Human Reproduction Update

Published by Oxford University Press (OUP)
Online ISSN: 1460-2369
Print ISSN: 1355-4786
Human development in the first 8 weeks is potentially one of the most exciting areas of biologic research. Beyond historic staging of fixed human embryos, it is also one of the least understood. In contrast, detailed information exists for the embryonic period of several other species, from which human development information is extrapolated. This period is also the most sensitive to system abnormalities generated by teratogens. This review combines the human embryo Carnegie stages, available online at UNSW Embryology (, with teratogen-sensitive information. Integrating this data with current molecular, imaging, and online tools will provide insights to this period.
PRISMA flow chart for identification of studies on PE and probability of pregnancy after IVF.
Meta-regression plot of PE threshold on the logit of PE rates for the studies included in this systematic review. Each study or study subgroup contributed data for only one PE threshold in this analysis. (circle size is proportional to the study weight) 
Forest plots of ORs for pregnancy achievement in women with PE when compared with those without PE (all studies analysed). CI, confidence interval.
BACKGROUND The aim of this meta-analysis was to evaluate the association of progesterone elevation (PE) on the day of hCG administration with the probability of pregnancy in fresh, frozen-thawed and donor/recipient IVF cycles.METHODSA literature search in MEDLINE, SCOPUS, COCHRANE CENTRAL and ISI Web of Science was performed aiming to identify studies comparing the probability of pregnancy in patients with or without PE after ovarian stimulation with gonadotrophins and GnRH analogues. Standard meta-analytic methodology was used for the synthesis of results and meta-regression for exploration of heterogeneity.RESULTSSixty-three eligible studies were identified evaluating 55 199 fresh IVF cycles, nine studies evaluating 7229 frozen-thawed cycles and eight studies evaluating 1330 donor/recipient cycles. In fresh IVF cycles, a decreased probability of pregnancy achievement was present in women with PE (when PE was defined using a threshold ≥0.8 ng/ml) when compared with those without PE. The pooled effect sizes were 0.8-1.1 ng/ml: odds ratio (OR) = 0.79; 1.2-1.4 ng/ml: OR = 0.67; 1.5-1.75 ng/ml: OR = 0.64; 1.9-3.0 ng/ml: OR: 0.68 (P < 0.05 in all cases). No adverse effect of PE on achieving pregnancy was observed in the frozen-thawed and the donor/recipient cycles.CONCLUSIONS Based on the analysis of more than 60 000 cycles, it can be supported that PE on the day of hCG administration is associated with a decreased probability of pregnancy achievement in fresh IVF cycles in women undergoing ovarian stimulation using GnRH analogues and gonadotrophins. On the other hand, an adverse effect of PE does not seem to be present in frozen-thawed and donor/recipient cycles.
Recently, one laser system has been introduced in IVF fulfilling all safety requirements, while achieving a high standard of reproducibility in terms of ablation diameter. This 1.48 microm wavelength indium-gallium-arsenic-phosphorus (InGaAsP) semiconductor laser offers a variety of laser applications to the embryologist. On the one hand, zona pellucida of oocytes or embryos can be manipulated in order to facilitate ICSI or biopsy and assist hatching, and on the other, spermatozoa may be paralysed or immobilized prior to usage. To conclude, the 1.48 microm diode laser provides a promising tool for the microdissection of subcellular targets. The diode laser stands out due to the rapidity, the simplicity and the safety of the procedure which is supported by healthy offspring after laser application.
Morphology of BG01 and HUES-7 human embryonic stem cell (hESC) lines cultured under the proprietary standard protocols ( A , B ) and published feeder- and conditioned medium-free conditions ( C–H ). BG01 ( A ) and HUES-7 ( B ) cultured under proprietary standard conditions on feeders (Mitalipova et al ., 2003; Cowan et al ., 2004). BG01 ( C ) and HUES-7 ( D ) cultured according to Sato et al . (2004). Cultures of both cell lines could not be maintained due to extensive cell death. BG01 ( E ) and HUES-7 ( F ) cultured according to Vallier et al . (2005). Cultures of both cell lines could not be maintained for more than 10 days under these conditions. BG01 ( G ) and HUES-7 ( H ) cultured according to Klimanskaya et al . (2005). BG01 cultures could be maintained for at least 10 passages, whereas the same culture conditions were not suitable for HUES-7 maintenance beyond the first passage. Bar, 100 μ m. All brightfield images were taken using Hoffman objective lenses. Magnification 160 × . 
The promise of human embryonic stem cell (hESC) lines for treating injuries and degenerative diseases, for understanding early human development, for disease modelling and for drug discovery, has brought much excitement to scientific communities as well as to the public. Although all of the lines derived worldwide share the expression of characteristic pluripotency markers, many differences are emerging between lines that may be more associated with the wide range of culture conditions in current use than the inherent genetic variation of the embryos from which embryonic stem cells were derived. Thus, the validity of many comparisons between lines published thus far is difficult to interpret. This article reviews the evidence for differences between lines, focusing on studies of pluripotency marker molecules, transcriptional profiling, genetic stability and epigenetic stability, for which there is most evidence. Recognition and assessment of environmentally induced differences will be important to facilitate the development of culture systems that maximize stability in culture and provide lines with maximal potential for safety and success in the range of possible applications.
The improved long-term survival of adolescents and young women treated for cancer has resulted in an increased focus on the effects of chemotherapy on ovarian function and its preservation. These women may seek advice and treatment regarding their reproductive status, including ways of preserving their fertility and preventing a premature menopause--factors that can have a profound impact on their quality of life. This article comprehensively reviews ovarian reserve testing (ORT) in general. Special emphasis is placed on patients with cancer, including the pathophysiology of gonadal damage following chemotherapy, fertility preservation and the potential role of ORT. Baseline parameters of ovarian reserve [FSH LH, estradiol, inhibin B and anti-Mullerian hormone (AMH)] have not yet performed sufficiently well in predicting poor outcome in assisted reproduction, but biochemical markers of ovarian reserve appear to be better than chronological age. Inhibin B and AMH show potential for future use. Dynamic testing appears to show much promise, especially stimulated levels of inhibin B and estradiol. The most promising tests of ovarian reserve are the biophysical markers, where total antral follicle count was found to be most discriminatory followed by ovarian volume. Combination of biochemical, biophysical and clinical markers of ovarian reserve may also improve predictive capacity. However, there is a lack of data pertinent to ORT in cancer. As yet there is no single clinically useful test to predict ovarian reserve accurately. Patients with cancer represent a distinct cohort who have particular concerns about their future fertility and the possibility of a premature menopause, they can benefit greatly from knowledge of their functional ovarian reserve. Large, prospective, randomized, adequately controlled studies specific to different geographical areas are required in a control population of comparable reproductive age to determine the potential role of ORT in clinical practice.
Developmental pathways in humans and other organisms are buffered against changes in genotype and environment. Therefore, it should not come as a surprise that most of the children conceived by assisted reproduction technology (ART) are healthy, although ART bypasses a lot of biological filters and subjects the gametes and the early embryo to environmental stress. If, however, the buffer breaks down, the development of certain tissues or organs may follow abnormal trajectories. We argue that both normal and abnormal development in children conceived by ART can be explained by epigenetic mechanisms, which control the establishment and maintenance of gene expression patterns in the placenta and fetus. Imprinted genes are of special importance in this respect. There is increasing evidence that genetic factors in infertile couples as well as environmental factors (hormones and culture media) can have adverse effects on epigenetic processes controlling implantation, placentation, organ formation and fetal growth. In addition, loss of epigenetic control may expose hidden genetic variation.
Monozygotic twinning arising from division in the ®rst 4 days of embryo development (adapted from Fox, 1978). 
Embryo division after the ®rst week of development demonstrates greater sharing of trophoblastic-derived tissue and amniotic sacs (adapted from Fox, 1978). 
Ultrasonic image of a diamniotic, dichorionic pregnancy at 8 weeks gestation. 
Ultrasonic image of a monoamniotic, monochorionic pregnancy at 10 weeks gestation. 
Monozygotic twinning is a relatively rare event in in-vivo conception, being estimated to occur in 0.42% of all births. The underlying mechanism for monozygotic twin formation is the division of the embryo early in its development. Separation of cells may theoretically occur before or after inner cell mass formation. In this analysis we report 11 cases of monozygotic twins resulting from IVF or intracytoplasmic sperm injection (ICSI) treatment cycles performed between 1991 and 1998 at St James's University Hospital, Leeds, and Bourn Hall Clinic, Cambridge, UK. Six cases (55%) followed treatment with ICSI and seven cases (64%) were in women aged > or = 35 years. This article also reviews the scientific literature discussing information pertaining to frequency, aetiology and potential complications of the monozygotic twinning phenomenon. We conclude that patients at risk of monozygotic twinning are those aged >35 years of age and those who had artificial opening in the zona pellucida by application of micromanipulation techniques. Women undergoing assisted conception treatment, particularly those with these two risk factors, must be informed of the low but definite risk of monozygotic twinning when counselled regarding the number of embryos to be transferred and chances of multiple births.
Forest plots for (a) facial defects, (b) eye defects only, and (c) cleft lip or palate. Studies are ranked according to size of the odds ratio.  
BACKGROUND There is uncertainty over whether maternal smoking is associated with birth defects. We conducted the first ever comprehensive systematic review to establish which specific malformations are associated with smoking. METHODS Observational studies published 1959–2010 were identified (Medline), and included if they reported the odds ratio (OR) for having a non-chromosomal birth defect among women who smoked during pregnancy compared with non-smokers. ORs adjusted for potential confounders were extracted (e.g. maternal age and alcohol), otherwise unadjusted estimates were used. One hundred and seventy-two articles were used in the meta-analyses: a total of 173 687 malformed cases and 11 674 332 unaffected controls. RESULTS Significant positive associations with maternal smoking were found for: cardiovascular/heart defects [OR 1.09, 95% confidence interval (CI) 1.02–1.17]; musculoskeletal defects (OR 1.16, 95% CI 1.05–1.27); limb reduction defects (OR 1.26, 95% CI 1.15–1.39); missing/extra digits (OR 1.18, 95% CI 0.99–1.41); clubfoot (OR 1.28, 95% CI 1.10–1.47); craniosynostosis (OR 1.33, 95% CI 1.03–1.73); facial defects (OR 1.19, 95% CI 1.06–1.35); eye defects (OR 1.25, 95% CI 1.11–1.40); orofacial clefts (OR 1.28, 95% CI 1.20–1.36); gastrointestinal defects (OR 1.27, 95% CI 1.18–1.36); gastroschisis (OR 1.50, 95% CI 1.28–1.76); anal atresia (OR 1.20, 95% CI 1.06–1.36); hernia (OR 1.40, 95% CI 1.23–1.59); and undescended testes (OR 1.13, 95% CI 1.02–1.25). There was a reduced risk for hypospadias (OR 0.90, 95% CI 0.85–0.95) and skin defects (OR 0.82, 0.75–0.89). For all defects combined the OR was 1.01 (0.96–1.07), due to including defects with a reduced risk and those with no association (including chromosomal defects). CONCLUSIONS Birth defects that are positively associated with maternal smoking should now be included in public health educational materials to encourage more women to quit before or during pregnancy.
Cancer registry studies 
Summary estimates of risk increase per year in three HT groups, stratified by region 
Knowledge about the impact of menopausal hormone therapy (MHT) on the risk of ovarian cancer (OvC) is insufficient, and studies are inconsistent. Mortality from OvC ranks highest among cancer sites in female reproductive organs. We performed meta-analyses to assess the impact of specified types of MHT on the risk of OvC in cohort studies (CS), case-control studies (CCS), randomized controlled trials (RCT) and cancer registry studies (CRS). We used data published 1966-2006 on estrogen therapy (ET), estrogen/progestin therapy (EPT) or MHT (unspecified regimen) identified by a structured, computerized and manual literature search. We identified 42 studies (30CCS, 7CS, 1 RCT and 4 CRS) with 12 238 cases. The risk of OvC (ever-use, annual risk) is increased 1.28-fold by ET [confidence interval (CI) 1.18-1.40] and 1.11-fold by EPT (CI 1.02-1.21) with a suggestion of greater risks with ET. There appears to be no differential impact of any therapy on histological subtypes. Risks were greater in European than North American studies for both ET and EPT. In conclusion, ET as well as EPT, are risk factors for OvC. Given the widespread use of MHT, known benefits should be weighed against the increased risk of OvC, and more studies are warranted, particularly on factors with the greatest apparent risks.
PP13: LGALS13 polymorphisms and localization in the placenta. (A) Like most galectins, the gene of PP13, LGALS13, is built of four exons (E1-E4) with the CRD on exon 4. The 5 ′ and 3 ′ ends are shown on the left and right side of the figure, respectively. The various mutations are marked by redlines and letters. Note particularly 221delT , the intron mutations between exons 2 and 3 that lead to the development of the spliced variant Dex2 and the promotor-98 A-to-C mutation. (B) Cloned recombinant wild-type PP13 (left), splice variant Dex-2 produced by intronic A-to-G replacement between exons 2 and 3 (middle) and the truncated version generated due to thymidine deletion in position 221 (221delT) (right). The three recombinant versions were generated by recombinant technology, expressed in E. coli and subsequently purified and separated with sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) (modified from Sammar et al., 2006). (C and D) PP13 immunostaining in a case of early pre-eclampsia (33 weeks of gestation). Note the disturbed surface of the syncytiotrophoblast showing evidence of necrotic release of subcellular fragments (arrows in C, D). (E and F) PP13 immunostaining in a normal term placenta. Note the smooth surface of the syncytiotrophoblast and the absence of staining in the cytotrophoblast (stars in F).
PP13 levels in pregnant women’s serum during pregnancy PP13 levels in maternal serum in normal pregnant women ( N 1⁄4 41; black line) and patients who developed late pre-eclampsia at term ( N 1⁄4 4; red line). Each woman has given 10 – 12 blood samples after enrolment during the first trimester. Data are represented as medians with 95% confidence interval of intervals of 2 weeks. Weeks are presented after last menstrual period. Accordingly, weeks 41 – 55 represent 1 – 15 weeks after delivery. The black and red lines at 40 weeks indicate mean week of delivery for both groups. The figure points to the presence of a ‘therapeutic window’ from early to midgestation, during which PP13 replenishment for keeping its level within the normal range may assist in proper placentation. Later in pregnancy, blocking the elevated PP13 level may be required to prevent complications. 
Background Pre-eclampsia affects 2-7% of all pregnant women and is a major cause of maternal and fetal morbidity and mortality. The etiology of pre-eclampsia is still unknown but it is well documented that impaired placentation is a major contributor to its development. One of the placenta-specific proteins is placental protein 13 (PP13). Lower first trimester levels of maternal serum PP13 and its encoding placental mRNA are associated with the development of both early and late-onset severe pre-eclampsia. In cases where this protein is mutated, the frequency of pre-eclampsia is higher.Methods19 out of 68 studies on PP13, published between January 2006 and September 2012, were used to evaluate the value of maternal blood PP13 as a marker of pre-eclampsia.ResultsA meta-analysis presented in this review shows that low serum levels of PP13 in the first trimester of pregnancy can predict the development of pre-eclampsia later in pregnancy. Although some functions of this protein have been assessed in in vitro experiments, the in vivo functions of PP13 are still unknown, especially when circulating in the maternal bloodstream. A recent pilot study has shown that in gravid rats PP13 causes significant vasodilatation, reduced blood pressure and increased maternal uterine artery remodeling.Conclusion Reviewing these effects of PP13, the authors propose the use of PP13 as a new drug candidate. Replenishing PP13 in those women with low serum levels early in pregnancy may help prepare their vasculature for pregnancy. This novel pharmacological approach to combat pre-eclampsia is presented as a new direction to transfer from individualized risk to personalized prevention.
BACKGROUND; Mitochondrial DNA (mtDNA) disorders are an important cause of human diseases. In view of the limitations of prenatal diagnosis and preimplantation genetic diagnoses, alternatives such as ooplasmic transfer (OT) and nuclear transfer (NT) have been proposed to prevent the transmission of mtDNA mutations. Both OT and NT are radical in the sense that they do not entail genetic selection, but genetic intervention to correct the genetic cause of the disease. After interviews with experts in the field, the relevant literature was searched and analyzed. Bioethical issues were divided into conceptual and normative points. OT is the transfer of normal mitochondria to a carrier's oocyte containing mutant mtDNA. In case of NT, a donated oocyte is enucleated and replaced with the nuclear DNA from a woman carrying a mtDNA mutation. NT can be performed both before and after in vitro fertilization, respectively, with the nucleus of an unfertilized oocyte, with the pronuclei of the zygote, or with the nucleus of a blastomere of an embryo. Conceptual questions regard whether these techniques amount to germ-line modification and human cloning. Normative questions concern, among others, the significance of intervening in the mtDNA, the implications of having 'three genetic parents', the ethics of oocyte donation and the health and safety risks for children conceived as a result of one of these techniques. Further interdisciplinary debate and research is needed to determine whether a clinical application of OT and NT can be morally justified, and if so, under which conditions.
Hormonal replacement therapy (HRT) is generally regarded as first choice for pharmacological prevention of osteoporosis in women. We reviewed recent studies of HRT regimens and selective oestrogen receptor modulators (SERMs), including controlled trials of at least one-year duration published since 1995 until February 2000 providing data on bone mineral density (BMD) or fractures. Natural and synthetic oestrogens exert a continuum of positive effects on BMD in a dose-dependent, though non-proportional, fashion independent of age and mode of administration. Bone loss may be largely prevented by 25 microg transdermal patch oestradiol, 0.3 mg conjugated equine or 0.3 mg esterified oestrogens. Progestogens neither attenuate nor augment the effect of oestrogens; sole use of tibolone prevents bone loss. Both the SERMs, tamoxifen and raloxifene, slightly increase BMD. There are no adequately powered fracture trials for any HRT regimen. Raloxifene 60 mg daily decreases the relative risk of vertebral fractures by at least 30%, as demonstrated by one 3-year fracture study of osteoporotic women. In conclusion, the recommendation to use oestrogen for postmenopausal osteoporosis, given both the lack of fracture trials and the rare trials on long-term use of HRT in (late) postmenopausal women, is not well supported. Fracture trials could overcome shortcomings of the current level of evidence.
The warning issued by the UK Committee on Safety Medicines in October 1995, followed by their 'Dear Doctor' letter of October 18, 1995, that oral contraceptive pills containing gestodene or desogestrel were associated with a higher risk of venous thromboembolism has had a negative impact on public heath. A significant number of women either switched brands or ceased contraception altogether following the announcement. National data suggest a strong association between the pill scare and a substantial increase in the number of unintended pregnancies, particularly significant among younger women, with use of oral contraception falling from 40 to 27% of under 16s between 1995-1996 and 1996-1997. The resulting cost of the increase in births and abortions to the National Health Service has been estimated at about Pound Sterling 21 million for maternity care and from Pound Sterling 46 million for abortion provision. The level of risk should, in future, be more carefully assessed and advice more carefully presented in the interests of public health.
Both the ability to freeze human spermatozoa and the possibility of pregnancy following intrauterine insemination have existed for >40 years. There have been a number of improvements during that time concerning the methods of freezing and thawing human spermatozoa. Initially, the use of the cryoprotective properties of glycerol allowed a major improvement; subsequently, changes were mainly empirical. It was a long time before specific cryobiological studies were undertaken. However, the necessity for these became apparent with the partial recovery or sometimes loss of motility after freezing either subfertile semen before chemotherapy or radiotherapy, or spermatozoa collected from non-physiological situations (epididymal or testicular spermatozoa). The main trends in improvement have defined end-points other than the percentage of motility recovery or the assessment of ultrastructural damage. More sensitive criteria of the objective assessment of motility, energy status, damage to the plasma membrane or to subcellular elements, chromatin stability and chromosomal damage have been proposed as complementary end-points to better assess sperm cryopreservation. A different approach was related to the biochemical environment and physical conditions imposed on spermatozoa during the freezing and thawing process. Biochemical changes were assessed following different combinations of various extenders which attempted either to better preserve some parameter or to avoid the tendency towards drastic increase in osmotic pressure. Analysis of physical conditions was linked to the rate of cooling, freezing and warming, and was based on cryobiological studies. Finally, even though such improvements are not negligible, many questions remain unanswered. The extensive use of frozen spermatozoa during assisted reproductive techniques, together with the development of assisted fertilization using surgically collected spermatozoa, creates the need for additional studies to improve the cryopreservation of human spermatozoa.
shows the process of study selection. We identified 79 studies, of which 45 described 1 or more molecular NIPT technique(s) of  
Process of study selection for the analysis of quality and outcome.  
BACKGROUND Research on noninvasive prenatal testing (NIPT) of fetal trisomy 21 is developing fast. Commercial tests have become available. To provide an up-to-date overview of NIPT of trisomy 21, an evaluation of the methodological quality and outcomes of diagnostic accuracy studies was made.METHODS We undertook a systematic review of the literature published between 1997 and 2012 after searching PubMed, using MeSH terms 'RNA', 'DNA' and 'Down Syndrome' in combination with 'cell-free fetal (cff) RNA', 'cffDNA', 'trisomy 21' and 'noninvasive prenatal diagnosis' and searching reference lists of reported literature. From 79 abstracts, 16 studies were included as they evaluated the diagnostic accuracy of a molecular technique for NIPT of trisomy 21, and the test sensitivity and specificity were reported. Meta-analysis could not be performed due to the use of six different molecular techniques and different cutoff points. Diagnostic parameters were derived or calculated, and possible bias and applicability were evaluated utilizing the revised tool for Quality Assessment of Diagnostic Accuracy (QUADAS-2).RESULTSSeven of the included studies were recently published in large cohort studies that examined massively parallel sequencing (MPS), with or without pre-selection of chromosomes, and reported sensitivities between 98.58% [95% confidence interval (CI) 95.9-99.5%] and 100% (95% CI 96-100%) and specificities between 97.95% (95% CI 94.1-99.3%) and 100% (95% CI 99.1-100%). None of these seven large studies had an overall low risk of bias and low concerns regarding applicability. MPS with or without pre-selection of chromosomes exhibits an excellent negative predictive value (100%) in conditions with disease odds from 1:1500 to 1:200. However, positive predictive values were lower, even in high-risk pregnancies (19.7-100%). The other nine cohort studies were too small to give precise estimates (number of trisomy 21 cases: ≤25) and were not included in the discussion.CONCLUSIONSNIPT of trisomy 21 by MPS with or without pre-selection of chromosomes is promising and likely to replace the prenatal serum screening test that is currently combined with nuchal translucency measurement in the first trimester of pregnancy. Before NIPT can be introduced as a screening test in a social insurance health-care system, more evidence is needed from large prospective diagnostic accuracy studies in first trimester pregnancies. Moreover, we believe further assessment, of whether NIPT can be provided in a cost-effective, timely and equitable manner for every pregnant woman, is required.
The pre-ovulatory surge of gonadotrophins triggers a marked and obligatory increase in follicular prostaglandin synthesis prior to ovulation, and the cyclooxygenase (COX) enzyme is a key rate-limiting step in the biosynthesis of prostaglandins. In the early 1990s, the pre-ovulatory rise in follicular prostaglandin synthesis was shown to result from the selective induction of a novel COX isoform, now referred to as COX-2. Differences in the time-course of COX-2 induction in species with a short versus a long ovulatory process suggest that the enzyme could be a molecular determinant that sets the alarm of the mammalian ovulatory clock. Some of the fine molecular mechanisms involved in the transcriptional activation of the COX-2 gene in granulosa cells have also been elucidated. The binding of trans-activating upstream stimulatory factors (USF) to a consensus E-box cis-element in the proximal region of the promoter was shown to play a predominant role in COX-2 transcription. Studies showed that COX-2 expression could also serve as a valuable marker for follicular commitment to ovulation during hyperstimulatory cycles. This paper presents a comprehensive review of the events that led to the characterization of COX-2 in pre-ovulatory follicles, updates current concepts on the control of COX-2 expression in pre-ovulatory follicles, and addresses the consequences of COX-2 inhibition to women fertility and potential implications of COX-2 expression in ovarian cancer.
BACKGROUND Approximately 10 years after the first publication introducing the motile sperm organelle morphology examination (MSOME), many questions remained about sperm vacuoles: frequency, size, localization, mode of occurrence, biological significance and impact on male fertility potential. Many studies have tried to characterize sperm vacuoles, to determine the sperm abnormalities possibly associated with vacuoles, to test the diagnostic value of MSOME for male infertility or to question the benefits of intracytoplasmic morphologically selected sperm injection (IMSI).METHODS We searched PubMed for articles in the English language published in 2001-2012 regarding human sperm head vacuoles, MSOME and IMSI.RESULTSA bibliographic analysis revealed consensus for the following findings: (i) sperm vacuoles appeared frequently, often multiple and preferentially anterior; (ii) sperm vacuoles and sperm chromatin immaturity have been associated, particularly in the case of large vacuoles; (iii) teratozoospermia was a preferred indication of MSOME and IMSI.CONCLUSION The high-magnification system appears to be a powerful method to improve our understanding of human spermatozoa. However, its clinical use remains unclear in the fields of male infertility diagnosis and assisted reproduction techniques (ARTs).
Congenital adrenal hyperplasias (CAH) are inherited defects of cortisol biosynthesis. More than 90% of CAH are caused by 21-hydroxylase deficiency (21-OHD), found in 1:10 000 to 1:15 000 live births. Females with 'classical' 21-OHD, being exposed to excess androgens prenatally, are born with virilized external genitalia. Potentially lethal adrenal insufficiency is characteristic of two-thirds to three-quarters of patients with the classical salt wasting (SW) form of 21-OHD. Non-SW 21-OHD may be diagnosed on genital ambiguity in affected females, and/or later on the occurrence of androgen excess in both sexes. Non-classical 21-OHD, detected in > or =1:100 of certain populations, may present as precocious pubarche in children or polycystic ovarian syndrome in young women. 21-OHD is caused by mutations in the CYP21 gene encoding the steroid 21-hydroxylase enzyme. More than 90% of these mutations result from intergenic recombination between CYP21 and the closely linked CYP21P pseudogene. The degree to which each mutation compromises enzymatic activity is strongly correlated with the clinical severity of the disorder. This close association between genotype and phenotype makes it possible to predict clinical outcome in affected subjects. The risk of SW and prenatal virilization can be estimated, and overtreatment can be avoided in mildly affected cases. Glucocorticoid and mineralocorticoid replacement therapies are the mainstays of treatment, but additional therapies are being developed. A first trimester prenatal diagnosis should be proposed in families in whom molecular studies have been performed previously. The state of heterozygotism can be predicted by hormonal testing and confirmed by molecular studies. Prenatal diagnosis by direct mutation detection in previously genotyped families permits prenatal treatment of affected females in order to avoid or minimize genital virilization. Neonatal screening by hormonal methods identifies affected children before SW crises develop, reducing mortality in this disorder.
Comparative regional distribution of IVF clinics per capita.  
Comparative regional distribution of IVF clinics per estimated numbers of infertile women.  
BACKGROUND Infertility is estimated to affect as many as 186 million people worldwide. Although male infertility contributes to more than half of all cases of global childlessness, infertility remains a woman's social burden. Unfortunately, areas of the world with the highest rates of infertility are often those with poor access to assisted reproductive techniques (ARTs). In such settings, women may be abandoned to their childless destinies. However, emerging data suggest that making ART accessible and affordable is an important gender intervention. To that end, this article presents an overview of what we know about global infertility, ART and changing gender relations, posing five key questions: (i) why is infertility an ongoing global reproductive health problem? (ii) What are the gender effects of infertility, and are they changing over time? (iii) What do we know about the globalization of ART to resource-poor settings? (iv) How are new global initiatives attempting to improve access to IVF? (v) Finally, what can be done to overcome infertility, help the infertile and enhance low-cost IVF (LCIVF) activism?
Determination of embryonic age groups or stages has been based on the Carnegie Institute collection started in 1887. Improved technology has enabled the building of a new collection of embryos of < 9 weeks gestation; these were then used to compare with the original Carnegie collection. The results suggest that in providing definitive stages that are rigidly bound by developmental events, limitations are placed on categorizing the embryo. Allocation of embryos to a specific stage can assist in identifying post-ovulatory age but overlaps between stages could lead to classification into an incorrect stage.
Transforming growth factor (TGF) is known to have the ability to modify mitogenic responses of tissues to other peptide growth factors and therefore may contribute to the rapid growth rate of an embryo. Throughout the TGF superfamily there is a similar fundamental molecular architecture. Included in this superfamily are inhibin A, activin A and activin B. It has been shown that activin is a powerful mesodermal inducing factor in the early embryo. The human embryo has shown localization of inhibin in the gonads after 16 weeks gestation but it has not been previously identified in earlier embryos. The inhibin-activin protein was found in a range of tissues including the liver stages 19-21 (alpha) and stages 19-22 (beta); oesophagus stages 19-22 (alpha and beta); stomach stages 21 and 22 (alpha and beta); gut stages 16-22 (alpha) and 21 and 22 (beta); pericardium stages 12-22 (alpha and beta); gonad stages 21 and 22 (beta) stage 22 (alpha); adrenal stages 19-22 (alpha and beta); urogenital system stages 21 and 22 (alpha and beta); yolk sac stage 12 (alpha and beta); mesenchyme stages 16-22 (alpha); surface ectoderm stages 13-22 (alpha) and stages 16-22 (beta a); notocord stages 13-22 (beta) and stages 21 and 22 (alpha); nasal, trachea and bronchi stages 19-22 (alpha and beta) leading to speculation of the role of both subunits.
Localization of the mRNA alpha and beta a subunits of inhibin has previously been reported in the human gonads during the second trimester. Adrenal inhibin has also been reported in the second trimester for the alpha, beta a and beta b subunits. Investigations showing localization by in-situ hybridization during the first trimester have not been reported. The results have shown hybridization of the alpha and beta a subunits, throughout the period of development studied, in a variety of tissues including the dorsal and thoracic aortas and pericardium stages 13-22 (beta a subunit); liver stages 19-21 (beta a) and stages 21-22 (alpha); mesonephos stages 21 and 22 (beta a); gonad stages (alpha and beta a); adrenal stages 19-22 (alpha); surface ectoderm stages 16-22 (beta a); mesenchyme stages 16-22 (beta a); amnion stages 13-16 (beta a); yolk sac stage 12 (alpha and beta a); cartilage stages 19-22 (beta a); and nasal proliferation stages 21 and 22 (beta a). When compared with distribution of the protein subunits it was noted that more immunostaining activity was found, suggesting that that probes were not sufficiently sensitive enough to detect all levels of mRNA expressed. It can be surmised, therefore, that the lack of visual hybridization of the mRNA cannot preclude the possibility that it is not being translated within the tissue even though hybridization was not apparent.
The developmental age of an embryo in the first trimester of pregnancy is generally determined by ultrasound scanning and/or by calculation from menstrual age. In the original studies, validation of the estimate of gestational age by ultrasound was not possible as the exact date of conception was unknown. Variation in growth rates of identically aged fetuses has previously been reported after assisted conception and with the use of ultrasound scanning. As these pregnancies were ongoing the accuracy of the scanning results could not be determined. Comparison of scanning and direct measurements after termination of pregnancy and menstrual age were carried out to determine the accuracy in fetal dating. The results suggest that the use of ultrasound scanning to determine gestational age is of less use than previously thought, and that the use of menstrual age is severely limited.
BACKGROUND For a number of reasons, the results of previous meta-analyses may not fully reflect the mental health status of the average woman suffering from polycystic ovary syndrome (PCOS), or the causes of this distress. Our objective was to examine emotional distress and its associated features in women with PCOS. METHODS A comprehensive meta-analysis of comparative studies reporting measures of depression, anxiety or emotional-subscales of quality of life (emoQoL) was performed. PubMed, Embase, PsychInfo and the Cochrane trial register databases were searched up to November 2011 (see Supplementary Data for PUBMED search string). Unpublished data obtained through contact with authors were also included. The standardized mean difference (SMD) of distress scores was calculated. Subgroup analyses and meta-regression analysis of methodological and PCOS-related features were performed. RESULTS Twenty-eight studies (2384 patients and 2705 control women) were included. Higher emotional distress was consistently found for women with PCOS compared with control populations [main outcomes: depression: 26 studies, SMD 0.60 (95% confidence interval (CI) 0.47–0.73), anxiety: 17 studies, SMD of 0.49 (95% CI 0.36–0.63), emoQoL: 8 studies, SMD −0.66 (95% CI −0.92 to −0.41)]. However, heterogeneity was present (I² 52–76%). Methodological and clinical aspects only partly explained effect size variation. CONCLUSIONS Women with PCOS exhibit significantly more emotional distress compared with women without PCOS. However, distress scores mostly remain within the normal range. The cause of emotional distress could only partly be explained by methodological or clinical features. Clinicians should be aware of the emotional aspects of PCOS, discuss these with patients and refer for appropriate support where necessary and in accordance with patient preference.
The transformation of endometrium into decidua is an essential feature of normal implantation and pregnancy. There is a close association with an unusual leukocyte population, uterine natural killer (NK) cells, and onset of decidualization. These uterine NK cells are seen in close contact with stromal cells ultrastructurally and are also seen encircling vessels and glands. The possibility that uterine NK cells in the late secretory phase and in early decidua may be important in initiating and maintaining decidualization is raised. In contrast, the death of uterine NK cells could be an early event in the onset of endometrial breakdown at menstruation. The period between implantation and menstruation (7-14 days after luteinizing hormone surge) is the time when implantation is known to be particularly vulnerable. In this review, the possibility that uterine leukocytes might influence the critical decision that the mid- to late secretory endometrium must make either to decidualize or to undergo menstruation is explored.
Patients with low sperm counts combined with normal concentrations of gonadotrophins, and in whom physical examination and post-ejaculatory urine analysis are normal, present a diagnostic dilemma. This situation can be caused by testicular failure or by ductal obstruction, which have very different clinical prognoses. Ductal obstruction might be correctable by microsurgical vasovaso/vasoepididymostomy, whereas this approach is of no use in primary testicular failure. A possible diagnostic step for these patients is a testicular biopsy to differentiate between hypospermatogenesis and a normal gonad. However, to date testicular biopsy is seldom performed because of its invasive character. An alternative accurate, non-invasive method to assess testicular function could be very helpful in the evaluation of idiopathic azoospermia or idiopathic oligozoospermia. During the past decade, magnetic resonance (MR) spectroscopy has been developed from a scientific tool into a non-invasive clinical diagnostic tool and has also been used to study testicular function. Recent studies have shown that 31P-MR spectroscopy, based upon differences in the ratio of peaks of phosphomonoester to beta-adenosinetriphosphate, is a non-invasive technique able to differentiate between groups of patients with testicular failure and ductal obstruction, and it correlates reasonably well with the averaged mean Johnsen score of testicular biopsy. The role for a non-invasive technique in the diagnosis of male infertility, such as 31P-MR spectroscopy, can be manifold. It serves not only as an alternative for biopsy but can also be used to assess obstruction as the cause of infertility in patients with subnormal sperm counts, and to predict the chances of pregnancy in patients planned for vasovasostomy to correct a prior vasectomy. However, the main limitation to MR spectroscopy becoming a universal clinical diagnostic technique is the limited availability of 1.5 Tesla MR scanners.
Adaptation of the maternal immune response to accommodate the semi-allogeneic fetus is necessary for pregnancy success, and disturbances in maternal tolerance are implicated in infertility and reproductive pathologies. T regulatory (Treg) cells are a recently discovered subset of T-lymphocytes with potent suppressive activity and pivotal roles in curtailing destructive immune responses and preventing autoimmune disease. A systematic review was undertaken of the published literature on Treg cells in the ovary, testes, uterus and gestational tissues in pregnancy, and their link with infertility, miscarriage and pathologies of pregnancy. An overview of current knowledge on the generation, activation and modes of action of Treg cells in controlling immune responses is provided, and strategies for manipulating regulatory T-cells for potential applications in reproductive medicine are discussed. Studies in mouse models show that Treg cells are essential for maternal tolerance of the conceptus, and that expansion of the Treg cell pool through antigen-specific and antigen non-specific pathways allows their suppressive actions to be exerted in the critical peri-implantation phase of pregnancy. In women, Treg cells accumulate in the decidua and are elevated in maternal blood from early in the first trimester. Inadequate numbers of Treg cells or their functional deficiency are linked with infertility, miscarriage and pre-eclampsia. The potency and wide-ranging involvement of Treg cells in immune homeostasis and disease pathology indicates the considerable potential of these cells as therapeutic agents, raising the prospect of their utility in novel treatments for reproductive pathologies.
Relation between age and blood pressure, adapted form Kaplan (1994). 
Relationship between incidence of type I (insulin-dependent diabetes mellitus; IDDM) and type II diabetes (non-insulin-dependent diabetes mellitus; NIDDM) and age. Adapted from Joslin's Diabetes Mellitus (1985). 
The objective of this report is to provide an update of our current knowledge about the impact of maternal age on pregnancy outcome. Pregnancy in women > or =35 years old is associated with a higher maternal and perinatal mortality. The older gravida also has a higher chance of being delivered by Caesarean section. Most of the complications associated with older age are caused by age-related confounders such as leiomyomas, type II diabetes, hypertension and multiparity. Diabetes and hypertension increase almost linearly with age. Pregnant women with diabetes or hypertension are at increased risk of adverse pregnancy outcome irrespective of age. The currently available literature indicates that premenopausal pregnant women of advanced age who are in good health do not need special care besides the normal obstetric practice. At present, establishing pregnancy in postmenopausal women is more an ethical than a medical issue, partly because the information reported on pregnancy in postmenopausal women is insufficient to determine a reliable risk profile. In these women cardiovascular ageing accelerates. Therefore, until proven otherwise, postmenopausal women should be considered particularly at increased risk for vascular complications during pregnancy. This risk is likely to increase progressively with the number of years elapsed since the onset of postmenopause.
The optimal ovarian stimulation dose to obtain the best balance between the probability of pregnancy and the risk of complications, while maximizing cost-effectiveness of in vitro fertilization (IVF) treatment, is yet to be established. A systematic search of the electronic databases PubMed, EMBASE and Cochrane library, from 1984 until October 2009 for randomized controlled trials comparing different doses of recombinant FSH in IVF, was performed. Ten studies (totaling 1952 IVF cycles) were included in the present meta-analysis, comprising patients younger than 39 years with regular menstrual cycle, normal basal FSH levels and two normal ovaries. Comparison was made between studies using a daily dose of 100 versus 200 IU recFSH, and between 150 versus 200 IU recFSH or higher. Although oocyte yield was greater in the >200 IU/day dose group, pregnancy rates were similar compared with lower dose groups. The risk of insufficient response to ovarian stimulation was greatest in the 100 IU/day dose group. The risk of developing ovarian hyperstimulation syndrome was greater in the >200 IU/day dose group. The number of embryos available for cryopreservation was lowest in the 100 IU/day group, but similar comparing the 150 IU/day and the >200 IU/day dose groups. This meta-analysis suggests that the optimal daily recFSH stimulation dose is 150 IU/day in presumed normal responders younger than 39 years undergoing IVF. Compared with higher doses, this dose is associated with a slightly lower oocyte yield, but similar pregnancy and embryo cryopreservation rates. Furthermore, the wide spread adherence to this optimal dose will allow for a considerable reduction in IVF costs and complications.
The use of ICSI has been a major breakthrough in the treatment of male infertility. Even azoospermic patients with focal spermatogenesis in the testis, may benefit from the ICSI technique in order to father a child. As ICSI use has become more common, centres have introduced infertility treatment for Klinefelter patients. To date, 34 healthy children have been born using ICSI without PGD, and the conception of one 47,XXY fetus has been reported. In view of the possible risk of an increased gonosome number in the spermatozoa of Klinefelter patients, a safer approach--offering these couples ICSI combined with PGD--has been used, and has resulted in the birth of three healthy children. Couples in which the male suffered from Klinefelter's syndrome were first treated in 1995; these patients were offered ICSI + PGD using FISH technology, notably to enumerate the X and Y chromosomes. ICSI + PGD was performed in 32 cycles of 20 couples with spermatozoa originating from a fresh ejaculate (n = 1), testicular biopsy (n = 21) or frozen-thawed testicular biopsy (n = 10). Normal fertilization occurred in 56.0 +/- 22.4% of the successfully injected oocytes. On day 3 of development, 119 embryos from 29 cycles were of sufficient quality to undergo biopsy and subsequent PGD; a positive result was obtained in 113 embryos. Embryos were available for transfer in 26 cycles, with a mean of 1.6 +/- 0.6 embryos per transfer. Eight pregnancies were obtained, and five resulted in a delivery. A total of 113 embryos from couples with Klinefelter's syndrome was compared with 578 embryos from control couples with X-linked disease where PGD was used to determine gender. A significant fall occurred in the rate of normal embryos for couples with Klinefelter's syndrome (54.0%) compared with controls (77.2%). Moreover, a significantly increased risk of abnormalities was observed for sex chromosomes and autosomes; for each autosome separately, this reached significance level for chromosomes 18 and 21 only. Hence, a cautious approach is warranted in advising couples with non-mosaic Klinefelter's syndrome. Moreover, the use of ICSI + PGD or prenatal diagnosis should be carefully considered.
High dose chemotherapy and radiotherapy have radically increased long-term survival of young cancer patients, but major side effects of these treatments are ovarian failure and infertility. Knowledge of the risks and probabilities of ovarian failure caused by treatment is crucial for patients and physicians in order to make informed choices that will best serve patients' interests. This review presents data on ovarian damage and failure following exposure to radiotherapy, chemotherapy and ablative therapy. The risk is evaluated from the published literature according to patient's age, treatment protocol and also according to the diagnosis of some common malignancies. Many of these patients will not be sterilized immediately following treatment, but will suffer from premature menopause. In order to prevent sterilization, ovarian transposition before pelvic irradiation is mandatory. Besides cryopreservation of ovarian tissue and embryos before administration of chemotherapy, the possible protective effect of pituitary-ovarian down-regulation is discussed. The mechanism of primordial follicles damage induced by radio/chemotherapy is presented as well as the role of apoptosis signalling pathways underlying destruction. Increased knowledge of these mechanisms could help to identify potential effective inhibitors that can block the path of primordial follicles destruction and reduce ovarian failure rate.
Duplicating sperm centriole at higher magnification (TEM)  
In a recent CD-ROM, we portrayed the microstructure of the pre-implantation human embryo (Sathananthan et al., 1999), which was a multimedia production with computer colour-enhanced electron micrographs of mainly monospermic embryos. This disk portrays light and electron micrographs of over 250 tripronuclear (3PN), dispermic, human embryos during pre-implantation development, viewed in thick and thin Araldite sections, as well as appearances of whole embryos flat embedded in Araldite blocks visualized with the light microscope. The 100 figures were computerized (IBM TIFF format), edited and labelled using Adobe Photoshop 5. Some of the figures were coloured on computer. The early development of 3PN embryos overtly resembles that of normal embryos but there are important differences in their microstructure which are portrayed in this presentation. This is a multicentric study involving researchers from four IVF centres.
Varicoceles are vascular lesions of the pampiniform plexus and are the most common identifiable abnormality found in men being evaluated for infertility. Despite the long history associated with varicoceles, there remains much controversy regarding their diagnosis and management. The purpose of this manuscript is to address three of the most pressing controversies: (i) the association of varicoceles with male infertility, (ii) whether varicoceles exert a progressive deleterious effect and (iii) the relationship of varicocele size and outcome following varicocele repair. The current literature is reviewed in an effort to answer these questions. Based upon this analysis, conclusions can be drawn regarding the best management of varicoceles in subfertile men, adolescents, young fertile men and men with subclinical varicoceles. Although there remain many controversies due to a paucity of data, there appears to be a significant difference between adults and adolescents with respect to a progressive deterioration of semen parameters and it is clear that subclinical varicoceles do not play a major role in male infertility.
Abdominal myomectomy (by laparotomy or by laparoscopy) enables all the myomata to be excised while maintaining reproductive function. The actual risk of recurrence after abdominal myomectomy is difficult to assess because of methodological problems. Studies using life-table analysis find a cumulative risk of clinically significant recurrence of approximately 10% at 5 years for myomectomy by laparotomy. This risk probably underestimates the true prevalence of myomata as assessed by systematic ultrasound investigation. After laparoscopic myomectomy there appears to be a greater risk of recurrence. In one third of cases, recurrence becomes the reason for a hysterectomy. The risk of recurrence increases when there is more than one myoma. The use of gonadotrophin-releasing hormone agonists preoperatively could increase the risk of recurrence. Persistence or recurrence of the myoma thus reduces the chances of conception or taking a pregnancy full term after the myomectomy. It is essential to obtain the most complete exeresis possible in order to reduce the risk of recurrence to a minimum. However, it is inevitable that small, undetectable nuclei will remain within the myometrium whatever approach is used (laparoscopy or laparotomy). It would be an advantage to know what the growth factors are and how to identify groups at high risk of recurrence so that the treatment strategies could be better adapted and appropriate prophylactic methods developed.
Temporal–spatial control of gene expression in (human) spermatogenesis. The generation of spermatozoa in humans takes ~ 74 days, with the pachytene stage of prophase I taking roughly 14 of those days. In comparison, the remainder of meiosis I and all of meiosis II requires <3 days to complete. During proliferation and meiosis, expression is mainly under transcriptional control as indicated by the green line. Transcriptional control also applies to the early haploid stages of spermatogenesis, represented by the blue line, and indeed, MI prophase is when many of the RNAs that will be translated post- meiotically are transcribed. Their translation corresponds with nuclear shutdown and is represented by the red line. The fate of these transcripts is suggested, with most of the pre-meiotic and early meiotic RNAs going into the residual bodies (blue and green line). Selected pre-meiotically transcribed RNAs and haploid-expressed transcripts are subsequently retained by the spermatozoa. This relationship has not been experimentally verified. 
Clinical application of spermatozoal RNA. Spermatozoal RNAs are transcribed in the testes during the production of spermatozoa. (A) As indicated by spermatogenic and Leydig cells in highlighted colours, perturbations in spermatogenic gene expression may occur. This altered gene expression can lead to the production of abnormal spermatozoa as shown in panel B. Changes in spermatogenic gene expression leading to subfertility or infertility should be detected by variations in spermatozoal fingerprints utilizing in this example, microarrays (panel C).
(A) Inter-sample comparisons to define the shared, fertile (core) set of transcripts using microarrays. (i) The same sample probed twice by an arbitrary microarray illustrating procedural noise; (ii) two samples illustrating shared and variant sets; (iii) the more comparisons we make, the higher the resolution obtained for defining the core set of transcripts panel; (B) (iv-vi) intra-sample comparisons resolved on discontinuous density gradients showing (iv), normozoospermic sample with three sperm population subsets comprising the pellet, cells of abnormal morphology and cells with poor motility (recovered from the fractionated semen). The same subsets are indicated for asthenozoospermc (v) and teratozoospermic (vi) samples showing expected variations in their respective sizes.
Research on spermatozoal RNA has made considerable progress since the original reports on its presence appeared in the late 1950s and early 1960s. Through the use of stringent procedures aimed at eliminating contamination artefacts, we now appreciate that a complex cohort of mRNAs persists in the ejaculate cell but that 80S (cytoplasmic) ribosomal complexes are not present in sufficient quantities to support cytoplasmic mRNA translation. Despite this, under certain conditions, at least some cytoplasmic mRNAs can apparently be translated de novo, possibly on mitochondrial polysomes. The detection of mRNA translation by mature spermatozoa essentially supports the earliest research reports on spermatozoal gene expression although the suggested relationship with protein turnover and capacitation is wholly unexpected. We also examine some alternative explanations and roles for RNA carriage, including the RNAs passive retention as a consequence of nuclear shutdown and a more active role in chromatin repackaging, genomic imprinting, gene silencing and post-fertilization requirements of essential paternal RNAs. The recent report of an RNA-mediated epigenetic alteration to phenotype that is likely to be sperm derived is of particular interest in this regard. We finally show that regardless of the biological role(s) of spermatozoal RNA, its utility in infertility studies, particularly when coupled with modern techniques in gene-expression analysis (e.g. microarrays), is obvious. As a wholly non-invasive proxy for the testis, this RNA offers considerable potential as a marker for fertility status and the genetic and environmental influences that could make all the difference between a fertile and an infertile phenotype.
Two oocytes at germinal-vesical stage. One is normal size and mononucleated; the other is much larger, with two nuclei. 
'Dominant blastomere' embryo, and fluorescence in-situ hybridization (FISH) on the large blastomere, showing that it is polyploid. 
Fluorescence in-situ hybridization (FISH) with X (blue), Y (white), 13 (yellow), 16 (green), 18 (pink) and 21 (red) specific probes on blastomeres from trisomic embryos. Figure 3 shows a blastomere which belonged to an embryo with all its cells trisomic 16, but endoreduplication of chromosome 13 has also occurred (nine copies). Figure 4 shows a blastomere from a trisomy 21 embryo; Figure 5 shows a blastomere from a trisomy 16 embryo; Figure 6 shows a blastomere from a trisomy X embryo and Figure 6 shows a blastomere from a trisomy 18 embryo. 
The presence of numerical chromosome abnormalities in human embryos was studied using fluorescence in-situ hybridization with four or more chromosome-specific probes. When most cells of an embryo are analysed, this technique allows differentiation to be made between aneuploidy, mosaicism, haploidy and polyploidy. Abnormal types of fertilization, such as unipronucleated, tripronucleated zygotes and zygotes with uneven pronuclei, were studied using this technique. We have found a strong correlation between some types of dysmorphism with chromosomal abnormalities. In addition, the more impaired the development of an embryo, the more chromosomal abnormalities were detected in those embryos. Maternal age and other factors were linked to an increase in chromosome abnormalities (hormonal regimes, temperature changes), but not to intracytoplasmic sperm injection.
Trisomy is the most commonly identified chromosome abnormality in humans, occurring in at least 4% of all clinically recognized pregnancies; it is the leading known cause of pregnancy loss and of mental retardation. Over the past decade, molecular studies have demonstrated that most human trisomies originate from errors at maternal meiosis I. However, Klinefelter syndrome is a notable exception, as nearly one-half of all cases derive from paternal non-disjunction. In this review, the data on the origin of sex chromosome trisomies are summarized, focusing on the 47,XXY condition. Additionally, the results of recent genetic mapping studies are reviewed that have led to the identification of the first molecular correlate of autosomal and sex chromosome non-disjunction; i.e. altered levels and positioning of meiotic recombinational events.
Inhibitory natural killer (NK) cell receptors for human leukocyte antigen (HLA) class I. KIR molecules contain two or three immunogloulin-like domains in the extracellular region. The two immunglobulin domains KIR recognize a polymorphism at positions 77 and 80 of the HLA-C heavy chain; some recognize HLACw4 and related alleles [possessing asparagine (N) at residue 77 and lysine (K) at residue 80], whereas others bind HLA-Cw3 and related alleles [possessing serine (S) at residue 77 and asparagine (N) at residue 80]. In the monomeric and dimeric forms, the three immunoglobulin domain KIR-3D recognize HLA-Bw4 and HLA-A3 respectively. CD94 and NKGA2 contain C-type lectin domains in the extracellular region. The ILT2 glycoprotein is characterized by an extracellular region of four immunoglobulin-like domains and, apparently, has a broad specificity for HLA class I molecules. 
Variables affecting spontaneous in-vitro fertilization (IVF)/embryo transfer pregnancy rate in endometriosis. 
The observation that natural killer (NK) cell activity is abnormally low in endometriosis patients and abnormally high in women with otherwise unexplained recurrent spontaneous abortion represents, at present, an intriguing curiosity. There is evidence suggesting that these conditions are associated with an opposite regulation of NK cell behaviour. This review discusses these observations and potential relationships.
Framework for studies on reproductive function in mouse models Knockout mice mated together are compared to wild-type breeders of the same genetic background. The flowchart enquires whether the parameters of fertility are the same between knockout and control breeders. If the answer is yes for all parameters, the knockout mice have normal fertility, however does not preclude a role for the cytokine in allogeneic matings or under pathogenic challenge. To evaluate reproductive function in male and female knockout mice caged together, the female is checked each morning for a mating plug to indicate a mating event. To determine the success of the mating event, the females are sacrificed during late pregnancy (e.g. day 17.5 post-coitus) and the uterus dissected. Number of implantation sites and resorptions are quantified, and fetal and placental weights can be determined to evaluate placental function. Pups are monitored post-partum for developmental and lactational defects. If these fertility parameters deviate from wild-type controls, knockout mice can be mated with wild-type mice to evaluate whether the defect is due to male or female reproductive disorders, or a fetal requirement of the cytokine. Male knockout mice mated with wild-type controls may have defects in hormone synthesis, sexual behaviour dysfunction, spermatogenesis or altered seminal plasma content causing downstream effects on embryo quality or maternal immune response to pregnancy (Johansson et al ., 2004). Female knockout mice may have defects in hormone synthesis, oocyte development, ovulation, implantation, placental development, parturition or lactation 
Cytokines identified as important in functioning of the hypothalamo-pituitary-gonadal axis, and the production of mature gametes, as determined by knockout mouse studies CSF-1, colony-stimulating factor
Cytokines identified as important in pregnancy, as determined by knockout mouse studies
Cytokines play many diverse and important roles in reproductive biology, and dissecting the complex interactions between these proteins and the different reproductive organs is a difficult task. One approach is to use gene ablation, or 'knockout', to analyse the effect of deletion of a single cytokine on mouse reproductive function. This review summarizes the essential roles of cytokines in reproductive biology that have been revealed by gene knockout studies, including development and regulation of the hypothalamo-pituitary-gondal axis, ovarian folliculogenesis, implantation and immune system modulation during pregnancy. However, successful utilization of this approach must consider the caveats associated with gene ablation studies, e.g. embryonic lethality, systemic effects of cytokine ablation on local reproductive processes and the limited exposure to pathogens in mice housed in laboratory conditions. New sophisticated technology that temporally or spatially regulates gene ablation can overcome some of these limitations. Discoveries on the roles of cytokines in reproductive function uncovered by gene ablation studies can now be applied to improve in vitro fertilization for infertile couples and in the development of contraceptive therapies.
Study selection process for the individual patient data meta-analysis to assess the effectiveness of laparoscopic uterosacral nerve ablation (LUNA) in chronic pelvic pain.
Mean pain scores over time for trials included in the analysis. Lower score indicates less pain.
There have been conflicting results in randomized trials of the effects of laparoscopic uterosacral nerve ablation (LUNA) in chronic pelvic pain. Our objective was to perform a meta-analysis using individual patient data (IPD) to provide the most comprehensive and reliable assessment of the effectiveness of LUNA. Electronic searches were conducted in the Medline, Embase, PsycInfo and Cochrane Library databases from database inception to August 2009. The reference lists of known relevant papers were searched for any further articles. Randomized trials comparing LUNA with no additional intervention were selected and authors contacted for IPD. Raw data were available from 862 women randomized into five trials. Pain scores were calibrated to a 10-point scale and were analysed using a multilevel model allowing for repeated measures. There was no significant difference between LUNA and No LUNA for the worst pain recorded over a 12 month time period (mean difference 0.25 points in favour of No LUNA on a 0-10 point scale, 95% confidence interval: -0.08 to 0.58; P = 0.1). LUNA does not result in improved chronic pelvic pain.
Adenomyosis persisting under endometrial ablation scar. Original magnification ×25. (Reprinted with permission from the American Journal of Obstetrics and Gynecology, 174, 1791, 1996.) 
Adenomyosis proliferating through endometrial ablation scar. Original magnification ×25. (Reprinted with permission from the American Journal of Obstetrics and Gynecology, 174, 1791, 1996.) 
Operative hysteroscopy is a relatively new technique that has significantly improved the diagnosis and therapy of abnormal uterine bleeding. At first, the success of operative hysteroscopy in controlling this bleeding seemed extremely high but, with long-term follow-up, a significant failure rate became evident requiring a repeat hysteroscopic procedure or a hysterectomy. Deep adenomyosis is a major cause of these failures. This paper describes three operative ablation techniques and relates many of their failures to deep adenomyosis. The definition and pathophysiology of adenomyosis are also explored. The possibility of delaying the diagnosis of endometrial cancer under an ablation scar is discussed. Ultimately the depth of adenomyosis seems to correlate with the outcome of endometrial ablation or resection. Patients without or with only minimal endometrial penetration of <2.5 mm (superficial adenomyosis) have good results from the ablation. Patients with deep endometrial penetration of >2.5 mm (deep adenomyosis) usually have persistent problems and should be offered hysterectomy over repeat ablation. Magnetic resonance imaging or ultrasound may be an appropriate pre-operative screening tool to determine the depth of ademomyosis.
Proportions of cytogenetically normal, abnormal and mosaic 
Numbers of mosaic 8-cell stage embryos at each level of mosaicism. A/N and A/A stand for all possible combinations of mosaicism: A 1 /N, A 1 /A 2 /N, ¼ A 1 /A 2 /A 3 /A 4 /A 5 /A 6 /A 7 /N and A 1 /A 2 , A 1 /A 2 /A 3 , ¼ A 1 / A 2 /A 3 /A 4 /A 5 /A 6 /A 7 /A 8. The 133 cases of mosaicism due to trisomic and monosomic zygote rescue consisting of the mosaicisms A (original)/N and A (original)/A (mitotically changed)/N are not separately indicated in the group of A/N mosaicism.
Probabilities of normal and abnormal results of 1-cell biopsies taken from 8-cell embryos with various levels of mosaicism and the 
Probabilities of the results (N/N, A/N, A/A) of 2-cell biopsies 
Assisted reproduction and preimplantation genetic diagnosis (PGD) involve various complicated techniques, each of them with its own problems. However, the greatest problem with PGD for chromosome abnormalities is not of a technical nature but is a biological phenomenon: chromosomal mosaicism in the cleavage stage embryo. Here, we present a hypothetical, quantitative model for the development of chromosomally normal, abnormal and mosaic embryos. The arising of mosaicism in 2-8-cell embryos was described by a binomial probability model on the occurrence of mitotic events inducing chromosomal changes in the blastomeres. This model converted the 'mean' rate of mosaicism found in cross-sectional studies (60%) into an equal rate of mosaic embryos at arrival at the 8-cell stage (59.8%). The disappearance of > 90% of the mosaic embryos or the mosaicism itself from surviving embryos during the morula stage was explained by mitotic arrest of most of the mitotically changed cells under increasing cell cycle control. In our model, 25.9 and 14.3% of the embryos at the 8-cell stage are normal and abnormal respectively. The remaining 59.8% of the embryo shows mosaicism: 34.6% of abnormal/normal cells and 25.2% of abnormal/abnormal cells. The high proportion of abnormal and mosaic embryos together explains the high rate of abnormal laboratory findings in PGD for chromosomal abnormalities and aneuploidy screening. The poor representation of a 1- or 2-cell biopsy for the 7- or 6-cell post-biopsy embryo in the case of mosaicism explains the high rate of false-negative and false-positive results.
Over the past 10 years there has been an upsurge of interest in the mechanisms underlying normal and disturbed menstrual bleeding. These studies have particularly focused on the mechanisms underlying the common problems of menorrhagia associated with ovulatory and anovulatory dysfunctional uterine bleeding (DUB) and of unpredictable breakthrough bleeding during hormonal contraceptive use. A wide range of abnormalities of endometrial morphology and function have been demonstrated, but it is still not clear how all the pieces of this complex jigsaw puzzle fit together. Ovulatory DUB is predominantly associated with decreased endometrial vasoconstriction and vascular haemostatic plug formation, leading to defective control of the volume of blood which is lost during menstruation. By contrast, breakthrough bleeding is associated with a wide range of molecular disturbances which appear to result in unpredictable vessel breakdown through disturbed endometrial angiogenesis, increased vascular fragility and loss of the integrity of the endothelial, epithelial and stromal supporting structures. Anovulatory DUB is very poorly understood, but may be associated with disturbed angiogenesis, fragile vessels and defective haemostatic processes. Little is known about the actual mechanisms of the common problem of abnormal bleeding associated with specific genital tract pathologies such as uterine myomata.
Mirror curves of concentration of TSH and hCG during gestation. Reproduced with permission from Glinoer et al ., 1990. ã 1990 The Endocrine Society. 
Familial gestational thyrotoxicosis. Schematic diagram of the thyrotrophin receptor (TSHR) showing the location of the mutated residue K183R in the extracellular domain. Increased sensitivity to hCG of the mutant TSHR (K183R) when compared to the wild-type TSHR, as shown by the accumulation of the second messenger cAMP in response to increasing doses of hCG. Reproduced with permission from Rodien et al ., 1998. ã 1998 Massachusetts Medical Society. All rights reserved. 
Proposed assessment of thyroid function during gestation. By plotting concentration of free thyroxine (fT4) against concentration of hCG, it should be possible to distinguish the normal population (grey ellipse), from patients affected by gestational transient thyrotoxicosis (hatched ellipse) and from cases of hypersensitivity to hCG or cases of hCG with increased thyrotrophic activity (black circle). Note that this last category will be distinguished from Graves' disease only by the absence of ophthalmopathy, goitre and thyroid antibodies. By the end of second trimester of pregnancy, gestational transient thyrotoxicosis will have resolved with decline of hCG, whereas cases of hypersensitivity or hyperactive hCG will still be in the hyperthyroid range. The horizontal dashed line represents the upper normal value for fT4. 
Pregnancy induces physiological alterations in thyroid function which may make difficult the interpretation of results of thyroid hormone measurement. A state of hyperstimulation of the thyroid gland is common in early pregnancy. In a few cases, thyroid hormone values will deviate from the normal range, which corresponds to the gestational transient thyrotoxicosis. This syndrome is closely associated with hyperemesis gravidarum. The relationship between the two syndromes, demonstrated by epidemiological studies, has been illustrated by an exceptional case of familial recurrent gestational thyrotoxicosis presenting as hyperemesis gravidarum due to hypersensitivity of the thyrotrophin receptor to hCG. However, the exact mechanisms of hyperemesis gravidarum have not yet been identified. Gestational transient thyrotoxicosis has to be distinguished from Graves' disease, because the latter is associated with potential maternal and fetal complications when thyrotoxicosis is not controlled, whereas the former has usually a favourable outcome. The existence of other cases of thyroid hypersensitivity or hCG endowed with abnormal thyrotrophic activity is suspected. They may be identified only by assessment of the thyroid function in cases of hyperemesis gravidarum. The identification of these cases would be helpful to understand the mechanisms of specificity of glycoprotein hormone receptors.
Human male infertility is often related to chromosome abnormalities. In chromosomally normal infertile males, the rates of chromosome 21 and sex chromosome disomy in spermatozoa are increased. Higher incidences of trisomy 21 (seldom of paternal origin) and sex chromosome aneuploidy are also found. XXY and XYY patients produce increased numbers of XY, XX and YY spermatozoa, indicating an increased risk of production of XXY, XYY and XXX individuals. Since XXYs can reproduce using intracytoplasmic sperm injection (ICSI), this could explain the slight increase of sex chromosome anomalies in ICSI series. Carriers of structural reorganizations produce unbalanced spermatozoa, and risk having children with duplications and/or deficiencies. In some cases, this risk is considerably lower or higher than average. These patients also show increased diploidy, and a higher risk of producing diandric triploids. Meiotic disorders are frequent in infertile males, and increase with severe oligoasthenozoospemia (OA) and/or high follicle stimulating hormone (FSH) concentrations. These patients produce spermatozoa with autosomal and sex chromosome disomies, and diploid spermatozoa. Their contribution to recurrent abortion depends on the production of trisomies, monosomies and of triploids. The most frequent sperm chromosome anomaly in infertile males is diploidy, originated by either meiotic mutations or by a compromised testicular environment.
Top-cited authors
Bart C J M Fauser
  • University Medical Center Utrecht
F.J.M. Broekmans
  • University Medical Center Utrecht
Robert John Norman
  • University of Adelaide
Ben W Mol
  • Monash University (Australia)
Lisa Moran
  • University of Adelaide