e. f.

Plan the growth journey of a business by: a. b.

3)

Lay out the strategy in terms of segments of consumers to be targeted and approached. Understand the products and services to be developed and offered to the market. Understand the timing of rollout of various products and services.

5)

What are the financial investments in setting up an in vitro fertilization (IVF)/infertility business? When will I break even and recover investments? When will I reach steady state and what will be the profit margins at that time?

Raise financing: a.

Investors such as banks like to see business plans before investing in a venture.

g. h.

Determine the cost of material used in delivering the product. Gain an understanding of the type of team to develop, lay out the organization’s structure and roles and the responsibilities at each level. Figure out the selling and marketing expenses involved in “creating the right buzz” with the “right customer set.” Outline corporate overheads and overall administrative expenses. Determine the cost of renting out a place. List the expenditures on utilities and business transport. Factor in the interest payout for any loan taken. List any other miscellaneous expenses the business may incur.

Thus, a business plan may be the most important document you ever prepare for your business. It will be the custodian of what the business aims to achieve in the present and in the future. Continuous revision and updating of a business plan are necessary as market conditions keep on changing, government regulations may change, competition may engage in a different fashion or consumer choices may evolve.

How to Get Information to Develop a Business Plan A business plan requires a clear understanding of the market environment and requires the business owner

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to make scientific, data-driven assumptions on how the business is likely to behave and operate. Market research and assessment is one of the key tools to understand how an industry operates and works. A market research study uncovers several aspects that are important to the business: 1) What are the various segments of consumers and how do they think and behave? 2) What are consumers’ stated/unstated, met/ unmet needs? 3) What are the emerging trends that one must be aware of? 4) What are my competitors doing to win the market? How much of the business are they clocking and from which set of consumers and geographies? 5) What are the critical capabilities required to “win” a market? 6) What other challenges/considerations does one need to keep in mind while operating in the market? Market research is done by conducting focused group discussions with potential consumers; mystery following of consumers; one-to-one in-depth customer surveys; speaking to all participants of an ecosystem, including competition, vendors, government officials, sales and administration employees of the competition; speaking to industry forum members etc. Since market research unravels a lot of information about consumers and the competition, adequate care must be taken while administering questions and to ensure that only the right information is sought in a legally correct fashion. It is also necessary and mandatory to provide the right context while interviewing people for ethical and fair business practice. Market research for businesses is usually done by consulting firms, specialized market research firms or freelancing consultants.

Understanding a Business Plan for an IVF Unit Setup Before we dive deeply into a business plan for an IVFrelated business, we take this section to develop an understanding of what a business plan is and some of the measures an investor/business owner needs to keep in mind while operating the business.

To this end, we use a demonstration business plan developed for an IVF/infertility business operating in steady state or maturity. For simplicity’s sake, we have taken only a year’s numbers and have made them common size (on a scale of 100 percent for easy understanding and removing any currency- or location-based biases). Table 41.1 Indicative High-Level Profit and Loss Account of a Mature IVF Lab

Revenue

Percent of revenue

100%

Cost of material

Percent of revenue

26%

Gross margin

Percent of revenue

74%

Sales and marketing

Percent of revenue

10%

Manpower

Percent of revenue

15%

Utilities cost

Percent of revenue

2%

Rentals

Percent of revenue

3%

Other fixed costs

Percent of revenue

1%

Miscellaneous expense

Percent of revenue

1%

Total operating expenses

Percent of revenue

33%

EBITDA or operating profit

Percent of revenue

41%

Depreciation

Percent of revenue

7%

Interest payout (assuming no debt)

Percent of revenue

0%

Earnings before taxes

Percent of revenue

34%

Taxes

Percent of earnings before taxes

13%

Net Profit

Percent of revenue

21%

Operating expenses

Financial Models in an IVF Practice

The preceding table is an indicative profit and loss account of an IVF clinic doing about 35–40 IVF cycles per month. The financial statement may vary across geographies as the cost of provision of services will be different and so will be the method to compute taxes. Each term is briefly explained before a detailed look into its constituents in the later section.

Understanding the Business Economics: Profit and Loss Statement Before we get into defining and understanding some of the key business terms, we introduce two key terms commonly used in business:

Fixed cost: As the name suggests, fixed cost is incurred by business irrespective of the volume of business done. Examples may include salaries, utilities, rentals etc. Variable cost: These costs vary by the amount of business done. Examples may include material cost, which is used in supplying the product or service. Now we go through the B-plan terms in detail, explaining each term: Thus, we see that a business plan clearly details the sources of income and the expenditure it will entail to generate that income for the business owner. It gives an idea of the overall profitability or sustainability of the business, which is important for the business to run on its feet.

Table 41.2 Profit and loss statement – commonly used terms

Business term

Definition

Formula

Range as percent of revenue

Revenue

Revenue is what the business generates through selling its products and services.

Price of services * Volume of services consumed by consumer

100%

For an infertility clinic, overall business may be two to three times revenue though IVF treatments. Cost of materials

Cost of materials is the direct cost of materials used in provision of services.

Volume of services consumed by consumer * Material cost of each service

24–28% of overall revenue

Gross Margins = Revenue – Cost of materials

75–77% of overall revenue

It is also called variable cost as it is directly proportional to the volume of goods sold. Eq: Cost of drugs in IVF treatment is proportional to number of IVF cycles done Gross Margins

Gross Margins = Revenue – Cost of materials

Operating costs (the costs to run the business on the ground) Sales and marketing

Sales and marketing costs that a company incurs to engage with consumers and make them aware about the brand

Sales and marketing budgets are usually fixed in initial days of business to create a buzz in the market and create awareness among consumers.

~10% of annual revenue

Salaries, wages to all employees working in the setup.

Manpower cost is largely fixed as people are required to be paid, irrespective of the volume of business done.

~15% of annual revenue

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Table 41.2 (cont.)

Business term

Definition

Formula

Range as percent of revenue

Will include payouts to all the doctors, specialists, medical assistants, front office staff etc.

Incentive models are built for some employees to boost performance.

Manpower cost

Cost of utilities (electricity, water etc.) for good functioning of the clinic

Utilities are also almost considered as fixed costs as they are incurred irrespective of volume of cycles.

~2% of annual revenue

Utilities cost

Most of the businesses don’t purchase their own land, but lease out space.

Rentals are typically fixed for a certain duration of time.

~1–2% of annual revenue

Rentals

Other fixed costs

Other fixed costs

Miscellaneous expenses such as logistics, transportation, printing, money spent during festivals, auditor and legal fees etc.

Factored as a percent of revenue. Could be higher in certain geographies.

~1–2% of annual revenue

Miscellaneous expense

EBITDA stands for earnings before interest, taxes, depreciation and amortization.

EBITDA = Revenue – Operating costs (the costs to run the business on the ground)

~40–42% of revenues

EBITDA or operating profit

EBITDA measures the operating profit of the business and is a good measure of profitability as it is revenue minus all the fixed and variable costs taken to product the services

Depreciation

Depreciation is an accounting method of allocating the cost of a tangible asset over its useful life.

Depreciation = Value of asset/ useful life

For a mature center, it will be 4–5% of revenue

~1–2% of annual revenue

So, for instance, if all the medical equipment purchased by a company cost 10 million pounds and depreciation is assumed to be a period of 10 years, then only onetenth of the cost of the asset will be counted as an expense for any particular year. Interest expenses

Interest expenses is the money paid out toward servicing debt each year.

Interest = Debt * Rate of interest

This varies and depends on the capital structure of the company. Earnings before taxes (EBT)

Earnings on which tax computations will be done

Earnings before taxes = EBITDA – Depreciation – Interest Expenses

Calculation value

Taxes

Taxes paid to government and varies by country

Taxes = EBT * tax rate

Calculation value

Financial Models in an IVF Practice

Table 41.2 (cont.)

Business term

Definition

Formula

Range as percent of revenue

Net Income

Net income is the amount of money made by the firm after paying out for all operating expenses, depreciation and interest expenses.

Net Income = EBT – Taxes

Calculation value

A healthy business should target to generate a return of 18–20% of net revenue.

This is the net amount of profit generated by the business and is a key measure of profitability.

Table 41.3 Primary Sources of Revenue

Revenue and Costs for an IVF/ Infertility Clinic Revenue from Services There are two key sources of revenue to an IVF/ infertility lab: • Primary sources of revenue: revenue accrued from core fertility services such as IVF, intrauterine insemination, frozen embryo transfers and surrogacy. As the name suggests, these procedures aid in getting the woman pregnant. • Secondary sources of revenue: a range of ancillary services that are required only for a certain section of patients and assist the primary procedures in fertility treatment. These services include intracytoplasmic sperm injection (ICSI), blastocyst implant, donor services, andrology treatment etc. In certain countries, complete/partial coverage may be through public/private insurance, which may impact overall clinic realizations. In several places, infertility treatments come under out-of-pocket expenditure and thus are incurred by the patients themselves.

Area

Definition

Consultation fee

Physician consultation charges typically charged on a per-visit basis During treatment, a female patient visits and consults her gynecologist 8 to 10 times

IUI treatment

Procedure fee for IUI, which involves taking washed sperm and putting it next to the egg Drugs and pharmacy charges toward treatment Charges towards radiology and pathology tests conducted

IVF treatment

Procedure fee of IVF, which involves extraction of eggs and implantation of embryos Drugs and pharmacy charges toward treatment Charges toward radiology and pathology tests conducted

Surrogacy treatment1

Surrogate fee: fee paid out to the surrogate Procedure fee for surrogacy, which is similar to that of the IVF procedure Drugs and pharmacy charges toward treatment Charges toward radiology and pathology tests conducted

1

This could vary by geography and location

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Table 41.4 Secondary sources of revenue

Table 41.5 Cost of consumables.

Area

Definition

Area

Definition

ICSI

Only a few patients require ICSI and labs may choose to charge patients separately for this

Pharmacy consumables

Cost of drugs administered by the patient

Medical consumables

Cost of medical consumables (bandages, swabs, sheets) etc. used in an infertility procedure

Pathology consumables

Cost of reagents used to perform a test

Donor eggs/ sperm

Donor services for both eggs and sperm are available and this includes charges for services related to: 1)

Finding an appropriate donor

2) 3)

Extraction of eggs/sperm Preservation of eggs/sperm

4)

Donor service fee

Egg/sperm freezing

Freezing charges for egg/ sperm for usage at a later date

Blastocyst transfer

Some labs grow the embryo to the fifth day and then implant it, which is called the blastocyst transfer, and choose to charge separately for the same

Laser-assisted hatching

Some labs use lasers for hatching embryos and choose to charge patients separately for the same

1

This could vary by geography and location

Cost of Running an Infertility Center There are two key costs incurred in running an infertility center. They are: 1) Cost of consumables used in provision of treatment 2) Other operating costs to run and establish a business

Table 41.6 Operation cost.

Area

Definition

Manpower cost

•

Doctors (IVF specialists)

•

Andrologists

• •

Embryologists Medical support staff

•

Front office staff

•

Other permanent staff

Salary/wage costs include:

Payout to provident fund/gratuity Variable/bonus payout to the team Sales and marketing

Sales and marketing expenses include: • •

Digital marketing Printing of marketing material and brochures

•

Awareness campaigns for consumers

•

Newspaper, radio and TV ads etc.

Financial Models in an IVF Practice

Setting Up Your Center and an Introduction to a Balance Sheet As you set up your infertility/IVF center, you will incur a few expenses that you need to keep in mind and that are discussed in the following section:

Financial Investments in Setting Up an IVF/ Infertility Business Key investments in setting up an IVF center are one time and they include setting up of: Table 41.7 Once off key investment in setting up an IVF buisness

Area

Definition

Infertility lab

Setting up of an infertility lab, which includes medical equipment (detailed list in Appendix)

Operation theater

Operation theaters are used to perform infertility procedures

Medical equipment

Medical equipment that required

Center set-up Cost of building up and setting a center costs OR the interior fit-outs of the place taken on lease

center and buying land, and this may happen through either taking a loan or the founders (promoters) providing their own capital. A balance sheet is a very important financial document that summarizes the ownership (assets) and the liabilities (owner’s capital and debt) at any point in time. A fair business practice will warrant that both the values match at all times. We open this section with a brief description of the key business terms a business owner must keep in mind: Table 41.8 Key business terms

Definition Explanation Assets

Assets could include IVF equipment, center and furniture, medical devices, cash and other monetary equivalents. Liability

This may also include furniture, development of patient waiting area etc. Typical build-up area of a mid-sized center is in the range of 4,000–5,000 square feet Land

This could vary by geography and location

Some of the medical equipment may be imported from abroad and therefore additional excise and custom duty charges may have to be paid. Also shipping costs, installation costs, start-up costs and training for the people involved will have to be accounted for. In several countries, land may be provided at discounted rates by the local government or tax rebates may be provided.

Liabilities = Owner’s equity + Debt of the business Owner’s equity

The amount of money put by business owners through their own resources into the business at the time

Debt of business

Money taken as loans, or borrowings from banks or financial institutions or relatives that is serviced through a defined interest payout

Cost of capital

Cost of funds for financing a business. It refers to the cost of equity if only equity capital is used or the cost of debt in case a debt (loan) is taken. Since a typical business will require both equity and debt, a weighted measure is used.

Introduction to a Balance Sheet When you set up an IVF/infertility clinic, you will spend money on buying equipment, setting up the

Liability is a company’s financial debt or obligations that arise during business operations. Liabilities are settled over time through the transfer of economic benefits including money, goods or services. Liabilities are typically funded by money provided by the owners of a business (owner’s equity), debt taken by a firm (debt) and other accrued expenses that have been paid out.

Land may be taken on lease and thus the owner may save himself from incurring fixed costs, which in many countries are quite significant (as IVF/ infertility centers are commonly located in densely populated neighborhoods)

1

An asset is a resource with economic value that an individual, corporation owns/ controls with the expectation that it will provide future benefit. Assets are reported on a company’s balance sheet, and they are bought or created to increase the value of a firm or benefit the firm’s operations.

Capital invested

Capital invested equals total assets or total liabilities.

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Building a Balance Sheet for an IVF/ Infertility Business

Thus we see in steady state that the assets of the firm equal the liabilities. Total capital invested in the firm is 100 of the local currency unit.

We try to build a balance sheet for an IVF business in steady state where debt and equity are in equal proportion. Please note that this is a dummy balance sheet and only shown for representative purposes.

Profitability Measures of an Infertility/ IVF Clinic

Table 41.9 Example of a dummy balance sheet

Assets

Value

Cash in hand

Local currency

10

Pharmacy and consumables stock in hand

Local currency

10

Account receivables

Local currency

5

Medical equipment

Local currency

40

Land

Local currency

10

Other capital investments

Local currency

25

Total Assets

Local currency

100

Account payables

Local currency

10

Owner’s equity

Local currency

45

Long-term assets

Liabilities

Debt

Local currency

45

Total Liabilities

Local currency

100

As an organization or an individual physician sets up an IVF/infertility business, there are several key questions for them to consider: 1) What are the financial investments in setting up an IVF/infertility business? 2) When will I break even and recover investments? 3) When will I reach steady state, and what will be the profit margins at that time? Answering these questions beforehand is imperative to plan the journey well, serve patients better and avoid the risks of failure.

Which Measure to Use While Setting Up a Business Three options are available to see whether a business should be set up: internal rate of return, payback method and net present value. Per the majority of leading financial experts, NPV and IRR are the most favored tool to decide whether to move ahead with setting up business and true measures of long-term business profitability.

Table 41.10 Profitability measures.

Area

Definition

Approximate values

Business profit

Revenue generated by operations – (minus) cost of producing services

Depends on the business state and stage

Returns generated on the business

Profit of business/Total capital employed (or total assets of the business or total liabilities of the business)

In our examples, business gave a return of ~20 in local currency while ~100 was the capital deployed.

Total capital employed is usually the total value of assets purchased at the start of business, including the cash/cash equivalents in hand

So total return is ~20%

Length of time required for an investment to recover its initial outlay through profits

Payback period for an IVF business is two to three years.

Payback period of business

Financial Models in an IVF Practice

Table 41.10 (cont.)

Area

Definition

Approximate values

Return on sales

Net profit generated by business divided by sales done. It is a key measure of profitability.

In our example, our business gave a return of ~21%.

IRR or internal rate of return (IRR)

Rate of return on the investment made by the business owner (through a mix of debt and equity), measured assuming the business was to run till eternity

IRR for IVF/infertility business is in the range of ~25–27%.

Net present value (NPV)

NPV is the present value of the future cash flows of the business at the required rate of return of your project compared to your initial investment.

NPV of a business must be positive for it to be feasible.

NPV considers the time value of money, translating future cash flows into today’s money as this is a concrete measure of return on investment (ROI)

Business owners are requested to not proceed with projects that have a negative NPV.

Operational/Strategic Choices (Options) while Setting Up an IVF Business There are several operational/strategic choices that an IVF/infertility clinic may make while operating in the market.

Table 41.11 Operating model choices

Choices

What that involves

Key benefits

Disadvantages

Hub-and-spoke model of geographic expansion

As the capital equipment required to set up an IVF lab is significant, several IVF clinics try to open only one large reference laboratory and open several outpatient clinics around it.

Minimum capital expenditure for the IVF institute while maximizing reach out for patients

Nothing in particular

This creates access to a wide variety of patients who want to visit the lab for treatment but are not able to do so as the distances are far. How does it work? IVF being a micro-market-driven industry, the outpatient clinics are used as gatekeepers to filter patients and for drug-based stimulation. IVF patients are sent to the main lab only for egg pickup and implantation of embryos.

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Table 41.12 Tactical and Strategic Options for IVF Centers

Choices

What that involves

Key benefits

Disadvantages

Purchasing land or take a building on rent

IVF clinics require a huge facility and take up nothing less than ~4,000 sq. ft. space in most geographies.

Purchasing land and constructing a facility provides several benefits to the business owner such as:

Purchasing a piece of land and constructing a building takes a lot of time and a lot of capital up front.

Clinics must choose early on whether to buy land and then build a facility or take a building on lease and then focus on delivering quality medical care through their premises

•

Peace of mind

•

Guaranteed own space

Most business owners have limited capital, want to hit the ground running and want easy and “lean” business that is easy to exit in case it doesn’t work.

•

Greater cash flow no rentals to be paid

Taking land on lease helps in reducing EBT and thereby the overall tax payout and improving profitability measures such as ROCE (return on capital employed).

For Further Reading https://hbr.org/2015/04/the-most-common-mistakepeople-make-in-calculating-roi&ab=Article-LinksEnd_of_Page_Recirculation https://hbr.org/2015/04/a-refresher-on-cost-ofcapital https://hbr.org/2014/11/a-refresher-on-net-presentvalue

Investopedia.com Entrepreneur.com Sba.gov Bplans.com Accountingcoach.com HBR.org Business-literacy.com

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42

The Essentials for Building a Great Fertility Team Take Five with Fiona Pringle Fiona Pringle and Ying Cheong

1. YING: Fiona, can you summarize your current role and position within IVI UK? FIONA: I am the director of nursing for the whole of the UK IVI, with the responsibility of building the teams within the clinic, setting

going to be the perfect scanner, not everybody is going to be brilliant in embryo transfer or taking bloods … all those things, but you want people [who] will complement each other, and support each other and allow people to grow and develop.

the standards, making sure that nursing is a high priority,

You also have to recognize that some people want to come to work and do a job, but don’t really want to progress further.

making sure that everyone is working to protocols, standards,

Some people are very happy just being good at what they are

trouble shooting, bit of everything really!

doing; they don’t want to progress, they don’t want to be pro-

Career wise, I spent the past 20 years working as a nurse manager in Oxford fertility, which is a large unit with satellite units attached, so I have a lot of experience managing a team … both in the clinical setting on site but also situated elsewhere with people working for different employers. They weren’t specifically your employees, but you were still managing them and needed to try to get them to work to the standard that you wanted them to. I also carried a clinical caseload then. 2. YING: What are the key elements to building a great fertility team?

moted, they just want to do their job, and you have to recognize that. 3. YING: How did your past clinical experience help you in building the fertility team you have now? FIONA: You do learn life skills as you get older, and I do think it’s important to treat people with respect. I would like to feel that I have treated all the people that I have managed the way I would like to be treated. I don’t ever ask somebody to do things that I won’t be prepared to do.

FIONA: You need the right personality of people; I would say the

I have drawn a lot of these skills through my past career

ability to communicate is the most important thing. You can teach people practical skills if they’ve got the right attitude and

experience. Also, of course, [through] my own upbringing, but

willingness to learn, so that they don’t all have to come from

own personality.

some of the skills that maybe I inherently have are related to my

a fertility background, if they can show me that they are inter-

I think you can teach people, and train people, and send them

ested in the specialty, if they’ve got skills, maybe outside fertility,

on courses to learn the theory, but in practice, it is much harder.

that would be good. My career experience taught me a lot of that. I spent eight

And I think sometimes actually having people [whom] you look up to or aspire to that they could mentor you is a much better

years in a dermatology department; someone took a chance on

way to impart that knowledge … people shadowing you, so that

me, and actually said to me, “You’ve shown in your current role you know skills of advance practice, being motivated and keen.

they see you doing it.

You are the right person, we are going to change you, and you

a consultant [who] mentored me, who is really pro nursing. It was

are going to be fertility nurse.”

at the time [when] NHS was looking at mentoring in a much

The first year in the post, I honestly thought this was the worst decision that I had ever made! You go from knowing everything

I have been mentored. I had a really good nurse, and actually

bigger way, part of a project that was in the Trust.

and being confident, to knowing nothing and struggling with the

It is good to look out of your own clinical area; to visit elsewhere to see how people work can be really useful. Not only does

whole idea that you don’t know. Actually, looking back, it’s

it bring back good ideas, but it also makes people think, hang on

a really good life lesson and you need to make sure [when] you build a team that they complement each other and that you …

a minute, we are doing this really well or better than, and fosters

build a team that has a broad set of skills. So not everybody is

things well!

a sense of pride and belonging – that actually you are doing

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4. YING: How do you deal with the non-team players? FIONA: It depends on what it is. I think you need to be very clear as the leader what you expect, in terms of behavior, what is acceptable and what is not acceptable. A team is like a family so that we are not going to get on with each other all the time, and we are going to whine each other up and irritate each other occasionally, and some of that actually there is no point in jumping in straightaway if a couple of members of the team have had a minor disagreement, but when it starts to affect how people are interacting in the team or patient care or the rest of the team, then you have to sit down and have a conversation with each individual involved; you have to find out what exactly is going on. You need to remember that a lot of what you are doing is not personal, it’s not about each individual, that you like them or don’t like them, or agree with them or not agree with them, you need to be objective about what you are doing. You can get on well with staff at work and you can be friends. I personally won’t socialize with people I work with unless it is a work do. I always try to keep my home life and my work life

slightly distant. When [I] do have to come down on somebody or talk to him or her, I don’t want them remembering that two months ago, we were out having a good time. I think there is a place for being one step removed; you have to look at the wider picture, what’s right for the clinic and ultimately what’s right for the patients. 5. YING: As a leader, are you a fighter, a fixer or a friend? FIONA: I think ultimately I am a fixer. I want things to be right, and I would problem-solve and I would work outside the box, trying to work around a problem. But I will fight for what I believe is right. If, say, in the clinic, someone said that we should do this, and we should implement this policy, and I don’t think it is the right thing to do, I will stand in my corner and fight for the reasons why I think it was not the right thing. I think it is really difficult sometimes, the better the leader you are, the easier you make it look, and the less they think you are “doing.” But I think to achieve that is a real skill!! YING: Thank you, Fiona, for your fabulous insight into how to build an excellent fertility team.

Chapter

43

The Future of IVF ART 3.0 Nicholas S. Macklon

Predicting the future is a dangerous business. Not only do most experts get it spectacularly and entertainingly wrong, but they are bound to have a view that upsets somebody. So writing a book chapter fraught with such risks demands a disclaimer at the outset. These are my personal and only partially informed views, penned in the summer of 2016. But perhaps they provide some pointers as to the risks and opportunities our exciting field faces, and what our future patients might expect from their carers in the years ahead. When we reflect on what has been achieved since the early days of in vitro fertilization (IVF), there is much of which we can be proud. The story of scientific endeavor and unshakeable belief in the moral imperatives underpinning the efforts of Edwards and Steptoe remains a source of inspiration, pride and guidance for all of us working in the field of infertility. In the parlance of computer software, the founders of “ART 1.0” laid the foundation of our field and yet while they were grappling with the challenges of getting IVF to work at all, they were already looking to the future. Many examples of Robert Edwards’s predictions of now-established technological advances such as stem cell techniques can be cited, but in the fascinating account of the work that led to the birth of Louise Brown in 1978 [1], a particularly prescient clinical example emerges. While discussing the difficulties encountered in preparing the endometrium to receive the embryo, Edwards records: “There is an alternative,” I said to Jean. “We could try freezing human embryos, and keep them in store until the effects of the fertility drugs have faded away and their menstrual cycles were back to normal. The womb would then be receptive, and capable of sustaining the growth of the fetus.” The idea suddenly excited me. We could provide the mother with a whole family spaced in the way she wished, just thawing out each embryo when desired.

Writing some 40 years after this conversation was first recorded, it is clear that Edwards was already defining “ART 2.0,” the practices that characterize how we offer fertility treatment to the second generation of assisted reproductive technology (ART) children. Again, there is much to be proud of (Table 43.1). ART 2.0 has not only seen a rise in success rates, and a more mature view as to what constitutes success in IVF – the healthy child rather than the positive pregnancy test – but has also seen the two most serious complications of IVF effectively dealt with. The burdens placed on individuals and health care systems by multiple pregnancies arising from transferring large numbers of embryos is rapidly becoming part of our folklore, and the misery and sometimes catastrophic impact on young women caused by severe ovarian hyperstimulation syndrome (OHSS) are now mercifully rare events. So, are we there? Is IVF done? Or will the coming third generation of ART parents and children benefit from more advances? What will ART 3.0 look like?

ART 3.0 Anyone visiting a major ART conference will be overwhelmed by the advances in embryo culture, analysis and selection techniques on display in the exhibition hall. The IVF industry is growing, it is lucrative and it drives innovation very effectively. It is likely that we will see the emergence and refinement of increasingly

Table 43.1 Key advances of ART 2.0

Milder stimulation regimens Fewer multiple pregnancies Reduced perinatal morbidity Blastocyst culture Efficient freezing OHSS disappearing

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sophisticated but less invasive means of selecting the best embryo for single transfer. The advances and techniques offered by preimplantation genetic screening will soon appear crude, disruptive and limited compared with the molecular fingerprinting techniques beginning to emerge. Rather than focusing on just genetic health, whether that be nuclear or mitochondrial, we will soon be able to access a more holistic and functional picture of embryo health. But however clever we become in choosing the embryo, the next challenge will be to make better embryos. Here clinicians and embryologists will work together to optimize the quality of the oocytes and sperm being passed into the care of the laboratory, and to produce the ideal environment for fertilization and preimplantation development. Oocyte and embryo “rejuvenation” techniques based on simply adding mitochondria are likely to be seen as sound in concept but simplistic and crude in practice. And the health of the offspring will remain a concern. These changes need not always rely on high-tech developments, but will focus our attention on the importance of preconceptional health and nutrition as determinants of gamete quality and offspring health. The hypothesis first enunciated by Barker during ART 1.0, which describes how health in later life is determined by early life events, has become now rooted in medical orthodoxy in ART 2.0 and will gain further currency in ART 3.0. The current concerns as to how synthetic culture media may be influencing development, perhaps by epigenetic mechanisms, will be addressed by a further understanding of the regulators of embryo programming, refinement of culture conditions and even a move to reengaging the woman as her own incubator. In vivo techniques for culturing embryos are now emerging and may offer an alternative way forward for ART 3.0. The focus will then turn increasingly from the embryo to the woman. Optimizing preconceptional health will expand our field to one of “periconceptional medicine” [2], in which clinicians will reverse the trend toward isolation from our colleagues in obstetrics and gynecology, and work with them at the exciting interface between conception, implantation and pregnancy. It is at this interface that the next advances in the management and prevention of obstetric complications will take place, and our colleagues will increasingly work with us to ensure optimal outcomes for the children and mother we help to create.

The key focus will be on the preimplantation intrauterine environment. The endometrium will no longer be seen as a matrix described in passive terms as “receptive” or “invaded,” but will be viewed as an active partner determining successful implantation. Recent work identifying the biosensor role of the endometrium, and the active migration of stromal cells in response to embryo quality [3, 4] has highlighted the need to change our attitudes to the endometrium, and an increased understanding of the biology of human implantation is likely to allow us to move away from the largely empirical interventions to improve implantation that characterize ART 2.0. Our means of interrogating the endometrium will develop from invasive tests of gene expression to less invasive approaches, such as molecular analysis of endometrial secretions [5]. We will go beyond looking for markers of “receptivity” but will be able to describe the selective and nutritional phenotypes of the preimplantation uterine environment through the new omics. The reproductive microbiome is currently attracting much attention, and novel interventions are likely to be identified that can optimize these new markers of reproductive health. Some challenges faced by ART 2.0 may be harder to solve. The size of the efforts made to identify effective treatments to improve the number and quality of eggs obtained from poor responders is matched only by their signal failure to improve outcomes. We are repeatedly reminded that if the eggs aren’t there, we can’t make them. Or can we? As we move to ART 3.0, fundamental scientific research, carried out in partnership with clinical expertise, is offering a more promising future than the “why not try this” empiricism of ART 2.0. The identification of the molecular pathways that regulate the transition of oocyte development from the primordial state to growth has already led to novel and effective means of obtaining eggs for IVF in women that previously were resistant to medical therapy [6]. This kind of work is not only leading to genuine advances in understanding of ovarian biology, follicle recruitment and stimulation, with potentially profound implications for women faced with ovarian failure, but promises to change the attitudes of medical research funders to IVF. There is no doubt that ART 2.0 has an image problem, and this represents one of the major challenges for ART 3.0. The “try it and see” empirical approach to improving outcomes that has

The Future of IVF: ART 3.0

Research funding difficult

Poor knowledge and understanding

Patient demand

IVF ‘Industry’

Empirical Treatment

Patient belief/ medical adoption Figure 43.1 The circle challenging progress in IVF research

characterized clinical research in IVF in recent years has been harmful not only to our patients but also to the reputation of IVF as a branch of medicine. It is, however, easy to understand how we have arrived at this situation. A combination of lack of knowledge of the fundamental processes that govern fertility and successful implantation with the need to offer something to desperate patients has resulted in a vicious circle that threatens to limit true clinical advances (Figure 43.1). ART 3.0 must emerge from this cycle, but to do so we will need to recognize the drivers for our current behaviors. And the key is the ascendency of the patient as a customer. The customer who is always right. And who must be served to prevent loss of their business. This short termism and commercial expediency is not serving our patients well. ART 3.0 needs to see a new covenant with our patients. A new covenant that creates an open, balanced, honest partnership that recognizes the vulnerability as well as the autonomy of people faced with the crisis of subfertility, or those who wish to use IVF to expedite the completion of their families. One that involves patients in acknowledging the limits of our knowledge and capabilities, but also engages them in finding the solutions.

As IVF moves increasingly into the private sector, and consolidation creates larger chains of clinics, new opportunities emerge for this covenant. Benchmarking for clinical excellence will increase between clinics operating under the same company organization. Research and development will, as in other industries, be seen as vital to company strategy. And successful companies will be in a position to fund the fundamental research on which true innovations will depend. Among the challenges then, there is much to look forward to. ART 3.0 awaits. Let’s get on with it.

References 1. R. G. Edwards. A Matter of Life: The Story of IVF. 2nd edn. Impression Publishing. 2011. 2. N. Macklon and Greer I. Steegers (eds.). Textbook of Periconceptional Medicine. Informa London. 2009. 3. N. Macklon and J. Brosens. The human endometrium as a sensor of embryo quality. Biol. Reprod. 2014;91:98. 4. B. Gellersen and J. J. Brosens. Cyclic decidualization of the human endometrium in reproductive health and failure. Endocr. Rev. 2014;35:851–905. 5. Y. Cheong, C. Boomsma, C. Heijnen and N. Macklon. Uterine secretomics: A window on the maternal-embryo interface. Fertil. Steril. 2013;15: 1093–9. 6. K. Kawamura, Y. Cheng, N. Suzuki, M. Deguchi, Y. Sato, S. Takae, C. H. Ho, N. Kawamura, M. Tamura, S. Hashimoto, Y. Sugishita, Y. Morimoto, Y. Hosoi, N. Yoshioka, B. Ishizuka and A. J. Hsueh. Hippo signaling disruption and Akt stimulation of ovarian follicles for infertility treatment. Proc Natl Acad Sci U S A. 2013;110: 17474–9 7. K. Kawamura, N. Kawamura and A. J. Hsueh. Activation of dormant follicles: a new treatment for premature ovarian failure? Curr Opin Obstet Gynecol. 2016 Jun;28(3):217–22

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Peering into the Future of the Fertility Business A Conversation with Sue Channon, CEO, Virtus Health Sue Channon and Ying Cheong

Sue Channon is the Group Chief Executive Officer (CEO) of Virtus Health, a recognized global health care company and the market-leading assisted reproductive services (ARS) provider in Australia and Ireland with a growing presence in Singapore and Denmark. Virtus Health was the first ARS provider in the world to be publicly listed, and it was listed on the Australian Securities Exchange (ASX) in June 2013. For more than 20 years, Sue has gained extensive experience and expertise in senior management positions for a number of Australian health care organizations. These positions include the state manager for NSW and ACT for Medical Imaging Australia, the national director of nursing for the Mayne Group (now part of Ramsay Health Care), the CEO of Kareena Private Hospital, the CEO of Castlecrag and Mosman Private Hospital, the CEO and director of nursing for Castlecrag Private Hospital and the CEO of IVF Australia Pty Ltd. In November 2010, she was appointed CEO of Virtus Health, a publicly listed ASX 200 world-class clinical, scientific, diagnostic and day surgery services provider for people on their path to parenthood. In this interview, we explore Sue’s views on salient topics relating to her career thus far and of the in vitro fertilization (IVF) business now and for the future.

Conversation Questions 1. How did you become CEO of Virtus? I have had a long and extensive career in health care and health care management. My career commenced in New Zealand with a hospital-based nursing degree, which meant that I studied while working full time within the hospital training program.

Working within health care for most of my career and health care management in the past 20 years has provided me with a clear appreciation of the day-to-day workings of clinical practice, including the tensions and competing priorities that can occur. This hands-on training and experience sharpens one’s ability to make decisions [that] impact all key stakeholders working at the coal face. With this background, my position and experience within the sector as CEO of IVF Australia, as well as the experience I had had in taking the organization through the partnership with Quadrant Private Equity, I felt I had the knowledge and ability to successfully apply for the role of CEO Virtus Health. 2. What do you love about your job? Leading an organization of passionate and committed people, who provide the broader community with the chance to achieve their dream of creating and/ or growing the family they desire, is meaningful and extremely rewarding work for us all. 3. Career challenge – what is the biggest and hardest challenge you have ever faced in your career, and how did you deal with it? I started my career as a nurse; I did not plan nor expect to be in the position of CEO of a public company. I have been challenged in many aspects through my career as I have changed my focus from bedside nursing to CEO with all the management, investor relations, corporate compliance and governance requirements of my current position. However, the biggest challenge has been taking Virtus Health through an IPO (Initial Public Offering) to a listing on the ASX. This demanded the use of skills I did not know I had and required the learning of a whole new language and set of skills. Managing this process whilst keeping our key

Peering into the Future of the Fertility Business

stakeholders engaged in the process was quite challenging. 4. What is the biggest challenge you think the IVF/ fertility sector will face in the future? Each country has its different challenges based on the local funding and regulatory environment. Generally, the introduction of new and costly technologies [that] ultimately improve outcomes for patients also increases the cost of providing the service to patients (and subsequently the cost to patients). This is making IVF less accessible for the everyday person.The sector needs to assess how the new technologies can automate current, less efficient manual processes, particularly within the laboratories, for a more efficient service provision with subsequent reduction in labor expenditure and keep the service affordable for patients. Technology should ultimately result in reduced costs for patients through efficiency and accuracy. From a business perspective, the challenge is to balance potential reductions in revenue per cycle with lower operational expenditure through these efficiencies for each cycle delivered. 5. What is the single most important element of a successful IVF business? Patient care and outcomes. Word of mouth is critical in the promotion of an IVF clinic. If patients are not receiving the highest level of care and outcomes, they will move on. Hence, ensuring a well-trained team of the highest caliber, from the administrative staff through all levels of the clinical service (nursing, counseling, specialists, scientists, etc.), is paramount. The patient’s experience needs to be one of empathy, care and the highest level of efficiency and accuracy. Patients need to have confidence in the service and that it is being provided by the highest-quality team. The organization needs to be committed to ongoing research and development in the area of reproductive medicine to ensure all possible techniques are available for patients to ensure the best possible outcomes.

Virtus has well-developed research policies and protocols and approval processes that ensure oversight of any research and innovative practice. This includes Research and Development Committees, Ethics Committees and, at the highest level, a Risk Committee comprised of Management, Medical representation and Board representation. Delegated authority is well documented. Having a broad representation and a hierarchy of approval allows a wide range of opinions and thoughts to be included in the decision-making and approval process. A robust internal audit process is also in place, undertaken by external consultants, who identify where and how risk is being managed across the whole organization. Risk management policies and procedures also assist in managing the risk and determining the level of risk the organization is prepared to take. Risk is being identified and monitored across the whole organization daily through the implementation of “Riskman,” an electronic incident management tool [that] staff at every level of the organization contribute to. Our risk matrix is also held within this tool, allowing management to view areas of weakness and apply appropriate risk mitigation activities. It is this combination of risk identification and management achieved in a proactive manner that allows innovation to thrive. 7. In your opinion, who will be the disruptive competitors in the future? This will likely be different in various geographies. In Australia, we are seeing disruption occur as cheap, repetitive, “one-size-fits-all” services are being operated by nonspecialists, which may not result in optimal treatment emerging into the market. Technology could well be another disrupter, with larger clinics having the ability to utilize expensive technologies and therefore capitalize more efficiently; size and scale may have an advantage over smaller standalone clinics.

6. Embracing risk and innovation – how to strike a balance?

8. What advice do you have for women on effective leadership?

Innovation and a commitment to research is critical for an IVF clinic to ensure that we can continue to provide the best possible outcomes for patients and maintain the highest possible reputation. It is also a significant motivator for the team as a whole.

Be authentic, always operate with the highest level of honesty and integrity and don’t compromise this. Don’t lose your passion for what you do. Develop good listening skills; always be ready to learn, you won’t always be right. Employ the right people.

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9. Will you plan for succession? If so, how? Succession planning in any organization is an obligation executive management needs to take seriously. Within Virtus we have an executive coaching and mentorship program in place to support our senior management team (Managing Directors). We utilize an external resource to manage this program. Our Board has this (succession planning) as a standing agenda item on one of its committees. A middle management development program supports our middle managers in developing business and commercial acumen and a set of competencies for management teams to aspire to is currently being planned. This will provide the skills required to develop the next wave of managers. From a clinical perspective, the organization supports the CREI (Certificate of Reproductive Endocrinology and Infertility) training program; this allows us to support the next generation of fertility specialists who will be the future of the organization.

The Medical Directors within the organization are also identifying opportunities to coach the next generation of specialists who have the potential to take on the roles of Medical Directors in the future. This is achieved through the role of Clinical Directors who take additional responsibilities for oversight of a specific clinic. Within this program additional training along the lines of Responsibilities of Directors, etc. is undertaken. The committee structure hierarchy also allows exposure of clinicians to business management and aspects of running an IVF clinic within corporate and regulatory frameworks that we need to adhere to. 10. What is your idea of luxury? Given the amount of local and international travel I do, I want my downtime to be as relaxing as it can be, so a tropical island/banana chair/cocktail and a good book. However, I also love just being at home with family and friends; some gardening, cooking and the simple things in life are pure luxury to me.

Index

abortion, adenomyosis and, 29 acupuncture, 46, 47–8 anaesthesia use with, 99–100 CPR and, 47 ET and, 47–8 adenomyosis abortion rates with, 29 clinical features of, 28 cystic, 29–30 defined, 28 diagnosis of, 28 with MRI, 28 discovery of, 28 endometriosis and, 29 incidence rates for, 28 infertility and, 28 IVF and, 29 failure of, 162 JZ hyperplasia, 28, 29, 30 pregnancy rates with, 29 treatment strategies, 29–30 management recommendations, 30 through surgery, 29–30 ultrasound for, 155 adequate follicular recruitment, 110–12 cycles of, 112 hCG levels, 111–12 management of, 112 progesterone bioavailability, 111–12 adjuvant therapies, 16–17 androgens in, 16–17 growth hormones in, 17 AFC. See antral follicle count aging, for males ART consultations influenced by, 32 paternity influenced by, 33 sperm quality and, 32–4 AH. See assisted hatching albumin, 95 alcohol consumption, IVF failure and, 159 alfentanil, 99 ALSPAC. See Avon Longitudinal Study of Pregnancy and Childhood AMH levels. See anti-Mullerian hormone levels amnion graft, 43

anaesthesia use acupuncture with, 99–100 advantages of, 99 clinical pregnancy rates, 98 disadvantages of, 99 guidelines for, 97 management of, 98–100 for general anaesthesia, 99 for local/regional anaesthesia, 99 under MAC, 98–9 OHSS and, 98, 100 postoperative care, 100 for fever management, 100 for home readiness, 100 for hypotension, 100 for nausea and vomiting, 100 pain management, 100 for urinary retention, 100 preoperative considerations, 97–8 anxiety issues, 98 preexisting illness, 98 subfertility causes, 97 androgens, in adjuvant therapies, 16–17 aneuploidy screening. See also preimplantation genetic screening poor embryo development and, 170–1 animal studies, for intrauterine fluid issues, 80 antagonist stimulation protocols, 11 antibiotics, for IUAs, 43 anti-Mullerian hormone (AMH) levels, 81 FSH, for poor responders to GnRH therapy, 15, 16 OHSS and, 92 antral follicle count (AFC), 156 anxiety, 98 Asherman’s syndrome. See also intrauterine adhesions thin endometrium and, 76 aspiration pumps. See electric oocyte aspiration pump; manual aspiration pump aspirin, 17, 77 for OHSS, 94 assisted hatching (AH), 160

assisted reproduction treatment (ART), 46–7. See also in vitro fertilization; transvaginal ultrasound; ultrasound culture media and, 63 future of, 209–11 key advances in, 209 intramural fibroids and, 8 IUI, 46–7 for older males age criteria for, 32 clinical reasons for, 32 equipment for, 34 information sheets for, 34 ‘must have’ protocols, 34 strategies for, 34 patient’s perspective on on egg collection day, 5 on ET, 5–6 IVF costs and, 6 Avanafil, 125 Avon Longitudinal Study of Pregnancy and Childhood (ALSPAC), 33 bifollicular recruitment, 110 bilateral endometrioma, 36 blastocyst transfer, 160 blastocyte-stage transfer, 64 blastomere biopsy, 149 body mass index (BMI), 118 IVF failure and, 159 Bologna Criteria, 14, 15 Bolt, Usain, 181 bromocriptine, 94 Brown, Louise, 1, 209 business plans. See financial models cabergoline, 94 calcium infusions, 95 CAM. See Complementary and Alternative medicine cancer patients fertility preservation methods, for boys, 26 fertility preservation methods, for girls, 24, 26 through ovarian tissue cryopreservation, 25

215

Index

fertility preservation methods, for men, 26 fertility preservation methods, for women, 24–6 with GnRH agonists, 24–5 through oocyte cryopreservation, 25–6 through ovarian tissue cryopreservation, 25 through ovary transposition, 25 Candida albicans, 144–5 CAT. See chlamydia antibody testing cervical dilatation, 120 cervix, ET and appearance of, 118 canal, 119 mucus and blood in, 118–19 Channon, Sue, 212–14 Chinese herbal medicine (CHM), 46, 48 IVF and, 46–7 WS-TCM, 48 chlamydia antibody testing (CAT), 60–1 PID and, 61 CHM. See Chinese herbal medicine clinical pregnancy rate (CPR), 47 clomiphene nitrate, 76 coasting, OHSS and, 12 COCP. See combined oral contraceptive pill co-enzyme Q, 17 COH. See controlled ovarian hyperstimulation combined oral contraceptive pill (COCP), 76 Complementary and Alternative Medicine (CAM), IVF and acupuncture, 46, 47–8 CPR and, 47 ET and, 47–8 CHM, 46, 48 IVF and, 46–7 WS-TCM, 48 intervention strategies, 48–9 follicle ripening, 49 oocyte retrieval, 49 ovarian follicle stimulation, 49 consent. See also informed consent for ovarian tissue cryopreservation, 21 controlled ovarian hyperstimulation (COH), 76 FF and, 167 Cook intrauterine balloon, 43 Cooke, Ian, 1–4 corifollitropin alfa, 16 Costa Rica, IVF in, 3 costs. See financial models

counselling, for poor responders, 17–18 for couples, 17 CPR. See clinical pregnancy rate cryopreservation. See also embryo cryopreservation; oocyte cryopreservation; ovarian tissue cryopreservation OHSS and, 12 PGD/PGS management and, 151 sperm, 124 cryptozoospermia, 165 culture media, human embryo in ART, 63 criteria for choice of, 64–5 culture environment, 64 embryo manipulation, 64–5 physical infrastructure, 64 technical infrastructure, 64 development approaches, 64–5 future prospects for, 65 historical development of, 63 KPI and, 72 poor development as result of, 171–2 sequential blastocyte-stage transfer, 64 defined, 63 single-step compared to, 63–4 single-step blastocyte-stage transfer, 64 defined, 63 sequential compared to, 63–4 cystic adenomyosis, 29–30 danazol, 29 data collection, by research nurses, 192–3 difficult ET, 118 cervical dilatation, 120 past embryo transfer risks, 120 predictions of, 120–2 prevention strategies, 121–2 documentation, for ovarian tissue cryopreservation, 22 audits of, 23 donor oocytes, 89 dosage regimes, for ovarian stimulation protocols, 12–13 for GnRH, 12–13 SOP guidelines, 12 down-regulation stimulation protocols, 16 Edwards, Robert, 209 egg collection. See also laparoscopic egg retrieval; oocyte retrieval; transvaginal ultrasounddirected oocyte retrieval patient’s perspective on, 5 through TVOR, 104, 114–15

with TVOR, 104 ejaculatory function normal, 123 retrograde, 125 risk factors for, 123 electric oocyte aspiration pump functions of, 108 installation and set-up, 107–9 machine failure actions, 109 possible issues/solutions for, 108–9 electroejaculation, 125 Elonva Flare, 16 embryo cryopreservation, 2 downward trends for, investigation of, 134 bias in, 134 data volume, 134 embryo demographics, 135 laboratory changes, 135 patient demographics, 134 upstream factors in, changes to, 135 in OHSS, 94 ovarian tissue cryopreservation compared to, 19 poor survival after corrective actions, 135–7 culture conditions and, 133 environmental issues and, 133 equipment issues and, 133 freezing/thawing issues, 132 human factors for, 133–4 investigation of, 132 operator protocols as factor in, 132–3 patient history as factor in, 132 preventative measures for, 135–7 processes and factors for, 133 reporting considerations for, 134 embryo development. See also culture media; poor embryo development with endometrioma, 38 normal, 169 embryo transfer (ET). See also ‘lost’ embryos acupuncture and, 47–8 difficult, 118 cervical dilatation, 120 past embryo transfer risks, 120 predictions of, 120–2 prevention strategies, 121–2 early history of, 118 endometrial patterns, 75 ET catheter selection, 120 firm/soft catheters, 120 factors for, 118–20 bladder filling, 119–20 BMI, 118 cervical appearance, 118

Index

cervical canal, 119 cervical mucus and blood, 118–19 UCA, 119 utero-cervical angle, 119 FET, 76–7 luteal phase support and, 89, 90 intrauterine fluid issues and, 79, 82, 83 IUAS and, 44 IVF failure and, 161 ‘lost’ embryos in, 138–42 from accidents, 141 causes of, 139 from failure to follow processes, 140, 141 from moving between dishes, 138–41 mock, 121 stylet use, 121 tenaculum use, 121 timing of, 121 patient’s perspective on, 5–6 personal preference for, 120 processes for, 118 under sedation, 121 TMET, 121 ultrasound guidance, 120 empty follicle syndrome, 105 EMT. See endometrial thickness endometrial perfusion therapies, 77 endometrial thickness (EMT), 74 clinical significance of, 74–5 TVS for, 80 endometrioma, endometriosis and adenomyosis and, 29 bilateral, 36 embryo development with, 38 management of, 36–8 oocyte retrieval with, 38 patient history with, 36 pregnancy and, 38 treatment options, 36–7 adverse effects of, 37 conservative, 36 ESHRE guidelines for, 36 individualisation of care, 37 medical, 36 with stimulation protocols, 37 with surgery, 36–7 TVOR and, 105 unilateral, 36 endometriotic cysts, 102 endometrium. See also thin endometrium dating of, 74 EMT, 74 clinical significance of, 74–5 with IUAs, 44 patterns, 74 clinical significance, 75–6

ET, 75 polyps, 161 regeneration of, 43–4 sonographic measurement of, 74 EQA programmes. See external quality assurance programmes EQC. See External Quality Control erectile function normal, 123 risk factors for, 123 Escherichia coli, 144–5 ESHRE. See European Society of Human Reproduction and Embryology estrogen therapy for IUAs, 43–4 in luteal phase support, 89 for thin endometrium, 77 ET. See embryo transfer ET catheters, 120 firm/soft catheters, 120 European Society of Human Reproduction and Embryology (ESHRE), 2 endometrioma treatment guidelines, 36 KPI recommendations, 66, 67 poor responders to GnRH, definitions of, 14 European Union Tissues and Cells Directive, 71 external quality assurance (EQA) programmes, 72 External Quality Control (EQC), 178 fallopian tubes, assessment of with 3D ultrasound, 155–6 HyCoSy and, 155–6 with hydrosalpinges, 156 for patency, 155–6 falloposcopy, 60 performance of, 60 risks with, 60 fentanyl, 99 fertilisation failure (FF) clinical definitions of, 165–6 COH, 167 incidence rates, 165 individualisation of care, 167 lab environment changes, 166–7 male factors, 167 multidisciplinary input for, 167 peri-conception factors, 167 PFF, 165 polar bodies and, 165 TFF, 165–6, 167 fertility preservation methods, for cancer patients for boys, 26 for girls, 24, 26

through ovarian tissue cryopreservation, 25 for men, 26 for women, 24–6 with GnRH agonists, 24–5 through oocyte cryopreservation, 25–6 through ovarian tissue cryopreservation, 25 through ovary transposition, 25 fertility research, 191. See also research nurses patient choice in, 194, 195 stakeholders in, 194 fertility/infertility centers. See financial models; marketing fertiloscopy, 59–60 performance of, 60 risks with, 60 FET. See frozen embryo transfer fever management, 100 FF. See fertilisation failure financial models, for fertility/infertility centers balance sheets, 203–4 building of, 204 definition of, 203 business plans, 197–9 development of, 197–8 profit and loss accounts, 198–9, 201 unit setup in, 198–9 for center setup, 203 costs of running centers, 203–3 for consumables, 202 operating costs, 202 investments in, 203 operational options, 205 profitability measures, 204, 205 revenues from services, 201 primary sources, 201 secondary sources, 202 tactical and strategic options, 206 Foley balloon therapy, 42 Foley catheter with balloon, 42 follicle ripening, 49 follicles. See primordial follicles follicular flushing, 104 follicular recruitment, limited, 110–12 adequate, 110–12 cycles of, 112 hCG levels, 111–12 management of, 112 progesterone bioavailability, 111–12 bifollicular, 110 cycles of, 110 cancellation of, no follicular recruitment and, 110 monofollicular, 110

217

Index

follicular stimulating hormone (FSH), 11–12 OHSS and, 93–4 for poor responders to GnRH therapy, 15, 16 frozen embryo transfer (FET), 76–7 luteal phase support and, 89, 90 FSH. See follicular stimulating hormone gamete donation, 163 gas supply failure, in incubators, 127–8 identification of, 127–8 prevention of, 127 resolution of, 128 gatekeeper principle, 194 GcSF. See granulocyte colony stimulating factor gentamycin, 143 globozoospermia, 165 gonadotrophin-releasing hormone (GnRH), 12–13. See also poor responders in luteal phase support, 89, 90 OHSS and, 92–4 thin endometrium management with, 77 for women with cancer, 24–5 granulocyte colony stimulating factor (GcSF), 78 growth hormones, in adjuvant therapies, 17 hCG. See human chorionic gonadotropin HES. See hydroxy-ethyl starch HF. See human factors home readiness, 100 hormones AMH levels, 81 OHSS and, 92 FSH, 11–12 OHSS and, 93–4 GcSF, 78 GnRH, 12–13. See also poor responders in luteal phase support, 89, 90 OHSS and, 92–4 thin endometrium management with, 77 for women with cancer, 24–5 growth, in adjuvant therapies, 17 hCG, 77 adequate follicular recruitment and, 111–12 luteal phase support and, 86, 89 OHSS, 93–4 LH, 86 in luteal phase support, 87 estrogen, 89

GnRH agonists, 89, 90 LH, 86 progesterone preparations, 86–9 protocols for, 89–90 progesterone, 86–9 adequate follicular recruitment and, 111–12 in luteal phase support, 86–9 synthetic, 86 HSG. See hysterosalpingogram human chorionic gonadotropin (hCG), 77 adequate follicular recruitment and, 111–12 luteal phase support and, 86, 89 OHSS, 93–4 human embryo culture media. See culture media human factors (HF) after-action review, 184 checklists, 183–4 double checking, 184 environmental influences, 181–2 errors, 181 incident analysis, 184 in teamwork, 182–3 authority and, 182–3 leadership and, 182–3 variabilities in, 181–3 Human Fertilisation and Embryology Act (1991), UK, 2 hyalobarrier, 43 hyaluronic acid, 43 HyCoSy. See hysterosalpingo-contrastsonography hydrosalpinges causes of, 51 defined, 51 diagnosis of, 51–2 fallopian tube assessment and, 156 future research on, 53 intrauterine fluid issues and, 79–81 IVF and, 51–2 IVF failure and, 162 clinical trials, 162 treatment strategies, 51, 52–3 audits before, 53 with reduced ovarian reserve, 52 salpingectomy as, 52 TVOR and, 105 hydroxy-ethyl starch (HES), 94 hypotension, 100 hysterosalpingo-contrast-sonography (HyCoSy), 59 fallopian tube assessment and, 155–6 performance of, 59 purpose and function of, 59 risks of, 59 hysterosalpingogram (HSG), 40

with OSCM, 57 performance of, 57 purpose and function of, 56–7 risks of, 57–8 SIN, 57 tubal flushing at, 57 with WSCM, 57 hysteroscopic proximal intratubal occlusion, 81 hysteroscopic surgery for IUAs, 41–2, 44 second-look, 44 hysteroscopy, 161 ICMART. See International Committee for Monitoring Assisted Reproduction ICSI. See intracytoplasmic sperm injection IMSI. See intracytoplasmic morphologically selected sperm injection in vitro fertilization (IVF). See also assisted reproduction treatment; Complementary and Alternative medicine; infection; intramural fibroids; marketing; quality management system adenomyosis and, 29 in ambulatory settings, 97 birth of Louise Brown, 1 CHM and, 46–7 Cooke and, personal history of, 1–4 in Costa Rica, 3 costs of, 6 embryo cryopreservation, 2 future of, 209, 211 under Human Fertilisation and Embryology Act, 2 ICSI in, 2 laparoscopic egg retrieval, 1–2 OHSS, 3 SET in, 2–3 in UK, 11 workflow chart, 70 in vitro fertilization (IVF) failures, management plans for alcohol consumption and, 159 BMI factors, 159 embryo quality and selection, improvements to, 160–1 through AH, 160 through blastocyst transfer, 160 through PGS, 160–1 endometrial receptivity and, 161 ET and, 161 gamete donation and, 163 hydrosalpinges removal and, 162 clinical trials, 162

Index

hysteroscopy for, 161 intra-cavity lesions, 161–2 endometrial polyps, 161 IUAs, 161–2 submucous fibroid, 161 uterine septum, 161 multi-disciplinary approach to, 159–60 through lifestyle changes, 159 ovarian stimulation protocol, 160 with sperm DNA fragmentation, 160 myometrial pathology, 162 adenomyosis, 162 intramural fibroids, 162 in poor responders, 160 smoking as factor in, 159 surrogacy and, 163 incubator failure equipment malfunction, 129–30 identification of, 129–30 prevention strategies, 129 resolution of, 130 flowchart for, 130 gas supply failure, 127–8 identification of, 127–8 prevention of, 127 resolution of, 128 power supply failure, 128–9 identification of, 128–9 prevention strategies, 128 resolution of, 129 infection, in IVF laboratories Candida albicans, 144–5 consequences of, 145 Escherichia coli, 144–5 identification of, 144–5 prevention of, 145–7 resolution of, 145, 146 through cleaning solutions, 145 sources of, 143–4 from bacteria, 144–5 from environmental contaminant, 143–4 from follicular aspirates, 144 from fungus, 144 microbial, 146 in seminal fluid, 143 from staff, 143 types of, 143, 144 infertility. See also hydrosalpinges adenomyosis and, 28 WHO task force on, development of, 1 infertility centers. See financial models; marketing informed consent, 192 inositol, 17 Internal Quality Control (IQC), 178

International Committee for Monitoring Assisted Reproduction (ICMART), 2 Intra Uterine Insemination (IUI), 46–7 intracytoplasmic morphologically selected sperm injection (IMSI), 171 intracytoplasmic sperm injection (ICSI), 2, 165–6 intramural fibroids ART influenced by, 8 health outcomes, 7 IVF failure and, 162 management approaches, in clinical practice, 9–10 mechanism of action for, 9 myomectomy for, 9 laparoscopy for, 9 laparotomy for, 9 NNT assessment for, 9 non-cavity distorting, 8 prevalence of, 7 review data on, 7–8 size of, 8–9 submucosal, 7, 9 intrauterine adhesions (IUAs) causes of, 40 clinical features, 40 defined, 40 diagnosis of, 40–1 with HSG, 40 with hysteroscopy, 41 with ultrasound, 40–1 ET and, 44 future pregnancies with, management of, 44 incidence rates for, 40 IVF failure and, 161–2 postoperative assessment of, 44 of endometrial function, 44 with second-look hysteroscopy, 44 prevention strategies, 42–3 with amnion graft, 43 Cook intrauterine balloon, 43 Foley catheter with balloon, 42 hyaluronic acid, 43 IUCDs, 42 thin endometrium from, 76, 77 treatment strategies, 41–2 with antibiotics, 43 with estrogen therapy, 43–4 with Foley balloon therapy, 42 with hysteroscopic surgery, 41–2, 44 regeneration of endometrium, 43–4 ultrasound for, 155 intrauterine contraceptive devices (IUCDs), 42

intrauterine fluid issues aetiology, 79 causes of, 79 clinical implications for, 79 epidemiology, 79 ET and, 79, 82, 83 management of, 81–3 with AMH, 81 through clinical trials, 83 health outcomes, 81 pathophysiology, 79–81 in animal studies, 80 with hydrosalpinges, 79–81 pelvic anatomy scans for, 81 TVS for, 80 3D TVS, 80 intrauterine synaechiaes, 40 IQC. See Internal Quality Control IUAs. See intrauterine adhesions IUCDs. See intrauterine contraceptive devices IUI. See Intra Uterine Insemination IVF. See in vitro fertilization; in vitro fertilization failures JZ hyperplasia, 28, 29, 30 ketoconazole, 95 key performance indicators (KPI) ACE, 68 control charts and, 66 control data, 67 EQA programmes, 72 ESHRE recommendations, 66, 67 establishment of, 66–70 IVF workflow chart, 70 management of, 66–70 for oocyte cryopreservation, 134 preparation process flowchart, 69–70 in QMS, 178 red-alerts, 70–2 consumables and, 72 culture medium, 72 general clinics and lab work, 71 laboratory equipment and, 71–2 new processes, 70–1 staff response to, 72 VOC levels, 71 sperm production processes, 69–70 TQI approach and, 67–9 troubleshooting process flowchart, 71 laparoscopic egg retrieval, 1–2 laparoscopic tubal ligation, 81 laparoscopy for myomectomy, 9 for ovarian tissue cryopreservation, 21, 22

219

Index

tissue replacement in, 22 laparotomy, for myomectomy, 9 L-Arginine, 77 letrozole, 26, 95 levonorgestrel, 29 LH. See luteinizing hormone limited follicular recruitment. See follicular recruitment long agonist stimulation protocols, 11 ‘lost’ embryos, 138–42 from accidents, 141 causes of, 139 from failure to follow processes, 140, 141 from moving between dishes, 138–41 luteal phase support audits for, 90 donor oocyte cycles, 89 FET and, 89, 90 historical background for, 86 hormones used as, 87 estrogen, 89 GnRH agonists, 89, 90 hCG, 86, 89 progesterone preparations, 86–9 protocols for, 89–90 LH, 86 mechanisms for, 86 OHSS and, 86–9, 94 research for, 90 luteinizing hormone (LH), 86 MAC. See monitored anaesthesia care magnetic activation cell sorting (MACS), 171 magnetic resonance imaging (MRI), of adenomyosis, 28 males aging for ART consultations influenced by, 32 paternity influenced by, 33 sperm quality and, 32–4 ART and age criteria for, 32 clinical reasons for, 32 equipment for, 34 information sheets for, 34 ‘must have’ protocols, 34 strategies for, 34 fertility preservation methods for boys, 26 for men, 26 TFF and, 165–6, 167 manual aspiration pump, 109 machine failure actions, 109 marketing, of IVF assessments of markets, 186 branding in, 186–7

buyer behaviour and, 189, 190 control evaluation in, 190 customer service and, 188–9 defined, 186 marketing plans, 186–7 clinic location, 187, 190 pricing strategies, 187 for products and services, 187 positioning in, 186–7 promotion strategies, 187–8 in digital media, 187–8 in written media, 188 segmentation in, 186 staff for, 188–9 target markets, 186 metformin, 94 methylprednisolone, 95 mobile ovaries, 104 mock ET, 121 stylet use, 121 tenaculum use, 121 timing of, 121 monitored anaesthesia care (MAC), 98–9 monofollicular recruitment, 110 Mortimer, David, 66 Mortimer, Sharon, 66 mosaicism, 150–1 MRI. See magnetic resonance imaging myomectomy, for intramural fibroids, 9 laparoscopy for, 9 laparotomy for, 9 NNT assessment for, 9 National Institute of Health Research (NIHR), 191 nausea and vomiting, 100 necrospermia, 165 next generation sequencing (NGS), 150 NIHR. See National Institute of Health Research number needed to treat (NNT), for myomectomy, 9 nurses. See research nurses OHSS. See ovarian hyperstimulation syndrome oil-soluble contrast media (OSCM), 57 oligoasthenoteratozoospermia, 165 oocyte cryopreservation downward trends for, investigation of, 134 bias in, 134 data volume, 134 embryo demographics, 135 laboratory changes, 135 patient demographics, 134 upstream factors in, changes to, 135

for female cancer patients, 25–6 KPI for, 134 letrozole and, 26 ovarian tissue cryopreservation compared to, 19 poor survival after corrective actions, 135–7 culture conditions and, 133 environmental issues and, 133 equipment issues and, 133 freezing/thawing issues, 132 human factors for, 133–4 investigation of, 132 operator protocols as factor in, 132–3 patient history as factor in, 132 preventative measures for, 135–7 processes and factors for, 133 reporting considerations for, 134 semen sample production failure and, 126 success rates for, 26 oocyte retrieval. See also transvaginal ultrasound-directed oocyte retrieval with CAM, 49 donor, 89 with endometrioma, 38 ‘lost’ embryos in, 138–42 from accidents, 141 causes of, 139 from failure to follow processes, 140, 141 from moving between dishes, 138–41 organizational guidelines for, 115 OSCM. See oil-soluble contrast media ovarian follicle stimulation, 49 poor response to, 110 management of, 111 ovarian hyperstimulation syndrome (OHSS), 3, 11, 92 AMH and, 92 anaesthesia use and, 98, 100 coasting and, 12, 94 controlled ovarian stimulation, 92–4 coasting in, 94 with FSH, 93–4 with GnRH antagonists, 92–4 with hCG, 93–4 in luteal phase support, 94 cryopreservation and, 12 of embryos, 94 luteal phase support and, 86–9 pharmacologic interventions, 94–5 albumin, 95 comparisons between, 95 dopamine agonists, 94 HES, 94 IV calcium infusions, 95

Index

low-dose aspirin, 94 metformin, 94 prediction of, 92 demographics in, 92 ovarian reserve markers, 92 ovarian response parameters, 92 preventative measures, 92–5 ovarian reserve. See also reduced ovarian reserve PGD/PGS management and, 151 ovarian reserve markers, 92 ovarian response parameters, 92 ovarian stimulation protocols antagonist, 11 dosage regimes, 12–13 for GnRH, 12–13 SOP guidelines, 12 FSH in, 11–12 IVF failure and, 160 long agonist, 11 protocol selection, 12 ovarian tissue cryopreservation for cancer patients for girls, 25 for women, 25 consent for, 21 criteria for, 20 fertility risk assessments, 21–2 patient selection, 20–2 development of, 19–20 documentation for, 22 audits of, 23 embryo cryopreservation compared to, 19 global availability of, 22 key features of, 19–20 laparoscopy for, 21, 22 tissue replacement in, 22 oocyte cryopreservation compared to, 19 options in, 19 primordial follicles in, 25 radiotherapy risks, 22 ovaries, assessment of, 156–7 AFC, 156 follicle count, 156–7 ovary transposition, 25 pain management, anaesthesia use and, 100 patient engagement teams (PETs), 195 patient’s perspective, on assisted reproduction on egg collection day, 5 on ET, 5–6 IVF costs and, 6 PB biopsy. See polar body biopsy pelvic anatomy scans, 81 pelvic inflammatory disease (PID), 61 TVOR and, 102

penile vibratory stimulation (PVS), 125 pentoxifylline, 77 peri-conception issues, 167 PETs. See patient engagement teams PFF. See previous fertilisation failure PGD. See preimplantation genetic diagnosis PGS. See preimplantation genetic screening PID. See pelvic inflammatory disease platelet-rich plasma (PRP), 78 polar bodies, FF and, 165 polar body (PB) biopsy, 148–9 poor embryo development, 169–73 aneuploidy oocytes and, 170–1 causes of, 170–2 culture media integrity, 171–2 definition of, 169–70 IMSI and, 171 MACS and, 171 reprotoxicity and, 171–2 sperm integrity and, 171 sperm selection methods, 171 in subsequent cycles, prediction of, 172–3 poor responders, to GnRH therapy adjuvant therapies for, 16–17 androgens in, 16–17 growth hormones in, 17 Bologna Criteria for, 14, 15 counselling for, 17–18 for couples, 17 drop-out rates for, 16 ESHRE definitions for, 14 etiology of, 14 FSH doses for, 15, 16 heterogeneity of, 14 IVF failure in, 160 PGD/PGS management and, 151 predictive tests for, 14–15 prevalence of, 14 previous cycle outcomes, 15 reduced ovarian reserve and, 15 stimulation protocols in, 15–16 corifollitropin alfa in, 16 down-regulation, 16 Elonva Flare in, 16 unexpected, 15 power supply failure, in incubators, 128–9 identification of, 128–9 prevention strategies, 128 resolution of, 129 pregnancy adenomyosis, 29 anaesthesia use and, 98 CPR, 47 with endometrioma, 38 IUAs and, 44

preimplantation genetic diagnosis (PGD) biopsy procedures, 148–9 blastomere, 149 PB, 148–9 trophectoderm, 149 diagnostic techniques, 149–50 for chromosome disorders, 150 for genetic disorders, 149–50 with NGS, 150 early history of, 148 indications of, 148 management and policy strategies, 151–2 for inconclusive results, 152 with ovarian reserve issues, 151 with PGS, 151–2 with poor responders, 151 with untested cryopreserved embryos, 151 mosaicism and, 150–1 non-invasive sampling, 149 quality control standards in, 151 preimplantation genetic screening (PGS) advanced maternal age and, 148 biopsy procedures, 148–9 blastomere, 149 PB, 148–9 trophectoderm, 149 diagnostic techniques, 149–50 for chromosome disorders, 150 for genetic disorders, 149–50 with NGS, 150 with PGS indications, 150 early history of, 148 indications of, 148 IVF failure and, 160–1 management and policy strategies, 151–2 for inconclusive results, 152 with ovarian reserve issues, 151 with PGD, 151–2 with poor responders, 151 with untested cryopreserved embryos, 151 mosaicism and, 150–1 non-invasive sampling, 149 quality control standards in, 151 preparation process flowcharts, KPI, 69–70 previous fertilisation failure (PFF), 165 primordial follicles, 25 Pringle, Fiona, 207–8 progesterone, 86–9 adequate follicular recruitment and, 111–12 synthetic, 86 propofol, 99

221

Index

PRP. See platelet-rich plasma PVS. See penile vibratory stimulation QMS. See quality management system quality control, 178 quality management system (QMS), in IVF, 175–9 accreditation, 179 audits of, 178 clinic documents, 176, 177 in clinical settings, 175 components of, 174 defined, 174–5 document control, 176–8 equipment in, 176–7 feedback in, 178–9 incident reporting in, 178 KPIs for, 178 manuals, 176 non-conformities in, 178 patient satisfaction for, 178–9 personnel and, 175–6 satisfaction of, 178–9 policy objectives of, 176 for premises, 176–7 problem analysis, 179 quality control, 178 third-party agreements and, 176 validation of, 178 quinagolide, 94 radiation-induced uterine damage, 76 radiotherapy risks, 22 randomised control trials (RCTs), 192 reprotoxicity, 171–2 reduced ovarian reserve, 15 hydrosalpinge treatment strategies with, 52 research nurses in case studies procedures, 192 research recruitment in, 193 during setup, 191–2 data collection by, 192–3 data input by, 193 in gatekeeper principle, 194 informed consent and, 192 in national research agenda, 191 potential barriers, 195 in RCTs, 192 in referral process, 193–5 role and responsibilities of, 191, 192 safety reporting by, 193 retrograde ejaculation, 125 revenues. See financial models safety reporting, 193 saline infusion sonography (SIS), 41 3D SIS, 41

salpingectomy, 52 for intrauterine fluid issues, 81 salpingitis isthmica nodosum (SIN), 57 salpingoscopy. See falloposcopy salpingostomy, 81 second-look hysteroscopy, 44 sedation, ET under, 121 semen samples, failure to produce, 123–4 aetiology of, 123–3 ejaculatory function and normal, 123 retrograde, 125 risk factors for, 123 erectile function and normal, 123 risk factors for, 123 management strategies, 124–6 with drug treatment, 125 with electroejaculation, 125 pre-treatment risk assessment, 124 with PVS, 125 sample production at home, 124 timing of intercourse, 124–5 prevention strategies, 124–6 audiovisual aids, 124 cycle cancellation, 126 oocyte cryopreservation, 126 partner support as part of, 124 sperm cryopreservation, 124 risk factors for, 123–3 SSR and, 125–6 urinary sperm retrieval, 125 Seprafilm, 43 sequential culture media blastocyte-stage transfer, 64 defined, 63 single-step compared to, 63–4 SET. See single embryo transfer sildenafil, 77, 125 SIN. See salpingitis isthmica nodosum single embryo transfer (SET), 2–3 single-step culture media blastocyte-stage transfer, 64 defined, 63 sequential compared to, 63–4 SIS. See saline infusion sonography Smith, Kevin, 34 smoking, IVF failure and, 159 sperm cryopreservation, 124 sperm DNA fragmentation, 160 sperm donors, age guidelines for, 33–4 sperm integrity, 171 sperm production processes, KPI, 69–70 sperm quality, aging and, 32–3. See also semen samples child health and development, 33–4 DNA damage and, 32–3

SSR. See surgical sperm recovery Steptoe, Patrick, 118 stimulation protocols, 15–16 corifollitropin alfa in, 16 down-regulation, 16 Elonva Flare in, 16 endometrioma treatment options and, 37 ovarian follicles, 49 stylets, 121 subfertility, causes of, 97 submucous fibroids, 161 surgical sperm recovery (SSR), 125–6 surrogacy early development of, 3 IVF failure and, 159 synthetic progesterone, 86 tadalafil, 125 tenaculums, 121 TFF. See total fertilisation failure thin endometrium. See also endometrial thickness Asherman’s syndrome and, 76 causes of, 76 iatrogenic, 76 idiopathic, 76 IUA, 76, 77 Turner’s syndrome, 76 COH and, 76 definition, 74 FET and, 76–7 incidence rates, 74 management of, 76–8 with endometrial perfusion therapies, 77 with estrogen, 77 with GcSF, 78 with GnRH agonists, 77 with HGC, 77 with PRP, 78 3D SIS. See three-dimensional saline infusion sonography 3D TVS. See three-dimensional transvaginal ultrasound 3D ultrasound fallopian tube assessment with, 155–6 HyCoSy and, 155–6 with hydrosalpinges, 156 for patency, 155–6 historical background of, 153 ovary assessment with, 156–7 AFC, 156 follicle count, 156–7 uterine assessment with, 153–5 of acquired anomalies, 155 of congenital anomalies, 154–5 of endometrium, 153–4 of sub-endometrium, 153–4

Index

three-dimensional saline infusion sonography (3D SIS), 41 three-dimensional transvaginal ultrasound (3D TVS), 41, 80 TMET. See transmyometrial embryo transfer total fertilisation failure (TFF), 165–6 Total Quality Improvement (TQI) approach, 67–9 transmyometrial embryo transfer (TMET), 121 transvaginal aspiration, 81 transvaginal hydrolaparoscopy. See fertiloscopy transvaginal ultrasound (TVS) for EMT, 80 for intrauterine fluid issues, 80 3D TVS, 80 2D TVS, 40–1 3D TVS, 41, 80 transvaginal ultrasound-directed oocyte retrieval (TVOR) aspiration procedures, 103–4 blood vessel identification, 104–5 complications from, 104 early, 115–17 for egg collection, 114–15 late, 117 patient selection as factor in, 114 prevalence of, 114 procedural, 114 rates of, 102 risk management in, 117 egg collection and, 104, 114–15 electric oocyte aspiration pumps and functions of, 108 installation and set-up, 107–9 machine failure actions, 109 possible issues/solutions for, 108–9 with empty follicle syndrome, 105 endometriosis and, 105 endometriotic cysts and, 102 follicles and, 104–5 follicular flushing and, 104 hydrosalpinx, 105 with large uterus, 105 manual aspiration pumps and, 109 machine failure actions, 109 ovary location and, 104 mobile ovaries, 104 above vaginal fornices, 104

PID and, 102 preliminary steps of, 102–3 comorbidities and, 102–3 transvaginal probe, 103 urination procedures, 103 temperature control with, 104 trophectoderm biopsy, 149 troubleshooting process flowchart, KPI, 71 tubal assessment, 58. See also hysterosalpingogram CAT in, 60–1 PID and, 61 comparisons between tests, 61 falloposcopy for, 60 performance of, 60 risks with, 60 fertiloscopy for, 59–60 performance of, 60 risks with, 60 HyCoSy for, 59 performance of, 59 purpose and function of, 59 risks of, 59 infertility rates and, 55 laparoscopy and dye for, 58–9 function and purpose of, 58 performance of, 58–9 risks of, 59 methods of, 56–61 patency, of Fallopian tubes, 55 timing of, 55–6 with no history of tubal assessments, 56 with tubal blockage risk factors, 55 with un-investigated tubal abnormalities, 55, 56 with uterine abnormalities, 55 tubal disease. See hydrosalpinges Turner’s syndrome, 76 TVOR. See transvaginal ultrasounddirected oocyte retrieval TVS. See transvaginal ultrasound two-dimensional transvaginal ultrasound scan (2D TVS), 40–1 UCA. See utero-cervical angle UK. See United Kingdom ultrasound. See also 3D ultrasound; transvaginal ultrasound

for adenomyosis, 155 early history of, 1 for ET, 120 for IUAs, 40–1, 155 SIS, 41 2D TVS, 40–1 3D TVS, 41 unilateral endometrioma, 36 United Kingdom (UK) Human Fertilisation and Embryology Act, 2 IVF centres in, 11 urinary retention, 100 urinary sperm retrieval, 125 utero-cervical angle (UCA), 119 uterus, assessment of IVF failure and, 161 with 3D ultrasound, 153–5 for acquired anomalies, 155 for congenital anomalies, 154–5 of endometrium, 153–4 of sub-endometrium, 153–4 TVOR and, 105 vaginal fornices, 104 vardenafil, 125 vitamin E, 77 volatile organic compounds (VOCs), 71 Warnock, Mary, 3 water-soluble contrast media (WSCM), 57 WHO. See World Health Organisation whole system Traditional Chinese Medicine (WS-TCM), 48 women, with cancer, fertility preservation methods for, 24–6 with GnRH agonists, 24–5 through oocyte cryopreservation, 25–6 through ovarian tissue cryopreservation, 25 through ovary transposition, 25 World Health Organisation (WHO), task force on infertility, development of, 1 WSCM. See water-soluble contrast media WS-TCM. See whole system Traditional Chinese Medicine

223