Clinicians' perspectives on diagnosing polycystic ovary syndrome in Australia: a qualitative study.

STUDY QUESTION: What are clinicians' views about the diagnosis of polycystic ovary syndrome (PCOS), and how do they handle any complexities and uncertainties in practice? SUMMARY ANSWER: Clinicians have to navigate many areas of complexity and uncertainty regarding the diagnosis of PCOS, related to the diagnostic criteria, limitations in current evidence and misconceptions surrounding diagnosis, and expressed concern about the risk and consequences of both under- and overdiagnosis. WHAT IS KNOWN ALREADY: PCOS is a complex, heterogeneous condition with many areas of uncertainty, raising concerns about both underdiagnosis and overdiagnosis. Quantitative studies with clinicians have found considerable variation in diagnostic criteria used and care provided, as well as a lack of awareness around the breadth of PCOS features and poor uptake of recommended screening for metabolic complications. Clinicians' views about the uncertainties and complexities of diagnosing PCOS have not been explored. STUDY DESIGN, SIZE, DURATION: Semi-structured telephone interviews were conducted with clinicians from September 2017 to July 2018 to explore their perceptions about the diagnosis of PCOS, including how they handle any complexities and uncertainties in practice. PARTICIPANTS/MATERIALS, SETTING, METHODS: A group of 36 clinicians (15 general practitioners, 10 gynaecologists and 11 endocrinologists) currently practicing in Australia, were recruited through advertising via professional organisations, contacting a random sample of endocrine and gynaecology teams across Australia and snowballing. Transcribed audio-recordings were analysed thematically using Framework analysis. MAIN RESULTS AND THE ROLE OF CHANCE: Clinicians expressed a range of uncertainties and complexities regarding the diagnosis of PCOS, which were organised into three areas: (i) establishing diagnosis (e.g. lack of standardisation regarding diagnostic cut-offs, risk of misdiagnosis), (ii) factors influencing the diagnostic process (e.g. awareness of limitations in evidence and consideration of the benefits and harms) and (iii) strategies for handling challenges and uncertainties (e.g. using caution and communication of uncertainties). Clinicians also varied in their concerns regarding under- and overdiagnosis. Overall, most felt the diagnosis was beneficial for women provided that it was the correct diagnosis and time was taken to assess patient expectations and dispel misconceptions, particularly concerning fertility. LIMITATIONS, REASONS FOR CAUTION: There is possible selection bias, as clinicians who are more knowledgeable about PCOS may have been more likely to participate. Clinicians' views may also differ in other countries. WIDER IMPLICATIONS OF THE FINDINGS: These findings underscore the vital need to first consider PCOS a diagnosis of exclusion and use caution before giving a diagnosis in order to reduce misdiagnosis, as suggested by clinicians in our study. Until there is greater standardisation of diagnostic criteria, more transparent conversations with women may help them understand the uncertainties surrounding the criteria and limitations in the evidence. Additionally, clinicians emphasised the importance of education and reassurance to minimise the potential harmful impact of the diagnosis and improve patient-centred outcomes. STUDY FUNDING/COMPETING INTEREST(S): The study was funded by the University of Sydney Lifespan Research Network and an NHMRC Program Grant (APP1113532). T.C. is supported by an Australian Government Research Training Program (RTP) Scholarship and a Sydney Medical School Foundation Scholarship, from the The University of Sydney, Australia. B.W.M. reports consultancy for ObsEva, Merck, Merck KGaA and Guerbet. No further competing interests exist. TRIAL REGISTRATION NUMBER: N/A.

The benefits and harms of receiving a polycystic ovary syndrome diagnosis: a qualitative study of women's experiences.

STUDY QUESTION: What are the benefits and harms of receiving a polycystic ovary syndrome (PCOS) diagnosis in a community sample of women, including impact on psychosocial wellbeing, lifestyle choices and behaviour? SUMMARY ANSWER: Although some women benefit considerably from the diagnosis, such as through increased awareness and reassurance, women with minimal symptoms may experience more harm than benefit, including long-lasting anxiety and altered life plans. WHAT IS KNOWN ALREADY: Disease labels can validate symptoms and play a vital role in understanding and coping with illness; however, they can also cause harm by evoking illness schemas about severity and permanence. Regarding PCOS, the diagnostic criteria have expanded over time to include women with milder phenotypes (such as those without signs of androgen excess). This has occurred despite limited investigation of the benefits and harms of the diagnosis and has increased the number of women diagnosed. STUDY DESIGN SIZE DURATION: Semi-structured interviews were conducted face-to-face or by telephone with 26 participants from April-July 2018 to explore women's experiences with the diagnosis, including the benefits and harms of receiving the diagnosis and the impact on their life. PARTICIPANTS/MATERIALS SETTING METHODS: In total, 26 women in the community self-reporting a diagnosis of PCOS (reporting mild to severe symptoms) made by a medical doctor, aged 18-45 years and living in Australia were recruited through social media. Data were analysed thematically using Framework analysis. MAIN RESULTS AND THE ROLE OF CHANCE: The study identified a range of both positive and negative effects of a PCOS diagnosis in the immediate, short and long-term, which were influenced by symptom severity, expectations and experience. For women with previously unexplained and bothersome symptoms, it was a relief to receive a diagnosis, and this resulted in an increased understanding about the importance of a healthy lifestyle. By contrast, women with milder symptoms often reported feeling shocked and overwhelmed by the diagnosis, consequently experiencing anxiety about the associated long-term risks. The majority of women, regardless of symptom severity, experienced prolonged worry and anxiety about infertility, resulting for some in risk taking with contraception, unintended pregnancies, pressure to conceive early or altered life plans. With time, many women developed positive coping strategies and perceived the diagnosis to be valuable, including those who felt they had experienced minimal benefit or even harm. LIMITATIONS REASONS FOR CAUTION: PCOS diagnosis was self-reported and the sample was highly educated. WIDER IMPLICATIONS OF THE FINDINGS: Fear of infertility was salient for many women, underscoring the need for accurate information, counselling and reassurance of fertility potential. Given the risk of significant consequences, health professionals should use a tailored approach to PCOS diagnosis to increase the benefits of appropriate and timely diagnosis for women affected by significant symptoms, while reducing the harms of unnecessarily labelling healthy women for whom the benefits of a diagnosis are small. STUDY FUNDING/COMPETING INTERESTS: The study was funded by the University of Sydney Lifespan Research Network and an NHMRC Program Grant (APP1113532). B.W.M. reports consultancy for ObsEva, Merck, Merck KGaA and Guerbet. No further competing interests exist. TRIAL REGISTRATION NUMBER: N/A.

A decrease in serum estradiol levels after human chorionic gonadotrophin administration predicts significantly lower clinical pregnancy and live birth rates in in vitro fertilization cycles.

BACKGROUND: Although close observation of serum estradiol (E2) levels remains a mainstay of assessing clinical response to controlled ovarian stimulation, the prognostic value of any change in E2 levels after administration of hCG remains unclear. The objective of this study is to evaluate the relationship between serum E2 response after hCG administration and the clinical pregnancy and live birth rates in fresh IVF cycles. METHODS: We conducted a retrospective cohort study of women aged 21-45 years undergoing their first IVF cycle from 1999 to 2008 at a single practice. We compared the post-hCG serum E2 level with values on the day of hCG trigger. IVF cycles were stratified by post-hCG E2 response and appropriate parametric and non-parametric statistics were performed. Clinical intrauterine pregnancy and live births were the primary outcomes of interest. Multivariable logistic regression models were created to identify predictive factors associated with outcomes while adjusting for potential confounders. RESULTS: Among the 1712 IVF cycles, 1065 exhibited a >10% increase (Group A), 525 had a plateau (± 10%, Group B) and 122 showed a >10% decrease (Group C) in post-hCG E2 levels. While the E2 levels on the day of hCG were similar across groups, Group C had more patients with diminished ovarian reserve, required higher gonadotrophin doses and had the lowest implantation rates. After adjusting for age, total gonadotrophin dose, infertility diagnosis, number of oocytes and number of transferred embryos, the associations between post-hCG E2 decline (Group C) and clinical pregnancy [adjusted odds ratio (aOR): 0.53; 95% confidence interval (CI): 0.33-0.84, P= 0.007] and live birth (aOR: 0.40; 95% CI: 0.22-0.71, P= 0.002) were significant. We also found significant associations between E2 plateau (Group B) and clinical pregnancy (aOR: 0.73; 95% CI: 0.57-0.94, P= 0.013) and live birth (aOR: 0.74; 95% CI: 0.56-0.97, P= 0.032) when adjusting for the same factors. CONCLUSIONS: In our study, >10% decrease in E2 levels after hCG administration was associated with 40-50% reduction in clinical pregnancy and live birth rates. Similarly, post-hCG E2 plateau (± 10%) lowered the clinical pregnancy and live birth rates by >25%. Our study suggests that the change in the post-hCG E2 level is another parameter that can be used by clinicians to counsel patients regarding their likelihood of success with assisted reproductive technologies prior to oocyte retrieval.

The tumour suppressor gene maspin is differentially regulated in cytotrophoblasts during human placental development.

Identification of factors that play a role in regulating the highly invasive ability of human placental cells throughout gestation will contribute to a better understanding of this unique developmental process. The aims of this study were to determine whether the tumour suppressor gene maspin is present in the human placenta and plays a putative role in the regulation of cytotrophoblast invasion during placental development. The data showed that the expression of maspin mRNA was maximum in term placentae compared to the first and second trimester tissues, and absent in the HTR-SVneo (immortalized extravillous cytotrophoblast), JEG-3 and JAR (choriocarcinoma) cell lines. Maspin protein, detected by Western blot analysis, was twofold higher in the second trimester and 4.4-fold higher in the third trimester compared to the first trimester. Maspin immunohistochemical staining was localized in cytotrophoblasts with increased and more diffuse staining in the second and third trimesters. Corresponding to the period of maximum maspin expression, cytotrophoblasts isolated from term placentae had significantly lower invasive ability as compared to first and second trimester cytotrophoblasts (P< 0.03). Further, addition of recombinant maspin significantly decreased cytotrophoblast invasion in vitro by 40-50 per cent in all three trimesters of gestation. This study provides the first evidence of the temporal expression of maspin during human gestation and suggests a putative role for maspin in regulating the invasive activity of cytotrophoblasts at term. The down-regulation of maspin expression may be critical at the time of implantation and early placental development, whereas upregulation of maspin may serve as a signal for the end of cytotrophoblast invasion and gestation.

Regulation of human cytotrophoblast morphogenesis by hepatocyte growth factor/scatter factor.

In vitro morphogenesis of epithelial cells to form tube-like structures is regulated by hepatocyte growth factor-scatter factor (HGF/SF). The placenta is a rich source of HGF/SF, and its absence in mice has been shown to lead to impaired placental growth and embryonic death. There is no information in the literature regarding in vitro morphogenesis of human cytotrophoblasts or the effect of HGF/SF on this process. In this study, cytotrophoblasts were isolated from human placentae obtained from all three trimesters of gestation and cultured on the recombinant basement membrane matrix (Matrigel). Under these conditions, cytotrophoblasts participated in morphogenetic events including formation of spheroid-like structures, radial linear processes with branching, and invaded Matrigel and formed large, tube-like structures. The presence of a developing lumen was documented in the linear projections arising from spheroids and in the tube-like structures by both confocal and transmission electron microscopy. Immunohistochemistry was used to characterize the phenotype of the cells, and staining with anti-cytokeratin and anti-E-cadherin antibodies confirmed the presence of cytotrophoblasts in both the spheroids and tube-like structures. Recombinant HGF (rHGF) significantly increased the invasive activity of cytotrophoblasts isolated from the first and second (P < 0.001) and third trimesters (P < 0.01). In addition, rHGF significantly increased the percentage of spheroids with branching processes in the first and second trimesters (P < 0.05). Anti-HGF antibody inhibited both these effects in a dose-dependent manner, indicating the specificity of the above findings. This study provides new evidence indicating that HGF/SF regulates invasion and branching morphogenesis of cytotrophoblasts throughout gestation, with maximum effects in the first and second trimester. These findings may help to elucidate the importance of the reduced expression of HGF/SF identified in placentae from women with preeclampsia or intrauterine growth restriction and suggest that HGF/SF may serve as an important candidate in therapeutic intervention strategies.

Bilateral undescended ovaries: association with infertility and treatment with IVF.

OBJECTIVE: To describe the clinical findings in a patient with bilateral undescended ovaries and infertility who was successfully treated by IVF. DESIGN: Case report. SETTING: University hospital. PATIENT(S): A 35-year-old woman with bilateral undescended ovaries. INTERVENTION(S): Hysterosalpingography, laparoscopy, and an IVF cycle. MAIN OUTCOME MEASURE(S): Anatomic description and pregnancy test. RESULT(S): This patient conceived and delivered a twin gestation after laparoscopic retrieval of oocytes and transfer of two blastocysts. CONCLUSION(S): Bilateral undescended ovaries is a rare condition that can be associated with infertility but can be successfully treated by IVF.

Secretion of inhibin B during ovarian stimulation is decreased in infertile women with endometriosis.

OBJECTIVE(S): To determine the levels and pattern of inhibin B secretion during ovarian stimulation in patients with endometriosis and to evaluate the correlation between inhibin B levels and variables of follicular and oocyte development. DESIGN: Retrospective cohort study. SETTING: IVF center. PATIENT(S): Women with endometriosis (n = 20) and tubal factor (n = 10) as the only cause of infertility. INTERVENTION(S): Inhibin B levels were measured during gonadotropin stimulation on day 1, days 8-11, day of hCG administration, and oocyte retrieval. MAIN OUTCOME MEASURE(S): Patterns and levels of inhibin B secretion were compared in the two groups. Other variables analyzed were age, days of gonadotropin stimulation and total ampules used, oocytes retrieved, and the fertilization and pregnancy rate. RESULT(S): Inhibin B levels and number of oocytes retrieved were significantly lower in patients with endometriosis than in the control group. Two patterns of inhibin B secretion were noted: It peaked in the midfollicular phase or on the day of hCG administration. In contrast, E2 levels peaked on the day of hCG administration. Inhibin B in the midfollicular phase in both groups correlated positively with number of oocytes retrieved. In addition, E2 and inhibin B levels in the midfollicular phase correlated positively. CONCLUSION(S): Decreased inhibin B secretion in patients with endometriosis suggests impaired granulosa cell function. Inhibin B may serve as an alternate marker to assess follicular development or to predict the number of oocytes retrieved.

Cornual heterotopic pregnancy: contemporary management options.

This review covers the clinical presentations, treatments, and outcomes of cornual heterotopic pregnancies reported in the literature. Infertile women with a history of ectopic pregnancy, tubal surgery, or disease are at increased risk for cornual heterotopic pregnancy when they undergo in vitro fertilization. Women who have undergone bilateral salpingectomy also seem to be predisposed to this condition when they undergo in vitro fertilization. We recommend that these patients be followed up closely after a successful in vitro fertilization cycle with monitoring of serum beta-human chorionic gonadotropin levels and serial transvaginal ultrasonography because of the high associated morbidity. Laparotomy remains the treatment of choice for rupture of a cornual heterotopic pregnancy. In the absence of cornual rupture, however, medical management is an option that eliminates the risk of surgery and anesthesia and results in outcomes similar to those associated with surgical treatment. Currently there is insufficient evidence to recommend any single treatment modality, and the decision should be based on such factors as clinical presentation, surgeon's expertise, side effects, overall cost, and the patient's preference.

Septate uterus associated with cervical duplication and vaginal septum.

A woman had an unusual müllerian anomaly with a septate uterus, cervical duplication, and longitudinal vaginal septum. Gynecologists should be aware of the possibility of cervical duplication associated with uterine septum and not didelphic uterus, as this disorder in a patient with infertility or recurrent miscarriages can be treated surgically by resection of the uterine and vaginal septum. Embryologic explanations may account for the occurrence of the anomaly.

Endometriosis and assisted reproductive technologies.

The association between endometriosis and infertility is well established, and different mechanisms have been postulated to cause infertility depending on the stage of endometriosis. Controlled ovarian hyperstimulation improves pregnancy outcome for minimal to mild endometriosis; however, there are no randomized controlled studies of this technique in patients with severe disease. Further, ART appears to have an overall beneficial effect for all stages of endometriosis. Based on the current staging system, several studies have not been able to demonstrate a significant difference in true outcomes for different stages of the disease. The limitations of these studies have been discussed above. Similarly, the effect of endometriomas when considered independent of the stage is still unclear. In addition to ART, pretreatment with GnRH analogues or surgical intervention may be beneficial and needs to be addressed in randomized controlled studies. It is evident that we urgently need large randomized studies to answer several questions about the treatment of endometriosis-associated infertility with ART. Although our current practice is based primarily on retrospective observational studies and small randomized studies, the overall impact on pregnancy outcome is beneficial.

Warfarin skin necrosis in a postpartum woman with protein S deficiency.

BACKGROUND: Warfarin-induced skin necrosis is a rare and dangerous complication affecting 0.01-0.1% of patients on warfarin. Deficiencies in protein C or protein S in association with other factors have been implicated in its etiology. No report has described this disorder in the immediate postpartum period in patients with protein S deficiency. CASE: A 1-week postpartum woman with known protein S deficiency presented with skin necrosis after a previously uneventful course of warfarin. CONCLUSION: Reduced levels of free protein S during the antepartum and immediate postpartum periods predispose protein S-deficient women to warfarin skin necrosis. Previously uncomplicated courses of warfarin do not obviate the possibility of skin necrosis with future warfarin administrations. Initiation of low-dose warfarin with heparin can reduce the likelihood of this disorder.

Embryotoxicity as a marker for recurrent pregnancy loss.

Some cases of unexplained recurrent pregnancy loss may be due to embryotoxic factors present in the mother's serum. In vitro assays have been devised to study the effect of these factors on pre-implantation, peri-implantation, and somite stage rodent embryos. Embryotoxic activity has been found in a high percentage of recurrent aborters but is also present in women with endometriosis and unexplained infertility, suggesting that there may be a range of factors acting at different stages of development. Embryotoxic activity may be predictive of pregnancy outcome and has been shown to be decreased after hormone and immunotherapy. However, its true value can only be assessed when the nature of the toxic factors is understood. Preliminary characterisation studies suggest that toxic activity may due to both immunological (antibodies or cytokines) and nutritional factors.

Does isolated teratozoospermia affect performance in in-vitro fertilization and embryo transfer?

The effect of the strict classification of spermatozoal morphology on the outcome of in-vitro fertilization and embryo transfer treatment cycles has been assessed in a retrospective analysis of 2144 consecutive cycles. The patients all had a standardized long protocol gonadotrophin-releasing hormone analogue cycle, with luteal phase start, to achieve down-regulation. All treatment cycles where the sperm density was abnormal (< 20 x 10(6)/ml), or where progressive motility was abnormal (< 40%), were excluded. The study excluded treatment cycles where the oocytes inseminated did not include at least one grade 1 or grade 2 oocyte. The percentage of couples achieving the normalized results of the clinic, including median fertilization rate per patient, insemination rates, numbers of embryos transferred, rates of 'spare embryo' blastocyst formation, cumulative pregnancy rates and pregnancy outcome, were calculated. No statistically significant difference arose between the two groups of patients with regard to the percentage of patients achieving the normalized median fertilization rate or higher (group 1 with > 14% normal forms, and group 2 with > or = 4%, < or = 14% normal forms). There was a statistically significantly lower chance of achieving this rate in patients of group 3 (< 4% normal forms) (P < 0.005), but 68.6% did achieve that fertilization rate or higher. There was no statistically significant difference in any of the other end points. In conclusion, a morphological classification may be appropriate as an indicator for counselling patients with regard to treatment expectations, but its use would be seem inappropriate as an index of fertilizing potential in clinical management.(ABSTRACT TRUNCATED AT 250 WORDS)

Micromanipulation of human embryos to assist hatching.

OBJECTIVE: To determine the effects of zona slitting and subsequent embryo transfer performed on different days of in vitro human embryonic development to allow appropriate application of assisted hatching techniques to clinical IVF-ET. DESIGN: A microsurgical technique was used to make a standardized slit in the zona pellucida of bipronucleate human embryos on day 2, 3, or 5 after oocyte retrieval. A mock ET was performed after the procedure and the rate of blastocyst formation and the process of hatching in each embryo was monitored. SETTING: All embryos used were donated for research by patients attending the IVF Unit at the John Radcliffe Hospital, Oxford, United Kingdom. MAIN OUTCOME MEASURES: The effects of the mock ET on the integrity of zona-slit embryos was documented. The embryos were cultured further and parameters such as blastocyst formation, initiation, and completion of hatching were monitored in all three groups of embryos. RESULTS: The mock ET did not have a detrimental effect on embryos as assessed by their morphology and subsequent blastocyst formation rate. The technique used in this study to make an opening in the zona increased the rate and number of blastocysts hatching in all three groups compared with nonmanipulated controls. However, the day on which the slit was made did not significantly alter the outcome. CONCLUSION: A gap in the zona pellucida of human embryos significantly increases the rate of hatching. Micromanipulative techniques such as zona slitting can be suitably used for this purpose without loss of blastomeres through the gap in the zona or trapping of the embryo during the process of hatching.

Human blastocyst grading: an indicator of developmental potential?

We have observed marked variation in the in-vitro development of individual human embryos to the blastocyst stage which was poorly associated with the grade of the embryo on the day of transfer (day 2). Therefore we also graded embryos at the blastocyst stage to determine if their development potential can be better predicted at this stage. A total of 41 blastocysts were categorized into three grades depending on their morphology. Various parameters such as hatching, adherence, growth and amount of human chorionic gonadotrophin (HCG) secreted were documented. The number of nuclei in another 34 blastocysts belonging to the three grades were measured. Also morula stage embryos with large vacuoles but no morphological differentiation were included in a separate group called 'vacuolated morulae'. Overall, there was no significant difference between grade 1 and 2 blastocysts as assessed by their development to day 14, cumulative HCG secretion and nuclei counts. Grade 3 blastocysts reflected 'poor' morphology, secreted significantly lower levels of HCG and had significantly fewer nuclei than the other two grades. Vacuolated morulae did not resemble blastocysts in their morphological development, secreted no HCG and had significantly fewer nuclei than grade 1 and 2 blastocysts. This study illustrates the need for selecting blastocysts for transfer to the uterus or for cryopreservation to allow a better assessment of the success of these techniques. In addition, the data will be useful for evaluating the effects of techniques such as co-culture with feeder cells on the development of embryos to the blastocyst stage and beyond.

Sera from women with unexplained infertility inhibit both mouse and human embryo growth in vitro.

OBJECTIVES: To compare the effect of sera from women with reproductive disorders on the in vitro growth of mouse and human embryos up to the blastocyst stage and to determine the influence on human pregnancy outcome. DESIGN: The growth of mouse embryos and in vitro fertilized human embryos up to the blastocyst stage was compared in sera from women with unexplained infertility, and these results were correlated with pregnancy outcome. Also the growth of mouse embryos in sera from women with a history of recurrent abortions was correlated with their pregnancy outcome. PATIENTS, SETTING: Women with unexplained infertility were attending the IVF Unit at the John Radcliffe Hospital, Oxford, United Kingdom. Women with a history of recurrent abortions were attending the high risk pregnancy unit at the same hospital. MAIN OUTCOME MEASURES: Human embryo growth was monitored by daily morphological assessment and mouse embryo growth by both morphological assessment and tritiated thymidine uptake. RESULTS: In 15 women with unexplained infertility, poor mouse embryo development correlated with poor human embryo quality and impaired blastocyst formation when cultured in the same serum, as well as failure to achieve a pregnancy. In 11 women with a history of recurrent abortion, inhibition of mouse embryo growth correlated with unsuccessful pregnancy outcome. DISCUSSION: Assessment of both morphological development and cell proliferation in mouse embryos may be a useful test to determine the suitability of maternal sera for human IVF embryo culture, predicting pregnancy outcome and for categorizing women with reproductive disorders for future clinical management.

A controlled study to assess the use of in vitro fertilization with donor semen after failed therapeutic donor insemination.

OBJECTIVE: To assess the outcome of in vitro fertilization and embryo transfer (IVF-ET) with donor semen in patients who have failed to achieve conception with therapeutic donor insemination (TDI). DESIGN: Retrospective, controlled study. SETTING: All patients were managed and treated at the in vitro fertilization unit, John Radcliffe Hospital, Oxford, United Kingdom. PATIENTS, PARTICIPANTS: Thirty-eight patients undergoing 63 cycles of IVF with donor semen were compared with a control group of patients undergoing the same treatment during the same time period for tubal disease (the subgroup with the best success rates in our program), matched for age, type of infertility (primary or secondary), and number of attempts at treatment. MAIN OUTCOME MEASURES: We compared number of follicles produced, number of oocytes retrieved, fertilization rates, number of embryos obtained, number of embryos transferred, quality of embryos transferred, blastocyst formation in sibling embryos produced, clinical pregnancy rates (PRs), pregnancy loss, and pregnancy outcome. STATISTICS: Chi-squared test statistic with Yates' correction for continuity. RESULTS: There was a statistically significant difference between the number of livebirths in the donor IVF-ET group (37) and the control group (18). The cumulative PR after four cycles for the donor IVF-ET group (83%) was statistically greater than that of the control group (59%). There was no significant difference in all other parameters compared. CONCLUSION: Patients undergoing IVF-ET with donor semen have an excellent outcome. With the decline in PR after six cycles of TDI, early recourse to IVF-ET should be considered in this group of patients.

Human trophectoderm biopsy and secretion of chorionic gonadotrophin.

We have previously developed a technique of trophectoderm biopsy to obtain cells from human blastocysts for preimplantation genetic diagnosis. To determine whether this technique affects the subsequent development of the blastocyst, 45 manipulated blastocysts were observed from days 3 to 14 in culture, the amount of human chorionic gonadotrophin (HCG) secreted by each embryo was measured and these results were compared with those of 26 non-manipulated controls documented in a previous study. A slit was made in the zona pellucida opposite the inner cell mass in 18 of the 45 blastocysts. This increased the rate of hatching but the other morphological changes up to day 14 were similar to those seen in the non-manipulated controls. There was no difference in the mean cumulative HCG secretion by these zona-slit controls (149.8 +/- 45.7 mIU/ml) compared to the non-manipulated controls (146.2 +/- 23.7 mIU/ml). A slit was also made in the zona pellicida of the other 27 blastocysts. Approximately 12-18 h later, in 18 blastocysts a biopsy of the herniating trophectoderm cells (5-30) was performed. The rate of hatching and adherence to the culture dish was similar to the non-manipulated controls. The mean cumulative HCG secretion decreased significantly (57.5 +/- 16.2 mIU/ml, P less than 0.01) after the biopsy procedure. However, if a small biopsy was performed (less than 10 cells removed) the decrease in HCG secretion (87.6 +/- 24.8 mIU/ml) was not significant, whereas when a large biopsy was performed (greater than 10 cells), HCG levels fell to 19.9 +/- 9.1 mIU/ml.(ABSTRACT TRUNCATED AT 250 WORDS)

An approach to preimplantation diagnosis of beta-thalassaemia.

Using the polymerase chain reaction (PCR), it was possible to amplify a single copy fragment of the beta-globin gene from 2-32 human embryonic cells obtained from arrested preimplantation embryos. For the detection of beta-thalassaemia mutations, allele specific priming of the PCR using nested primers was employed using approximately 10 pg of DNA from individuals known to carry these mutations. This approach was successful in detecting the presence or absence of five Asian Indian beta-thalassaemia mutations that were selected for this study. In spite of meticulous precautions against contamination, false-positive amplification was observed, a problem that will have to be overcome before this approach can be used in clinical practice.

The human blastocyst: morphology and human chorionic gonadotrophin secretion in vitro.

Micromanipulation of human oocytes and embryos has provided new opportunities for both the treatment of infertility and the preimplantation diagnosis of genetic disease. It is important to determine whether manipulated embryos develop normally in vitro, as an indication of their suitability for transfer. However, at present there is little information on the development of non-manipulated embryos in vitro for comparison. We have therefore monitored morphological changes and human chorionic gonadotrophin (HCG) secretion in 36 non-manipulated human embryos, including 26 blastocysts and 10 cavitating morulae, daily from day 3 to day 14 of culture. Hatching was observed in 10 (38.5%) blastocysts and five of these adhered to the culture dish and appeared viable until day 14. The secretion of HCG was first detected on day 8, peaked at day 10 (51.11 +/- 8.7 mIU/ml) and then declined but was still detectable in four blastocysts on day 14. There was no overall difference in HCG secretion by hatched blastocysts and those which remained within the zona. However, those hatched blastocysts which showed adherence had significantly increased (P less than 0.05) HCG secretion. For individual blastocysts, the pattern of HCG secretion correlated well with the assessment of morphology. These data provide the basis for comparative studies of morphological changes and HCG secretion in manipulated embryos.