Showing papers in "Human Reproduction in 1998"
TL;DR: It is shown that 2D:4D in right and left hands has a sexually dimorphic pattern and is probably established in utero, which raises the possibility that patterns of digit formation may relate to spermatogenesis and hormonal concentrations.
Abstract: The differentiation of the urinogenital system and the appendicular skeleton in vertebrates is under the control of Hox genes. The common control of digit and gonad differentiation raises the possibility that patterns of digit formation may relate to spermatogenesis and hormonal concentrations. This work was concerned with the ratio between the length of the 2nd and 4th digit (2D:4D) in humans. We showed that (i) 2D:4D in right and left hands has a sexually dimorphic pattern; in males mean 2D:4D = 0.98, i.e. the 4th digit tended to be longer than the 2nd and in females mean 2D:4D = 1.00, i.e. the 2nd and 4th digits tended to be of equal length. The dimorphism is present from at least age 2 years and 2D:4D is probably established in utero; (ii) high 2D:4D ratio in right hands was associated with germ cell failure in men (P = 0.04); (iii) sperm number was negatively related to 2D:4D in the right hand (P = 0.004); (iv) in men testosterone concentrations were negatively related to right hand 2D:4D and in women and men LH (right hand), oestrogen (right and left hands) and prolactin (right hand) concentrations were positively correlated with 2D:4D ratio and (v) 2D:4D ratio in right hands remained positively related to luteinizing hormone and oestrogen after controlling for sex, age, height and weight.
1,322 citations
TL;DR: Weight loss should be considered as a first option for women who are infertile and overweight, and the cost savings of the programme were considerable.
Abstract: Obesity affects ovulation, response to fertility treatment, pregnancy rates and outcome. In this prospective study, a weight loss programme was assessed to determine whether it could help obese infertile women, irrespective of their infertility diagnosis, to achieve a viable pregnancy, ideally without further medical intervention. The subjects underwent a weekly programme aimed at lifestyle changes in relation to exercise and diet for 6 months; those that did not complete the 6 months were treated as a comparison group. Women in the study lost an average of 10.2 kg/m2, with 60 of the 67 anovulatory subjects resuming spontaneous ovulation, 52 achieving a pregnancy (18 spontaneously) and 45 a live birth. The miscarriage rate was 18%, compared to 75% for the same women prior to the programme. Psychometric measurements also improved. None of these changes occurred in the comparison group. The cost savings of the programme were considerable. Prior to the programme, the 67 women had had treatment costing a total of A$550,000 for two live births, a cost of A$275,000 per baby. After the programme, the same women had treatment costing a total of A$210,000 for 45 babies, a cost of A$4600 per baby. Thus weight loss should be considered as a first option for women who are infertile and overweight.
729 citations
TL;DR: The ability to transfer just two blastocysts while maintaining high pregnancy rates will help to eliminate high order multiple gestations and improve the overall efficiency of human IVF.
Abstract: The effectiveness of blastocyst culture and transfer in human in-vitro fertilization (IVF) was evaluated in a prospective randomized trial in patients having a moderate to good response to gonadotrophin stimulation. Embryos were transferred either on day 3 after culture to around the 8-cell stage in Ham's F-10 medium supplemented with fetal cord serum, or on day 5 after culture to the blastocyst stage in the sequential serum-free media G 1.2 and G 2.2. The pregnancy rates after transfer on day 3 or day 5 were equivalent, 66 and 71% respectively; however, significantly more embryos were transferred on day 3 (3.7) than on day 5 (2.2). The number of blastocysts transferred did not affect the implantation rate, and pregnancy rates when either two or three blastocysts were transferred were 68 and 87% respectively. The implantation rate of the blastocysts (50.5% fetal heart beat) was significantly higher compared to the cleavage stage embryos transferred on day 3 (30.1%). The percentage of blastocyst development was not affected by the number of 2-pronuclear embryos, or by maternal age. Irrespective of the number of blastocysts formed, pregnancy rates were similar. Furthermore, the pregnancy rate following blastocyst transfer in patients with 10 or more follicles at the time of human chorionic gonadotrophin administration was not affected by patient age. More than 60% of patients having blastocyst culture and transfer had supernumerary embryos for cryopreservation. The establishment of a pregnancy following thaw and transfer confirmed the viability of cryopreserved blastocysts cultured in the absence of serum or co-culture. The ability to transfer just two blastocysts while maintaining high pregnancy rates will therefore help to eliminate high order multiple gestations and improve the overall efficiency of human IVF.
698 citations
TL;DR: Similarities in VEGF content were observed in the glandular epithelium of the eutopic endometrium of women with endometriosis and red lesions, suggesting that endometiosis probably arises from the peritoneal seeding of viable endometrial cells during retrograde menstruation and that red lesions can be considered as the first stage of implantation.
Abstract: Angiogenesis is likely to be involved in the pathogenesis of endometriosis. According to the transplantation theory, when the exfoliated endometrium is attached to the peritoneal layer, the establishment of a new blood supply is essential for the survival of the endometrial implant and development of endometriosis. From the known angiogenic factors, vascular endothelial growth factor (VEGF) has emerged as a pivotally important regulator of normal angiogenesis and pathological neovascularization. The VEGF protein was evaluated immunohistochemically in the eutopic endometrium of 10 women without endometriosis (group I) at laparoscopy and the eutopic endometrium and peritoneal endometriotic lesions of 43 women with endometriosis (group II). VEGF histological scores were 9.7 +/- 4.3 and 4.0 +/- 2.6 respectively in the epithelium and stroma of the eutopic endometrium of group I women, and 10.3 +/- 2.3 and 3.6 +/- 2.3 respectively in women of group II. In red lesions, the VEGF scores were 11.1 +/- 3.0 in the epithelium and 5.1 +/- 3.0 in the stroma, and in black lesions were 8.6 +/- 2.7 and 1.6 +/- 1.6, respectively. Significantly lower values were observed in black lesions as compared with eutopic endometrium and red lesions, the values of which were similar. Scores were also evaluated according to the phase of the cycle. In eutopic as well as ectopic endometrium, no significant cyclic variations were observed throughout the cycle. However, VEGF content was found to be higher in the eutopic glandular epithelium of women with endometriosis during the late secretory phase, possibly suggesting a more likely tendency to implant. In contrast, significantly higher VEGF content was noted in red lesions as compared with black lesions. During all phases of the cycle, the VEGF content in stromal cells of red lesions was higher than in black lesions. Similarities in VEGF content were observed in the glandular epithelium of the eutopic endometrium of women with endometriosis and red lesions, suggesting that endometriosis probably arises from the peritoneal seeding of viable endometrial cells during retrograde menstruation and that red lesions can be considered as the first stage of implantation. After the attachment phase, the high VEGF levels could provoke an increase in the subperitoneal vascular network and facilitate implantation and viability in the retroperitoneal space. Lower VEGF levels in black lesions explain the decrease in both stromal vascularization, followed by fibrosis and inactivation of the implant.
498 citations
TL;DR: The results indicate that high frequency UC on the day of embryo transfer hinder IVF-embryo transfer outcome, possibly by expelling embryos out of the uterine cavity, and the negative correlation between UC frequency and progesterone concentrations supports the uterusine relaxing properties of progester one.
Abstract: To investigate the possible consequences of uterine contractions (UC) as visualized by ultrasound (US) on in-vitro fertilization (IVF)-embryo transfer outcome, we studied prospectively 209 infertile women undergoing 220 cycles of controlled ovarian stimulation. Inclusion criteria were age 5.0 (n = 74) UC/min respectively. Patients, controlled ovarian hyperstimulation and embryology characteristics were comparable in all groups. A stepwise decrease in clinical and ongoing pregnancy rates as well as in implantation rates occurred from the lowest to the highest UC frequency groups (53, 36, 21; 46, 32, 20; 23, 19, 10; and 14, 11, 4%; P < 0.001). Plasma progesterone and UC frequency were negatively correlated (r = -0.34, P < 0.001). Direction of UC did not affect embryo transfer outcome. As this study was controlled strictly for confounding variables and UC were assessed objectively by a computerized system, its results indicate that high frequency UC on the day of embryo transfer hinder IVF-embryo transfer outcome, possibly by expelling embryos out of the uterine cavity. The negative correlation between UC frequency and progesterone concentrations supports the uterine relaxing properties of progesterone.
461 citations
TL;DR: It is concluded that there is a direct relationship between increased H2O2 concentration and apoptosis, and that further studies should be undertaken to confirm these findings.
Abstract: In human in-vitro fertilization (IVF)-embryo transfer, the in-vitro culture environment differs from in-vivo conditions in that the oxygen concentration is higher, and in such conditions the mouse embryos show a higher concentration of reactive oxygen species (ROS) in simple culture media. ROS are believed to cause damage to cell membranes and DNA fragmentation in somatic cells. This study was conducted to ascertain the level of H 2 O 2 concentration within embryos and the morphological features of cell damage induced by H 2 O 2 . A total of 62 human oocytes and embryos (31 fragmented, 15 non-fragmented embryos, 16 unfertilized oocytes) was obtained from the IVF-embryo transfer programme. The relative intensity of H 2 O 2 concentrations within embryos was measured using 2',7'-dichlorodihydrofluorescein diacetate by Quanti cell 500 fluorescence imaging and DNA fragmentation was observed with transmission electron microscopy and an in-situ apoptosis detection kit. The H 2 O 2 concentrations were significantly higher in fragmented embryos (72.21 ± 9.62, mean ± SEM) compared to non-fragmented embryos (31.30 ± 3.50, P < 0.05) and unfertilized oocytes (30.75 ± 2.67, P < 0.05). Apoptosis was observed only in fragmented embryos, and was absent in non-fragmented embryos. Electron microscopic findings confirmed apoptotic bodies and cytoplasmic condensation in the fragmented blastomeres. We conclude that there is a direct relationship between increased H 2 O 2 concentration and apoptosis, and that further studies should be undertaken to confirm these findings.
459 citations
TL;DR: The evidence presented in this review is compelling because of the consistency of effect across different study designs, sample size and types of outcome, however, continued reassurance is needed that the calculated overall effect is not in fact due to confounding variables.
Abstract: The high prevalence of smoking among women in their reproductive years continues to be a matter of concern. The negative effects of smoking on general health are well known, but smoking may also affect fertility. The objective of the present study was to perform a systematic review of the literature to determine whether there is an association between smoking and risk of infertility in women of reproductive age, and to assess the size of this effect. In the 12 studies used for this meta-analysis, the overall value of the odds ratio (OR) for risk of infertility in women smokers versus non-smokers was 1.60 [95% confidence interval (CI) 1.34-1.91]. Studies of subfertile women undergoing in-vitro fertilization (IVF) treatment also show a reduction in fecundity among women smokers. A meta-analysis of nine studies found an OR of 0.66 (95% CI 0.49-0.88) for pregnancies per number of IVF-treated cycles in smokers versus non-smokers. Despite the potential limitations of meta-analyses of observational studies, the evidence presented in this review is compelling because of the consistency of effect across different study designs, sample size and types of outcome. However, continued reassurance is needed that the calculated overall effect is not in fact due to confounding variables.
456 citations
TL;DR: ROS can cause an increase in DNA fragmentation and pretreatment with antioxidants can reduce DNA damage, and the addition of antioxidants significantly decreased the amount of DNA damage induced by ROS generation.
Abstract: The objective of this study was to evaluate the effect of the generation of reactive oxygen species (ROS) on the integrity of the DNA of human spermatozoa, and to determine if pretreatment with antioxidants can reduce DNA damage. Samples were obtained from 47 men undergoing infertility investigation. ROS were generated in the samples by the addition of xanthine/xanthine oxidase (X/XO) with or without antioxidants. After incubation at timed intervals (0-2 h) with X/XO, the percentage of spermatozoa with DNA fragmentation was determined using the method of TdT-mediated DNA end-labelling (TUNEL). Time intervals were selected to mimic the clinical situation in which spermatozoa are held for a period of time after swim-up while the oocytes are prepared for ICSI. A significant increase in sperm DNA damage was evident when samples were incubated in the presence of ROS for intervals of 1 and 2 h, but not when incubated with ROS for <1 h (P = 0.0001). The addition of antioxidants significantly decreased the amount of DNA damage induced by ROS generation (P < 0.04). ROS can cause an increase in DNA fragmentation and pretreatment with antioxidants can reduce DNA damage.
449 citations
TL;DR: The World Health Organization has proposed a scheme for the diagnostic classification of male infertility, based upon a standardized approach to clinical assessment and to the assessment of semen quality, which is now controversial.
Abstract: Infertility is a common problem, affecting perhaps one couple in six, the majority of whom now seek medical care. Although diagnostic problems make it difficult to establish the extent of the male partner's contribution with certainty, a number of studies suggest that male problems represent the commonest single defined cause of infertility. The World Health Organization has proposed a scheme for the diagnostic classification of male infertility, based upon a standardized approach to clinical assessment and to the assessment of semen quality. Some of these classifications are now controversial, and many are descriptive, rather than aetiological. Increasingly, the importance of occupation, environmental and particularly genetic factors in the causation of male infertility is being recognized.
416 citations
TL;DR: For cases involving excessive intracellular ROS generation, albumin appears to be an important means of neutralizing lipid peroxide-mediated damage to the sperm plasma membrane and DNA.
Abstract: Exposure of human spermatozoa to nicotinamide adenine dinucleotide phosphate (NADPH) resulted in the dose dependent generation of reactive oxygen species (ROS) which, at a critical level of intensity, induced lipid peroxidation, DNA damage and a dramatic decline of sperm motility. This system was then used as a model for screening the ability of different antioxidants to combat oxidative stress created through the excessive intracellular generation of toxic oxygen products of metabolism. A variety of antioxidants that has previously been shown to be protective against extracellularly derived oxidants (e.g. superoxide dismutase, catalase, vitamin E, hypotaurine) were ineffective in this system. Albumin, however, could provide complete protection against NADPH induced oxidative stress via mechanisms that did not involve the suppression of the lipid peroxidation cascade but rather the inactivation of lipid peroxides generated during this process. Albumin did not protect against DNA damage induced by NADPH but was extremely effective at preventing DNA fragmentation arising from the suppression of glutathione peroxidase activity with mercaptosuccinate. These studies emphasize that the design of clinically effective antioxidant treatments will depend, critically, upon the source of the oxidative stress. For cases involving excessive intracellular ROS generation, albumin appears to be an important means of neutralizing lipid peroxide-mediated damage to the sperm plasma membrane and DNA.
388 citations
TL;DR: The data show that oocyte quality and pronuclear embryo morphology are related to implantation and that pronuclear embryos can be successfully selected for embryo transfer.
Abstract: A retrospective analysis of results from 114 initiated in-vitro fertilization cycles utilizing pronuclear embryo transfer is presented. Patients were unselected for age or infertility criteria, constituted a continuous series and were grouped according to response to stimulation (Group 1, ideal; Group 2, suboptimal) or ovarian reserve (Group 3, poor). At 16-18 h post-insemination, embryos were scored for alignment of pronuclei and nucleoli and the appearance of the cytoplasm, generating an embryo score (ES). Transfers were performed 24-26 h post-insemination using two to six embryos with the highest ES. A corrected score was calculated (total score/number of embryos; CS). A total of 114 initiated cycles resulted in 97 oocyte retrievals with 38 clinical pregnancies (39%; 15% implantation). Pregnancy rates were significantly different between the three groups; 37 pregnancies in Group 1 (55% clinical pregnancy; 20% implantation), none in Group 2 and one in Group 3 (6%; 2% implantation: P 15 resulted in a 28% implantation; 65% delivery rate. CS <14 resulted in four pregnancies, one delivered. The data show that oocyte quality and pronuclear embryo morphology are related to implantation and that pronuclear embryos can be successfully selected for embryo transfer.
TL;DR: The results indicate that human sperm chromatin becomes cross-linked under conditions of oxidative stress and exhibits increased DNA strand breakage, yet the rate of pronucleus formation is no different from that of untreated control cells.
Abstract: We present the first evidence that genetically damaged human spermatozoa are able to form normal pronuclei in oocytes after intracytoplasmic sperm injection (ICSI). The role of reactive oxygen species (ROS) as a cause of chromatin and DNA damage is well recognized. The same class of molecule can be found in the semen of males with severe infertility, who remained infertile until the advent of ICSI. In this study we have investigated the role of ROS in the induction of chromatin damage, DNA strand breakage and the subsequent ability of spermatozoa to decondense and form pronuclei after ICSI. Spermatozoa from normozoospermic men participating in our research programme were exposed to oxidizing environments created by co-incubation with hydrogen peroxide, reduced nicotinamide adenine dinucleotide phosphate (NADPH) or activated white cells. The subsequent ability of the spermatozoa to decondense in vitro was examined using sequential incubations in EDTA, dithiothreitol and sodium dodecyl sulphate, and the amounts of DNA strand breakage were assessed using an in-situ nick translation protocol. Finally, cells exposed to hydrogen peroxide, NADPH and activated leukocytes were microinjected into hamster oocytes, and their ability to decondense and form normal pronuclei was determined. The results indicate that human sperm chromatin becomes cross-linked under conditions of oxidative stress and exhibits increased DNA strand breakage, yet the rate of pronucleus formation is no different from that of untreated control cells. The ability of genetically damaged spermatozoa to achieve normal fertilization following ICSI has implications for the practice of this form of assisted conception therapy.
TL;DR: It is shown that FSH is required for follicle growth beyond the two-layer stage, although growth initiation is independent of gonadotrophin stimulation, and xenograft models are potentially useful for studying human follicle development.
Abstract: In contrast to the many detailed studies of Graafian follicles, the biology of small follicles in the human ovary is poorly understood and the trigger for follicular growth initiation remains unknown. No practical model exists to study preantral follicle growth in the human because of their slow growth rate and lack of an effective culture system. We therefore tested ovarian xenografts as a new strategy to study the early stages of ovarian follicular growth in vivo. Mice homozygous for severe combined immunodeficiency (SCID) and hypogonadism (hpg) received human ovarian xenografts under their kidney capsules. Follicle growth was assessed by morphology and proliferating cell nuclear antigen (PCNA) immunostaining. The grafts were recovered after 11 (short-term) and 17 weeks (long-term), and serially sectioned. During the last 6 weeks of long-term grafting, mice were randomized to receive either placebo or 1 IU of purified follicle stimulating hormone (FSH) s.c. on alternating days. After 11 weeks of grafting, the most advanced follicles had a maximum of two granulosa cell layers. In the absence of FSH administration, follicles did not progress beyond the two-layer stage even after 17 weeks of grafting, and the oestradiol levels remained undetectable. In the FSH-treated long-term grafts, follicles had grown to antral stages and resulted in oestradiol levels as high as 2070 pmol/l. Growth initiation indices did not differ between control and FSH-treated grafts. This study demonstrates that follicles can survive and grow in human ovarian tissue grafted under the renal capsules of immunodeficient mice for at least 17 weeks, and indicate that xenograft models are potentially useful for studying human follicle development. Using this physiological model, we showed that FSH is required for follicle growth beyond the two-layer stage, although growth initiation is independent of gonadotrophin stimulation.
TL;DR: A multicentre study was carried out in seven top French centres for laparoscopic gynaecological surgery, in which 29,966 diagnostic and operative laparoscopies were performed, with a parallel and statistically significant increase in the rate of urological complications.
Abstract: A multicentre study was carried out in seven top French centres for laparoscopic gynaecological surgery. This series covers a period of 9 years, in which 29 966 diagnostic and operative laparoscopic operations were performed. The risk of complications has been assessed according to the complexity of the laparoscopic procedure in question. The means of diagnosis and treatment of the complications have been analysed, together with the importance of the surgeon's degree of experience. The mortality rate was 3.33 per 100 000 laparoscopies. The overall complication rate was 4.64 per 1000 laparoscopies (n = 139). The rate of complications requiring laparotomy was 3.20 per 1000 (n = 96). The complication rate was significantly correlated with the complexity of the laparoscopic procedure (P = 0.0001). One in three complications (34.1%; n = 43) occurred while setting up for laparoscopy, and one in four (28.6%) were not diagnosed during the operation. As new indications for laparoscopic surgery in gynaecology have appeared, there has been a parallel and statistically significant increase in the rate of urological complications (P = 0.001). Increased experience by the surgeons has had three consequences: a statistically significant drop in the number of bowel injuries (P = 0.0003), a drop in the rate of complications requiring laparotomy for those laparoscopic surgical procedures that are well defined (P = 0.01), and a change in the way complications are treated, with a significant increase in the proportion of incidents treated by laparoscopy (P = 0.0001).
TL;DR: The acute stress resulting from a great natural catastrophe can be a cause of a low sex ratio at birth 9 months later.
Abstract: We investigated the possible association between the Kobe earthquake (January 1995) and the sex ratio among live-born infants after the catastrophe. A significant decline in the sex ratio (0.501) of Hyogo Prefecture in October 1995 was observed 9 months after the Kobe earthquake as compared with an expected value of 0.516 in the period from January 1993 to January 1996 (P = 0.04; one-tailed). Simultaneously, a reduction in fertility of approximately 6% was also observed, compared with the month of October 2 years previously. Thus, the acute stress resulting from a great natural catastrophe can be a cause of a low sex ratio at birth 9 months later.
TL;DR: The finding of attenuated placental 11beta-HSD2 activity in IUGR suggests that glucocorticoids may, in part, contribute to impaired fetal growth and that this is closely controlled in normal gestation through placental 12beta- HSD2 expression.
Abstract: The type 2 isoform of 11beta-hydroxysteroid dehydrogenase (11beta-HSD2), which inactivates cortisol (F) to cortisone (E), has been suggested to play a role in the ontogeny of the fetal pituitary-adrenal axis and also protect the developing fetus from the deleterious effects of circulating maternal glucocorticoids. The abundance of 11beta-HSD2 in the placenta and other fetal tissues was inferred from the F/E ratio in 17 term deliveries in both umbilical arterial (1.73 +/- 0.24, mean +/- SE) and umbilical venous blood (1.16 +/- 0.14) compared with adult peripheral venous blood (7.76 +/- 0.57, n = 70). Using sensitive assays for 11beta-HSD2 and an in-house human 11beta-HSD2 antibody, the expression and activity of this enzyme in fresh frozen human placenta increased progressively from first (8-12 weeks, n = 16) and second (13-20 weeks, n = 9) to third trimester (term) pregnancies (39-40 weeks, n = 50). Placental 11beta-HSD2 activity was significantly reduced in deliveries complicated by intrauterine growth restriction (IUGR) [25-36 weeks, n = 12, activity 380 pmol/mg/h median (225-671; 95% confidence interval)], compared with the term deliveries [888 (725-1362)] and with appropriately grown pre-term deliveries [27-36 weeks, n = 14, activity 810 (585-1269)], P < 0.05. In human pregnancy placental 11beta-HSD2 activity increases markedly in the third trimester of pregnancy at a time when maternal circulating levels of glucocorticoid are rising. The finding of attenuated placental 11beta-HSD2 activity in IUGR suggests that glucocorticoids may, in part, contribute to impaired fetal growth and that this is closely controlled in normal gestation through placental 11beta-HSD2 expression.
TL;DR: It is shown that early cleavage to the two-cell stage is not influenced by the timing of fertilization, and is more likely due to intrinsic factors within the oocyte or embryo that promote embryo cleavage after fertilization.
Abstract: In-vitro fertilization (IVF) embryos are selected for transfer on the basis of morphology and rate of development. However, when a number of embryos have similar characteristics, the selection of the best embryos is left to chance. Recently, we proposed a simple, novel method to overcome this problem, based on pre-selection of embryos cleaving early to the two-cell stage. In this study we have adopted the same method to choose embryos fertilized after intracytoplasmic sperm injection (ICSI). Fertilized embryos that had cleaved to the two-cell stage by 27 h post-injection were designated as 'early cleavage' embryos, while those that had not yet reached the two-cell stage were designated as 'no early cleavage'. In all cases, the early cleavage embryos were transferred when available. Early cleavage was observed in 54 (61.4%) of the 88 cycles assessed. There were significantly (P = 0.04) more clinical pregnancies in the early cleavage group, 14/54 (25.9%), compared with the no early cleavage group 2/34 (3.2%). No differences between the groups were found when comparing key parameters (age, stimulation protocol and semen characteristics) of the couples. Using the ICSI technique, we have shown that early cleavage to the two-cell stage is not influenced by the timing of fertilization, and is more likely due to intrinsic factors within the oocyte or embryo that promote embryo cleavage after fertilization.
TL;DR: The knowledge about the origin and mechanisms of non-disjunction in human autosomal trisomies 8, 13, 15, 16, 18, and 21, accumulated during the last decade is summarized by using DNA polymorphism analysis.
Abstract: Chromosomal aneuploidy is one of the major causes of pregnancy wastage. In this review we summarize the knowledge about the origin and mechanisms of non-disjunction in human autosomal trisomies 8, 13, 15, 16, 18, and 21, accumulated during the last decade by using DNA polymorphism analysis. Maternal meiosis I non-disjunction is the most important single class, but chromosome-specific patterns exist. For the acrocentric chromosomes 15 and 21, meiosis I errors predominate among the maternal errors, in contrast to trisomy 18 where meiosis II errors predominate. For trisomy 16, virtually all cases are due to maternal meiosis I non-disjunction. Postzygotic (mitotic) non-disjunction constitutes 5-15% of cases of trisomies 15, 18, and 21, whereas for trisomy 8 and trisomy 8 mosaicism the majority of cases are due to mitotic non-disjunction. For paternal non-disjunction of chromosomes 18 and 21, meiosis II or mitotic errors predominate. There is aberrant meiotic recombination associated with maternal meiotic non-disjunction in all trisomies studied in detail so far. Advanced maternal age remains the only well documented risk factor for maternal meiotic non-disjunction, but there is, however, still a surprising lack of understanding of the basic mechanism(s) behind the maternal age effect.
TL;DR: The regimen appears as effective, in terms of high complete abortion rate and low continuing pregnancy rate, as any published alternative, and has the benefit of being less costly as the dose of mifepristone is 67% lower and misoprostol is substantially less expensive than gemeprost.
Abstract: A combination of the anti-progesterone mifepristone and gemeprost provides an effective non-surgical method for the induction of abortion at gestations up to 63 days, achieving complete abortion rates of over 95%. We report our experience with an alternate regimen, comprising a reduced dose of mifepristone in combination with vaginal misoprostol. A consecutive series of 2000 women requesting early medical abortion at gestations up to 63 days was studied retrospectively. Each woman received mifepristone 200 mg orally, followed 36-48 h later by misoprostol 800 microg vaginally. Of the 2000 women, 39 (2.0%) aborted completely following administration of mifepristone alone and a further 1912 experienced complete abortion following administration of misoprostol (a complete abortion rate of 97.5%). Surgical intervention was required in 49 women (2.5%): for incomplete abortion in 27 (1.4%), for missed abortion in seven (0.4%), for continuing pregnancy in 11 (0.6%) and to exclude ectopic pregnancy in four (0.2%). The surgical intervention rate was significantly higher among women at gestations > or = 49 days than among those at < or = 49 days (3.3 versus 1.5%, P = 0.0193). The regimen appears as effective, in terms of high complete abortion rate and low continuing pregnancy rate, as any published alternative. This regimen has the benefit of being less costly as the dose of mifepristone is 67% lower and misoprostol is substantially less expensive than gemeprost. Additionally, misoprostol does not require special transport or storage requirements. As such, the combination of mifepristone and misoprostol may be preferable to mifepristone and gemeprost.
TL;DR: It is postulate that an age- related decline in the process of folliculogenesis results in reduced oocyte quality and that the well characterized age-related increase in meiotic non-disjunction is one symptom of compromised oocyte growth.
Abstract: The human oocyte appears to be particularly prone to meiotic errors, and the incidence of these errors is strongly influenced by maternal age. We have initiated studies of human oocytes from unstimulated ovaries and have observed age-related effects on the meiotic process in oocytes from unselected antral follicles. Specifically, in oocytes obtained from donors over the age of 35 years, the majority of oocytes that extruded a first polar body in culture and arrested at second meiotic metaphase had aberrations in spindle formation and chromosome alignment. Similarly, observations of a limited number of oocytes at first meiotic metaphase suggest disturbances at this stage of meiosis as well. Finally, preliminary results of non-disjunction studies suggest that the frequency of errors in chromosome segregation at the first meiotic division is influenced by donor age in in-vitro matured oocytes as it is in oocytes undergoing meiotic maturation in vivo. These data provide direct evidence that the meiotic competence of oocytes from unstimulated ovaries declines with donor age. Similarly, studies of in-vitro fertilization (IVF) pregnancies in older women indicate that the developmental competence of the human oocyte declines with age. Since both meiotic and developmental competence are acquired during the late stages of oocyte growth, we postulate that an age-related decline in the process of folliculogenesis results in reduced oocyte quality and that the well characterized age-related increase in meiotic non-disjunction is one symptom of compromised oocyte growth.
TL;DR: Results show that spare ICSI embryos have significantly lower rates of development to the blastocyst stage compared with those developed after routine IVF and that a greater understanding of the molecular basis of male infertility is needed to broaden knowledge on the effect that abnormal spermatozoa have on fertilization and embryo development.
Abstract: In the first part of this report we investigate whether chromatin anomalies in human spermatozoa can influence fertilization after intracytoplasmic sperm injection (ICSI). We have examined the sperm chromatin packaging quality using the chromomycin A3 (CMA3) fluorochrome and the presence of DNA damage in spermatozoa using in-situ nick translation. When comparing the spermatozoa of patients undergoing in-vitro fertilization (IVF) and ICSI distinct differences are evident in that ICSI males have a higher CMA3 fluorescence, indicating spermatozoa with loosely packed chromatin, and more spermatozoa containing endogenous DNA nicks. When examining the unfertilized oocytes of ICSI patients we found that men who had a high percentage of anomalies in their chromatin, i.e. > 30% CMA3 fluorescence and > 10% nicks, had more than double the number of unfertilized oocytes containing spermatozoa that had remained condensed. The observation that failed fertilized oocytes, injected with spermatozoa from patients with a higher percentage of sperm nuclear anomalies, contain more condensed spermatozoa indicates that a selection process against these spermatozoa may be in place at the time of fertilization. In the second part of the study we show that spare ICSI embryos have significantly lower rates of development to the blastocyst stage compared with those developed after routine IVF. These results show that a greater understanding of the molecular basis of male infertility is therefore needed to broaden our knowledge on the effect that abnormal spermatozoa have on fertilization and embryo development.
TL;DR: In this paper, the effects of antioxidant supplementation in vitro on endogenous DNA damage in spermatozoa using the single cell gel electrophoresis (comet) assay and, second, to assess the effect of antioxidant supplements given prior to X-ray irradiation on induced DNA damage.
Abstract: The integrity of sperm DNA is crucial for the maintenance of genetic health. A major source of damage is reactive oxygen species (ROS) generation; therefore, antioxidants may afford protection to sperm DNA. The objectives of the study were, first, to measure the effects of antioxidant supplementation in vitro on endogenous DNA damage in spermatozoa using the single cell gel electrophoresis (comet) assay and, second, to assess the effect of antioxidant supplementation given prior to X-ray irradiation on induced DNA damage. Spermatozoa from 150 patients were prepared by Percoll centrifugation in the presence of ascorbic acid (300, 600 microM), alpha tocopherol (30, 60 microM), urate (200, 400 microM), or acetyl cysteine (5, 10 microM). DNA damage was induced by 30 Gy X-irradiation. DNA strand breakage was measured using the comet assay. Sperm DNA was protected from DNA damage by ascorbic acid (600 microM), alpha tocopherol (30 and 60 microM) and urate (400 microM). These antioxidants provided protection from subsequent DNA damage by X-ray irradiation. In contrast, acetyl cysteine or ascorbate and alpha tocopherol together induced further DNA damage. Supplementation in vitro with the antioxidants ascorbate, urate and alpha tocopherol separately has beneficial effects for sperm DNA integrity.
TL;DR: It appears that the predictive value of embryo morphology on day 3 for subsequent blastocyst formation is limited.
Abstract: The transfer of blastocysts has been associated with a very high implantation rate. However, not all embryos achieve the blastocyst stage. Our study was set up to demonstrate whether embryo morphology on day 3 predicts subsequent blastocyst formation. A prospective study was carried out in 48 patients with a mean of 2.9 failed in-vitro fertilization (IVF) attempts. In this new cycle, the morphology of the embryos on day 3 was noted. After pre-selection of the embryos which would have been transferred on day 3, all embryos were cultured individually and allowed to develop further until transfer on day 5. The clinical pregnancy rate per transfer was 46%, and the overall implantation rate was 24%. When only blastocysts were transferred the pregnancy rate was 53% with an implantation rate of 30%. Thirty-nine per cent of all embryos reached the blastocyst stage on day 5; 47% of class 1 and 2 embryos (good quality) in contrast to 21% of class 3 and 4 embryos (poor quality). Respectively 45% of class 1 and 2 embryos and 69% of class 3 and 4 embryos arrested in development or degenerated. Only 51% of the embryos that were transferred on day 5 had been pre-selected for transfer on day 3. In conclusion, it appears that the predictive value of embryo morphology on day 3 for subsequent blastocyst formation is limited.
TL;DR: In this article, the authors investigated the effect of supplementation of the medium with gonadotrophins, oestradiol and epidermal growth factor (EGF) on the nuclear and cytoplasmic maturation of immature oocytes.
Abstract: In-vitro maturation (IVM) of oocytes is a promising technique to reduce the costs and avert the side-effects of gonadotrophin stimulation for in-vitro fertilization (IVF). The pregnancy rates from oocytes matured in vitro are much lower than those of in-vivo stimulation cycles indicating that optimization of IVM remains a challenge. Therefore, we investigated the effect of supplementation of the medium with gonadotrophins, oestradiol and epidermal growth factor (EGF) and the effect of retaining or removing the cumulus cells on nuclear and cytoplasmic maturation of immature oocytes. Human germinal vesicle (GV) oocytes obtained after gonadotrophin stimulation for intracytoplasmic sperm injection (ICSI) were cultured in a complex defined medium either supplemented with gonadotrophins, oestradiol and physiological concentrations of EGF (2 ng/ml) or gonadotrophins and oestradiol alone. The cumulus cells were either removed or kept intact. In GV stage oocytes cultured without cumulus (group I) significantly more oocytes reached the metaphase II (MII) stage at 30 h in media supplemented with EGF (64.3 versus 33.9%, P < 0.003). For oocytes cultured with intact cumulus (group II), more oocytes reached MII at 30 h than in group I, but there was no difference in medium with or without EGF supplementation (81.8 and 79.8% respectively). Cytoplasmic maturation of MII oocytes was judged from their capability to activate and fertilize after ICSI. In group I, the rates of activation and normal fertilization were similar. However, in group II, significantly more oocytes underwent normal fertilization in the EGF-supplemented than the unsupplemented group (71.7 versus 45.6%, P < 0.05). The cleavage rates of the fertilized oocytes were similar in the sibling oocyte subgroups cultured with or without EGF supplementation, but the overall cleavage rates were higher in cumulus-intact compared to cumulus-denuded oocytes (88.9 versus 47.8%, P < 0.001). Thus, supplementation of the maturation medium with EGF and maintenance of the cumulus during culture improve the nuclear and cytoplasmic maturation of human oocytes in vitro.
TL;DR: As the implantation rate of blastocysts is higher than that of the cleavage stage embryo, fewer embryos will be required for transfer in order to establish a successful pregnancy, thereby reducing the number of multiple gestations and increasing the overall efficiency of human IVF.
Abstract: In human in-vitro fertilization (IVF), embryos are routinely transferred to the uterus on either day 2 or day 3 of development, resulting in a 10-15% implantation rate. However, in other mammalian species, the transfer of cleavage stage embryos, which normally reside in the oviduct, to the uterus results in a significantly lower implantation rate compared with blastocysts. It is therefore proposed that, in order to increase implantation rates in human IVF, one has to move to extended culture and transfer at the blastocyst stage. The transfer of blastocysts will not only help synchronize the embryo with the female tract but will facilitate the identification of those embryos with little or no developmental potential. In order to culture viable blastocysts it is important to use more than one culture medium to cater for the changing requirements of the preimplantation embryo as it develops and differentiates. If sequential culture media are not used, one can obtain blastocysts but their resultant viability is low. The use of sequential serum-free media in human IVF has resulted in > 50% of embryos becoming blastocysts with an implantation rate of approximately 50%. Further advances in human embryo culture should come from the replacement of protein with the glycosaminoglycan hyaluronate, which is more suitable than albumin in supporting implantation in the mouse, and which will eliminate biological variation and possible contamination from blood products. With the routine culture of human blastocysts will come the introduction of non-invasive tests of embryo viability, capable of identifying those blastocysts most likely to develop from a given cohort. As the implantation rate of blastocysts is higher than that of the cleavage stage embryo, fewer embryos will be required for transfer in order to establish a successful pregnancy, thereby reducing the number of multiple gestations and increasing the overall efficiency of human IVF.
TL;DR: It is anticipated that further improvements in embryo development and the selection of viable embryos can be achieved using this simple and effective culture system.
Abstract: A cell-free culture system was designed for human embryo development to the blastocyst stage by testing a range of culture conditions in a series of protocols. The culture system that was evolved has a brief 1 h exposure to spermatozoa and then culture of the pronucleate zygote for 2 days in IVF-50 medium. Two or three embryos were cultured together in 20 microl microdrops of medium under oil. Embryos were then regrouped and two or three at a similar stage were cultured together in 50 microl microdrops of Gardner's G2 medium under oil from days 3 to 5. Embryos were transferred to fresh G2 medium on day 5 and cultured for a further 1 or 2 days (day 6 or 7). No serum was used in any of the cultures. The embryo transfer medium and G2 medium were supplemented with human serum albumin. The zonae of all blastocysts to be transferred to patients were completely removed enzymatically. Using this protocol, 52% of zygotes developed to blastocysts and 34 out of 35 patients treated received 82 blastocysts and 11 morulae on day 5 or 6. Twenty-one fetal sacs with positive heartbeats (23% implantation rate) were detected in 13 ongoing pregnancies (38% pregnancy rate/transfer or 37%/patient treated). We anticipate that further improvements in embryo development and the selection of viable embryos can be achieved using this simple and effective culture system.
TL;DR: Survival, assessed by the morphology, was higher in 4-cell embryos on day 2 and 8-cell embryo on day 3 than in 2-3- cell embryos onday 2 or 2-7-cell babies on day3 and higher survival rates were obtained with more permeating conditions, suggesting that these cryoprotectants are considerably toxic.
Abstract: Experiments were conducted to find a suitable cryoprotectant and suitable procedure for vitrification of 8-cell mouse embryos. The method was then applied clinically to the cryopreservation of human embryos in our assisted reproduction programme. Mouse embryos were vitrified with 30 or 40% 1,2-propanediol (PROH), dimethylsulphoxide (DMSO), ethylene glycol, glycerol, or acetamide, each diluted with a solution containing 30% Ficoll plus 0.5 M sucrose. Embryos were exposed to the solutions for 0.5 or 2 min at 20 or 25 degrees C, cooled in liquid nitrogen and warmed rapidly. Embryo survival was assessed by in-vitro development. In PROH-, DMSO- and acetamide-based solutions, higher survival rates (29-82%) were obtained with less permeating conditions, suggesting that these cryoprotectants are considerably toxic. In glycerol- and ethylene glycol-based solutions, however, higher survival rates (74 and 92% respectively) were obtained with more permeating conditions, suggesting that these cryoprotectants are less toxic. Human embryos on days 2-3 were vitrified in an ethylene glycol-based solution (EFS40). Survival, assessed by the morphology, was higher in 4-cell embryos on day 2 and 8-cell embryos on day 3 than in 2-3-cell embryos on day 2 or 2-7-cell embryos on day 3. From 18 transfers, one ended with the delivery of healthy twin babies.
TL;DR: It is demonstrated that a large percentage of idiopathic SCOS may be genetically determined and an Y-related region that seems to possess one or more still unknown genes essential for spermatogenesis is identified.
Abstract: Idiopathic Sertoli cell-only syndrome (SCOS) is characterized by azoospermia, small testes, absence of germ cells in the testes, elevated follicle stimulating hormone and normal testosterone concentrations. The Y-chromosome is involved in the regulation of spermatogenesis and in the pathogenesis of a fraction of idiopathic male infertility. An azoospermia factor (AZF) is present on the Y-chromosome long arm euchromatic region (Yq11) and two gene families (DAZ and RBM) have been identified within this region. The aim of this study was to investigate whether a specific pattern of Yq11 microdeletions may be associated with idiopathic SCOS. Eighteen idiopathic subjects showing a testicular cytological picture of bilateral SCOS were selected and tested by polymerase chain reaction for a set of 29 Y-specific sequence-tagged sites (STS). We found Yq microdeletions in 10 out of 18 patients (55.5%) while the fathers or brothers of six out of 10 patients deleted for Yq were shown to carry an intact Y-chromosome. These deletions may therefore be considered as de-novo deletions and the cause of SCOS. The analysis of the microdeletions allowed us to identify two homogeneous regions that have a high incidence of deletion. The smallest deletion, common to all patients, is located in Yq interval 5. We therefore speculate that there is a relationship between specific, well-characterized Yq11 microdeletions and a testicular picture of SCOS, identifying an Y-related region frequently deleted in this syndrome. In conclusion, the findings of this study demonstrate that a large percentage of idiopathic SCOS may be genetically determined and identify an Y-related region that seems to possess one or more still unknown genes essential for spermatogenesis.
TL;DR: This study confirms previous reports that have postulated that abnormal spermatozoa may manifest a negative paternal effect on preimplantation embryo development.
Abstract: The success of intracytoplasmic sperm injection (ICSI) warrants further study on the role of paternal factors in early human embryogenesis. To investigate whether poor sperm parameters can influence embryo development, we examined the development of ICSI-fertilized embryos to the blastocyst stage. We present results of blastocyst development from supernumerary ICSI embryos after co-culture on monkey kidney epithelial cells. In addition, we compare the development of supernumerary embryos to the blastocyst stage after ICSI and in-vitro fertilization (IVF). Of 168 supernumerary ICSI embryos, 45 (26.8%) developed to blastocysts. Sperm concentration and morphology did not influence blastocyst development. In contrast, blastocysts arose from spermatozoa that had a significantly higher (P = 0.015) forward progressive motility compared with spermatozoa from those patients who failed to produce blastocysts (42.7% versus 28.2%, respectively). Overall the rate of embryo development to the blastocyst stage after ICSI was lower (26.8%) than that after IVF (47.3%). When the rate of blastocyst development was calculated for patients with three or more supernumerary embryos, it remained significantly higher for the IVF patients than for the ICSI patients (45.6% versus 30.0%). There was no significant difference in the mean cell number and quality of the supernumerary embryos between the IVF and ICSI patients. This study confirms previous reports that have postulated that abnormal spermatozoa may manifest a negative paternal effect on preimplantation embryo development.
TL;DR: Results are interpreted as evidence that matching for the entire 16-locus haplotype and/or alleles at an HLA-B-linked locus confers significant risk for fetal loss.
Abstract: The role that maternal and fetal human leukocyte antigen (HLA) genes play in pregnancy is unknown, but it has been suggested that fetuses whose HLA alleles do not differ from maternal alleles (i.e. histocompatible fetuses) are more likely to be aborted than fetuses with HLA alleles that differ from maternal alleles (i.e. histoincompatible fetuses). To elucidate the role of HLA compatibility in pregnancy, we tested the hypothesis that couples who match for HLA alleles or haplotypes would have reduced fertility because only these couples could produce histocompatible fetuses. We conducted a 10 year prospective study of HLA matching and pregnancy outcome in 111 Hutterite couples, providing information on 251 pregnancies. A logistic regression analysis was performed to determine the effects of HLA matching at HLA regions and loci on pregnancy outcome (fetal loss versus delivery). Significantly increased fetal loss rates were observed among couples matching for the entire 16-locus haplotype (P = 0.002). Among the individual loci, loss rates were increased among couples matching for HLA-B (P = 0.019), HLA-C (P = 0.033) and the complement component, C4 (P = 0.043). We interpret these results as evidence that matching for the entire 16-locus haplotype and/or alleles at an HLA-B-linked locus confers significant risk for fetal loss.