Gregory Horne

Semen cryopreservation, utilisation and reproductive outcome in men treated for Hodgkin's disease

Between 1978 and 1990, 122 men underwent semen analysis before starting sterilising chemotherapy for Hodgkins disease. Eighty-one (66%) had semen quality within the normal range, 25 were oligospermic (<20×106 sperm per ml) and five were azoospermic (no sperm in the ejaculate). Semen from 115 men was cryopreserved and after a median follow-up time of 10.1 years, 33 men have utilised stored semen (actuarial rate 27%) and nine partners have become pregnant resulting in 11 live births and one termination for foetal malformation. Actuarial 10 year rates of destruction of semen before death or utilisation and death before utilisation are 19% and 13% respectively. This retrospective cohort study demonstrates that approximately one-quarter of men utilising cryopreserved semen after treatment for Hodgkins disease obtain a live birth. The high non-utilisation rate is intriguing and warrants further investigation.

Use of in-vitro fertilisation embryos cryopreserved for 5 years or more

Human embryos cryopreserved after in-vitro fertilisation can be stored initially for 5 years, and the storage period may be extended to a maximum of 10 years. Of 1344 embryos cryopreserved between 1988 and 1994 at two centres in Manchester, 67% (904 embryos) have had to be destroyed at the end of the first 5-year interval, even if the couples involved remain childless.

Reducing risk in the IVF laboratory: Implementation of a double witnessing system

prises an in-vitro system designed to mimic events which normally take place in the body (i.e. fertilization of the egg by sperm), and the interface between that system and the couple being treated. Clearly risk can be present both in vitro and at the interface. The management of risk associated with the latter is considered by Kennedy in this issue.1 This article deals with risk arising specifically in the IVF laboratory. In any in-vitro system the environment is responsible for any differences from the in-vivo situation. These differences generally entail a level of risk. In the case of IVF, environmental hazards can cause damage to sperm, eggs and embryos including for example errors in fertilization, damage to sperm DNA, or bacterial, viral or fungal contamination of embryos. These can be a source of risk to any resulting fetus, but also to the mother, as contaminated embryos may carry a risk of infection and abnormal embryos a risk of miscarriage. A second major source of risk in the IVF laboratory is of course the potential loss of gametes or embryos, particularly those held in frozen storage in liquid nitrogen banks. Finally, perhaps the most high profile risk is that of confusion or ‘mix-ups’ in the IVF laboratory between gametes or embryos of different couples. INTRODUCTION There is an extensive literature on the effect of the laboratory environment on gamete and embryo integrity and development, and the risks involved therein.2,3 The issues of quality assurance in maintaining cryobanks of sperm and embryos and a general risk management approach to IVF laboratories are also dealt with in detail elsewhere.4,5 The focus of this review will therefore be the risk of mix-ups between gametes and embryos in IVF and the introduction of systems for reducing this risk.

Waiting for in vitro fertilization treatment: Spontaneous and ART live births

This study analysed the live birth rates in 760 couples referred in 1994 to St Marys Hospital, Manchester, a non-fee-paying National Health Service (NHS) centre, who had waited for up to 4 years for IVF treatment. These live birth rates were compared with those of 199 couples referred at a similar time to Manchester Fertility Services, a fee-paying unit, where they received IVF treatment shortly after referral. The waiting time was advantageous in that 17.8% (135 of 760) of the couples referred to St Marys Hospital conceived without IVF treatment, 60% within one year of referral. However, the waiting time was detrimental to women aged 30–34 in whom treatment was delayed by 3–4 years. Only 26.8% (204 of 760) of couples originally referred eventually received NHS-funded IVF treatment at St Marys. A waiting time not exceeding 18 months would allow most spontaneous conceptions and reduce the adverse effect of prolonged waiting on the take-up rate for treatment and on the chance of success in the older women.

Who needs ICSI? A nationwide UK survey on ICSI use

Background: Intracytoplasmic sperm injection (ICSI) has been a milestone in the treatment of male factor infertility. However ICSI is more expensive, demands more expertise, and involves more risk than conventional in vitro fertilisation (IVF). Currently there are large nationwide differences in ICSI usage, with some centres using ICSI for 21% of their IVF cycles and others for more than 80%. This is, most likely, due to differences in ICSI selection criteria but there are limited data on the criteria used. We have therefore carried out a national survey in the UK, the first, as far as we are aware, to examine different criteria used and their effect on ICSI usage and treatment outcomes. Methods: Centres which offer ICSI were identified using the Human Fertilisation and Embryology Authority (HFEA) website. Questionnaires were then posted to all centres which offer the procedure. Each centre received the questionnaire twice; the first was sent to the HFEA person responsible and a month later, a follow-up questionnaire was sent to the centre’s lead embryologist. Data were also extracted from the HFEA website. Results: 71 centres were identified and questionnaires returned from 43 (61%). When deciding to use ICSI, 43 (100%) of centres used sperm count, 93% used sperm motility, 76% used sperm morphology and 72% used anti-sperm antibodies. All centres stated that they would offer ICSI after failed fertilisation with conventional IVF and 38% of centres offered ICSI on patient request. No centres reported using other criteria for selection. The absolute values chosen for each criterion varied hugely between centres. Compared with the 2010 World Health Organization (WHO) guidelines of normal semen analyses, 32% of centres used a higher count, 50% a higher motility and 59% a higher morphology. Based on the WHO criteria, 27% of centres would use ICSI for sperm that were normal by all WHO criteria. Between centres, no significant difference in ICSI fertilisation rates was found. However, there was a significant negative correlation between increased ICSI usage and fertilisation rates by conventional IVF (p = 0.0058). Data obtained from the HFEA website failed to demonstrate an increase in live birth rate in centres using ICSI more frequently. Conclusion: ICSI usage varied widely, due to large differences in the ICSI selection criteria used, with many centres using ICSI for patients with normal semen parameters. Centres which used more ICSI did not report higher live birth rates. No evidence was found to suggest that higher ICSI usage increased overall fertilisation rates. These findings highlight the need for guidelines on when to use ICSI.

Implantation, clinical pregnancy and miscarriage rates after introduction of ultrasound-guided embryo transfer

The purpose of this study was to compare the reproductive outcome of ultrasound-guided (USG) embryo transfers versus clinical touch embryo transfers. A statistically powered retrospective analysis of women undergoing fresh and frozen embryo transfers in a National Health Service-based tertiary referral centre in the Department of Reproductive Medicine, St Marys Hospital, Manchester was carried out. A total of 1723 embryo transfers were included in the analysis. The implantation rate was significantly higher in the USG embryo transfer group compared with the non-USG embryo transfer group (fresh: 19.9 versus 9.5%, P < 0.0001; frozen: 13.1 versus 7.3%, P < 0.0004). The clinical pregnancy rate was also significantly higher in the former group (fresh: 26.9 versus 12.5%, P < 0.0001; frozen: 15.6 versus 8.9%, P < 0.0015). For the frozen embryos, the miscarriage rate was significantly elevated among the USG embryo transfer group [unadjusted rate ratio (RR) = 1.65, 95% CI: 1.04, 2.62], but this was of borderline significance when the model was adjusted for the potential confounders (adjusted RR = 1.56, 95% CI: 0.997, 2.45). There was no difference in the ectopic pregnancy rates between the two groups. The findings of this study show that the practice of USG embryo transfer is associated with statistically higher implantation and clinical pregnancy rates in IVF.

Alkylation of sperm DNA is associated with male factor infertility and a reduction in the proportion of oocytes fertilised during assisted reproduction

Approximately one-third of IVF cases in the UK are attributed to male factor infertility and in the majority of cases the origin of male infertility is unknown. The integrity of sperm DNA is important both for the success of assisted reproduction and the implications for the off-spring. One type of DNA damage that has not been investigated with respect to fertility outcomes is the adduct N7-methyldeoxyguanosine (N7-MedG), a biomarker for exposure to alkylating agents. A prospective cohort of couples attending for IVF had their N7-MedG levels in sperm measured using an immunoslot blot technique to examine whether sperm N7-MedG levels are associated with male factor infertility, semen quality measures or assisted reproduction outcomes. Sufficient DNA for analysis was obtained from 67/97 couples and N7-MedG was detected in 94% of sperm samples analysed. Men diagnosed with male factor infertility had significantly higher mean levels of N7-MedG in their sperm DNA (P=0.03). Logistic regression analysis showed that N7-MedG levels were significantly negatively associated with the proportion of oocytes successfully fertilised irrespective of the method of fertilisation used (IVF or intra-cytoplasmic sperm injection; ICSI, P<0.001). Therefore exposure to DNA alkylating agents is significantly associated with male infertility and the proportion of oocytes fertilised during assisted reproduction. Reducing such exposure may improve male fertility but further work is required to determine the relative importance of exogenous and endogenous sources of exposure.

FSH receptor genotype does not predict metaphase-II oocyte output or fertilization rates in ICSI patients

The objective of this study was to assess the role of the variant p.Asn680Ser in the FSH receptor gene (FSHR) in determining oocyte maturity. It also assessed the relationship between this FSHR variant with metaphase-II oocyte output rate (MOR) and the fertilization rate. This was a prospective observational study based at a tertiary referral centre for reproductive medicine. Women (n=212) undergoing their first cycle of ovarian stimulation for IVF with intracytoplasmic sperm injection (ICSI) were included in the study. Baseline pelvic ultrasound and blood tests were taken on day 2 or 3 of the cycle for assessment of baseline hormones and for DNA extraction. Genotypes for FSHR p.Asn680Ser was determined using TaqMan allelic discrimination assay. The outcome measures were the total dose of exogenous gonadotrophins used, antral follicle count (AFC), number of mature (metaphase-II) oocytes retrieved, MOR and fertilization rate. No statistically significant differences were found between the number of mature oocytes retrieved, MOR or fertilization rates among the patients with different p.Asn680Ser FSHR genotypes. No significant difference was noted in the clinical pregnancy rates per transfer. There is no evidence that the p.Asn680Ser FSHR genotype predicts oocyte maturity.

Testing for hypersensitivity to seminal fluid-free spermatozoa

Abstract Hypersensitivity to human semen (HHS) is an increasingly reported condition with symptoms manifested locally and systemically, which in some cases may result in anaphylaxis. This report describes four cases of HHS all with positive allergy skin prick tests to partners whole semen. None of the patients elicited a response to seminal fluid-free washed spermatozoa. In cases of high risk of anaphylaxis, we recommend avoiding exposure to semen. By carrying out an allergy skin prick test on seminal fluid-free washed spermatozoa, the risk of immunogenic reaction to the spermatozoa could be eliminated.

The recruitment of oocyte donors

exceed 46% resulting in a detection rate of 1%. During the study year, 32% of amniocenteses in the higher risk group were in women less than 35 years old. In the year before serum screening was introduced, 154 amniocenteses were performed against 150 in the study year, Summary data for Wales, however, suggested an increase in amniocenteses due to serum screening. In our study, the overall prevalence of Down’s syndrome in screened women was 1 in 600. For those with a higher risk result, it was 1 in 42 and for those with a low risk result, 1 in 1160: a 28-fold difference. This study was not designed to test the statistical hypotheses of screening, but we were interested to note that our detection rate using two analytes was similar to contemporary studies using the triple test (Haddow etal. 1992; Wald etal. 1992). Before serum screening became available, concerns about Down’s syndrome were not raised with women below the age of 35. At UHW an extra 10 min for the midwife and clinic doctor were required to discuss the test before screening, and for those women with a higher risk result, at least another 30 min were necessary. We were particularly concerned about potential psychological effects in the women with false positive and negative results; our experience emphasised the need for sound primary counselling. One case demonstrated multiple benefits for a woman who did not want a termination. Our impression is that at present evaluation of the serum screen for Down’s syndrome is incomplete. The participants believe that amniocentesis or other diagnostic tests should remain available for women aged 35 and over. This is especially so for women aged 38 and over where age is the dominant risk factor and serum screening often only introduces a time delay before amniocentesis is carried out (Reynolds er al. 1993).