Heat shock protein 90 plays critical roles in client protein maturation, signal transduction, protein folding and degradation, and morphological evolution; however, its function in human sperm is not fully understood. Therefore, our objective in this study was to elucidate the mechanism by which heat shock protein 90 exerts its effects on human sperm function. By performing indirect immunofluorescence staining, we found that heat shock protein 90 was localized primarily in the neck, midpiece, and tail regions of human sperm, and that its expression increased with increasing incubation time under capacitation conditions. Geldanamycin, a specific inhibitor of heat shock protein 90, was shown to inhibit this increase in heat shock protein 90 expression in western blotting analyses. Using a multifunctional microplate reader to examine Fluo-3 AM-loaded sperm, we observed for the first time that inhibition of heat shock protein 90 by using geldanamycin significantly decreased intracellular calcium concentrations during capacitation. Moreover, western blot analysis showed that geldanamycin enhanced tyrosine phosphorylation of several proteins, including heat shock protein 90, in a dose-dependent manner. The effects of geldanamycin on human sperm function in the absence or presence of progesterone was evaluated by performing chlortetracycline staining and by using a computer-assisted sperm analyzer. We found that geldanamycin alone did not affect sperm capacitation, hyperactivation, and motility, but did so in the presence of progesterone. Taken together, these data suggest that heat shock protein 90, which increases in expression in human sperm during capacitation, has roles in intracellular calcium homeostasis, protein tyrosine phosphorylation regulation, and progesterone-stimulated sperm function. In this study, we provide new insights into the roles of heat shock protein 90 in sperm function.

Purpose The study was designed to evaluate whether cumulus cell removal 4 h post-insemination could influence fertilization and embryo quality. Methods The study included 61couples undergoing standard long down regulation protocol from July 2011 to May 2012. Sibling oocytes of each patient were randomly assigned to either the 4 h group or the 20 group. For the 4 h group, cumulus cells were removed 4 h after gamete coincubation; for the 20 group, cumulus cells removal was performed 20 h after insemination. Fertilization rate, embryo quality, pregnancy rate and implantation rate were assessed. Results A total of 801 sibling cumulus-oocyte complexes (COCs) were randomized to the 4 h group (421 COCs) or 20 h group (380 COCs). There was no difference in the two pronuclei, one pronucleus and grade 1-2 embryo rate. Three pronuclei rate was significantly higher in the 4 h group compared to the 20 h group (12.6 % vs. 8.2 %, P=0.041). Comparison of embryo transfer cycles in which either embryos from the 4 h group or 20 h group were transferred did not reveal any statistically significant differences in pregnancy or implantation rates. Conclusion The results of the present study indicate that cumulus cell removal 4 h post-insemination may increase the percentage of tripronuclear zygotes. However, normal fertilization rate, embryo development, clinical pregnancy rate and implantation rates are not influenced by the timing of cumulus cell removal.

Purpose This study aimed to compare slow freezing (SF) and vitrification (VT) techniques for day 3 embryo cryopreservation in infertile couples. Methods This retrospective cohort study enrolled 5613 infertile patients, with 7862 frozen-thawed day 3 embryos and 3845 vitrified-warmed day 3 embryos, from 2010 to 2014, at a single center. The rates of embryo survival, pregnancy, implantation, miscarriage, live birth, and live birth weight were compared between the two groups. Results A total of 5613 cycles with 5520 transfers were analyzed. Using SF, the rates of overall embryo survival and fully intact blastomeres were lower than those in VT (91.5 vs. 97.4 % and 68.7 vs. 92.3 %, respectively). The rate of good quality embryos after thawing/warming was lower in SF than in VT. In single frozen embryo transfer cycles (FETs), the pregnancy and implantation rates were similar between the two groups (35.0 vs. 40.8 % and 34.6 vs. 35.9 %, respectively). In double FETs, the pregnancy rate per cycle was also similar between the groups (58.8 vs. 58.4 %). The implantation rate per embryo transfer was significantly higher with SF than with VT (38.8 vs. 34.6 %). With adjustment for maternal age and the number of good quality embryos, differences in implantation rate remained significant (adjusted P value, SF vs. VT P<0.05). No independent effect was found for the method of cryopreservation on the pregnancy rate. No significant differences in the rates of miscarriage, live birth, and live birth weight were observed between the two techniques. Conclusions Despite the significantly low embryo survival rate, fully intact blastomere rate, and good quality embryo rate in SF, the pregnancy and implantation rates were not adversely affected in single and double FETs. SF yielded an equivalent miscarriage rate, live birth rate, and live birth weight compared with VT. The SF protocol to cryopreserve day 3 embryos still should be considered.

This pilot study evaluated the effects of coadministration of metformin with clomiphene citrate (CC) and human menopausal gonadotrophin (HMG) in women with CC-resistant polycystic ovary syndrome (PCOS). Sixty women with PCOS were randomly assigned to receive 3 months' treatment with metformin or placebo together with CC and HMG. Transvaginal ultrasound was used to monitor follicular development and ovulation was induced by human chorionic gonadotrophin (HCG). The number of dominant follicles, the oestradiol level on the day HCG was given and the amount of HMG required were significantly lower in the metformin group than in the placebo group, whereas the mono-ovulatory rate and pregnancy rate in the third cycle were significantly higher. The cumulative pregnancy rate in the metformin group (43.3%) was higher than in the placebo group (20.0%), but this difference did not reach statistical significance. In conclusion, coadministration of metformin with CC and HMG reduced the amount of HMG required and increased the mono-ovulatory rate and pregnancy rate.

Purpose The study was designed to investigate the effect of vitrification and slow freezing for the cryopreservation of human day 3 embryos on serum β-hCG levels in pregnancies established after frozen embryo transfer (FET). Methods Of the 1384 FET cycles initiated, 1131 embryo transfers met study criteria and assigned to one of two groups: 797 slow-freezing embryo transfers or 334 vitrified embryo transfers. Median values of β-hCG and outcome of all pregnancies were compared between the two groups. Predictive values of serum β-hCG on day 12 after embryo transfer for establishing ongoing pregnancy and pregnancy failure were determined by receiver operating characteristic (ROC) curve analysis. Results In the slow-freezing group, 383 ongoing pregnancies occurred (48.1 %), and transfers of vitrified embryos resulted in 154 pregnancies (46.1 %). Median β-hCG values (279.2 IU/L) for ongoing pregnancies after transfer of vitrified embryos were significantly lower than that of slow frozen embryos (320.5 IU/L). The median values of β-hCG for singleton in the two groups was statistically significant (P<0.05). For slowfreezing embryo transfers, the cut-off value of β-hCG in predicting ongoing pregnancy was 147 IU/L (sensitivity 88.3 %, specificity 80.7 %). For vitrified embryo transfers, the value was 135 IU/L (sensitivity 84.4 %, specificity 76.3 %). Conclusions Day 12 β-hCG levels after FET are significantly affected by the methods of embryo cryopreservation for ongoing pregnancies. Furthermore, when using β-hCG cutoff value to assess pregnancy outcome, the cryopreservation methods should be taken into account.

S100P was originally isolated from the placenta, and is expressed in very high levels in trophoblast cells, but its role on trophoblast cells proliferation has not yet been studied. In this study, we aimed to investigate the potential role of S100P in human placental development, and the impact of its expression regulation on cellular function as well as molecular mechanisms involved in trophoblast-like cells. We found that the expression of S100P in first trimester placenta was significantly reduced in spontaneous abortion patients with respect to normal pregnant women. Up-regulation of S100P in JAR cells promoted JAR cells proliferation, and increased the expression of phosphorylated P38 (p-P38) mitogen-activated protein kinase (MAPK) and p-ERK MAPK. However, the effects of S100P on JAR cells proliferation were prevented by P38 inhibitor-SB203580, but not by ERK inhibitor-PD98059. These results showed that S100P may have a physiological role in normal pregnant development, and regulate trophoblast-like cell proliferation via modulating the P38 MAPK pathway.

Purpose The study was designed to evaluate the relationship between serum progesterone (P4) response after hCG administration and the number of oocytes retrieved and the embryo quality in fresh IVF cycles. Methods We conducted a retrospective cohort study of women aged 24-43 years who underwent first fresh IVF cycle from 2011 to 2013 at a single practice. We compared the post-hCG serum P4 level with values on the day of hCG trigger. Patients were analyzed in long and short protocols independently. In addition, patients were stratified by post-hCG P4 response. Number of oocytes retrieved and embryo quality were the primary outcomes of interest. Ordinary least square regression models and logistic regression analysis models were created to identify predictive factors associated with embryological outcomes while adjusting for potential confounders. Results Among the 2,978 IVF cycles, 2,484 patients were in long protocols, and 494 patients were in short protocols. After adjusting for patient age, rFSH duration, and basal FSH levels, the associations between P4 response after hCG administration and number of oocytes retrieved (P<0.001) remained statistically significant in both long and short protocols. Additionally, mature oocyte rate, fertilization rate, good quality embryo rate, pregnancy rate and implantation rate were not significantly associated with the P4 increase when adjusting for the same factors. However, pregnancy rate and implantation rate from frozen-thawed cycles increased gradually across the seven groups.Conclusions Post-hCG P4 levels were positively associated with the number of oocytes retrieved, but did not affect oocyte or embryo quality. Our study suggests that the change in the posthCG P4 level is another parameter that can be used by clinicians to assess the number of oocytes retrieved, and may further to estimate the pregnancy rate and live birth rate indirectly.

This retrospective study determined the efficacy of the 'freeze-all' embryo strategy in poor ovarian responders undergoing ovarian stimulation for in vitro fertilization (IVF). A total of 559 poor responders who met Bologna criteria between January 2012 and December 2014 were included in this study: 256 in the fresh embryo transfer group and 303 in the freeze-all group. Vitrification and warming of day 3 embryos were performed using the Cryotop method. The poor responders treated with fresh embryo transfer and those treated with freeze-all strategy showed similar live birth rates per cycle (12.1% vs. 16.2%, p = .172) and per transfer (15.9% vs. 20.9%, p = .182). Multivariate logistic regression analysis showed that maternal age at retrieval (odds ratio, 0.919; 95% confidence interval, 0.865-0.977; p = .006) and number of good-quality embryos transferred (odds ratio, 1.953; 95% confidence interval, 1.346-2.835; p < .001) were significantly associated with the live birth rate. Freeze-all cycle is an acceptable treatment in poor ovarian responders, and it should be suggested by physicians as an alternative to cycle cancelation in case in which a fresh transfer would not be advantageous.