Genetic regulation of ovulation rate and multiple births.

Ovulation rate in many mammalian species is controlled to regulate the numbers of offspring and maximise reproductive success. Pathways that regulate ovulation rate still respond to genetic and environmental factors and show considerable variation within and between species. Genetic segregation, positional cloning, and association studies have discovered numerous mutations and genetic risk factors that contribute to this variation. Notable among the discoveries has been the role of mutations in bone morphogenetic protein 15 (BMP15 ), growth differentiation factor 9 (GDF9 ) and bone morphogenetic protein receptor type 1B (BMPR1B ) from the intra-ovarian signalling pathway contributing to the evidence that signalling from the oocyte is the key driver in follicle regulation rather than circulating gonadotrophin concentrations. Multiple variants in different domains of BMP15 and GDF9 result in partial or complete loss of function of the proteins providing insights into their functional roles and differential regulation contributing to species differences in ovulation rate. Early success encouraged many more studies in prolific strains of sheep, cattle and goats providing a valuable catalogue of genetic variants of large effect increasing ovulation rate and litter size. More recently, genetic association studies are beginning to identify genetic risk factors with smaller effects. Most genes implicated are from pathways with defined roles in regulation of the ovarian function. However, some genomic regions suggest regulation by novel genes. Continuing genetic and related functional studies will add further to our understanding of the detailed regulation of ovulation rate and litter size with implications for health and animal production systems.

Association of genomic prediction for twin pregnancies with the incidence of double ovulation in primiparous dairy cows.

Twin pregnancies compromise the health and well-being of dairy cattle. A recent genomic prediction model for twin pregnancies has been developed based on twin calving or abortion. However, the incidence of double ovulation is significantly higher than that of twin births. This study aimed to evaluate whether genomic prediction values for twin pregnancies are associated with the incidence of double ovulation in primiparous dairy cows. Factors influencing the double ovulation rate were analysed using binary logistic regression on 676 cows: 475 (70.3%) inseminated at spontaneous estrus and 201 following one of two different estrus synchronization protocols for fixed-time artificial insemination (FTAI). The odds ratio for double ovulations was 0.92 (p = .002) per unit increase in prediction value and 2 (p = .01) for cows subjected to an FTAI protocol. Our findings suggest that genomic prediction values for twin pregnancies can effectively identify the risk of double ovulation at the herd level.

Reduction in multiple pregnancy rate in donor oocyte-recipient gestational carrier (GC) in vitro fertilization (IVF) cycles in the USA with single-embryo transfer and preimplantation genetic testing.

PURPOSE: To evaluate the utilization of single-embryo transfer (SET) and preimplantation genetic testing (PGT) in gestational carrier IVF cycles in the USA with donor oocyte and examine the impact on live birth and multiple gestation. METHODS: Retrospective cohort study using the Society of Assisted Reproductive Technology (SART) clinic database of 4776 donor oocyte-recipient IVF cycles in which a GC was used. The cycles were separated into 4 groups by use of PGT and number of embryos transferred as follows: (1) PGT and single-embryo transfer (PGT-SET); (2) PGT and multiple embryo transfer (PGT-MET); (3) no PGT and SET (NoPGT-SET); (4) no PGT and MET (NoPGT-MET). Primary outcomes were live birth rate (LBR) and multiple pregnancy rate (MPR). RESULTS: More than one blastocyst was transferred in 48.7% (2323/4774) of the cycles. When ≥1 blastocyst was transferred, with or without the use of PGT, the MPR was 45.5% and 42.0%, respectively. In comparison, in the PGT-SET and NoPGT-SET groups, the MPR was 1.4% (8/579) and 3.3% (29/883), respectively. Live birth rates increased with the use of PGT-A and with MET. CONCLUSION: This study shows that SET, with or without PGT, is associated with a significantly reduced MPR in donor oocyte-recipient GC IVF cycles while maintaining high LBR. It also demonstrates that many infertility centers in the USA are not adhering to ASRM embryo transfer guidelines. Our findings highlight an opportunity to increase GC safety, which ultimately may lead to widened access to this increasingly restricted service outside the USA.

Adding new ingredients to the recipe for successful embryo transfers.

The success of a pregnancy and the birth of a heathy baby following embryo transfer are conditioned by many factors, including embryo quality and the uterine environment. While we keep looking for more indicators of embryo quality, it also is critical to understand what constitutes a favorable uterine environment leading to a successful pregnancy and birth. This issue of JARG offers new insights on both components-so called by some "the seed and the soil"-and also highlights the critical interactions between the two. Collectively, these publications are contributing to a better understanding of basic embryology and reproductive biology. They could lead to multiple applications to mitigate infertility issues; however, our knowledge base remains rudimentary when it comes to sorting out the 'soil or seed' dilemma. The call from all authors for more research in their respective areas resonates within the ART community. Recognizing the practical and ethical limitations of studies in human patients also reemphasizes the need for solid research in multiple animal models to better understand what constitutes the best recipe for successful embryo transfer outcomes.

Risks of nonchromosomal birth defects, small-for-gestational age birthweight, and prematurity with in vitro fertilization: effect of number of embryos transferred and plurality at conception versus at birth.

PURPOSE: Excess embryos transferred (ET) (> plurality at birth) and fetal heartbeats (FHB) at 6 weeks' gestation are associated with reductions in birthweight and gestation, but prior studies have been limited by small sample sizes and limited IVF data. This analysis evaluated associations between excess ET, excess FHB, and adverse perinatal outcomes, including the risk of nonchromosomal birth defects. METHODS: Live births conceived via IVF from Massachusetts, New York, North Carolina, and Texas included 138,435 children born 2004-2013 (Texas), 2004-2016 (Massachusetts and North Carolina), and 2004-2017 (New York) were classified by ET and FHB. Major birth defects were reported by statewide registries within the first year of life. Logistic regression was used to estimate adjusted odds ratios (AORs) and 95% CIs of the risks of a major nonchromosomal birth defect, small-for-gestational age birthweight (SGA), low birthweight (LBW), and preterm birth (≤36 weeks), by excess ET, and excess ET + excess FHB, by plurality at birth (singletons and twins). RESULTS: In singletons with [2 ET, FHB =1] and [≥3 ET, FHB=1], risks [AOR (95% CI)] were increased, respectively, for major nonchromosomal birth defects [1.13 (1.00-1.27) and 1.18 (1.00-1.38)], SGA [1.10 (1.03-1.17) and 1.15 (1.05-1.26)], LBW [1.09 (1.02-1.13) and 1.17 (1.07-1.27)], and preterm birth [1.06 (1.00-1.12) and 1.14 (1.06-1.23)]. With excess ET + excess FHB, risks of all adverse outcomes except major nonchromosomal birth defects increased further for both singletons and twins. CONCLUSION: Excess embryos transferred are associated with increased risks for nonchromosomal birth defects, reduced birthweight, and prematurity in IVF-conceived births.

High-quality Cleavage Embryo versus Low-quality Blastocyst in Frozen-thawed Cycles: Comparison of Clinical Outcomes.

This study compared the clinical outcomes of the frozen-thawed cycles of high-quality cleavage embryos with low-quality blastocysts to provide a reference for the choice of frozen-thawed embryo transfer schemes and to improve clinical pregnancy rates. A retrospective analysis was performed on the clinical data of patients undergoing frozen-thawed embryo transfer at the Reproductive Medicine Center of Tongji Hospital of Tongji Medical College of Huazhong University of Science and Technology from 2016 to 2017. In total, 845 cases were divided into a high-quality cleavage embryo group (group A) and a low-quality blastocyst group (group B). Each group was further divided into subgroups based on the number of transplants. Group A was categorized into two subgroups comprising of 94 cases in subgroup A1 (1 high-quality 8-cell group) and 201 cases in subgroup A2 (2 high-quality 8-cell group). Group B was divided into four subgroups consisting of 73 cases in subgroup B1 (D53BC group), 65 cases in subgroup B2 (D54BC group), 110 cases in subgroup B3 (D63BC group), and 282 cases in subgroup B4 (D64BC group). The pregnancy outcomes and neonatal outcomes between the groups were compared. The clinical pregnancy rates (56.72% and 60.00%) and live birth rates (47.76% and 46.15%) in subgroups A2 and B2 showed no significant differences, but these rates were significantly higher in subgroups A2 and B2 than in the rest subgroups (P<0.05). The multiple birth rate (26.32%) in the subgroup A2 was significantly higher than that in the rest subgroups (P<0.05). There were no statistically significant differences in the abortion rates among all groups (P>0.05). In terms of neonatal outcomes, there were no statistically significant differences in the proportion of premature births, sex ratios, and birth defects among the low-weight and gigantic infants (P>0.05). Transplanting two high-quality cleavage embryos during the frozen-thawed embryo transfer cycles could significantly increase clinical pregnancy rates and live birth rates, but at the same time, it also increased the risks of multiple births and complications to mothers and infants. The D54BC subgroup had the most significant advantages among all groups (P<0.05). The rest low-quality blastocysts had clinical outcomes similar to the single high-quality cleavage embryo group.

Establishment of day 7 blastocyst freezing criteria using blastocyst diameter for single vitrified-warmed blastocyst transfer from live birth outcomes: a single-center, large cohort, retrospectively matched study.

PURPOSE: To establish blastocyst freezing criteria for day 7 blastocyst (day 7 BL) for single vitrified-warmed blastocyst transfer (SVBT) by examining the diameter of blastocysts. METHODS: Patients who underwent day 7 BL transfer cycles (1143 cycles, mean age: 38.5 ± 3.5) and randomly selected patients after 1:1 matching who underwent day 6 BL transfer cycles and day 2-single-embryo transfer (SET) cycles were used for analysis. Comparison of the miscarriage (per clinical pregnancy) and live birth rates were made among day 2-SET, day 7 BL, and day 6 BL. These blastocyst groups were stratified into six groups based on blastocyst diameter, namely, 180 µm, 190 µm, 200 µm, 210 µm, over 220 µm, and hatched, for making the freezing criteria. RESULTS: For each diameter, 180 µm, 190 µm, 200 µm, 210 µm, over 220 µm, and hatched, the live birth rates of day 7 BL after SVBT were 9.0%, 11.9%, 11.5%, 15.6%, 20.0%, and 19.9%, respectively. Compared with the 14.6% live birth rate of the day 2-SET group, the live birth rate of 220 µm day 7 BL was significantly higher (P < 0.05) and was around the same in other diameter groups. CONCLUSION: Our study demonstrates that sufficient live birth rates can be obtained after SVBT even from blastocysts on day 7 when blastocysts were vitrified at expanded blastocyst stage of over 180 µm of diameter or at hatched blastocyst stage and were transferred at the optimal time. This is the first study to establish a day 7 blastocyst freezing criteria using blastocyst diameter, which is an objective assessment way.

Naturally monozygotic quadruplets in a Braford cow confirmed by DNA analysis: A case report.

Cattle are a monotocous species, despite naturally conceived multiple births are sometimes observed. Although the number of twins has consistently increased, triplet and quadruplet pregnancies represent 0.015% and 0.004% of the total births, respectively. Multiple births are the result of multiple ovulation and/or the spontaneous cleavage of one fertilized oocyte, which is known as monozygotic (MZ) twinning. In cattle, approximately 5% to 14% of all twin births are MZ, and births with more than two MZ calves are extremely rare. Monozygotic animals are genetically identical, and those derived from two or more zygotes are genetically different. Furthermore, the presence of placental vascular anastomosis can result in foetal chimerism. Notwithstanding, animals born as single calves can be chimeras when one of the foetal twins dies undetected in utero. Here, we used DNA testing to study the zygotic condition of an unusual female quadruplet born from a Braford cow bred in a multi-sire natural mating system without hormone stimulation. Two tissues with different embryological origin were sampled to test zygosity and possible chimerism. The results showed an identical genotype, confirming they all originated in an MZ pregnancy and suggesting the lack of chimerism in all animals. The use of MZ twins in breeding and selection systems provides an alternative to the conventional progeny testing. Some works have suggested a genetic background of MZ twins in humans. This female and her daughters could be the founders of a lineage to study the possible inheritance of MZ multiple births in cattle.

Maternal physical activity and sedentary behaviour before and during in vitro fertilization treatment: a longitudinal study exploring the associations with controlled ovarian stimulation and pregnancy outcomes.

PURPOSE: To evaluate the association of objectively measured physical activity (PA) and sedentary behaviour before and during in vitro fertilization (IVF) with controlled ovarian stimulation (COS) and pregnancy outcomes. METHODS: This longitudinal study involved 107 infertile women undergoing IVF treatment. PA and sedentary behaviour were measured for 14 consecutive days using accelerometry as follows: (1) before IVF treatment, (2) during IVF at the implantation time, immediately after embryo transfer, and (3) after positive pregnancy test. Total screen time was assessed by questionnaires. COS results were measured as the number of oocytes and embryos obtained, and the study outcomes included positive hCG, clinical pregnancy, and live birth. RESULTS: Compared with baseline activity levels, women significantly reduced their PA and increased sedentary behaviour during IVF (p ≤ 0.001). Higher average PA, light PA, and ratio between breaks in every ≥ 30-min blocks of sedentary time showed positive associations, while sedentary time, number, and time accumulated in blocks of ≥ 30 min of sedentary time associated negatively with oocyte and embryo counts (all p < 0.05). Women with high total screen time during non-work days (≥ 7 h) obtained 4.7 oocytes (p = 0.005) and 2.8 embryos (p = 0.008) less in COS. PA and sedentary behaviour before and during IVF did not affect the positive hCG, clinical pregnancy, and live birth outcomes. CONCLUSION: Our study results suggest that higher time spent in PA and lower time spent in sedentary behaviour before entering assisted reproduction is associated with better COS outcomes, while activity levels before and during IVF do not affect the implantation, pregnancy, and live birth outcomes.

Breast Milk for Preterm Multiples: More Proteins, Less Lactose.

Exclusive breastfeeding is currently recommended until at least 6 months of postnatal age, due to maternal breast milk (BM) unique composition and beneficial properties. In fact, BM modifies itself according to gestational age (GA) at birth, adapting its composition to neonatal requests during lactation. Multiple births represent about 3% of the whole pregnancies; such neonates result more vulnerable than full-term newborns, due to lower GA and birth weight (BW) and the higher incidence of perinatal complications. Although an adequate nutrition is fundamental for twins and other multiples, studies on this topic are lacking. We collected and analyzed BM from mothers of 19 twins and 5 triplets showing GA < 33 weeks and BW < 1500 g, comparing it to a control group of 28 preterm singletons. As a result, at GA ≤ 28 weeks, we observed that protein content is higher in BM for multiples (1.53 vs. 1.29 g per 100 ml), lactose concentration is greater in BM for singletons (6.72 vs. 6.34 g per 100 ml) and GA results the most relevant factor influencing BM protein composition. BM for multiples results higher in proteins and lower in lactose, if compared with singleton's samples; this could promote and sustain growth and organ development in this vulnerable category. BM from multiples shows a trophic and immunologic role, since these neonates often show lower GA and BW instead of singletons. These findings could help in optimizing nutritional strategies and improving BM individualized fortification.

Morphological and ultrasound findings in multiple pregnancy placentation.

The incidence of multiple pregnancy has significantly increased over the past decades, reaching different statistics to double, triple, or even overcome these numerical orders globally. Zygosity and chorionicity are the key elements in the multiple pregnancy but the placentation issue should be correlated primarily with zygosity, unlike chorionicity that should be correlated with the outcome and complications of multifetal gestation. Multiple pregnancy is by itself a special maternal-fetal condition, and the monochorionic one, moreover, due to specific complications. These aspects make early assessment of chorionicity and amnionicity a priority. Ultrasound is essential in pregnancy but pathological placental examination after delivery is complementary, in order to have a complete overview of potential mechanisms and pathogenesis affecting twin gestation. In this review, we highlight both ultrasound aspects specific to multifetal placentation, complemented by macro and microscopic morphological aspects, which underpin the obstetric imaging.

Screening for Aneuploidy in Multiple Gestations: The Challenges and Options.

As the incidence of multiple gestations increases, the need for effective aneuploidy screening grows. There are unique challenges associated with aneuploidy screening in twins, and the data supporting its use are limited. This article summarizes the risk of aneuploidy for twin gestations and the available screening methods, including nuchal translucency, first-trimester and second-trimester serum screening, and cell-free DNA screening.

Seasonality in Multiple Maternities.

In the 19th century, a series of international statistical congresses introduced common rules for the national demographic registers. This activity contributed to the genesis of statistical research. During the history of twin research, Hellin's law has played a central role because it is an approximately correct association between the rates of multiple maternities. However, it has been mathematically proven that Hellin's law cannot hold exactly. The majority of all studies of Hellin's law are based on empirical rates of multiple maternities. Such studies can never confirm the law, but only identify errors too large to be characterized as random. It is of particular interest to examine why the rates of higher multiple maternities are sometimes too high or too low when Hellin's law is used as a benchmark. However, divergences from the law are often difficult to explain and/or eliminate. Different improvements to the law have been proposed. In this article, we study the seasonality of multiple maternities. We apply Hellin's law to compare the seasonality of twin and triplet rates.

Studies of the Seasonal Pattern of Multiple Maternities.

The seasonality of population data has been of great interest in demographic studies. When seasonality is analyzed, the population at risk plays a central role. In a study of the monthly number of births and deaths, the population at risk is the product of the size of the population and the length of the month. Usually, the population can be assumed to be constant, and consequently, the population at risk is proportional to the length of the month. Hence, the number of cases per day has to be analyzed. If one studies the seasonal variation in twin or multiple maternities, the population at risk is the total number of monthly confinements, and the study should be based on the rates of the multiple maternities. Consequently, if one considers monthly twinning rates, the monthly number of birth data is eliminated and one obtains an unaffected seasonality measure of the twin maternities. The strength of the seasonality is measured by a chi-squared test or by the standard deviation. When seasonal models are applied, one must pay special attention to how well the model fits the data. If the goodness of fit is poor, it can erroneously result in a statement that the seasonality is slight, although the observed seasonal fluctuations are marked.

[DNA profiling as a method of zygosity determination in multiple pregnancy]. / Profilowanie DNA jako metoda okreslania zygotycznosci w ciazy wieloraczej.

The aim of the report was to present a method of zygosity determination in multiple pregnancy. The study wascarried out on same-sex neonates born as a result of spontaneous quadruplet pregnancy. Zygosity was determinedby DNA profiling. The pregnancy was confirmed to be polyzygotic. DNA profiling may be used as a method ofzygosity determination in multiple pregnancy.

Screening and Testing in Multiples.

The choice of screening or invasive procedure in twin pregnancies is a personal choice of whether the patient wishes to take a small risk of having a baby with a serious disorder versus a small risk of having a complication because she wishes to avoid that. How to interpret such risks has profound effects on the perceived value of techniques, either leading to a decision to screening or going directly to chorionic villus sampling. There are profound issues surrounding the data and the interpretation of the data. No single short review can exhaustively examine all of the issues.

Multiple-embryo transfer for studying very early maternal-embryo interactions in cattle.

In the present paper, we highlight the need to study very early maternal-embryo interactions and discuss how these interactions can be addressed. Bovine species normally carry one or, less frequently, two embryos to term; there are very rare cases of triplets or higher-order multiple pregnancies in which all the offspring are born alive. Multiple-embryo transfer (MET) in cattle allows for the detection of endometrial responses in scenarios where single-embryo transfer would not. Although MET is non-physiological, the present study shows that at the very early embryonic stages, a uterus carrying zona-enclosed embryos does not exhibit non-physiological reactions. On the contrary, MET should be considered the sum of multiple individual effects triggered by developing embryos. We provide arguments to support our hypothesis that describe a rationale for current work with MET, and we discuss alternative hypotheses. Using cattle as a model, we describe how technical approaches to analyzing zona-enclosed early embryo-maternal interactions (i.e., transcriptomics, proteomics, and endometrial cell culture) can help identify molecular changes that may be difficult to observe when only a single embryo is present. We conclude that MET can be used for studying very early maternal-embryo interactions in vivo in monotocous species. Free Spanish abstract: A Spanish translation of this abstract is freely available at http://www.reproduction-online.org/content/150/2/R35/suppl/DC1.

Reproducción Asistida: ¿Debemos solo transferir un embrión? / Assisted Reproduction: Should be transferred only one embryo?

OBJETIVO: Comparar resultados obstétricos y perinatales de embarazos únicos espontáneos con los obtenidos por reproducción asistida, tanto únicos como gemelares. MATERIAL Y MÉTODO: Estudio descriptivo observacional retrospectivo. Se analizan un total de 1 096 embarazos de pacientes que acuden a control obstétrico en una clínica privada y se clasifican en tres grupos según el modo de concepción y número de fetos: Grupo 1, embarazo único espontáneo; Grupo 2, embarazo único por técnica de reproducción asistida (TRA) y, Grupo 3, embarazo gemelar por reproducción asistida. Se analizan los resultados obstétricos y perinatales entre los tres grupos. Se considera que hay diferencia estadística con un valor menor a <0.01 utilizando la prueba de chi cuadrado. RESULTADOS: La tasa de diabetes gestacional fue mayor en el grupo de embarazo único por TRA (23,78%) comparada a la de embarazo único espontáneo (6,66%) y gemelar por TRA (0%), presentando diferencia estadísticamente significativa. Hubo mayor presencia de anemia gestacional en el grupo de embarazo único espontáneo (76,66%) que en los grupos de TRA (19,51% y 31,25% para único y gemelar respectivamente) con diferencia estadística. Se presenta diferencia significativamente estadística a favor del grupo de embarazo gemelar por TRA en cuanto a un mayor número de casos de amenaza de parto pretérmino (25 %), una mayor ganancia de peso materno a final del embarazo (12,33 ± 3,2 Kg), un mayor número de nacimientos por cesárea (81,25%), menor edad en semanas de gestación al parto (35,28 ± 3,6), menor peso del recién nacido (2 388,69 ± 378 g) y un mayor número de casos que requieren ingreso a unidad de cuidados intensivos neonatales (18,75%). CONCLUSIONES: El embarazo gemelar tiene un peor pronóstico perinatal que el embarazo único sea espontáneo o por TRA

OBJETIVE: To determine the correlation between the semen parameters (concentration, motility, morphology and vitality) and sperm DNA integrity, using the test Halosperm. Material And METHODS: Prospective study conducted at the Laboratory of Andrology of Assisted Reproduction Laboratory FERTILAB, Lima - Peru, from August 2012 to March 2013. 282 patients were analyzed. We have established the DNA fragmentation index (IFA) from patients with semen sample normozoospermic and patients with semen sample altered in some parameters. We used two types of threshold (18% and 30%) and were correlated with semen parameters. In the total population was determined the Spearman correlation between sperm parameters and the IFA. RESULTS: It was determined that the value of the median the IFA of the patients with semen sample normozoospermic was significantly lower than that patients with semen sample altered in some parameters (12.8% vs 19.0, P=0.000). Using threshold of 18% was determined the there significant difference between the median of the two groups (Group 1: ≤ 18% y Group 2: > 18%) in age (37 years vs 40 years, P = .002), sperm concentration (82.30 X 106/ml vs 58.00 X 106/ml, P = 0.046), progressive motility (45.80% vs 27.40, P = 0.000), normal morphology (12.50% vs 9.00%, P = 0.000) and sperm vitality (89.0% vs 78.0%, P = 0.000). Using the threshold of 30 % was found significant difference between the median of the two groups (Group 1: ≤ 30% y Group 2:> 30%) in age (39 years vs 44 years, P=0.000), sperm concentration ( 78.00 X 106/ml vs. 36.75  X 106/ml, P = 0.015 ), motility progressive (40.85% vs 22.38 %, P=0.000 ), normal morphology (12.0 % vs 6.0 %, P=0.004) and sperm vitality (85.0 % vs 72.5%, P=0.000). Also identified an inverse correlation between the IFA and concentration (r=- 0.219 P=0.000), progressive motility (r=-0.452 P=0.000), normal morphology (r=-0.322 P=0.000) and vitality (r = -0.452 P = 0.000) in the total population. We determined a significant positive correlation between age and the IFA (r=0.267 P = 0.000). CONCLUSIONS: The results indicate that the level of DNA damage in sperm patients with semen sample altered in some parameters is significantly higher than in patients with semen sample normozoospermic. It has been shown that the semen parameters (concentration, motility, morphology and vitality) are highly negatively correlated with sperm DNA fragmentation index. It showed a positive correlation between age and the IFA

Associations between Individual and Combined Polymorphisms of the TNF and VEGF Genes and the Embryo Implantation Rate in Patients Undergoing In Vitro Fertilization (IVF) Programs.

BACKGROUND: A multiple pregnancy is now considered to be the most common adverse outcome associated with in vitro fertilization (IVF). As a consequence, the identification of women with the best chances of embryo implantation is a challenge in IVF program, in which the objective is to offer elective single-embryo transfer (eSET) without decreasing the pregnancy rate. To date, a range of hormonal and clinical parameters have been used to optimize eSET but none have significant predictive value. This variability could be due to genetic predispositions related to single-nucleotide polymorphisms (SNPs). Here, we assessed the individual and combined impacts of thirteen SNPs that reportedly influence the outcome of in vitro fertilisation (IVF) on the embryo implantation rate for patients undergoing intracytoplasmic sperm injection program (ICSI). MATERIALS AND METHODS: A 13 gene polymorphisms: FSHR(Asn680Ser), p53(Arg72Pro), AMH(Ile49Ser), ESR2(+1730G>A), ESR1(-397T>C), BMP15(-9C>G), MTHFR1(677C>T), MTHFR2(1298A>C), HLA-G(-725C>G), VEGF(+405G>C), TNFα(-308A>G), AMHR(-482A>G), PAI-1(4G/5G), multiplex PCR assay was designed to genotype women undergoing ICSI program. We analyzed the total patients population (n = 428) and a subgroup with homogeneous characteristics (n = 112). RESULTS: Only the VEGF(+405G>C) and TNFα(-308A>G) polymorphisms impacted fertilization, embryo implantation and pregnancy rates. Moreover, the combined VEGF+405.GG and TNFα-308.AG or AA genotype occurred significantly more frequently in women with high implantation potential. In contrast, the VEGF+405.CC and TNFα-308.GG combination was associated with a low implantation rate. CONCLUSION: We identified associations between VEGF(+405G>C) and TNFα(-308A>G) polymorphisms (when considered singly or as combinations) and the embryo implantation rate. These associations may be predictive of embryo implantation and could help to define populations in which elective single-embryo transfer should be recommended (or, conversely, ruled out). However, the mechanism underlying the function of these polymorphisms in embryo implantation remains to be determined and the associations observed here must be confirmed in a larger, more heterogeneous cohort.

Embryo number and periconceptional undernutrition in the sheep have differential effects on adrenal epigenotype, growth, and development.

Exposure to poor maternal nutrition around the time of conception results in an early prepartum activation of the fetal pituitary-adrenal axis and in increased adrenal growth and stress response after birth associated with epigenetic changes in a differentially methylated region (DMR) of adrenal IGF2/H19. We have determined the effects of maternal undernutrition during the periconceptional period (PCUN: 70% of control intake from 60 days before until 6 days after conception) and early preimplantation period (PIUN: 70% of control intake for 6 days after conception) on fetal plasma ACTH and cortisol concentrations and fetal adrenal ACTHR, StAR, 3ßHSD, CYP11B, CYP17, TGFß1, IGF1, IGF1R, IGF2, and IGF2R mRNA expression and the methylation level of sites within the DMRs of IGF2/H19 and IGF2R in the adrenal of twin and singleton fetuses at 136-138 days gestation. Being a twin resulted in a delayed prepartum increase in fetal ACTH and in a lower cortisol response to CRH in the control but not PCUN and PIUN groups. PCUN, but not PIUN, resulted in an increase in adrenal weight and CYP17 expression in singletons, a decrease in adrenal IGF2 expression in singletons, and an increase in adrenal IGF2R expression in both twins and singletons. IGF2/H19 and IGF2R DMR methylation levels and ACTHR expression were lower in the twin adrenal. Thus, exposure of the oocyte and embryo to maternal undernutrition or to the environment of a twin pregnancy have differential effects on epigenetic and other factors that regulate fetal adrenal growth and IGF2 and IGF2R expression.