Reproductive BioMedicine Online - Articles in Press

Management of deep endometriosis - Accepted Manuscript

J. Nassif, P. Trompoukis, S. Barata, A. Furtado, B. gabriel, A. wattiez — 2010-09-09

Abstract: Deep endometriosis is still a challenging disease in terms of diagnosis and treatment. About 10–12% of women of reproductive age will have a form of endometriosis. This can affect pelvic as well as extra pelvic locations. Risk of malignant transformation has been studied over a long period of time. Medical and surgical treatments can be proposed to patients for endometriosis-associated pain depending on the severity of symptoms and location of the disease. Results and outcomes are different according to different publications. Understanding of the benefit of surgical treatment on fertility is increasing. The place of medical and surgical treatment in recurrent symptoms or disease is also of interest. Here is presented a review on the management of endometriosis in the light of recent data. Further investigations in many fields of endometriosis are still required.

Is another meta-analysis on the effects of intramural fibroids on reproductive outcomes needed? - Accepted Manuscript

Mostafa Metwally, Cynthia M. Farquhar, Tin Chiu Li — 2010-09-07

Abstract: Intramural fibroids when encountered in women undergoing fertility treatment present a clinical dilemma. Despite recent studies that have suggested a negative outcome for intramural fibroids on fertility outcomes, the evidence remains far from conclusive. The current study presents a systematic review and meta-analysis of the currently available evidence. Relevant articles were identified in MEDLINE and EMBASE. Ten studies reported the effects of intramural fibroids on assisted conception treatment including one study reporting the effect of myomectomy for these fibroids. Combined analysis of the included studies, after taking into account possible confounding factors and including only studies with low risk of bias, showed no evidence of a significant effect for intramural fibroids on clinical pregnancy rate (OR 0.74, 95% CI 0.50–1.09), live birth rate (OR 1.17, 95% CI 0.62–2.22) or miscarriage rate (OR 1.61, 95% CI 0.61–4.20). There was also no evidence for a significant effect for myomectomy on the clinical pregnancy rate (OR 1.88, 95% CI 0.57–6.14) or the miscarriage rate (OR 0.89, 95% CI 0.14–5.48). These findings highlight the current deficiency in the literature and suggest that evidence is insufficient to draw any conclusions regarding the effect of intramural fibroids on reproductive outcomes.

Fate of fertilized human oocytes - Accepted Manuscript

Giuseppe Benagiano, Manuela Farris, Gedis Grudzinskas — 2010-09-07

Abstract: Establishing the proportion of fertilized oocytes and early human embryos that proceed to term may help policy makers in their evaluation of when the life of a new human individual begins and in determining the nature of protection to be accorded to it. The rate of spontaneous abortions, although increasing with age, overall does not exceed 15%. However, abortion rates refer only to ‘clinical pregnancy’, whereas early embryonic loss is more common than generally believed. Evidence of such wastage comes from many sources. Human fecundity rarely exceeds 35% and may be decreasing due to deterioration in semen quality. Embryological studies show that 50% of randomly recovered preimplantation embryos have severe anomalies. The study of sensitive markers of pregnancy, such as human chorionic gonadotrophin, indicates early embryo wastage in the order of 50%. Pregnancy wastage may be a function of the time lapse between ovulation and implantation as the implantation window extends between menstrual cycle days 20 and 24. Finally, data obtained with natural IVF cycles also indicate major losses, with an overall pregnancy rate of 7.2% per cycle and 15.8% per transfer. These data, however, are biased by a high cancellation rate and low oocyte retrieval in natural IVF cycles.

Egg sharing for assisted conception: a window on oocyte quality - Accepted Manuscript

Malcolm Faddy, Roger Gosden, Kamal Ahuja, Kay Elder — 2010-09-06

Abstract: The steep decline in both natural fertility and success after assisted reproduction treatment with increasing maternal age is universally recognized. Large variations in the developmental competence of oocytes collected are seen during assisted cycles and a link between the biological competence of oocytes retrieved and age has been confirmed. Patients who require donated oocytes can benefit from egg sharing programmes, in which a proportion of oocytes collected from selected patients aged ⩽35 years undergoing conventional assisted reproduction treatment are shared with a matched recipient. The reproductive outcomes of the egg provider and recipient can thus be compared with quantify the significance of oocyte quality. Data gathered from two comparable treatment centres resulted in 285 pairs of egg sharing providers and their recipients that could be analysed statistically. The chief finding was donor pregnancy as a predictor of recipient pregnancy given embryo transfer (odds ratio 2.15, 95% confidence interval 1.33–3.46, P ≈ 0.002), despite an appreciably higher mean age of the recipients. The probability of a recipient pregnancy increased by almost 0.2. Such results strongly indicate the key importance of oocyte quality for a successful clinical outcome in egg sharing practices and assisted reproduction treatment more generally.

Risk charts to identify low and excessive responders among first-cycle IVF/ICSI standard patients - Accepted Manuscript

2010-09-06

Abstract: Ovarian stimulation carries a risk of either low or excessive ovarian response. The aim was to develop prognostic models for identification of standard (ovulatory and normal basal FSH) patients’ risks of low and excessive response to conventional stimulation for IVF/intracytoplasmic sperm injection. Prospectively collected data on 276 first-cycle patients treated with 150 IU/day recombinant FSH (rFSH) in a long agonist protocol were analysed. Logistic regression analysis was applied to the outcome variables: low (seven or less follicles) and excessive (20 or more follicles) response. Variables were woman’s age, menstrual cycle length, weight or body mass index, ovarian volume, antral follicle count (AFC) and basal FSH. The predictive performance of the models were evaluated from the prediction error (Brier score,%) where zero corresponds to a perfect prediction. Model stability was assessed using 1000 bootstrap cross-validation steps. The best prognostic model to predict low response included AFC and age (Brier score 7.94) and the best model to predict excessive response included AFC and cycle length (Brier score 15.82). Charts were developed to identify risks of low and excessive ovarian response. They can be used for evidence-based risk assessment before ovarian stimulation and may assist clinicians in individual dosage of their patients.

Individual demands of human embryos on IVF culture medium: influence on blastocyst development and pregnancy outcome - Accepted Manuscript

B. Wirleitner, P. Vanderzwalmen, A. Stecher, M.H. Zech, M. Zintz, N.H. Zech — 2010-09-02

Abstract: The elucidation of the metabolic requirements of human embryos in vivo or in vitro remains, despite being intensively investigated, a work in progress. The adoption of extended embryo culture to the blastocyst stage during the last decade has entailed new challenges. With the increased attention to culture media formulations, more evidence on the sensitivity of embryos to their early environmental conditions is accumulating which might affect phenotype and developmental potential. A retrospective study was conducted that comprised 286 IVF cycles to evaluate the effect of two different culture media on blastocyst development and pregnancy outcome. Embryos were either cultured in a one step or a sequential medium. Higher fertilization rates and augmented blastocysts as well as higher implantation rates were observed when embryos were cultured in one step medium (P < 0.05). Interestingly, the transfer of two embryos where one embryo was cultured in either medium resulted in a significantly higher rate of twin pregnancies. Although multiple pregnancies should be avoided in assisted reproduction treatment to reduce risks for offspring and mother, this higher frequency of twin pregnancies resulting from the transfer of embryos derived from different culture media suggests that each embryo makes individual demands on its early environment.

Response: Efficacy of ultraviolet sterilization of liquid nitrogen - Accepted Manuscript

Vladimir Isachenko — 2010-09-02

Female gender pre-selection by maternal diet in combination with timing of sexual intercourse – a prospective study - Accepted Manuscript

A.M. Noorlander, J.P.M. Geraedts, J.B.M. Melissen — 2010-09-01

Abstract: Natural sex selection methods have been applied for several decades, but their use and effectiveness are still a matter of debate. Therefore, this study assessed the efficacy of a maternal diet low in sodium and high in calcium, in combination with timing of intercourse well before ovulation as a method to improve the chances of conceiving a girl. A total of 172 couples wanting a girl participated in the study. For the 150 couples that actually started, compliance with diet was assessed through mineral analyses of blood and timing of intercourse relative to ovulation was determined by ovulation tests. Based on mineral blood values and timing of intercourse of 28 participants, a prediction rule for conceiving a girl was constructed and was tested prospectively for validity on a subsequent group of 50 women. In this group, 21 women satisfied the criteria of the prediction rule and 16 gave birth to a daughter. It is concluded that the combination of maternal diet with timing of intercourse is capable of increasing the probability of conceiving a girl (P = 0.005). The observed percentage of female babies for all 32 women satisfying the prediction rule was 81% (95% confidence interval 68–95%).

Mutation analysis of three genes in patients with maturation arrest of spermatogenesis and couples with recurrent miscarriages - Accepted Manuscript

K. Stouffs, D. Vandermaelen, H. Tournaye, I. Liebaers, W. Lissens — 2010-09-01

Abstract: The primary aim of this study was to gain more insight into maturation arrest of spermatogenesis (MA) and its relationship with mutations in genes essential for meiosis. The study also investigated the possibility that mutations in human meiosis genes cause a milder phenotype and that, in such cases, meiosis could potentially be completed with the production of mature germ cells having an abnormal chromosomal constitution causing miscarriage. Among 40 patients with MA, five changes were observed that also predicted alterations at the amino acid level. However, since these changes were also present in men with normozoospermia in equal frequencies, it was assumed these changes to be single nucleotide polymorphisms. Among 46 patients with recurrent miscarriages, two additional changes were detected predicting an alteration at the amino acid level. One change was detected in controls. However, the second heterozygous change, detected in a conserved functional domain of the SYCP3 gene, was absent in >200 controls. These preliminary results stress the need to further investigate the relationship between abnormalities in meiosis genes and the formation of gametes with abnormal chromosomal constitution. More research is also necessary to determine the impact and frequency of such changes before implementing mutation screening in genetic counselling.

Vascular endothelial growth factor gene polymorphisms in North Indian patients with recurrent miscarriages - Accepted Manuscript

2010-09-01

Abstract: The association of four common polymorphisms of vascular endothelial growth factors (VEGF) with recurrent miscarriages (RM) was evaluated in North Indian women for 200 patients with RM and 200 controls. The subjects were genotyped for the polymorphisms –2578C/A, –2549 18-bp I/D, –1154G/A and +936C/T. Association of VEGF genotypes, alleles and haplotypes with recurrent miscarriage were evaluated by Fisher’s exact test. –1154G/A and +936C/T modified the risk of RM. –1154A and +936T significantly increased the risk of RM (OR = 1.485, P = 0.0210, 95% CI 1.072–2.057 and OR = 1.869, P = 0.0054, 95% CI 1.214–2.876, respectively) and predisposed to RM (OR = 2.0, P = 0.0310, 95% CI 1.068–3.747 and OR = 1.716, P = 0.0293, 95% CI 1.058–2.784, respectively). However, no association was found between –2578C/A or –2549 18-bp I/D and RM. Four haplotypes, AIAC, ADAC, CIAT and ADGT, were found to predispose to RM while the haplotypes CIAC, CDGT and ADGC were found to show protective effect. In conclusion, two common polymorphisms of the VEGF gene, –1154G/A and +936C/T, increase the risk of RM in North Indian women. RM is also predisposed in the presence of haplotypes AIAC, ADAC, CIAT and ADGT.

Efficacy of ultraviolet sterilization of liquid nitrogen - Accepted Manuscript

Lodovico Parmegiani, Graciela Estela Cognigni, Marco Filicori — 2010-09-01

A universal carrier test for the long tail of Mendelian disease - Corrected Proof

2010-08-23

Abstract: Mendelian disorders are individually rare but collectively common, forming a ‘long tail’ of genetic disease. A single highly accurate assay for this long tail would allow the scaling up of the Jewish community’s successful campaign of population screening for Tay–Sachs disease to the general population, thereby improving millions of lives, greatly benefiting minority health and saving billions of dollars. This need has been addressed by designing a universal carrier test: a non-invasive, saliva-based assay for more than 100 Mendelian diseases across all major population groups. The test has been exhaustively validated with a median of 147 positive and 525 negative samples per variant, demonstrating a multiplex assay whose performance compares favourably with the previous standard of care, namely blood-based single-gene carrier tests. Because the test represents a dramatic reduction in the cost and complexity of large-scale population screening, an end to many preventable genetic diseases is now in sight. Moreover, given that the assay is inexpensive and requires only a saliva sample, it is now increasingly feasible to make carrier testing a routine part of preconception care.

Birth of a journal: a personal memoire - Accepted Manuscript

Fiona Bennett — 2010-08-23

Rare chromosomal complement of trisomy 21 in a boy conceived only by IVF - Accepted Manuscript

2010-08-17

Rare chromosomal complement of trisomy 21 in a boy conceived only by IVF - Accepted Manuscript

Carmen Orellana — 2010-08-17

Possible role of NK and NKT-like cells in implantation failure after IVF - Accepted Manuscript

2010-08-09

Abstract: During implantation, maternal immunoactivation and tolerance are not only limited to the decidua but are also observed in the periphery, predominantly affecting the innate immune system. Since unexplained female infertility, as well as recurrent spontaneous abortion and implantation failure, are thought to be associated with pathological maternal immunotolerance mechanisms, this study focused on immune profile analysis of IVF candidates. Previous studies on peripheral natural killer (NK) cell characteristics of IVF patients have been limited to the comparison of blood samples taken prior to the IVF procedure. This study performed a follow-up study and compared patient’s data obtained on the day of oocyte collection with the data 1 week after embryo transfer. The aim was to investigate phenotypic (subpopulations, CD69, T-cell immunoglobulin mucin 3 and NK-activating receptor expression) and functional (perforin and CD107a expression) changes in the peripheral NK and NK T (NKT)-like cell populations. During this short period of time around the IVF procedure, women with failed IVF reflected unfavourable Th1-oriented changes of NK and NKT-like cells. In comparison the follow-up data for women with successful conception remained principally constant. The observed peripheral changes during early pregnancy in the same individual may also have importance in successful embryo implantation.

Effects of vitrification solutions and equilibration times on the morphology of cynomolgus ovarian tissues - Corrected Proof

2010-08-02

Abstract: This study assessed the effects of vitrification solutions and equilibration times on morphology of cynomolgus ovarian tissues. Ovarian cortical sections (0.1–0.2cm thickness) of seven cynomolgus monkeys were randomly allocated to either a control group or one of six vitrification groups. Ovarian tissue sections were vitrified ultra-rapidly by placing them directly into liquid nitrogen using two different vitrification solutions (VSEGP: 5.64mol/l ethylene glycol+5% (w/v) polyvinylpyrrolidone+0.5mol/l sucrose; and VSED: 3.22mol/l ethylene glycol+2.56mol/l dimethylsulphoxide+0.5mol/l sucrose) after three different exposure times (5–20min). After warming, follicle morphology was analysed using light and transmission electron microscopy. The proportion of morphologically normal follicles vitrified using VSED after a 5-min exposure was lower (P<0.05) than those vitrified by other conditions. The proportion of normally structured mitochondria in oocytes of preantral follicles vitrified after a 5-min exposure to VSED (56%) was lower (P<0.01) than those vitrified by other conditions (78–88%). Following tissue vitrification with VSED, the surface ratio of lysosome was increased compared with non-vitrified oocytes (1.64% versus 1.11%; P<0.05). These results indicate that VSEGP can support the morphology of vitrified preantral follicles and oocytes.Cryopreservation of preantral follicles in ovarian tissues has been expected to be an effective measure for preserving fertility in young women who need to undergo cytotoxic therapy. However, a cryopreservation protocol has not yet been well established. This paper revealed that only 5-min exposure time to a cryoprotectant containing a combination of ethylene glycol and polyvinylpyrrolidone yields better follicle and oocyte morphology at an ultra-structural level in mammalian ovarian tissues after ultra-rapid vitrification using a new ultra-rapid vitrification device compared with a cryoprotectant containing a combination of ethylene glycol and dimethylsulphoxide. Survival rates of cancer patients are increasing. However, patients requiring chemotherapy and/or radiotherapy for cancer, leukaemia or other benign pathologies are likely to experience premature ovarian failure and loss of fertility as a consequence of these potentially gonadotoxic treatments. Use of the procedure reported in this study may increase the chance of fertility preservation.

Cardiovascular and metabolic characteristics of infertile Chinese women with PCOS diagnosed according to the Rotterdam consensus criteria - Corrected Proof

2010-08-02

Abstract: Polycystic ovary syndrome (PCOS) is strongly associated with metabolic abnormalities in Western women. However, data from other populations and geographical regions are scarce. This study evaluated cardiovascular and metabolic risk factors in Chinese infertile women diagnosed with PCOS using the 2003 Rotterdam consensus criteria. A total of 615 women representing the four PCOS phenotypes (oligo- or anovulation (AO)+hyperandrogenism (HA)+polycystic ovaries (PCO), AO+HA, AO+PCO and HA+PCO) underwent standardized metabolic screening including a 75g oral glucose tolerance test. All groups presented with similar reproductive characteristics, with the only difference being a significantly higher Ferriman–Gallwey score for hirsutism (P=0.01) in the subgroup characterized by HA+PCO. Overall, the prevalence of metabolic syndrome was 6.4%, with no difference among the four groups (range of 2.3–12.2%). Metabolic syndrome was associated with body mass index (P<0.001), waist/hip ratio (P=0.002), index of insulin resistance (P=0.005) and fasting insulin (P=0.009) in multivariate analysis. Compared with Caucasians and Chinese women in Westernized societies, mainland Chinese women with PCOS have a low risk of metabolic syndrome and its presence does not vary across the specific PCOS phenotypes.Polycystic ovary syndrome (PCOS) is the most common endocrine disorder in women of reproductive age. PCOS is diagnosed if at least two of three of the following characteristics are present: lack of ovulation (oligo- or anovulation), specific ultrasound characteristics of the ovaries (enlarged ovary and/or increased number of antral follicles), and increased production of androgen hormones and/or its clinical manifestations such as hirsutism and acne. Therefore, four presentations of PCOS can be distinguished. PCOS is also associated with the metabolic syndrome, a clustering of from three to five risk factors of cardiovascular disease. Unfortunately, knowledge about PCOS and its metabolic associations for the Asian region is scarce. The objective of this study was to evaluate cardiovascular and metabolic risk factors in a group of 615 Chinese infertile women with PCOS representing the four PCOS subgroups, from mainland China. Overall, women in the four PCOS subgroups had similar physical, hormonal and biochemical characteristics, with only more hirsutism in the subgroup characterized by overproduction of androgen hormones and ovarian ultrasound characteristics. Of the total group, 6.4% had metabolic syndrome with no differences among subgroups (range 2.3–12.2%). We found that the presence of metabolic syndrome was associated with higher body mass index, higher waist/hip ratio, and a higher index of insulin resistance (HOMA-IR) and higher fasting insulin concentrations. We conclude that in contrast to Caucasians and Chinese women in Westernized societies, mainland Chinese women with PCOS have a low risk of metabolic syndrome and that its presence does not vary among the four PCOS phenotypes.

Pregnancy and live birth following the transfer of vitrified–warmed blastocysts derived from zona- and corona-cell-free oocytes - Corrected Proof

Yimin Shu, Wenlin Peng, John Zhang — 2010-08-02

Abstract: This study reports two clinical pregnancies and one live birth following the transfer of vitrified blastocysts developed from oocytes with neither zona pellucida nor corona cells. Two zona-free oocytes obtained from two patients of advanced maternal age undergoing minimal stimulation were normally fertilized after intracytoplasmic sperm injection. In case 1, all four blastomeres of the zona-free embryo were loosely associated and inserted back into ruptured zona on day 2. Zona-free embryo from case 2 had tight contacts between blastomeres and was cultured without zona. Both embryos derived from zona-free oocytes progressed to blastocyst stage and were cryopreserved by vitrification. When patients came back for a cryopreserved embryo transfer, both vitrified blastocysts survived warming. In case 1, transfer of a warmed blastocyst with reconstructed zona resulted in a clinical pregnancy that ended in a spontaneous abortion at 22weeks. In case 2, live birth was achieved with a normal healthy baby (male) weighing 2381g at 40weeks’ gestation. This report emphasizes the importance of maintenance of normal cell arrangement on the subsequent embryonic development for a zona-free oocyte. Zona-free oocytes may provide a valuable source of embryos for infertility patients, especially for those with a limited number of oocytes.In this study, we report two clinical pregnancies and one live birth following the transfer of vitrified blastocysts developed from oocytes with neither zona pellucida nor corona cells. Two zona-free oocytes obtained from two patients of advanced maternal age undergoing minimal stimulation were normally fertilized after intracytoplasmic sperm injection. In case 1, all four blastomeres of the zona-free embryo were loosely associated and inserted back into ruptured zona on day 2. Zona-free embryo from case 2 had tight contacts between blastomeres and was cultured without zona. Both embryos derived from zona-free oocytes progressed to blastocyst stage and were cryopreserved by vitrification. When patients came back for a cryopreserved embryo transfer, both vitrified blastocysts survived warming. In case 1, transfer of a warmed blastocyst with reconstructed zona resulted in a clinical pregnancy that ended in a spontaneous abortion at 22 weeks. In case 2, live birth was achieved with a normal healthy baby (male) weighing 2381 g at 40 weeks’ gestation. Our report emphasizes the importance of maintenance of normal cell arrangement on the subsequent embryonic development for a zona-free oocyte. Zona-free oocytes may provide a valuable source of embryos for infertility patients, especially for those with a limited number of oocytes.

Are US results for assisted reproduction better than the rest? Is it a question of competence or policies? - Accepted Manuscript

Hossam I. Abdalla — 2010-08-02

Abstract: A comparison of nationally published 2006 data from the USA, UK and Australia and New Zealand (ANZ) was performed. Although live births/cycle was higher in USA, live birth/embryo transferred was significantly higher in ANZ (18.2%) compared with both USA and UK (13.8%) (P < 0.001). The multiple rates were significantly lower in ANZ (12.0%) compared with USA (30.7%) and UK (25.2%) (P < 0.001). The incidence of oocyte donation was significantly higher in the USA (11.1%) than in ANZ (2.8%) and UK (3.9%) (P < 0.001). There was significantly higher cycle cancellation in USA (11.5%) compared with the UK (6.8%) and ANZ (9.5%) (P < 0.001). The incidence of frozen embryo transfer cycles was significantly higher in ANZ (59%) compared with both UK (24%) and USA (22%) (P < 0.001). The total live birth rate from fresh and frozen cycles for the same year was significantly higher in ANZ at 32.0% compared with the UK at 28.8% (P < 0.001) with half the multiple rate. It is argued that the USA’s higher success rates are explained by policy (transferring higher number of embryos) and selection issues (cancelling or avoiding poor responders) rather than being a matter of clinical competence.

Can Doppler ultrasound-guided oocyte retrieval improve IVF safety? - Corrected Proof

Francisco Rísquez, Edmond Confino — 2010-08-02

Abstract: Transvaginal ultrasound-guided oocyte retrieval has gained universal acceptance with an excellent safety record overall. However, even with contemporary ultrasound resolution, the aspiration needle can injure adjacent pelvic organs and blood vessels and result in external and internal bleeding. Although the idea that Doppler ultrasound might reduce the risk of blood vessel injury during follicular aspiration seems to be plausable, measurement of peritoneal blood loss and the validity of this opinion has never been appropriately tested. Using a proposed classification method in an IVF programme, it was estimated that a significant peritoneal bleeding occurred in 56/898 (6%) of IVF patients. Although Doppler ultrasound was routinely used in all patients, it did not predict 24/53 (45%) of the patients with moderate peritoneal bleeding. In 8/53 cases (15%) with moderate peritoneal bleeding, vaginal bleeding was also detected and correctly predicted during oocyte aspiration using colour Doppler vaginal vessel imaging. Colour Doppler ultrasound guidance is an easily accessible technology with a theoretical promise to improve IVF safety and, with proper usage, has the potential to reduce haemorrhagic complications.

Chromosome abnormality rates in human embryos obtained from in-vitro maturation and IVF treatment cycles - Corrected Proof

2010-08-02

Abstract: The aim of this retrospective study was to compare the incidence of chromosomal abnormality in embryos from in-vitro maturation (IVM) and IVF cycles. The copy numbers of chromosomes 13, 15, 16, 18, 21, 22, X and Y were assessed with fluorescence in-situ hybridization (FISH) in single blastomeres biopsied from cleavage stage embryos. Spare embryos that were not transferred or cryopreserved were also analysed in full. IVM and IVF groups comprised six and 30 couples, with mean±SD embryos with FISH result of 8.0±4.4 and 11.7±3.8, respectively. The incidence of chromosomal abnormality per FISH result was similar in IVM and IVF embryos (58.7% versus 57.4%, respectively). When embryos were categorized based on maturation time of oocytes in IVM cycles, embryos derived from oocytes that matured 48h after collection had a higher chromosomal abnormality rate compared with embryos derived from in-vivo matured oocytes and to embryos derived from oocytes that matured in the first 24h after collection.

Fertility preservation for breast-cancer patients using IVM followed by oocyte or embryo vitrification - Corrected Proof

2010-08-02

Abstract: Unstimulated in-vitro maturation (IVM) cycles are considered for fertility preservation in breast cancer due to avoidance of ovarian stimulation and shortened time to oocyte retrieval. This study evaluated the efficacy of this approach in a retrospective cohort analysis of 66 patients with breast cancer. Immature oocytes were collected and matured in vitro and then either vitrified or fertilized and preserved as vitrified embryos. In group 1 (vitrified oocytes, n=35), the average number of oocytes retrieved was 11.4±8.8, the maturation rate was 64.2% and an average of 7.9±6.6 oocytes were vitrified per patient treated. The median duration from the first evaluation to oocyte retrieval was 8days. In group 2 (vitrified embryos, n=31) the average number of oocytes retrieved was 9.7±6.4, the maturation rate was 53.2% and an average of 5.8±2.7 mature oocytes were available for fertilization/patient. The fertilization rate was 77.8%, resulting in 4.5±2.7 vitrified embryos/patient. The median duration from the first evaluation to oocyte retrieval was 13days. Calculated pregnancy rates per vitrified oocyte and embryo were 3.8% and 8.1%, respectively. IVM can be considered a useful option for fertility preservation in breast-cancer patients.Breast cancer represents about 30% of malignant tumours occurring in women of childbearing age. Approximately 10–15% of breast cancers are diagnosed in women of reproductive age. Over the past two decades, earlier diagnosis and highly effective systemic therapies have led to reductions in mortality. However, women undergoing treatment for breast cancer may suffer from fertility problems in the future. A number of methods are currently used to preserve fertility in young breast cancer patients including ovarian freezing, IVF and preservation of oocytes and embryos and the use of drugs to protect the ovary during chemotherapy. A newly described method, in-vitro maturation, was used in our study. Freezing the eggs of breast cancer patients using this technique was found to be a useful treatment option due to two advantages: (i) no hormones were used and (ii) short duration of treatment. Our study evaluates the efficacy of this treatment approach for patients suffering from breast cancer.

Follicular-fluid anti-Müllerian hormone concentration is similar in patients with endometriosis compared with non-endometriotic patients - Corrected Proof

2010-08-02

Abstract: This cross-sectional prospective study assessed follicular-fluid anti-Müllerian hormone (AMH) concentrations in infertile patients with mild/minimal endometriosis during natural IVF. Thirty-two women participated in the study. Patients were divided into two groups: tubal obstruction without endometriosis (control group) and minimal/mild endometriosis (study group). All patients underwent laparoscopy for assessment of infertility; at the same time, any foci of endometriosis found were cauterized. AMH concentration was measured in the follicular fluid of a single follicle when it achieved pre-ovulatory maturation by ultrasensitive enzyme-linked immunosorbent assay. Likewise, AMH, FSH and inhibin B content in serum was also measured. Age (30±1.3 and 32±0.8years) and body mass index (22±0.6 and 22±0.5kg/m2), day-3 antral follicle count (11.3±1.7 and 10.7±1.5), serum FSH concentrations (5.4±0.6 and 5.0±0.3IU/ml) and follicular-fluid AMH concentrations (1.8±0.3 and 1.5±0.1ng/ml, study and control group, respectively; mean difference 0.33, 95% CI −0.21 to 0.88) were similar in both groups. This study shows that infertile patients with minimal/mild endometriosis have a similar concentration of AMH in the follicular fluid after natural IVF as compared with control subjects.The purpose of this prospective study was to assess the follicular-fluid environment measuring anti-Müllerian hormone in infertile patients with mild/minimal endometriosis, which is a disease in which endometrial uterine tissue invades sites outside the uterus, during natural IVF. Measurement of anti-Müllerian hormone was chosen as it controls the formation of primary follicles and, therefore, has a role in folliculogenesis. Thirty-two women submitting to natural IVF cycles participated from the study. Patients were divided into two groups: tubal obstruction without endometriosis (control group), and minimal/mild endometriosis (study group). All patients underwent laparoscopy for assessment of infertility; at the same time, any foci of endometriosis found were cauterized. Anti-Müllerian hormone concentration was measured in the follicular fluid of a single follicle when it achieved pre-ovulatory maturation. Likewise, anti-Müllerian hormone content in serum was also measured, as well as FSH and inhibin B. Average age, approximately 30 years, was not statistically different between groups. In addition, day-3 antral follicular count (resting follicles at the beginning of each menstrual cycle), and serum FSH concentrations were also not statistically different between groups. Moreover, follicular-fluid AMH concentrations were also similar in both groups. This study shows that infertile patients with minimal/mild endometriosis have a similar concentration of AMH in the follicular fluid as compared with control subjects.

Analysis of oocyte cryopreservation in assisted reproduction: the Italian National Register data from 2005 to 2007 - Corrected Proof

G. Scaravelli, V. Vigiliano, J.M. Mayorga, S. Bolli, R. De Luca, P. D’Aloja — 2010-08-02

Abstract: This paper reports on oocyte cryopreservation efficacy in Italy with respect to successful IVF from 2005 to 2007, presenting data from 193 centres collected by the Italian National Register. Post-thawing survival rates, number of transferred embryos, implantation rates and clinical pregnancy rates per transfer with respect to frozen/vitrified oocytes (FVO) were analysed. These numbers were compared with those obtained using frozen embryos or fresh oocytes. A total of 121,708 cycles were initiated, of which, 7.1% (8682) were FVO cycles and 2.4% (2952) were frozen embryo cycles. Of the 81,786 FVO, 52.5% (42,917) were thawed and 26.9% (22,005) inseminated. Of those inseminated, 68.0% (14,966) yielded good embryos. These numbers were significantly lower than those using fresh oocytes in which 77.9% (197,242; fresh oocytes versus FVO P<0.001) of inseminated oocytes generated good embryos. Implantation rate using FVO was 6.9%, which was significantly lower than that using fresh oocytes (13.5%; P<0.001) and frozen embryos (8.8%; P<0.001). Pregnancy rate per transfer using FVO was 12.5% and significantly lower than that using fresh oocytes (24.9%; P<0.001) or frozen embryos (16.4%; P<0.001). There were 505 deliveries after IVF with FVO and 582 babies.

Normal serum concentrations of anti-Müllerian hormone in women with regular menstrual cycles - Corrected Proof

2010-08-02

Abstract: Anti-Müllerian hormone (AMH) has become the ‘molecule of the moment’ in the field of reproductive endocrinology. Indeed, it is valuable as a means of increasing understanding of ovarian pathophysiology and for guiding clinical management across a broad range of conditions. However, no normative values have been established for circulating AMH in healthy women. In this cross-sectional study, 277 healthy females (aged 18–50 years) were included. AMH was measured by commercial enzyme-linked immunosorbent assay. Serum AMH concentrations show a progressive decline with female ageing. The age-related changes in AMH were best fitted by a polynomial function. Mean AMH concentrations were not modified by past use of oral contraceptive and were independent of parity of women. Age-specific normative values for circulating AMH concentration were established. AMH concentrations seem to be independent of the reproductive history of the patient.Anti-Müllerian hormone (AMH) has become the ‘molecule of the moment’ in the field of reproductive endocrinology. Indeed, it is valuable as a means for increasing our understanding of ovarian pathophysiology, and for guiding clinical management across a broad range of conditions. However, no normative values have been established for circulating AMH in healthy women. In this cross-sectional study, 277 healthy females (aged 18–50) were included. AMH was measured by commercial enzyme-linked immunosorbent assay. Serum AMH concentrations show a progressive decline with female ageing. The age-related changes in AMH were best fitted by a polynomial function. Mean AMH concentrations were not modified by past use of oral contraceptive and were independent of parity of women. We established age-specific normative values for circulating AMH. AMH concentrations seem to be independent of the reproductive history of the patient.

Blastocysts from patients with polycystic ovaries exhibit altered transcriptome and secretome - Corrected Proof

2010-08-02

Abstract: Polycystic ovaries (PCO) is a common phenotype of women presenting for infertility treatment. This study investigated whether blastocysts derived from women with PCO have an altered molecular signature which could be a causative factor contributing to reproductive failure. Morphologically similar blastocysts derived from women with PCO and donor oocyte cycles were analysed for transcription and protein secretion. Unsupervised hierarchical clustering demonstrated that the transcriptome profiles of blastocysts derived from PCO patients and control blastocysts were markedly different with complete branch separation. Statistical analysis revealed 829 genes with significantly different expression: 784 decreased (94.6%) and 45 increased (5.4%) in blastocysts derived from women with PCO compared with controls (P<0.05). Functional annotation of these genes revealed predominant gene ontology biological processes including protein metabolism (30%), transcription (22%), signal transduction (15%), biosynthesis (15%) and cell cycle (14%). Proteomic profiling identified 12 biomarkers that displayed significant decrease in expression in blastocysts derived from women with PCO compared with controls (P<0.05). These data indicate molecular alterations in human blastocysts derived from PCO patients, potentially demonstrating for the first time a link between patient aetiology/phenotype and subsequent embryo development, which in part may explain the observed reduction in reproductive capacity.Lay summaryPolycystic ovaries (PCO) are common among women of reproductive age and, in particular, among women presenting for infertility treatment. This study examined embryos derived from women with PCO in an effort to better understand the underlying mechanisms involved with this phenotype. Embryonic gene expression and protein secretion were investigated and directly compared between embryos derived from women with PCO and embryos derived from donor oocyte IVF cycles (fertile controls). Results indicated significant differences between the two groups for both gene expression (n=829 genes) and protein secretion (n=12 proteins). Further investigation into the function of these differentially expressed genes revealed important affected biological processes including protein metabolism (30%), transcription (22%), signal transduction (15%), biosynthesis (15 %) and the cell cycle (14%). These data reveal molecular modifications in embryos derived from women with PCO who are undergoing infertility treatment. Therefore, potentially demonstrating a link between patient phenotype and subsequent embryo development may, in part, elucidate the underlying mechanisms associated with observed reduction in reproductive capacity.

Implication of apoptosis in sperm cryoinjury - Corrected Proof

Tamer M. Said, Aarti Gaglani, Ashok Agarwal — 2010-08-02

Abstract: Apoptosis is an ongoing physiological phenomenon that has been documented to play a role in male infertility, if deregulated. Caspase activation, externalization of phosphatidylserine, alteration of mitochondrial membrane potential and DNA fragmentation are markers of apoptosis found in ejaculated human spermatozoa. These markers appear in excess in subfertile men and functionally incompetent spermatozoa. Sperm cryopreservation is a widely used procedure in the context of assisted reproductive techniques. Cryopreservation and thawing is a procedure that inflicts irreversible injury on human spermatozoa. The damage is manifested by a decrease in recovery of viable spermatozoa with optimum fertilization potential. This review describes the implication of apoptosis as one of the possible mechanisms involved in sperm cryoinjury. Evidence shows significant increase in some apoptosis markers following cryopreservation and thawing. On the other hand, the increase in sperm DNA fragmentation following cryopreservation and thawing requires further investigation. Specific technical measures should be applied to minimize the induction of apoptosis in human spermatozoa during cryopreservation and thawing. These include standardization of freezing protocols and cryoprotectant use. Selection of non-apoptotic spermatozoa may also prove to be of benefit.Apoptosis, also termed selective cell death, is a phenomenon that affects cell viability. The process has been documented in human spermatozoa and is implicated in the failure of fertilization following assisted reproductive techniques. There are several markers of apoptosis that present in ejaculated human spermatozoa, especially in those from infertile men. Freezing and thawing of human spermatozoa is commonly used nowadays in conjunction with assisted reproductive techniques such as intrauterine insemination and IVF. Sperm freezing is also used to preserve fertility in men undergoing cancer treatment or before undergoing vasectomies. Therefore, it is important to ensure that frozen–thawed spermatozoa will maintain its fertilizing capability. Despite recent methodological advances, freezing exerts detrimental effects on spermatozoa that lead to significant decreases in sperm viability and motility and ultimately in fertilization potential. The process of freezing and thawing increases the amount of apoptotic spermatozoa, which in turn is expected to decrease the success rates of assisted reproductive techniques. Therefore, current protocols for sperm freezing and thawing should be carefully evaluated to ensure minimizing the extent of apoptosis induction. The integration of novel techniques that isolate non-apoptotic spermatozoa also increases sperm survival following freezing and thawing.

IVF treatment should not be postponed for patients with high basal FSH concentrations - Accepted Manuscript

2010-08-02

Abstract: This study determined the influence of inter-cycle variation of basal FSH concentrations on IVF treatment results, in patients with a history of high basal FSH. Patients underwent at least two IVF cycles, one with basal serum FSH ⩾10 IU/l and the other at least 3 IU/l lower (interval between cycles being <1 year when the second cycle had the elevated FSH). A subanalysis was performed in patients with exceptionally large differences in values (⩾16 IU/l and ⩽12 IU/l). IVF outcomes were compared according to basal FSH concentrations in two consecutive cycles. Seventy-six patients met the inclusion criteria. Mean basal serum FSH were 15.0 ± 3.6 IU/l in the ‘high FSH’ group (range 12–24 IU/l) and 9.0 ± 3.0 IU/l in the ‘low FSH’ group (range 5–14 IU/l). Patient age, oestradiol at HCG administration, number of collected oocytes, fertilization and clinical pregnancy rates were similar for all cycles compared. Analysis of the subgroup with exceptionally large differences of basal FSH concentration yielded similar results. Neither high nor low basal serum FSH values were associated with IVF outcome in patients with reduced ovarian reserve and previously determined high basal FSH concentrations. Ovarian stimulation need not be delayed until FSH declines.

Possible role of endometriosis in the aetiology of spontaneous miscarriage in patients with septate uterus - Corrected Proof

2010-08-02

Abstract: A recent study found a significant correlation between endometriosis and non-obstructive forms of Müllerian anomalies. Other studies described an increased miscarriage rate in patients with endometriosis. This study assessed the effect of endometriosis on pregnancy outcome in a group of patients with endometriosis and septate uterus. Spontaneously achieved pregnancies were taken into consideration. The outcome of 179 infertile women who underwent surgery for septate uterus was analysed in a retrospective study. Stage I or II endometriosis was found by laparoscopy in 36 patients. The pregnancy outcomes, before and after metroplasty, of the group of 36 patients with septum and endometriosis were compared with the pregnancy outcomes of 143 patients with septate uterus with no endometriosis. Before metroplasty the incidence of pregnancy loss was 67% in patients without endometriosis and 75% in patients with endometriosis and the difference was not significant. After metroplasty, no significant differences have been found between the two groups, suggesting that endometriosis could be an occasional finding not influencing pregnancy outcome.Septate uterus is a relatively common minor anomaly. The septum may be complete, which is rare, or incomplete. In incomplete cases the uterus is divided in two cavities or, more often, hemi cavities by a midline indentation of the fundus. Endometriosis is described as a cause of increased miscarriage rate. It is characterized by the presence of the mucosa covering the internal part of the uterus outside the uterus, most of times in the abdominal cavity in the form of peritoneal implants, like hundreds of pinpoints, or in the form of ovarian (chocolate) cysts. The connection between obstructive forms of Müllerian anomalies and endometriosis is commonly accepted, according to the theory of retrograde reflux of menstrual blood throughout the tubes. Few studies found an increased incidence of endometriosis even in women with non-obstructive forms of Müllerian anomalies such as septate uterus. The aim of our study was to assess the hypothesis that, even in patients with endometriosis, uterine septum is the cause that leads to miscarriage. Before surgical correction of septate uterus, the incidence of miscarriage was 67% in patients without endometriosis and 75% in patients with endometriosis. After surgery, a clear reduction of miscarriage rate was obtained, but no difference between the two groups was found. In our study, increased abortion rate depends more likely on uterine malformations than on endometriosis. Endometriosis seems to be an occasional finding not influencing pregnancy outcome.

Fragile X syndrome prenatal diagnosis: parental attitudes and reproductive responses - Corrected Proof

2010-08-02

Abstract: Fragile X syndrome (FXS) is the most common inherited form of mental retardation. It is caused by a CGG repeat expansion, which results in hypermethylation and silencing of the FMR1 gene. The results from 213 FXS prenatal diagnoses performed in the study centre were reviewed. Family history of FXS or undiagnosed mental retardation (MR) were the reasons for referral and 64% of mothers were not aware of their status so prenatal and mother tests were performed at the same time. Among those women referred for family history of unknown MR, 17.6% were found to be FXS carriers. The attitudes and perceptions of the syndrome of 52 FXS carriers were also evaluated. Most of them had been diagnosed as carriers when the child was already born and the most common feeling was sadness, followed by impotence and guilt. The majority of them had received genetic counselling and they considered it useful. Regarding reproductive options, prenatal diagnosis was chosen by 40.5% of women. Prenatal diagnosis for FXS is a good reproductive option and it should be carried out whenever family history of MR is present. A high percentage of FXS carriers are detected following this approach.Fragile X syndrome (FXS) is the most common inherited form of mental retardation. It is caused by a CGG repeat expansion, which results in silencing of the FMR1 gene. Individuals can be considered as normal (<55 repeats), carriers (55–200) and affected (>200) for the FMR1 gene. FXS prenatal diagnosis is feasible and we present the results from 213 FXS prenatal diagnoses performed in our centre. Reasons for referral were family history of FXS or undiagnosed mental retardation (MR). It is worth mentioning that 64% of mothers were not aware of their status before the test, so prenatal and mother tests were performed at the same time. Among those women referred for family history of unknown MR, 17.6% were FXS carriers. We also evaluated the attitudes and perceptions of the syndrome of 52 FXS carriers. Most of them had been diagnosed as carriers when the child was already born and the most common feeling was sadness, followed by impotence and guilt. The majority of them had received genetic counselling and they considered it useful. Regarding reproductive options, prenatal diagnosis was chosen by 40.5% of women. Prenatal diagnosis for FXS is a good reproductive option, and it should be carried out whenever family history of MR is present. A high percentage of FXS carriers are detected following this approach.

Case report: first successful application of preimplantation genetic diagnosis for hereditary angiooedema - Accepted Manuscript

2010-08-02

Abstract: Hereditary angiooedema is an autosomal dominant disease caused by mutations in the SERPING1 gene. It is characterized by oedemas in different parts of the body, being particularly dangerous when swelling involves the upper airway. Preimplantation genetic diagnosis (PGD) was performed in a couple where the woman carries a deletion of 2.9 Kb that includes exon 4 of the SERPING1 gene. Four polymorphic short tandem repeat markers were tested in order to establish the disease-bearing haplotype and three of them were fully informative. Amplification efficiency at the preclinical work up ranged from 71–100% for each locus and allele drop out rates were between 0–20% for the polymorphic markers. The couple underwent PGD using fluorescent multiplex heminested polymerase chain reaction. Six embryos were biopsied and five of them were diagnosed as healthy. Two embryos were transferred and a singleton pregnancy was achieved, resulting in the birth of a healthy boy.

Intracytoplasmic sperm injection outcome versus intracytoplasmic morphologically selected sperm injection outcome: a meta-analysis - Corrected Proof

2010-08-02

Abstract: The development of a modified intracytoplasmic sperm injection (ICSI), called intracytoplasmic morphologically selected sperm injection (IMSI), demonstrated that a profound morphological investigation of the spermatozoon, under the magnification of 6600×, enables outcome improvement. The aim of this study was to compare ICSI outcome with IMSI outcome. The meta-analysis results demonstrated no significant difference in fertilization rate between ICSI and IMSI groups. However, a significantly improved implantation (odds ratio (OR) 2.72; 95% confidence interval (CI) 1.50–4.95) and pregnancy rate (OR 3.12; 95% CI 1.55–6.26) was observed in IMSI cycles. Moreover, the results showed a significantly decreased miscarriage rate (OR 0.42; 95% CI 0.23–0.78) in IMSI cycles as compared with ICSI cycles. This is the first meta-analysis of published data to evaluate the potential benefits of IMSI. The pooled data of IMSI cycles demonstrate a statistically significant improvement in implantation and pregnancy rates and a statistically significant reduction in miscarriage rates. However, more randomized controlled trials are needed to confirm these results.The development of a modified intracytoplasmic sperm injection (ICSI) method, called intracytoplasmic morphologically selected sperm injection (IMSI), has enabled the improvement of pregnancy rates. The aim of this study was to perform a systematic review of the literature and compare the fertilization, implantation, pregnancy and miscarriage rates in ICSI and IMSI cycles. Our meta-analysis results demonstrated no significant difference in fertilization between ICSI and IMSI groups. However, implantation and pregnancy rates were almost 3-fold higher in IMSI cycles. Moreover, the miscarriage rate was more than 40% lower in IMSI cycles as compared with ICSI cycles. This meta-analysis demonstrates a significant improvement in implantation and pregnancy rates, and a significant reduction in miscarriage rates. However, further studies are needed to confirm our results.

The source and implications of progesterone rise during the follicular phase of assisted reproduction cycles - Corrected Proof

Richard Fleming, Julian Jenkins — 2010-08-02

Abstract: Moderate elevations in serum progesterone concentrations are observed following the use of gonadotrophin-releasing hormone agonists during ovarian stimulation. The clinical significance of this phenomenon has been investigated, but findings have been inconclusive. This commentary proposes that progesterone concentrations are indeed important in endometrial advancement and oocyte/embryo development, which, may lead to asynchrony between endometrial and embryo development. Based on the two-cell, two-gonadotrophin model, this commentary proposes a hypothesis to describe how progesterone concentration increases during ovarian stimulation and three factors influencing this during ovarian stimulation are identified: the number of follicles, the FSH drive and the LH activity. It also suggests how differences in gonadotrophin preparations used for ovarian stimulation may have differential effects on progesterone synthesis. It remains to be tested whether routine measurement of late follicular progesterone concentrations may prove beneficial as suitable assay methods are now available. However, strategies that reduce follicular recruitment in high-responding women and gonadotrophins that contain LH activity may reduce the degree of progesterone elevation prior to luteinization.

Religion, reproduction and public policy: disentangling morality from Catholic theology - Accepted Manuscript

Edgar Dahl — 2010-08-02

Abstract: Many people look to religion to help resolve the serious moral and legal issues associated with assisted reproductive technologies. Doing so presupposes that religion is the cornerstone of ethics, but this assumption is not well founded. While various faiths are entitled to articulate their views on matters of human reproduction, the contradictions involved in doing so make it unwise to rely on religion in the formulation of law and policy. These contradictions – such as the indeterminacy about what revealed truths means – make moral secular philosophy a better guide for the protection of human welfare.

Impact of the Spanish Fertility Society guidelines on the number of embryos to transfer - Accepted Manuscript

2010-08-02

Abstract: The multiple pregnancy rate in assisted reproduction treatment cycles depends, fundamentally, on the number of embryos transferred. It is essential that patients and professionals should have good practical guidelines on the best number of embryos to be transferred in each cycle in order to obtain high pregnancy rates with minimal risk of multiple pregnancies. This study analysed the impact made by the Spanish Fertility Society (SEF) guidelines on the number of embryos to be transferred, as regards the policies adopted at clinics in Spain and the resulting financial repercussions. Data were collected from the assisted reproduction treatment register of the SEF and compared over three periods of time: 2002–2003, with no legal regulation and no SEF guidelines; 2004, with only legal regulation; and 2005–2006, with legal regulation and SEF guidelines. The acceptance of SEF guidelines varies according to the IVF technique. The guidelines have led to a reduction in multiple pregnancy rates, especially concerning triplets, in patients’ own-egg and with donor-egg cycles. Even without full implantation, these results validate the clinical utility of the SEF guidelines. They constitute a useful tool to reduce the incidence of the principal adverse effect of treatment cycles: multiple pregnancies.

Single nucleotide polymorphisms in the anti-Müllerian hormone signalling pathway do not determine high or low response to ovarian stimulation - Accepted Manuscript

2010-08-02

Abstract: There is substantial variability in ovarian response to exogenous gonadotrophins in women undergoing ovarian stimulation for IVF. Genetic variation in signalling pathways of the ovary could influence ovarian stimulation outcome. Studies have shown a correlation between the serum concentration of anti-Müllerian hormone (AMH) and ovarian stimulation outcome. This paper present a retrospective case-controlled genetic association study designed to test the association between single nucleotide polymorphisms (SNP) in the AMH signalling pathway and two clinically important outcomes of ovarian stimulation: low and high response. Blood samples from 53 high responders, 38 low responders and 100 controls were analysed for eight SNP of interest. Odds ratios and 95% confidence intervals were estimated by a binary logistic regression model adjusting for age and body mass index. As far as is known, this is the first report on the influence of these SNP, present in approximately 19% of women, on ovarian stimulation outcome. No statistically significant association was found between any of the SNP studied and high or low response to ovarian stimulation. It seems unnecessary to detect these SNP when applying the serum concentration of AMH as a predictor of ovarian response to stimulation.

Redefining advanced maternal age as an indication for preimplantation genetic screening - Accepted Manuscript

2010-08-02

Abstract: In this retrospective study, the utility of preimplantation genetic screening (PGS) in patients with advanced maternal age is evaluated. The patient population consisted of women aged 38–44 years and included in a regular IVF programme with or without PGS analysis. Transfer rate, ongoing implantation rate and ongoing pregnancy rate were the main outcome parameters measured. A trend of better ongoing pregnancy rate per oocyte retrieval was observed in patients aged 38 and 39 years in the non-PGS group when compared with PGS groups, but better ongoing pregnancy rate per oocyte retrieval was observed in patients 41–44 years old in the PGS group. When patients with a low ovarian response accumulated oocytes in several stimulation cycles, clinical outcomes were comparable to those of normal-responder patients. These results show that, although PGS does not benefit patients less than 40 years of age, reproductive success increases more than two-fold in patients over 40 years, especially in patients with more than six metaphase II oocytes, as a result of a good ovarian response or gamete accumulation, suggesting a redefinition of advanced maternal age as indication for PGS.

Erythropoietin receptor-like immunostaining on human spermatozoa - Accepted Manuscript

2010-08-02

Abstract: This study aimed to demonstrate the presence of erythropoietin (EPO) receptor on spermatozoa. Whole ejaculates of four healthy volunteers were incubated with polyclonal rabbit anti-EPO receptor and subsequently stained with a Cy-3 labelled secondary antibody. Four slides per subject were analysed, no staining was observed in slides incubated with either primary or secondary antibody alone. EPO receptor staining was positive in 92 ± 8% of EPO pre-treated and 91 ± 4% of non-treated sperm cells. The results suggest that spermatozoa express EPO receptor on plasma membrane, which might act to protect these cells from damage after ejaculation.

Simplified PGD of common determinants of haemoglobin Bart’s hydrops fetalis syndrome using multiplex-microsatellite PCR - Accepted Manuscript

2010-08-02

Abstract: The high incidence of double-gene deletions in α-thalassaemia increases the risk of having pregnancies with homozygous α 0-thalassaemia, the cause of the lethal haemoglobin (Hb) Bart’s hydrops fetalis syndrome. Preimplantation genetic diagnosis (PGD) has played an important role in preventing such cases. However, the current gap-PCR based PGD protocol for deletional α -thalassaemia requires specific primer design for each specific deletion. A universal PGD assay applicable to all common deletional determinants of Hb Bart’s hydrops fetalis syndrome has been developed. Microsatellite markers 16PTEL05 and 16PTEL06 within the α -globin gene cluster were co-amplified with a third microsatellite marker outside the affected region in a multiplex-PCR reaction and analysed by capillary electrophoresis. Eight informed couples at risk of having Hb Bart’s hydrops fetalis were recruited in this study and all patients underwent standard procedures associated with IVF. A total of 45 embryos were analysed. Three pregnancies were achieved from three couples, with the births of two healthy babies and one ongoing pregnancy. This work has successfully adapted an earlier protocol and developed a simple and reliable single-cell assay applicable to PGD of Hb Bart’s hydrops fetalis syndrome regardless of type of deletion.

Analysis of factors associated with multiple pregnancy in an oocyte donation programme - Accepted Manuscript

2010-08-02

Abstract: The aim of this study is to identify the factors associated with multiple pregnancy in an oocyte donation programme. A retrospective study (2000–2007) of 945 synchronous cycles was performed. Two embryos were transferred in all cycles on day 2 after oocyte retrieval. All variables (egg donor and recipient age, number of inseminated oocytes, fertilized oocytes, cleaved embryos, good-quality embryos available, good-quality embryos transferred and frozen embryos) were analysed in relation to the clinical pregnancy rate per transfer (PR) and the multiple pregnancy rate (MPR). The donor age was 26.8 4.5 years and recipient age was 41.0 5.4. The number of good-quality embryos per recipient was 3.1 2.5. The PR was 55.1% and the MPR 36.5%. The number of good-quality embryos transferred (2 versus 0) was significantly associated (P < 0.05) with the PR (60.6% versus 43.5%). The relationship between the MPR and the number of good-quality embryos transferred was adjusted by donor and recipient’s age. For those patients who received 2 versus 0 good-quality embryos, the odds ratio of a multiple pregnancy was 2.1 (95% CI 1.121–3.876). The only predictive factor for multiple pregnancies in an oocyte donation programme is the quality of the transferred embryos.

Polymorphisms of nucleotide-excision repair genes may contribute to sperm DNA fragmentation and male infertility - Accepted Manuscript

2010-08-02

Abstract: The nucleotide-excision repair (NER) system is crucial for the removal of bulky DNA adducts during spermatogenesis. Dysfunction of its repair capacity is likely related to the increased susceptibility to DNA damage. In this study, four polymorphisms in NER pathway (XPA(–4) G/A, ERCC1 C8092A, XPD Lys751Gln and XPF Ser835Ser) were selected to evaluate their potential impact on sperm DNA damage and male infertility. Genotypes were determined by PCR-restriction fragment length polymorphism. Sperm DNA damage was evaluated by TdT-mediated dUDP nick-end labelling assay. A case-only study of 620 infertile men found a significant association between XPA(–4) G/A polymorphism and sperm DNA damage. Individuals with the XPA(–4) A allele showed more sperm DNA damage and lower sperm concentration than G allele carriers. Further analysis, including 620 patients and 385 controls, revealed a 1.52-fold risk (95% CI 1.08–2.02) of developing male infertility in the XPA(–4) AA carriers compared with noncarriers. Luciferase assay verified that the promoter with the XPA(–4) A allele had a lower transcriptional activity than that with the G allele. These data provide the first evidence that –4 G/A polymorphism in XPA promoter alters its transcriptional activity and, thus, might contribute to sperm DNA damage and male infertility.

Hypo-osmotic swelling test identifies individual spermatozoa with minimal DNA fragmentation - Accepted Manuscript

James D Stanger, Long Vo, John L Yovich, Ghanim Almahbobi — 2010-08-02

Abstract: One concern during intracytoplasmic sperm injection (ICSI) is that selected spermatozoa may have increased levels of DNA damage; however, the available testing for this is largely destructive in nature and therefore unsuitable as a tool for sperm selection. One alternative selection process that has previously achieved pregnancies is the hypo-osmotic swelling test (HOST). This study reports that low HOST values of neat semen samples were significantly (P < 0.001) associated with increased DNA damage identified by the DNA fragmentation index (DFI) from the sperm chromatin structure assay as well as the TdT-mediated dUTP nick-end labelling (TUNEL) assay. The HOST value was highly predictive of an abnormal DFI value by receiver operating characteristic curve analysis (P < 0.001). Furthermore, when individual spermatozoa were assessed for both HOST status and DNA fragmentation by TUNEL, the key HOST-induced tail-swelling grades D, E and F were most commonly associated with high HOST values and were significantly (P < 0.001) associated with minimal DNA damage regardless of the DNA status of the ejaculate. The application of HOST may be a valuable tool in the routine identification and selection of viable, DNA-intact individual spermatozoa for ICSI after further research to demonstrate its efficacy and safety.

Development of humanized culture medium with plant-derived serum replacement for human pluripotent stem cells - Accepted Manuscript

2010-08-02

Abstract: For human embryonic stem cells (ESC) to be used in cell replacement therapies, they must be grown under good manufacturing conditions in a chemically defined medium that lacks animal proteins. This study examined the ability of a newly designed medium containing the plant-derived serum replacement VegetaCell and other reagents of human origin to support undifferentiated growth and pluripotency of human ESC. This medium was tested in several culture systems, using human fibroblasts as a feeder layer or Matrigel in a feeder-free culture. Even under the most stringent feeder-free conditions without conditioned medium, human ESC exhibited an undifferentiated morphology, expressed markers of undifferentiated cells, demonstrated high alkaline phosphatase activity and multilineage differentiation and retained a normal karyotype. Compared with human ESC grown in standard culture conditions, human ESC maintained in humanized VegetaCell medium show longer cell cycles and decreased cell death. The availability of an animal protein-free medium supplemented with the low-cost VegetaCell reagent expands the repertoire of media for culturing human ESC as well as induced pluripotent stem cells for drug testing and cell replacement therapy.

Septate, subseptate and arcuate uterus decrease pregnancy and live birth rates in IVF/ICSI - Accepted Manuscript

2010-08-02

Abstract: A retrospective matched-control study to evaluate the effect of uterine anomalies on pregnancy rates after 2481 embryo transfers in conventionally stimulated IVF/intracytoplasmic sperm injection (ICSI) cycles. The study group of 289 embryo transfers before and 538 embryo transfers following hysteroscopic resection of a uterine septum was compared with two consecutive embryo transfers in the control group. Groups were matched for age, body mass index, ovarian stimulation, embryo quality, IVF or ICSI and infertility aetiologies. Number of embryos transferred, embryo quality and absence of uterine anomalies significantly predicted the pregnancy rates in the study group: odds ratios (OR) 1.7, 2.6 and 2.5, respectively (P < 0.001). Pregnancy rates after embryo transfer before hysteroscopic metroplasty were significantly lower, both in women with subseptate and septate uterus and in women with arcuate uterus compared with controls. If two or three embryos with at least one best-quality embryo were transferred, the differences were 9.6% versus 43.6%, OR 7.3 (P < 0.001) and 20.9% versus 35.5%, OR 2.1 (P < 0.03), respectively. Differences in terms of live birth rates were even more evident: 1.9% versus 38.6%, OR 32 (P < 0.001) and 3.0% versus 30.4%, OR 14 (P < 0.001). After surgery, the differences disappeared.

New Turkish legislation on assisted reproductive techniques and centres: a step in the right direction? - Accepted Manuscript

Bulent Urman, Kayhan Yakin — 2010-08-02

Abstract: The Ministry of Health of Turkey has just passed new legislation on assisted reproduction techniques. The new legislation covers significant changes in the regulation of opening new assisted reproduction treatment centres, funding of the treatment, number of embryos to be transferred, cryopreservation and donation. It takes a brave step towards reducing the incidence of multiple pregnancies. However, it also makes the Turkey one of the strictest countries in the world. The strict statements on gamete cryopreservation and donation will have social and clinical consequences.

LH (as HCG) and FSH surges for final oocyte maturation: sometimes it takes two to tango? - Accepted Manuscript

Shahar Kol, Peter Humaidan — 2010-08-02

Abstract: Until now, clinicians have been relying solely on LH activity-dependent triggering of final oocyte maturation and thus taken it for granted that the natural midcycle FSH surge is biologically redundant. However, it is time to question this paradigm. Evidence from clinical studies hint that in a yet-to-be-defined subset of patients, dual LH and FSH surge is advantageous compared with LH-only surge in the form of human chorionic gonadotrophin (HCG) trigger. Dual surge can be triggered by a bolus of gonadotrophin-releasing hormone agonist causing a flare-up of both endogenous LH and FSH, resembling the natural midcycle surge of gonadotrophins. HCG given in parallel secures adequate exposure to LH activity. Further research is needed to characterize the patients in whom FSH surge is needed for proper resumption of the oocyte meiotic process.

Pharmacokinetics and follicular dynamics of corifollitropin α versus recombinant FSH during ovarian stimulation for IVF - Accepted Manuscript

2010-08-02

Abstract: A single injection of corifollitropin α can replace seven daily injections of recombinant FSH (rFSH) using a gonadotrophin-releasing hormone antagonist protocol in ovarian stimulation prior to IVF or intracytoplasmic sperm injection. This double-blind randomized controlled trial assessed the pharmacokinetics and pharmacodynamics of 150 μg corifollitropin α versus daily 200 IU rFSH in 1509 patients. Comparative analyses were performed on serum concentrations of FSH immunoreactivity (pharmacokinetics), and the number and size of growing follicles, and inhibin B and oestradiol concentrations as biomarkers of ovarian response (pharmacodynamics). The rate of follicular development was similar in both treatment groups. By stimulation day 8, 33% of patients treated with corifollitropin α reached the criterion for human chorionic gonadotrophin (HCG) injection. The number of follicles ⩾11 mm was slightly higher after corifollitropin α compared with daily rFSH at stimulation day 8 (difference, 1.2; 95% confidence interval (CI) 0.5–1.8; P < 0.01) and on the day of HCG injection (difference, 2.1; 95% CI 1.4–2.8; P < 0.01). The rise of inhibin B and oestradiol concentrations was similar in both treatment groups. Although the pharmacokinetics of corifollitropin α and rFSH are quite different their pharmacodynamic profiles at the dosages used are similar

Minimal ovarian stimulation (mini-IVF) for IVF utilizing vitrification and cryopreserved embryo transfer - Corrected Proof

John Zhang, Lyndon Chang, Yoshie Sone, Sherman Silber — 2010-08-02

Abstract: Gentle ovarian stimulation protocols, such as ‘mini-IVF’, have several potential advantages over conventional IVF protocols, including less medication and fewer injections, producing fewer eggs, but eggs of higher quality. The particular ‘mild’ stimulation protocol called ‘mini-IVF’ is described. This protocol requires a reliable and cheap method for embryo cryopreservation such as vitrification, because of the negative impact of clomiphene citrate on the endometrium and since cryopreserved embryo transfers with this protocol have yielded much higher pregnancy rates than fresh transfers. In this series, patients were not denied treatment based on their day-3 FSH value or ovarian reserve. Yet very acceptable pregnancy rates were achieved (20% for fresh embryo transfers and 41% for cryopreserved embryo transfers). These results strengthen the argument for a mini-IVF protocol and vitrification as an alternative to standard conventional IVF stimulation protocols. Now a randomized control trial with cryopreserved single-embryo transfer is required.Gentle ovarian stimulation protocols have several potential advantages over conventional IVF protocols, including less medication and fewer injections, producing fewer eggs, but eggs of higher quality. ‘Mini-IVF’ is safe, patient friendly and physiologically more natural. It may be more cost effective if results are comparable to conventional protocols. Vitrification of embryos allows the transfer of warmed embryos in subsequent cycles when the endometrium is more receptive. In this series, patients were not denied treatment based on their day-3 FSH value or ovarian reserve. Yet very acceptable pregnancy rates were achieved (20% for fresh embryo transfers and 41% for cryopreserved embryo transfers). These results strengthen the argument for gentle stimulation protocols and vitrification in preference to standard conventional IVF stimulation protocols. Now a randomized control trial with cryopreserved single-embryo transfer is required.

Salpingotomy or salpingectomy in tubal ectopic pregnancy: what do women prefer? - Accepted Manuscript

2010-08-02

Abstract: There is an ongoing debate whether tubal ectopic pregnancy should be treated by salpingotomy or salpingectomy. It is unknown which treatment women prefer in view of the potentially better fertility outcome but disadvantages of salpingotomy. This study investigated women surgically treated for tubal ectopic pregnancy and subfertile women desiring pregnancy and their preferences for salpingotomy relative to salpingectomy by means of a web-based discrete choice experiment consisting of 16 choice sets. Scenarios representing salpingotomy differed in three attributes: intrauterine pregnancy (IUP) chance, risk of persistent trophoblast and risk of repeat ectopic pregnancy. An ‘opt out’ alternative, representing salpingectomy, was similar for every choice set. A multinomial logistic regression model was used to analyse relative importance of the attributes. This study showed that the negative effect of repeat ectopic pregnancy was 1.6 times stronger on the preference of women compared with the positive effect of the spontaneous IUP rate. For all women, the risk of persistent trophoblast was acceptable if compensated by a small rise in the spontaneous IUP rate. The conclusion was that women preferred avoiding a repeat ectopic pregnancy to a higher probability of a spontaneous IUP in the surgical treatment of tubal ectopic pregnancy.