Expression of granulosa cell microRNAs, AVEN and ATRX are associated with human blastocyst development.

A greater understanding of the key molecules associated with embryo development during human-assisted reproduction is imperative for the development of advanced diagnostics. Previous studies have shown that follicular microRNAs (miRNAs) are reliable markers of the polycystic ovarian syndrome (PCOS). Leveraging the utility of miRNAs in PCOS, the aim of this study was to identify miRNAs in human granulosa cells that may be indicative of blastocyst development. Granulosa cells and oocytes were collected from the first follicle aspirated from patients undergoing oocyte retrieval for in vitro fertilization or intracytoplasmic sperm injection. The development of isolated oocytes was recorded, and granulosa cell samples in this study were separated as follows. Group 1-BLAST: granulosa cells from follicles containing an oocyte that fertilized and developed into a blastocyst, and Group 2-FERT: granulosa cells from oocytes that fertilized but failed to reach blastocyst. A panel of 84 miRNAs, related to development and cellular differentiation, was assessed between the two groups using a miScript PCR array. Fourteen miRNAs and one snoRNA were differentially expressed between the groups. In addition, two downstream candidate protein biomarkers, ATRX and AVEN, were also found to be differentially expressed between the groups. The findings of this pilot study reveal follicular abnormalities on a molecular level, which may affect oocyte competence and its potential to develop successfully as an embryo. We encourage additional studies to confirm and expand on our findings and to determine the usefulness of granulosa-borne miRNAs, ATRX, and AVEN as biomarkers.

Review of clinical indicators, including serum anti-Müllerian hormone levels, for identification of women who should consider egg freezing.

OBJECTIVE: To identify women, on the basis of clinical history and serum anti-Müllerian hormone (AMH) levels, who are at risk of premature ovarian insufficiency and thereby guide appropriate early referral for egg freezing. METHODS: In a retrospective study, data were reviewed from women attending two fertility clinics in Dublin, Ireland, between August 2011 and December 2012. Case histories of women aged 35 years or younger were assessed to identify risk factors for reduced ovarian reserve, including endometriosis, ovarian surgery, and family history of premature ovarian failure (POF). RESULTS: Among 490 women aged 35 years or younger, 195 (39.7%) had an AMH level below 10 pmol/L, 94 (19.2%) had an AMH below 5 pmol/L, and 21 (4.3%) had an AMH below 1 pmol/L. Among 104 women aged 30 years or younger, the respective numbers were 28 (26.9%), 15 (14.4%), and 9 (8.7%). Among the 490 women, significantly lower AMH levels were observed for those with endometriosis (P=0.017) and a family history of POF (P=0.006). However, 53 (56.4%) of 94 women aged 35 years or younger with low AMH levels had no clinical risk factors. CONCLUSION: Universal AMH screening should be considered for all women in their 30s who are not ready to try to conceive; clinical risk factors will only identify approximately 50% of women at risk of low ovarian reserve.

A retrospective study of the effect of increasing age on success rates of assisted reproductive technology.

OBJECTIVE: To demonstrate the effect of increasing age on the outcome of assisted reproductive technology, particularly among women aged 40 years or older. METHODS: A retrospective analysis was conducted using prospectively collected data for all in vitro fertilization and intracytoplasmic sperm injection cycles among women aged 30-35 years or 40-44 years conducted at Merrion Fertility Clinic, Dublin, Ireland, between January 1, 2010, and December 31, 2014. The relationship between age and treatment outcome was assessed. RESULTS: Among women aged 30-35 years, 726 cycles led to 281 (38.7%) clinical pregnancies and 242 (33.3%) live births. By contrast, among women aged 40-44 years, 433 cycles led to 102 (23.6%) clinical pregnancies and 64 (14.8%) live births (both P<0.001). The live birth rate was particularly low after cycles among the women aged 42 years (5/89 [5.6%]) or 43 years (2/30 [6.7%]). CONCLUSION: The success rates of assisted reproductive technology are decreased among women aged older than 40 years. Fertility clinics have a responsibility to fully inform this group about the limitations of assisted reproductive technology.

Maternal Age and Contractility of Human Myometrium in Pregnancy.

There is controversy as to whether maternal age exerts an influence on the contractility of human myometrium in pregnancy. The aim of this study was to examine a series of functional contractile parameters of human myometrium in vitro, over a broad range of maternal ages. Myometrial tissue specimens were obtained at cesarean delivery from 32 women with maternal ages ranging from 28 to 52 years. Using in vitro recordings, a number of contractile parameters including maximal amplitude, mean contractile force, time to maximal amplitude, maximum rate of rise, and occurrence of simple and complex (biphasic and multiphasic) contractions were examined for spontaneous and induced contractile activity. The relationship between maternal age and individual parameters was evaluated using linear regression analysis. For all contractile parameters examined, for both spontaneous and induced contractions, no significant correlation was observed with maternal age between 28 and 52 years. The mean maximum amplitude values for spontaneous and oxytocin-induced contractions were 23 ± 3 and 43 ± 5 mN, respectively. The mean contractile forces for spontaneous and oxytocin-induced contractions were 1.5 ± 0.2 and 6.5 ± 0.9 mN, respectively. There was no variation in the proportion of biphasic or multiphasic contractions with maternal age. These results indicate there is no significant functional impairment of uterine contractility and no lack in responsiveness of myometrium in vitro, in the older mother. These findings do not support the concept that there may be a biological basis for dysfunctional labor or increased cesarean delivery rates in older parturients.

Human uterine lower segment myometrial cell and nuclear volume at term: influence of maternal age.

Little is known about the cytoarchitecture of human myometrial cells in pregnancy, and whether or not this may be influenced by maternal characteristics such as age, parity and body mass index (BMI). The aim of this study was primarily to evaluate human myometrial smooth muscle cell (SMC) and nuclear volume in the third trimester of human pregnancy, and secondarily to investigate if these parameters are altered in relation to the maternal characteristics outlined above. Myometrial biopsies were obtained from 30 women undergoing elective caesarean delivery at term. One-micrometer sections were prepared for light microscopy and 100-nm sections for electron microscopy. The nucleator technique was used to assess nuclear volume from the light microscopy images. Point-counting methodology was used on transmission electron micrographs to assess the percentage of the cell volume occupied by the nucleus. Cell volume was calculated from these measurements. The euchromatin to heterochromatin (Eu/Het) ratio was determined to ascertain whether differences in nuclear volume were due to an increased range of genes being transcribed. The mean (±â€…SEM) nuclear volume was 175 ±â€…10 µm(3) , the nucleus occupied 1.5 ±â€…0.1% of the SMC and the mean cell size was 14 047 ±â€…1352 µm(3) . The Eu/Het ratio was 7.54 ±â€…0.4. The mean volume of heterochromatin and euchromatin in the nucleus was 21.5 ±â€…1.7 and 149 ±â€…9 µm(3) , respectively. A multivariate regression analysis revealed that advanced maternal age was associated with an increase in the percentage of the cell occupied by nucleus (R(2)  = 0.32, P = 0.004). There were no other significant effects of maternal age, BMI or parity on the measured parameters. These findings provide reliable volumes for human myometrial cells and their nuclei at term gestation, and show that nuclear volume fraction may be influenced by maternal age.

The influence of smooth muscle content and orientation in dissected human pregnant myometrial strips on contractility measurements.

This study examined the hypothesis that the force generated by myometrial strips from pregnant women is influenced by the smooth muscle content and fibre orientation of the strips and that correcting for these structural variables will provide a more accurate measure of contractility. Myometrial strips (n=72) were contracted by exposure to KCl, oxytocin, U44619 and phenylephrine and maximum responses were recorded. Morphological techniques were used to determine the cross-sectional area of the strips, the area occupied by smooth muscle and the area occupied by smooth muscle longitudinal in the strip. Maximum responses to contractile agents were expressed in terms of these three variables. The mean cross sectional area of strips was 2.01 ± 0.06 mm(2), of which 50% was smooth muscle, and 18% was smooth muscle longitudinal in the strip (n=72). There was much heterogeneity in responses, smooth muscle content and fibre orientation. Correction for morphological variability did not improve the heterogeneity in responses where coefficients of variation among strips from the same donor ranged from 43% to 63% when expressed in relation to longitudinal smooth muscle cross-sectional area. The standard method of preparation of myometrial strips for in vitro recording results in samples that are not structurally uniform. Correcting for the known structural variables does not provide a more accurate measure of maximum contractile responses. Because of the heterogeneity shown here, experiments that are dependent upon accurate estimation of maximum contractile responses require a large number of replicates to reach meaningful conclusions.

The effects of extracellular calcium-sensing receptor ligands on the contractility of pregnant human myometrium in vitro.

Ligands for extracellular calcium-sensing (CaS) receptors inhibit oxytocin-induced contractions of the rat's uterus. In this study, we investigated whether the CaS receptor ligands calindol, cinacalcet, and calhex 231 have similar effects on pregnant human myometrium. We compared their effects to those of the calcium-channel blocker nifedipine. In conventional concentration-effect experiments, both the mean contractile force (MCF) and the maximum amplitude of contractions induced by 1 nmol/L oxytocin were inhibited by nifedipine. Calindol and cinacalcet were ineffective as inhibitors, while calhex-231 produced partial inhibition. When single 10 µmol/L doses were applied calhex-231 produced a slowly developing inhibition, reducing the MCF to 38%, and amplitude to 34%, of vehicle controls after 1 hour. In similar experiments, calindol was ineffective while cinacalcet weakly inhibited only the amplitude. Immunohistochemistry revealed sparse expression of CaS receptors in pregnant human myometrium.

The reliability of foetal blood sampling as a test of foetal acidosis in labour.

OBJECTIVES: To establish whether foetal blood sampling for pH is a reliable test of foetal acidosis in labour by comparing paired foetal blood samples taken at a single procedure. STUDY DESIGN: We conducted a prospective study assessing 293 consecutive attempts at foetal blood sampling in labour over a four month period from February to May 2012. A total of 100 paired samples were suitable for analysis. We compared the consistency of pH results of paired foetal blood samples, evaluated cases where inconsistent results would result in conflicting clinical decisions, and explored factors associated with discordant results. RESULTS: There was a statistically significant difference between the mean pH of the two samples: 7.297 (SD 0.065) versus 7.315 (SD 0.059), p<0.0005. Of the 100 paired samples, 43 had a difference greater than the laboratory acceptable maximum analytical difference of 0.038. There was discordance between the samples in 16 cases with results crossing a decision threshold, and in 11 cases (69%) delivery was by emergency caesarean section. Inconsistent results were not associated with specific clinical factors and occurred more often with senior operators. CONCLUSION: Foetal blood sampling is considered by many as the gold standard in assessing intrapartum foetal wellbeing. We have demonstrated inconsistency of paired foetal blood pH results which suggests that foetal blood sampling should not be considered infallible.

17 alpha-hydroxyprogesterone caproate vehicle, castor oil, enhances the contractile effect of oxytocin in human myometrium in pregnancy.

OBJECTIVE: The possibility exists that the vehicle for 17-alpha-hydroxyprogesterone caproate, castor oil, exerts an effect on human uterine contractility. The aim of this study was to evaluate its effects on contractility of myometrial preparations that were obtained during pregnancy. STUDY DESIGN: Myometrial strips were suspended under isometric conditions. Contractility was induced with oxytocin. Strips were incubated in castor oil or physiologic salt solution and suspended for a further oxytocin challenge. Contractile integrals were compared between both groups. RESULTS: Strips that were exposed to castor oil demonstrated increased contractile activity that was elicited by oxytocin (mean contractility value, 165.53%+/-17.03%; n=8; P=.004), compared with control strips (mean contractility value, 72.57%+/-7.48%; n=8; P=.003). There was a significant increase in contractile activity of the castor oil-exposed strips, compared with those that were exposed to physiologic salt solution (n=8; P<.001). CONCLUSION: Exposure of human myometrial preparations to castor oil results in enhanced oxytocin-induced contractility.