Predictive models of recurrent implantation failure in patients receiving ART treatment based on clinical features and routine laboratory data.

STUDY QUESTION: The objective was to construct a model for predicting the probability of recurrent implantation failure (RIF) after assisted reproductive technology (ART) treatment based on the clinical characteristics and routine laboratory test data of infertile patients. A model was developed to predict RIF. The model showed high calibration in external validation, helped to identify risk factors for RIF, and improved the efficacy of ART therapy. WHAT IS KNOWN ALREADY: Research on the influencing factors of RIF has focused mainly on embryonic factors, endometrial receptivity, and immune factors. However, there are many kinds of examinations regarding these aspects, and comprehensive screening is difficult because of the limited time and economic conditions. Therefore, we should try our best to analyse the results of routine infertility screenings to make general predictions regarding the occurrence of RIF. STUDY DESIGN, SIZE, DURATION: A retrospective study was conducted with 5212 patients at the Reproductive Center of the First Affiliated Hospital of USTC from January 2018 to June 2022. PARTICIPANTS/MATERIALS, SETTING, METHODS: This study included 462 patients in the RIF group and 4750 patients in the control group. The patients' basic characteristics, clinical treatment data, and laboratory test indices were compared. Logistic regression was used to analyse RIF-related risk factors, and the prediction model was evaluated by receiver operating characteristic (ROC) curves and the corresponding areas under the curve (AUCs). Further analysis of the influencing factors of live births in the first cycle of subsequent assisted reproduction treatment in RIF patients was performed, including the live birth subgroup (n = 116) and the no live birth subgroup (n = 200). MAIN RESULTS AND THE ROLE OF CHANCE: (1) An increased duration of infertility (1.978; 95% CI, 1.264-3.097), uterine cavity abnormalities (2.267; 95% CI, 1.185-4.336), low AMH levels (0.504; 95% CI, 0.275-0.922), insulin resistance (3.548; 95% CI, 1.931-6.519), antinuclear antibody (ANA)-positive status (3.249; 95% CI, 1.20-8.797) and anti-ß2-glycoprotein I antibody (A-ß2-GPI Ab)-positive status (5.515; 95% CI, 1.481-20.536) were associated with an increased risk of RIF. The area under the curve of the logistic regression model was 0.900 (95% CI, 0.870-0.929) for the training cohort and 0.895 (95% CI, 0.865-0.925) for the testing cohort. (2) Advanced age (1.069; 95% CI, 1.015-1.126) was a risk factor associated with no live births after the first cycle of subsequent assisted reproduction treatment in patients with RIF. Blastocyst transfer (0.365; 95% CI = 0.181-0.736) increased the probability of live birth in subsequent cycles in patients with RIF. The area under the curve of the logistic regression model was 0.673 (95% CI, 0.597-0.748). LIMITATIONS, REASONS FOR CAUTION: This was a single-centre regression study, for which the results need to be evaluated and verified by prospective large-scale randomized controlled studies. The small sample size for the analysis of factors influencing pregnancy outcomes in subsequent assisted reproduction cycles for RIF patients resulted in the inclusion of fewer covariates, and future studies with larger samples and the inclusion of more factors are needed for assessment and validation. WIDER IMPLICATIONS OF THE FINDINGS: Prediction of embryo implantation prior to transfer will facilitate the clinical management of patients and disease prediction and further improve ART treatment outcomes. STUDY FUNDING/COMPETING INTEREST(S): This work was supported by the General Project of the National Natural Science Foundation of China (Nos. 82,201,792, 82,301,871, 81,971,446, and 82,374,212) and the Natural Science Foundation of Anhui Province (No. 2208085MH206). There are no conflicts of interest to declare. TRIAL REGISTRATION NUMBER: This study was registered with the Chinese Clinical Trial Register (Clinical Trial Number: ChiCTR1800018298 ).

Association between MTHFR c.677C>T variant and erectile dysfunction among males attending fertility clinic.

ABSTRACT: Genetic risk factors have been shown to contribute to the development of sexual dysfunction. However, the role of methylenetetrahydrofolate reductase (MTHFR) gene variants in the risk of erectile dysfunction (ED) remains unclear. In this study, we recruited 1254 participants who underwent ED assessed by the International Index of Erectile Function-5. The MTHFR c.677C>T variant was also measured by fluorescence polymerase chain reaction (PCR). No significant difference in the genotypic frequency of the MTHFR C677T polymorphism (CC, CT, and TT) was observed between men from the ED and non-ED groups. In addition, on binary logistic regression analysis, both crude and adjusted models showed that the risk of ED was not significantly associated with the C677T polymorphism. Interestingly, a significantly higher frequency of the 677TT polymorphism was found in severe and moderate ED (P = 0.02). The positive correlation between the MTHFR 677TT polymorphism and severe ED was confirmed by logistic regression analysis, even after adjusting for potential confounders (odds ratio [OR] = 2.46, 95% confidence interval [CI]: 1.15-5.50, P = 0.02). These findings suggest a positive correlation between the MTHFR 677TT polymorphism and the risk of severe ED. Identification of MTHFR gene polymorphisms may provide complementary information for ED patients during routine clinical diagnosis.

Safety of early cumulus cell removal combined with early rescue ICSI in the prevention of fertilization failure.

RESEARCH QUESTION: What are the clinical outcomes and safety implications of early cumulus cell removal after short-term insemination combined with early rescue intracytoplasmic sperm injection (ICSI) in preventing fertilization failure? DESIGN: In this retrospective study, a total of 14,360 cycles were divided into four groups based on insemination method and fertilization ability: conventional IVF group (n = 5519); early cumulus cell removal group (n = 4107); conventional ICSI group (n = 4215); and early rescue ICSI group (where failed or low fertilization was predicted, n = 519). Fertilization outcomes, pregnancy outcomes, neonatal outcomes and birth defects were analysed by comparing the early cumulus cell removal group with the conventional IVF group, and the early rescue ICSI group with the conventional ICSI group. RESULTS: There were no significant differences in the outcomes of fertilization, pregnancy, neonates or birth defects between the conventional IVF group and the early cumulus cell removal group (P > 0.05). When compared with the conventional ICSI group, the early rescue ICSI group had similar rates of two pronuclei (2PN) at fertilization, clinical pregnancy, miscarriage, ectopic pregnancy, live birth, sex, mean gestational age, very low birthweight, macrosomia and birth defects (P > 0.05) but a higher polyploidy rate, lower high-quality embryo rate (both P < 0.001), lower twin pregnancy rate (P < 0.01), lower rate of low birthweight, and a higher rate of normal birthweight (both P = 0.024). CONCLUSIONS: Early cumulus cell removal combined with early rescue ICSI led to good pregnancy and neonatal outcomes without an increase in birth defects. This approach could therefore be an effective and safe method for patients with fertilization failure in conventional IVF.

Association between semen collection time and semen parameters: an observational study.

The process of semen collection plays a key role in the quality of semen specimens. However, the association between semen collection time and semen quality is still unclear. In this study, ejaculates by masturbation from 746 subfertile men or healthy men who underwent semen analysis were examined. The median (interquartile range) semen collection time for all participants was 7.0 (5.0-11.0) min, and the median time taken for semen collection was lower in healthy men than that in subfertile men (6.0 min vs 7.0 min). An increase in the time required to produce semen samples was associated with poorer semen quality. Among those undergoing assisted reproductive technology (ART), the miscarriage rate was positively correlated with the semen collection time. After adjusting for confounders, the highest quartile (Q4) of collection time was negatively associated with semen volume and sperm concentration. A longer time to produce semen samples (Q3 and Q4) was negatively correlated with progressive and total sperm motility. In addition, there was a significant negative linear association between the semen collection time and the sperm morphology. Higher risks of asthenozoospermia (adjusted odds ratio [OR] = 2.06, 95% confidence interval [CI]: 1.31-3.25, P = 0.002) and teratozoospermia (adjusted OR = 1.98, 95% CI: 1.10-3.55, P = 0.02) were observed in Q3 than those in Q1. Our results indicate that a higher risk of abnormal semen parameter values was associated with an increase in time for semen collection, which may be related to male fertility through its association with semen quality.

Association of sexually transmitted infection with semen quality in men from couples with primary and secondary infertility.

This study aims to compare the prevalence of sexually transmitted infections (STIs) with semen quality in men from couples with primary and secondary infertility. Semen samples were collected from 133 men who requested fertility evaluation. Seminal tract infection with Ureaplasma spp. (UU), Mycoplasma hominis (MH), Mycoplasma genitalium (MG), Chlamydia trachomatis (CT), Neisseria gonorrhoeae (NG), and herpes simplex virus-2 (HSV-2) was assessed by PCR-based diagnostic assays. Among all patients, the prevalence of STIs was higher in men from couples with primary infertility than that in men from couples with secondary infertility (39.7% vs 21.7%, P = 0.03). The prevalence of UU was 28.8% and 13.3% in men from couples with primary and secondary infertility, respectively. Men from couples with primary infertility were more likely to be positive for UU than men from couples with secondary infertility (P = 0.04). Regarding the UU subtype, the prevalence of Ureaplasma urealyticum (Uuu) and Ureaplasma parvum (Uup; including Uup1, Uup3, Uup6, and Uup14) did not differ between the two groups. No associations between the prevalence rates of MH, MG, and CT were found in men from either infertility group. A lower sperm concentration was associated with STI pathogen positivity in men with primary infertility according to the crude model (P = 0.04). The crude and adjusted models showed that semen volume (both P = 0.03) and semen leukocyte count (both P = 0.02) were independently associated with secondary infertility. These findings suggest the importance of classifying the type of infertility during routine diagnosis of seminal tract infections.

Does the elevated estradiol level on the day of human chorionic gonadotropin administration affect embryo quality among women undergoing in vitro fertilization?

OBJECTIVE: To evaluate the association between estradiol (E2 ) levels on the day of human chorionic gonadotropin (hCG) administration and embryo quality during in vitro fertilization (IVF) cycles. METHODS: A retrospective study of 6676 IVF cycles among women treated at the Reproductive Center of The First Affiliated Hospital of USTC, Hefei, China, from June 2014 to May 2017. E2 levels on hCG day were divided into four groups by 25th percentile: 0-1763 pg/mL (group I), 1763-3692 pg/mL (group II), 3692-4800 pg/mL (group III), and higher than 4800 pg/mL (group IV). Analysis of variance and multiple linear regression were used to test associations. RESULTS: There were significant differences in the frequency of high-quality embryos between group I (51.6 ± 1.1%) and groups II (65.6 ± 0.8%), III (62.1 ± 0.7%), and IV (62.3 ± 0.7%). Using E2 as a dummy variable and group II as a control, multiple linear regression showed that E2 levels were associated with the frequency of high-quality embryos obtained (P < 0.05). CONCLUSION: Serum E2 on hCG day had an impact on embryo quality. Higher E2 levels did not produce the most high-quality embryos; the highest frequencies were achieved for E2 levels within 1763-3692 pg/mL.

Identification of microRNAs that Regulate the MAPK Pathway in Human Cumulus Cells from PCOS Women with Insulin Resistance.

Polycystic ovary syndrome (PCOS) is one of the most common gynaecological endocrine disorders, and more than 60% of PCOS patients have varying degrees of insulin resistance (IR). The regulatory role of microRNAs (miRNAs) at post-transcriptional levels in human cumulus cells relating to IR in PCOS remains unclear. In this case-control study, 26 PCOS patients with IR (PCOS-IR) and 24 patients without IR (PCOS-control) were enrolled. We determined the differentially expressed miRNA and mRNA using next-generation sequencing technology, and these miRNAs and mRNAs were validated by quantitative real-time polymerase chain reaction (PCR). These miRNA regulating pathways (e.g., MAPK pathway) were analysed by bioinformatics analysis, and the Rap1b was demonstrated to be targeted by miR-612 based on quantitative real-time PCR, western blot and luciferase activity assay. A total of 59 known miRNAs and 617 differentially expressed genes were identified that differentially expressed between PCOS-IR and PCOS-control cumulus cells. Moreover, the potential regulating roles of miRNAs and their targeting genes in pathophysiology of IR and PCOS were analysed by gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway annotation, and several key processes were enriched, such as MAPK activity. Furthermore, Rap1b, a regulator of the MAPK pathway, was demonstrated to be suppressed directly by miR-612 in PCOS-IR cumulus cells based on negative expression correlation validation, dual luciferase activity assay and reduction of Rap1b expression after miR-612 mimics transfection. Our results suggested that miRNAs and their targeted pathways in ovarian cumulus cells may play important roles in the aetiology and pathophysiology of PCOS with IR.

IL-7/IL-7R signaling pathway might play a role in recurrent pregnancy losses by increasing inflammatory Th17 cells and decreasing Treg cells.

PROBLEM: We aim to investigate a possible role of IL-7/IL-7R signaling pathway in recurrent pregnancy losses (RPL). MATERIAL AND METHODS: Using the abortion-prone (AP) and non-abortion-prone (NP) mice model, fetal resorption rates (FRR), Th17 and Treg cells-related factors, and the effect of IL-7 and IL-7R antagonist were investigated by flow cytometry, quantitative real-time PCR, and immunohistochemistry. IL-7 and IL-7R expressions in human decidua were investigated by immunohistochemistry. RESULTS: In the AP mice, IL-7R antagonist treatment significantly decreased FRR by downregulating Th17 and upregulating Treg-related factors. When the NP mice were treated with IL-7, FRR was significantly increased by upregulating Th17 and downregulating Treg-related factors. In decidual stromal cells of women with RPL, increased IL-7 and decreased IL-7R expressions were present when compared to normal controls. CONCLUSION: IL-7/IL-7R signaling pathway plays a possible role in RPL by upregulating Th17 immunity, meanwhile downregulating Treg immunity. Regulation of IL-7/IL-7R may be a new therapeutic strategy for RPL.

Expression Profile of microRNAs and Their Targeted Pathways in Human Ovaries Detected by Next-Generation Small RNA Sequencing.

Recently, post-transcriptional gene regulation by microRNAs (miRNAs) has been reported to play a key role during ovary development and differentiation. However, there are no published studies identifying miRNA profiles of human ovarian tissues directly using next-generation sequencing technology. In the human ovary, a total of 762 known and 21 novel human miRNAs were detected, indicating that human ovaries have a complex population of small RNAs. To confirm the miRNA profile in human ovaries, quantitative real-time polymerase chain reaction was used to validate the expression of known miRNAs and novel miRNAs. The potential regulating roles of miRNA in physiological function of ovaries were analyzed by gene ontology and Kyoto encyclopedia of genes and genomes pathway annotation, and several important processes were identified to be targeted by the most abundantly expressed miRNAs, for example, antral ovarian follicle growth, ovarian follicle rupture, and fertilization. Our current findings extend the knowledge of the regulatory role of miRNAs and their targeted processes in human ovaries, suggesting that miRNAs play important roles in development and physiological function of ovaries. In this study, we provide a useful resource for further research of the regulatory role of miRNAs in the ovaries, which may also provide novel candidates for molecular biomarkers or treatment targets in the research of female infertility.

Oocyte quality is decreased in women with minimal or mild endometriosis.

Endometriosis, a pathological condition in which the endometrium grows outside the uterus, is one of the most common causes of female infertility; it is diagnosed in 25-40% of infertile women. The mechanism by which endometriosis affects the fertility of females remains largely unknown. We examined the ultrastructure of oocytes from patients with minimal or mild endometriosis and control females undergoing in vitro fertilization (IVF) treatment by transmission electron microscopy (TEM) to investigate the physiological significance of oocyte quality for patients with minimal or mild endometriosis. The TEM results revealed that the oocytes from women with minimal or mild endometriosis exhibited abnormal mitochondrial structure and decreased mitochondria mass. Quantitative real time PCR analysis revealed that the mitochondrial DNA copy number was significantly reduced in the oocytes from women with minimal or mild endometriosis compared with those of the control subjects. Our results suggest that decreased oocyte quality because of impaired mitochondrial structure and functions probably an important factor affecting the fertility of endometriosis patients.

Characterization of microRNA profile in human cumulus granulosa cells: Identification of microRNAs that regulate Notch signaling and are associated with PCOS.

CONTEXT: Polycystic ovary syndrome (PCOS), a complex and heterogeneous endocrine condition, is characterized by polycystic ovaries, hyperandrogenism, insulin resistance and chronic anovulation. Cumulus granulosa cells surrounding the oocyte are involved in different aspects of PCOS pathology. Several studies suggested that miRNAs play an important regulatory role at the post-transcriptional level in cumulus granulosa cells. OBJECTIVE: Our objective was to describe the altered miRNA expression profiles and miRNA targeted signaling pathways in PCOS. DESIGN: Case-control study that involved 21 women with PCOS and 20 women without the disease (controls). The miRNA expression profiles of human cumulus granulosa cells were determined using next generation sequencing by Illumina Hiseq 2000. The differentially expressed miRNAs and novel miRNAs were validated by quantitative real-time PCR. The Notch3 and MAPK3 were demonstrated to be targeted by miR-483-5p based on quantitative real-time PCR, western blot and luciferase activity assay. RESULTS: Compared with controls, a total of 59 known miRNA were identified that differentially expressed in PCOS cumulus granulosa cells, including 21 miRNAs increase and 38 miRNAs decrease. Moreover, the novel miRNAs were predicted in PCOS and control cumulus granulosa cells. The potential regulating roles of miRNA in pathophysiology of PCOS were analyzed by GO and KEGG pathway annotation, and several important processes were identified to be targeted by the differentially expressed miRNAs, such as Notch signaling, regulation of hormone, and energy metabolism. Furthermore, Notch3 and MAPK3, the members of Notch signaling and ERK-MAPK pathway, were demonstrated to be regulated by miR-483-5p based on negative expression correlation validation and detection of Notch3/MAPK3 expression after miR-483-5p mimics transfection. Dual luciferase activity assay suggested that Notch3 and MAPK3 were directly targeted by miR-483-5p. CONCLUSION: Our data suggested that miRNAs and their targeted pathways (e.g. Notch signaling pathway) play important roles in the etiology and pathophysiology of PCOS, and provides novel candidates for molecular biomarkers or treatment targets in the research of female infertility associated with PCOS.

Research resources: comparative microRNA profiles in human corona radiata cells and cumulus oophorus cells detected by next-generation small RNA sequencing.

During folliculogenesis, cumulus cells surrounding the oocyte differentiate into corona radiata cells (CRCs) and cumulus oophorus cells (COCs), which are involved in gonadal steroidogenesis and the development of germ cells. Several studies suggested that microRNAs (miRNAs) play an important regulatory role at the post-transcriptional level in cumulus cells. However, comparative miRNA profiles and associated processes in human CRCs and COCs have not been reported before. In this study, miRNA profiles were obtained from CRCs and COCs using next generation sequencing in women undergoing controlled ovarian stimulation for IVF. A total of 785 and 799 annotated miRNAs were identified in CRCs and COCs, while high expression levels of six novel miRNAs were detected both in CRCs and in COCs. In addition, different expression patterns in CRCs and COCs were detected in 72 annotated miRNAs. To confirm the miRNA profile in COCs and CRCs, quantitative real-time PCR was used to validate the expression of annotated miRNAs, differentially expressed miRNAs, and novel miRNAs. The miRNAs in the let-7 family were found to be involved in the regulation of a broad range of biological processes in both cumulus cell populations, which was accompanied by a large amount of miRNA editing. Bioinformatics analysis showed that amino acid and energy metabolism were targeted significantly by miRNAs that were differentially expressed between CRCs and COCs. Our work extends the current knowledge of the regulatory role of miRNAs and their targeted pathways in folliculogenesis, and provides novel candidates for molecular biomarkers in the research of female infertility.

Oocytes with a dark zona pellucida demonstrate lower fertilization, implantation and clinical pregnancy rates in IVF/ICSI cycles.

The morphological assessment of oocytes is important for embryologists to identify and select MII oocytes in IVF/ICSI cycles. Dysmorphism of oocytes decreases viability and the developmental potential of oocytes as well as the clinical pregnancy rate. Several reports have suggested that oocytes with a dark zona pellucida (DZP) correlate with the outcome of IVF treatment. However, the effect of DZP on oocyte quality, fertilization, implantation, and pregnancy outcome were not investigated in detail. In this study, a retrospective analysis was performed in 268 infertile patients with fallopian tube obstruction and/or male factor infertility. In 204 of these patients, all oocytes were surrounded by a normal zona pellucida (NZP, control group), whereas 46 patients were found to have part of their retrieved oocytes enclosed by NZP and the other by DZP (Group A). In addition, all oocytes enclosed by DZP were retrieved from 18 patients (Group B). No differences were detected between the control and group A. Compared to the control group, the rates of fertilization, good quality embryos, implantation and clinical pregnancy were significantly decreased in group B. Furthermore, mitochondria in oocytes with a DZP in both of the two study groups (A and B) were severely damaged with several ultrastructural alterations, which were associated with an increased density of the zona pellucida and vacuolization. Briefly, oocytes with a DZP affected the clinical outcome in IVF/ICSI cycles and appeared to contain more ultrastructural alterations. Thus, DZP could be used as a potential selective marker for embryologists during daily laboratory work.

Human oocyte vitrification with corona radiata, in autologous follicular fluid supplemented with ethylene glycol, preserves conventional IVF potential: birth of four healthy babies.

The aim of this study was to examine the effects of vitrification with autologous follicular fluid (AFF) supplemented with ethylene glycol (EG) and sucrose on human oocytes with corona radiata. A total of 182 human oocytes with corona radiata from fifteen infertile patients were vitrified using either equilibration solutions (ES) and vitrification solution (VS) consisting of AFF, EG and sucrose (AFF group, n=67) or commercial ES and VS (control group, n=115). All oocytes were thawed in the next cycle, with surviving oocytes being inseminated by conventional IVF. The clinical outcome of vitrified-warmed oocytes by both vitrification methods was analysed retrospectively. In the AFF group, six patients received embryo transfer, with three couples taking four healthy babies home. In the control group, nine patients received embryo transfer, with four couples taking five healthy babies home. There was no significant difference in the survival rate (91.0 vs 92.2%), two pronuclei (2PN) fertilisation rate (73.8 vs 73.6%), cleavage rate (100 vs 100%), top-quality embryo rate (62.2 vs 59.2%), clinical pregnancy rate (50.0 vs 44.4%), implantation rate (33.3 vs 25%) or take-home baby rate (50.0 vs 44.4%) between the AFF group and the control group, respectively. These results show that AFF supplemented with EG and sucrose is an efficient, cost-effective cryoprotectant for human oocyte cryopreservation. A corona radiata on vitrified-warmed oocytes retains the oocytes' fertilisation capability in conventional IVF.

A retrospective study on IVF/ICSI outcome in patients with anti-nuclear antibodies: the effects of prednisone plus low-dose aspirin adjuvant treatment.

BACKGROUND: Anti-nuclear antibodies (ANA) are suspected of having relevance to adverse reproductive events. METHODS: This study aims to investigate the potential effect of ANA on IVF/ICSI outcome and the therapeutic role of prednisone plus low-dose aspirin (P + A) adjuvant treatment in ANA + patients. The first IVF/ICSI cycles without P + A of sixty-six ANA + women were enrolled as the ANA + group, and the 233 first IVF/ICSI cycles of matched ANA- women served as the ANA- group. The ANA + group was divided into the Titre < =1:320 subgroup and the Titre > 1:320 subgroup. Twenty-one ANA + women with adverse outcomes in their first cycles (ANA + cycles without P + A) received P + A adjuvant treatment for three months before the second IVF/ICSI cycle (ANA + cycles with P + A). The clinical characteristics and the IVF/ICSI outcomes were compared, respectively, between 1) the ANA + group and the ANA- group, 2) the Titre < =1:320 subgroup and the Titre > 1:320 subgroup, and 3) the ANA + cycles without P + A and the ANA + cycles with P + A. RESULTS: No significant differences were observed between each of the two-group pairs in the clinical characteristics. The ANA + group exhibited significantly lower MII oocytes rate, normal fertilisation, pregnancy and implantation rates, as well as remarkably higher abnormal fertilisation and early miscarriage rates. The Titre < =1:320 subgroup's IVF/ICSI outcomes were as poor as those of the Titre > 1:320 subgroup. After the P + A adjuvant treatment, the number of two pro-nuclei, perfect embryos and available embryos, and the implantation rate increased significantly. CONCLUSIONS: These observations suggest that ANA could exert a detrimental effect on IVF/ICSI outcome that might not be titre-dependent, and P + A adjuvant treatment could be useful for ANA + patients. This hypothesis should be verified in further prospective randomised studies.

The optimum number of oocytes in IVF treatment: an analysis of 2455 cycles in China.

STUDY QUESTION: What is the association between the number of oocytes retrieved and the live birth rate (LBR) following first IVF treatment cycles in China? SUMMARY ANSWER: In first IVF treatment cycles, the LBR after fresh transfer was maximal in the groups with 6-10 or 11-15 oocytes and reduced in the groups with 0-5 or >15 oocytes. Despite this, the cumulative LBR after including frozen embryo transfer cycles increased with an ovarian response. WHAT IS KNOWN ALREADY: There is a strong association between oocyte number and IVF outcome; however, this is a comprehensive analysis conducted to investigate the relationship between oocyte number and fresh cycle as well as cumulative LBRs in first IVF treatment cycles. STUDY DESIGN, SIZE AND DURATION: This is a large retrospective cohort study (n = 2455); patients were categorized into four groups according to the number of oocytes retrieved (0-5, 6-10, 11-15 and >15 oocytes). The fresh embryo transfer LBR and cumulative LBR were evaluated by group. Univariate analysis was performed to identify factors that predict the chance of LBR. Multivariate logistic regression was used to assess the association between oocyte number and LBR after adjusting for confounding factors that were identified as significant in the univariate analysis. PARTICIPANTS/MATERIALS, SETTING AND METHODS: A total of 2455 women who were undergoing their first IVF treatment cycle at the Reproductive Medicine Center of Anhui Provincial Hospital, P.R. China from April 2007 to December 2011 were identified and reviewed. All patients had normal menstrual cycles and were stimulated with a long GnRH agonist protocol. Associations between oocyte number and LBR and miscarriage rate as well as the rate of moderate-severe ovarian hyperstimulation syndrome (OHSS) were analyzed. MAIN RESULTS AND ROLE OF CHANCE: The fresh embryo LBR per started cycle increased with the number of retrieved oocytes up to Groups 2 and 3 (6-10 and 11-15 oocytes) and then decreased, because of the high number of cycles with all embryos being cryopreserved, in order to avoid moderate-severe OHSS in group 4 (>15 oocytes). However, the cumulative LBR per started cycle continued to increase with oocyte number, as did the incidence of moderate-severe OHSS. There was no significant difference in the miscarriage rates among the patient groups. LIMITATIONS, REASONS FOR CAUTION: As a retrospective study, our analysis depends on previously recorded data; therefore, certain variables could not be collected. Our findings may be influenced by our young and thin patient population and the inability to control for certain markers of ovarian reserve such as historical maximum serum FSH, antral follicle count and serum anti-Mullerian hormone. WIDER IMPLICATIONS OF THE FINDINGS: Our data suggest that in IVF cycles stimulated with a long protocol, the optimal number of oocytes for achieving a live birth is somewhere between 6 and 15. The balance between maximum treatment success and minimum risk of complications, especially OHSS, should be further investigated. STUDY FUNDING/COMPETING INTERESTS: This study was supported by the Medical Scientific Research Plan Project of Anhui Provincial Department of Health (13ZC014) and the Natural Science Foundation of Anhui Higher Education Institutions (KJ2013Z132).

[Ultra-rapid cryopreservation of human spermatozoa with different concentrations of sucrose].

OBJECTIVE: To explore the feasibility of ultra-rapid freezing of human spermatozoa in the cryogenic vial with different concentrations of sucrose solution. METHODS: We divided 40 normal semen samples prepared with the routine swim-up technique into 6 aliquots, 1 as the control and the other 5 cryopreserved with sucrose solution at the concentrations of 0.15, 0.20, 0.25 and 0.30 mol/L, respectively. After thawing, we determined and compared the motility, progressive motility and plasma membrane integrity of the sperm among the 6 groups. RESULTS: The motility, progressive motility and plasma membrane integrity of the sperm were significantly lower after thawing than before cryopreservation ([96.2 +/- 1.8]%, [93.8 +/- 2.8]% and [99.0 +/- 0.8 ]%) (P<0.05). Post-thawing sperm motility was (55.5 +/- 6.3)% in the 0.20 mol/L sucrose group, significantly higher than in the 0.15, 0.25 and 0.30 mol/L groups ([45.9 +/- 6.6]%, [50.4 +/- 9.4]% and [45.5 +/- 11.2]%) (P<0.05), and it was (53.6 +/- 5.0)% in the conventional freezing group, with no statistically significant difference from the 0.20 and 0.25 mol/L sucrose cryopreservation groups (P> 0.05), but remarkably higher than in the 0.15 and 0.30 mol/L groups (P<0.05). Post-thawing progressive sperm motility exhibited no statistically significant differences between the 0.20 mol/L sucrose and conventional freezing groups ([44.4 +/- 7.4]% vs [42.3 +/- 8.1]%, P>0.05), but markedly higher in both than in the 0.15, 0.25 and 0.30 mol/L sucrose groups ([37.1 +/- 8.3 ]%, [33.1 +/- 9.2]% and [22.0 +/- 9.1]%) (P<0.05). Post-thawing plasma membrane integrity was significantly higher in the 0.20 mol/L sucrose cryopreservation group ( [70.1 +/- 6.9]%) than in either the conventional freezing group ([63.1 +/- 6.8]%) or the 0.15, 0.25 and 0.30 mol/L sucrose groups ([57.7 +/- 8.3]%, [63.5 +/- 10.7]% and [57.8 +/- 12.9]%) (P<0.05). CONCLUSION: As a simple, safe and effective method, ultra-rapid freezing with sucrose solution at the final concentration of 0.20 mol/L can be used for the cryopreservation of human spermatozoa.

Fertilization rates are improved after IVF if the corona radiata is left intact in vitrified-warmed human oocytes.

BACKGROUND: Before human MII oocytes are vitrified they are usually denuded from their cumulus cells. In this study we wanted to investigate the effects of an intact corona radiata on the vitrification and fertilization of human oocytes. METHODS: The study comprised two different parts. In Part 1, 36 MII stage oocytes, from 6 patients, were randomly assigned into a control group, a group of vitrified-warmed oocytes without a corona radiata and a group of vitrified-warmed oocytes with an intact corona radiata. In each group of 12, 6 oocytes were used for evaluation of the zona pellucida solubility (hardening) and another 6 oocytes were used for the analysis of their ultrastructure. In addition, six polyspermically fertilized oocytes were used as positive controls for zona pellucida hardening. In Part 2, 16 patients in total produced 107 fresh and 98 vitrified-warmed oocytes, with or without an intact corona radiata. All oocytes were fertilized via conventional IVF and embryos were transferred according to our standard ET routines. The oocyte survival and fertilization rates, embryo quality and pregnancy and implantation rates were evaluated. RESULTS: There were no differences in oocyte survival, zona pellucida solubility (hardening) or the number of cortical granules between the vitrified-warmed and fresh oocytes. There were also no differences in the zona pellucida solubility and the number of cortical granules between vitrified-warmed oocytes with or without an intact corona radiata. However, the oocytes with an intact corona radiata had a higher fertilization rate after conventional IVF insemination. No differences were seen in the survival and cleavage rates, the percentage of high-quality embryos or the clinical outcome. CONCLUSIONS: Zona hardening and ultrastructural damage do not seem to occur in vitrified human oocytes. An intact corona radiata in vitrified-warmed oocytes retains their fertilization capacity in conventional IVF, but does not improve the embryo quality. Poor fertilizing capacities of vitrified-warmed oocytes without an intact corona radiata seem to have been due to the complete removal of the cumulus cells.

Chronic unpredictable stress decreases expression of brain-derived neurotrophic factor (BDNF) in mouse ovaries: relationship to oocytes developmental potential.

BACKGROUND: Brain-derived neurotropic factor (BDNF) was originally described in the nervous system but has been shown to be expressed in ovary tissues recently, acting as a paracrine/autocrine regulator required for developments of follicles and oocytes. Although it is generally accepted that chronic stress impairs female reproduction and decreases the expression of BDNF in limbic structures of central nervous system, which contributes to mood disorder. However, it is not known whether chronic stress affects oocytes developments, nor whether it affects expression of BDNF in ovary. METHODS: Mice were randomly assigned into control group, stressed group, BDNF-treated group and BDNF-treated stressed group. The chronic unpredictable mild stress model was used to produce psychosocial stress in mice, and the model was verified by open field test and hypothalamic-pituitary-adrenal (HPA) axis activity. The methods of immunohistochemistry and western blotting were used to detect BDNF protein level and distribution. The number of retrieved oocytes, oocyte maturation, embryo cleavage and the rates of blastocyst formation after parthenogenetic activation were evaluated. RESULTS: Chronic unpredictable stress decreased the BDNF expression in antral follicles, but didn't affect the BDNF expression in primordial, primary and secondary follicles. Chronic unpredictable stress also decreased the number of retrieved oocytes and the rate of blastocyst formation, which was rescued by exogenous BDNF treatment. CONCLUSION: BDNF in mouse ovaries may be related to the decreased number of retrieved oocytes and impaired oocytes developmental potential induced by chronic unpredictable stress.