NLRP4E regulates actin cap formation through SRC and CDC42 during oocyte meiosis.

BACKGROUND: Members of the nucleotide-binding oligomerization domain, leucine rich repeat and pyrin domain containing (NLRP) family regulate various physiological and pathological processes. However, none have been shown to regulate actin cap formation or spindle translocation during the asymmetric division of oocyte meiosis I. NLRP4E has been reported as a candidate protein in female fertility, but its function is unknown. METHODS: Immunofluorescence, reverse transcription polymerase chain reaction (RT-PCR), and western blotting were employed to examine the localization and expression levels of NLRP4E and related proteins in mouse oocytes. small interfering RNA (siRNA) and antibody transfection were used to knock down NLRP4E and other proteins. Immunoprecipitation (IP)-mass spectrometry was used to identify the potential proteins interacting with NLRP4E. Coimmunoprecipitation (Co-IP) was used to verify the protein interactions. Wild type (WT) or mutant NLRP4E messenger RNA (mRNA) was injected into oocytes for rescue experiments. In vitro phosphorylation was employed to examine the activation of steroid receptor coactivator (SRC) by NLRP4E. RESULTS: NLRP4E was more predominant within oocytes compared with other NLRP4 members. NLRP4E knockdown significantly inhibited actin cap formation and spindle translocation toward the cap region, resulting in the failure of polar body extrusion at the end of meiosis I. Mechanistically, GRIN1, and GANO1 activated NLRP4E by phosphorylation at Ser429 and Thr430; p-NLRP4E is translocated and is accumulated in the actin cap region during spindle translocation. Next, we found that p-NLRP4E directly phosphorylated SRC at Tyr418, while p-SRC negatively regulated p-CDC42-S71, an inactive form of CDC42 that promotes actin cap formation and spindle translocation in the GTP-bound form. CONCLUSIONS: NLRP4E activated by GRIN1 and GANO1 regulates actin cap formation and spindle translocation toward the cap region through upregulation of p-SRC-Tyr418 and downregulation of p-CDC42-S71 during meiosis I.

Multigenerational paternal obesity enhances the susceptibility to male subfertility in offspring via Wt1 N6-methyladenosine modification.

There is strong evidence that obesity is a risk factor for poor semen quality. However, the effects of multigenerational paternal obesity on the susceptibility to cadmium (a reproductive toxicant)-induced spermatogenesis disorders in offspring remain unknown. Here, we show that, in mice, spermatogenesis and retinoic acid levels become progressively lower as the number of generations exposed to a high-fat diet increase. Furthermore, exposing several generations of mice to a high fat diet results in a decrease in the expression of Wt1, a transcription factor upstream of the enzymes that synthesize retinoic acid. These effects can be rescued by injecting adeno-associated virus 9-Wt1 into the mouse testes of the offspring. Additionally, multigenerational paternal high-fat diet progressively increases METTL3 and Wt1 N6-methyladenosine levels in the testes of offspring mice. Mechanistically, treating the fathers with STM2457, a METTL3 inhibitor, restores obesity-reduced sperm count, and decreases Wt1 N6-methyladenosine level in the mouse testes of the offspring. A case-controlled study shows that human donors who are overweight or obese exhibit elevated N6-methyladenosine levels in sperm and decreased sperm concentration. Collectively, these results indicate that multigenerational paternal obesity enhances the susceptibility of the offspring to spermatogenesis disorders by increasing METTL3-mediated Wt1 N6-methyladenosine modification.

NLRC5 potentiates anti-tumor CD8+ T cells responses by activating interferon-ß in endometrial cancer.

OBJECTIVES: NLR family CARD domain containing 5 (NLRC5) could promote major histocompatibility complex class I (MHC-I)-dependent CD8+ T cell-mediated anticancer immunity. In this study, the immunosurveillance role and underlying mechanisms of NLRC5 in endometrial cancer (EC) were characterized. METHODS: CD8+ T cells were separated from healthy women's peripheral blood by using magnetic beads. The effect of NLRC5 and interferon-ß (IFN-ß) on immunosurveillance of EC were examined through a mouse tumor model and a CD8+ T cell-EC cell coculture system after NLRC5 overexpression and IFN-ß overexpression or depletion. The effect of NLRC5 on IFN-ß expression was examined with gain- and loss-of-function experiments. RESULTS: NLRC5 overexpression in the EC cell and CD8+ T cell coculture system inhibited EC cell proliferation and migration and promoted EC cell apoptosis and CD8+ T cell proliferation. In vivo, NLRC5 overexpression increased the proportion of CD8+ T cells and inhibited EC progression. Furthermore, IFN-ß overexpression in the EC cell and CD8+ T cell coculture system activated CD8+ T cell proliferation; however, genetic depletion of IFN-ß exerted the opposite effects. In addition, NLRC5 could negatively regulate IFN-ß expression in EC cells. Mechanistically, NLRC5 potentiated the antitumor responses of CD8+ T cells to EC by activating IFN-ß. CONCLUSIONS: Taken together, our findings demonstrated that NLRC5 potentiates anti-tumor CD8+ T cells responses by activating interferon-ß in EC, suggesting that genetically escalated NLRC5 and IFN-ß may act as potential candidates for the clinical translation of adjuvant immunotherapies to patients with EC.

METTL3 facilitates immunosurveillance by inhibiting YTHDF2-mediated NLRC5 mRNA degradation in endometrial cancer.

BACKGROUND: N6-methyladenosine (m6A) methylation is the most abundant chemical posttranscriptional modification of mRNA, and it is associated with the regulation of the immune response to tumors. However, the function of m6A modification in the immune response to endometrial cancer (EC) remains unknown. Our study investigated the immunological role of methyltransferase-like 3 (METTL3) in EC and the underlying molecular mechanism. METHODS: We investigated the correlation between the expression of METTL3 and CD8 by using an endometrial tissue microarray cohort. Next, we investigated the role and mechanism of METTL3 in the immune response to EC using a mouse tumor model and a CD8+ T cell-EC cell coculture system after METTL3 overexpression or depletion. Additionally, RNA immunoprecipitation (RIP), methylated RIP, and RNA stability experiments were used to investigate the mechanism underlying the function of METTL3 in immunosurveillance of EC. RESULTS: METTL3 levels were downregulated in EC patients, low levels of METTL3 were correlated with poor prognosis in EC patients. There was a positive correlation between METTL3 expression and CD8 expression. Overexpression of METTL3 in the EC cell and CD8+ T cell coculture system inhibited EC cell proliferation, migration, and promoted CD8+ T-cell proliferation, and in vivo, METTL3 overexpression increased CD8+ T cell proportions and inhibited EC progression; however, genetic depletion of METTL3 exerted the opposite effects. NLR family CARD domain-containing 5 (NLRC5) was identified as a target of METTL3-mediated m6A modification. The degradation of NLRC5 was increased by YTH domain-containing family 2 (YTHDF2). CONCLUSIONS: Overall, METTL3, YTHDF2, and NLRC5 have potential to be the diagnostic and prognostic biomarkers for EC. METTL3 facilitated the m6A modifications of NLRC5 and inhibited its degradation through a YTHDF2-dependent mechanism in EC. Genetic overexpression of METTL3 attenuated the immune evasion of EC by promoting NLRC5-mediated immunosurveillance, suggesting that the METTL3/YTHDF2/NLRC5 axis is a promising target of immunotherapy in EC.

Recent Insights into Noncoding RNAs in Primary Ovarian Insufficiency: Focus on Mechanisms and Treatments.

CONTEXT: Primary ovarian insufficiency (POI) is a heterogeneous disease with an unknown underlying trigger or root cause. Recently many studies evaluated noncoding RNAs (ncRNAs), especially microRNAs (miRNAs), long noncoding RNA (lncRNAs), circular RNAs (circRNAs), and small interfering RNAs (siRNAs) for their associations with POI. EVIDENCE ACQUISITION: In this review, we outline the biogenesis of various ncRNAs relevant to POI and summarize the evidence for their roles in the regulation of disease occurrence and progression. Articles from 2003 to 2022 were selected for relevance, validity, and quality from results obtained in PubMed and Google Scholar using the following search terms: noncoding RNAs; primary ovarian insufficiency; premature ovarian failure; noncoding RNAs and primary ovarian insufficiency/premature ovarian failure; miRNAs and primary ovarian insufficiency/premature ovarian failure; lncRNAs and primary ovarian insufficiency/premature ovarian failure; siRNAs and primary ovarian insufficiency/premature ovarian failure; circRNAs and primary ovarian insufficiency/premature ovarian failure; pathophysiology; and potential treatment. All articles were independently screened for eligibility by the authors. EVIDENCE SYNTHESIS: This review summarizes the biological functions and synthesis of miRNAs, lncRNAs, siRNAs, and circRNAs in POI and discusses the findings of clinical and in vitro and in vivo studies. Although there is variability in the findings of individual studies, overall the available literature justifies the conclusion that dysregulated ncRNAs play significant roles in POI. CONCLUSION: The potential of ncRNAs in the treatment of POI requires further investigation, as ncRNAs derived from mesenchymal stem cell-secreted exosomes play pivotal roles and have considerable therapeutic potential in a multitude of diseases.

Resilience of infertile families undergoing in vitro fertilization: An application of the double ABC-X model.

BACKGROUND: Family resilience plays a crucial role in protecting the mental health and family stability of infertile patients. However, information associated with infertile families resilience is scarce. The double ABC-X model provides a roadmap for this, helps organize knowledge, and lays the foundation for knowledge development. AIMS: To describe the current situation of family resilience of infertile women, and to test the predictive theoretical model of family resilience based on infertility stigma, individual resilience, coping style, and posttraumatic growth. DESIGN: A cross-sectional study. METHODS: A convenience sample of 372 infertile women undergoing in vitro fertilization were recruited between April and August 2020. The Chinese-Family Resilience Assessment Scale, Infertility Stigma Scale, Simplified Coping Style Questionnaire, Chinese version of Connor-Davidson Resilience Scale, and Chinese version of Post Traumatic Growth Inventory were used to measure family resilience, infertility stigma, individual resilience, coping style, and posttraumatic growth. Structural equation models were used to analyze the relationship among these variables. RESULTS: The results showed that family resilience was related to infertility stigma, positive coping, and individual resilience. Moreover, the path analysis indicated that positive coping and individual resilience mediated the effects of infertility stigma on family resilience. CONCLUSIONS: A high level of stigma among infertile women should be identified. Interventions for targeting positive coping and individual resilience might ultimately increase their family resilience.

Acute exposure of triclocarban affects early embryo development in mouse through disrupting maternal-to-zygotic transition and epigenetic modifications.

Triclocarban (TCC) is a broad-spectrum antibacterial agent used globally, and high concentrations of this harmful chemical exist in the environment. The human body is directly exposed to TCC through skin contact. Moreover, TCC is also absorbed through diet and inhaled through breathing, which results in its accumulation in the body. The safety profile of TCC and its potential impact on human health are still not completely clear; therefore, it becomes imperative to evaluate the reproductive toxicity of TCC. Here, we explored the effect of TCC on the early embryonic development of mice and its associated mechanisms. We found that acute exposure of TCC affected the early embryonic development of mice in a dose-dependent manner. Approximately 7600 differentially expressed genes (DEGs) were obtained by sequencing the transcriptome of 2-cell mouse embryos; of these, 3157 genes were upregulated and 4443 genes were downregulated in the TCC-treated embryos. GO and KEGG analysis revealed that the enriched genes were mainly involved in redox processes, RNA synthesis, DNA damage, apoptosis, mitochondria, endoplasmic reticulum, Golgi apparatus, cytoskeleton, peroxisome, RNA polymerase, and other components or processes. Moreover, the Venn analysis showed that the zygotic genome activation (ZGA) was affected and the degradation of maternal effector genes was inhibited. TCC induced changes in the epigenetic modification of 2-cell embryos. The level of DNA methylation increased significantly. Further, the levels of H3K27ac, H3K9ac, and H3K27me3 histone modifications decreased significantly, whereas those of H3K4me3 and H3K9me3 modifications increased significantly. Additionally, TCC induced oxidative stress and DNA damage in the 2-cell embryos. In conclusion, acute exposure of TCC affected early embryo development, destroyed early embryo gene expression, interfered with ZGA and maternal gene degradation, induced changes in epigenetic modification of early embryos, and led to oxidative stress and DNA damage in mouse early embryos.

Role of EGFR expressed on the granulosa cells in the pathogenesis of polycystic ovarian syndrome.

Polycystic ovarian syndrome (PCOS) is one of the most common endocrinological disorders affecting between 6 to 20% of reproductive aged women. However, the etiology of PCOS is still unclear. Epidermal growth factor receptor (EGFR) plays a critical role in the growth and development of ovarian follicles. In our previous study, we showed that the expression level of EGFR was significantly higher in the cumulus granulosa cells from women with PCOS than that of normal women, suggesting that EGFR may play a potential role in the pathogenesis of PCOS. The present study further evaluated the association between EGFR and PCOS through both in clinical observation and animal experiments. We firstly validated the differential expression of EGFR in cumulus granulosa cells between PCOS patients and normal subjects by qRT-PCR and immunofluorescence staining. Then we generated a mouse model (n=20) of PCOS by injecting dehydroepiandrosterone (DHEA). The PCOS mice were then injected with an E corpus GFR inhibitor (AG1478) (n=10), which significantly improved the sex hormone levels in the estrous cycle stage, and the serum levels of LH, FSH and testosterone were compared with the PCOS mice without EGFR inhibitor treatment (n=10). Decreasing the expression level of EGFR in the PCOS mice also improved the ovulatory function of their ovaries which was indicated by the multifarious follicle stage in these mice as compared with the PCOS mice without EGFR inhibitor treatment. Also, the number of corpopa lutea were higher in the control group and the EGFR inhibitor treated group than in the PCOS group. The sex hormone levels and reproductive function were not significantly different between the control mice and the PCOS mice treated with the EGFR inhibitor. Our results demonstrated that EGF/EGFR signaling affected the proliferation of cumulus granulosa cells, oocyte maturation and meiosis, and played a potential role in the pathogenesis of PCOS. Therefore, the selective inhibition of EGFR may serve as a novel strategy for the clinical management of PCOS.

Sexual Function, Self-Esteem, and Quality of Life in Infertile Couples Undergoing in vitro Fertilization: A Dyadic Approach.

Purpose: Infertility has adverse effects on the quality of life (QoL) of infertile couples. Previous studies found important associations between sexual function, self-esteem and QoL, but mainly focused on one individual's approach rather than both partners. This study adopted a dyadic approach to evaluate the relationship between sexual function and QoL in couples with infertility through mediation and improving self-esteem. Patients and Methods: Between October 2020 and January 2021, 428 couples with infertility (n=856) undergoing in-vitro fertilization (IVF) at a tertiary hospital in Hefei, China, were registered for the current descriptive cross-sectional research. The dyads' sociodemographic and clinical features, as well as their sexual function, self-esteem, and QoL were evaluated. The Fertility quality of life scale (FertiQoL), Rosenberg Self-Esteem Scale (RSES), Female Sexual Function Index (FSFI), and International Index of Erectile Function-15 (IIEF-15) were used to evaluate the participants. The Actor-Partner Interdependence Mediation Model (APIMeM) was utilized to examine data from the dyadic relationships. Results: According to the APIMeM analysis, sexual function of individuals with infertility was directly and indirectly connected with their QoL, mediated through their self-esteem. The women's sexual function was found to be positively associated with their partner's QoL, with the women's self-esteem acting as a complete mediator. The men's sexual function was found to be positively associated with partner's QoL, with the men's self-esteem acting as a complete mediator. Conclusion: The findings suggest that boosting participants' self-esteem can help them and their partners have a better QoL. Also, therapies aimed at improving and sustaining self-esteem of couples with infertility could help mitigate the negative influence of low sexual function on their QoL.

NLRC5 Might Promote Endometrial Cancer Progression by Inducing PD-L1 Expression.

Aims: The NOD-like receptor (NLR) family, caspase recruitment (CARD) domain containing 5 (NLRC5) was dysregulated in endometrial cancer (EC). However, the potential regulatory mechanisms of NLRC5 in EC remained unclear. We aimed to explore whether NLRC5 could regulate the programmed cell death protein ligand 1 (PD-L1) in EC. We also investigated the related molecular which led to the inactivation of NLRC5 in EC. Methods: The expressions of NLRC5 and PD-L1 in endometrium tissue microarray were detected by immunohistochemistry. Pearson's correlation analysis was performed to detect the expression correlation between NLRC5 and PD-L1. Immunofluorescence staining, western blotting, and quantitative real-time PCR (qRT-PCR) were used to detect the role of NLRC5 in PD-L1 in EC cell lines. The somatic mutation in EC patients was detected by whole-exome sequencing (WGS). Results: NLRC5 was downregulated in the endometrium of EC patients when compared to those in the normal endometrium. The level of PD-L1 in the endometrium of EC patients was higher when compared to those in the normal endometrium. There was a negative expression correlation between NLRC5 and PD-L1. NLRC5 could promote the expression of PD-L1 in EC cell lines. The mutations of ANKRD20A2, C2orf42, ADGRB3, AVPR2, GOLGA6C, and IPPK may lead to the downregulation of NLRC5 in EC patients. Conclusion: NLRC5 could inhibit the activation of PD-L1 in EC. Mutations of ANKRD20A2, C2orf42, ADGRB3, AVPR2, GOLGA6C, and IPPK may lead to the downregulation of NLRC5 in EC patients. Future study should investigate the mechanism of NLRC5 in PD-L1, as well as the mechanism of ANKRD20A2, C2orf42, ADGRB3, AVPR2, GOLGA6C, and IPPK in NLRC5.

Genetic pathogenesis of acephalic spermatozoa syndrome: past, present, and future.

Acephalic spermatozoa syndrome (ASS) is one of the most severe spermatogenic failures of all infertility in men. The cognition of ASS has experienced a tortuous process. Over the past years, with the in-depth understanding of spermatogenesis and the emergence of new genetic research technologies, the unraveling of the genetic causes of spermatogenic failure has become highly active. From these advances, we established a genetic background and made significant progress in the discovery of the genetic causes of ASS. It is important to identify pathogenic genes and mutations in ASS to determine the biological reasons for the occurrence of the disease as well as provide genetic diagnosis and treatment strategies for patients with this syndrome. In this review, we enumerate various technological developments, which have made a positive contribution to the discovery of candidate genes for ASS from the past to the present. Simultaneously, we summarize the known genetic etiology of this phenotype and the clinical outcomes of treatments in the present. Furthermore, we propose perspectives for further study and application of genetic diagnosis and assisted reproductive treatment in the future.

Increased ApoE Expression in Follicular Fluid and the ApoE Genotype Are Associated With Endometriosis in Chinese Women.

More than 10% of women suffer from endometriosis (EMT) during their reproductive years. EMT can cause pain and infertility and requires further study from multiple perspectives. Previous reports have indicated that an increase inapolipoprotein E (ApoE) may be associated with a lower number of retrieved mature oocytes in older women, and an association between ApoE and spontaneous pregnancy loss may exist in patients with EMT. The purpose of this study was to investigate the existence of an increase in ApoE in follicular fluid (FF) and the possible relationship between ApoE and EMT in Chinese women. In the current study, 217 Chinese women (111 control subjects and 106 EMT patients) were included. The ApoE genotypes were identified by Sanger sequencing. We found that ApoE expression in FF was higher in patients with EMT than in the control group. In addition, a significant difference in ApoE4 carriers (ϵ3/ϵ4, ϵ4/ϵ4) was found between the control subjects and the patients with EMT. Furthermore, a nonparametric test revealed significant differences in the numbers of blastocysts and high-quality blastocysts, but not the hormone levels of FSH, LH, and E2, between the two groups. We also established a multifactor (BMI, high-quality blastocysts, and ϵ4) prediction model with good sensitivity for identifying patients who may suffer from EMT. Our results demonstrate that ApoE expression in FF is increased in EMT, the ApoE-ϵ4 allele is significantly linked to EMT, and a combined analysis of three factors (BMI, high-quality blastocysts, and ϵ4) could be used as a predictor of EMT.

Association analysis between the tag single nucleotide polymorphisms of DENND1A and the risk of polycystic ovary syndrome in Chinese Han women.

BACKGROUND: The DENND1A gene is one of the most important sites associated with polycystic ovary syndrome (PCOS). We attempted to analyze the correlation between five single nucleotide polymorphisms (SNPs) in the DENND1A gene and the development of PCOS. METHODS: A total of 346 PCOS patients and 225 normal ovulatory women were involved in the case-control study. Clinical variables and hormones were recorded. According to the Hap Map database, five tagging SNPs (rs2479106, rs2768819, rs2670139, rs2536951 and rs2479102) in the DENND1A gene were identified. The TaqMan probe and the PCR-RFLP (restriction fragment length polymorphism) methods were used for revealing these genotypes. TaqMan Genotype Software was used to analyze the alleles of the five SNPs. RESULTS: Linkage disequilibrium and the gene frequency analysis demonstrated that the CCGGG haplotype might increase the risk of PCOS (P = 0.038, OR = 1.89, 95% CI = 1.027-3.481). Significant differences were found in genotypic and allelic distributions at the rs2536951 and rs2479102 loci between PCOS women and controls (P <  0.001). The LH levels and LH/FSH ratios were higher in PCOS patients than in the control group. A detailed analysis revealed that for the rs2479106 locus, these two values were significantly different in the control subjects who had AA, AG and GG genotypes (P = 0.013 and P = 0.007, respectively), and for the rs2468819 locus, these two values were significantly different among the PCOS patients with AA, AG and GG genotypes (P = 0.013 and 0.002, respectively). CONCLUSIONS: The tagging SNPs rs2479106 and rs2468819 in the DENND1A gene are associated with PCOS in the Chinese population, whereas rs2670139, rs2536951 and rs2479102 are not correlated with PCOS in the same population.

Melatonin maintains mitochondrial membrane potential and decreases excessive intracellular Ca2+ levels in immature human oocytes.

AIMS: Previous reports have demonstrated that melatonin exists in multiple extrapineal sites, and higher amounts of melatonin are present in human follicular fluid than in serum, which indicates that it might play key roles in human oocyte maturation and subsequent embryonic development. Melatonin has been shown to be a potent antioxidant and might be beneficial to human oocytes during in vitro maturation (IVM). However, the underlying mechanisms of melatonin action during IVM have not been thoroughly investigated. MAIN METHODS: Immunofluorescence staining, western blotting, and ELISA were applied to investigate whether melatoninergic components are expressed in the cultured human ovarian cumulus cells. TMRE staining and Fluo-4 AM staining were performed to detect the mitochondrial membrane potential and intracellular Ca2+ levels of immature human oocytes respectively. KEY FINDINGS: First, cultured human ovary cumulus cells synthesized melatonin in vitro, and it expressed serotonin (the precursor of melatonin) and the two key enzymes, i.e. N-acetyltransferase (NAT) and hydroxyindole-O-methyltransferase (HIOMT). Additionally, the results suggest that melatonin maintains the mitochondrial membrane potential and decrease excessive Ca2+ levels in immature human oocytes during IVM. SIGNIFICANCE: In conclusion, we provide evidence that the melatoninergic components were expressed in cultured human ovarian cumulus cells, and melatonin might reduce oxidative stress of human oocytes by ameliorating mitochondrial function. In view of the significant clinical value that immature human oocytes have in assisted reproductive technology (ART), our findings highlight a potential treatment strategy of using melatonin to improve mitochondrial function and to enhance the quality of human oocytes during IVM.

Delivery of folic acid-modified liposomal curcumin for targeted cervical carcinoma therapy.

Introduction: In this study, novel folic acid (FA)-modified curcumin (CUR) liposomes (LPs) were developed and evaluated for their antitumor activity in vitro and in vivo. Methods: Characterization of the LPs, including transmission electron microscopy, morphology, particle size, and zeta potential studies, was carried out. Drug entrapment efficiency, drug-loading capacity, and release properties in vitro were tested. The in vitro growth inhibition activity, cellular uptake efficiency, and cell apoptosis of FA-modified CUR LPs were also investigated by a cervical cancer HeLa cell model. Results: The optimized distearoyl-l-a-phosphatidylethanolamine (DSPE)-PEG2000-FA-LPs/CUR formed spherical vesicles of nanometer sizes and had particle sizes of 112.3±4.6 nm, polydispersity index of 0.19±0.03, and zeta potential of -15.3±1.4 mV. In addition, the EE% and DL% of (DSPE)-PEG2000-FA-LPs/CUR were 87.6% and 7.9%, respectively. Compared with the free drug, FA-modified CUR LPs had sustained-release properties in vitro. In vivo, a strong green fluorescence was observed in the cytoplasmic region after incubation of (DSPE)-PEG2000-FA-LPs/CUR for 2 hrs. Conclusion: (DSPE)-PEG2000-FA-LPs/CUR showed a superior antiproliferative effect on HeLa cells and had a better antitumor effect in vivo than the non-modified LPs. These results indicated that (DSPE)-PEG2000-FA-LPs/CUR was a promising candidate for antitumor drug delivery.

Stimulative role of ST6GALNAC1 in proliferation, migration and invasion of ovarian cancer stem cells via the Akt signaling pathway.

BACKGROUND: Ovarian cancer is known as one of the most common cancers in the world among women. ST6GALNAC1 is highly expressed in cancer stem cells (CSCs), which correlates to high tumor-initiating, self-renewal and differentiation abilities. This present study aims to investigate how ST6GALNAC1 affects ovarian cancer stem cells (OCSCs). METHODS: In order to identify the differentially expressed genes related to ovarian cancer, microarray-based gene expression profiling of ovarian cancer was used, and ST6GALANC1 was one of the identified targets. After that, levels of ST6GALNAC1 in OCSCs and ovarian cancer cells were examined. Subsequently, an Akt signaling pathway inhibitor LY294002 was introduced into the cluster of differentiation 90+ (CD90+) stem cells, and cell proliferation, migration and invasion, levels of CXCL16, EGFR, CD44, Nanog and Oct4, as well as tumorigenicity of OCSCs were examined. RESULTS: By using a comprehensive microarray analysis, it was determined that ST6GALNAC1 was highly expressed in ovarian cancer and it regulated the Akt signaling pathway. High levels of ST6GALNAC1 were observed in OCSCs and ovarian cancer cells. Silencing ST6GALNAC1 was shown to be able to reduce cell proliferation, migration, invasion, self-renewal ability, tumorigenicity of OCSCs. In accordance with these results, the effects of ST6GALNAC1 in OCSCs were dependent on the Akt signaling pathway. CONCLUSIONS: When taken together, our findings defined the potential stimulative roles of ST6GALNAC1 in ovarian cancer and OCSCs, which relied on the Akt signaling pathway.

Novel DPY19L2 variants in globozoospermic patients and the overcoming this male infertility.

Globozoospermia has been reported to be a rare but severe causation of male infertility, which results from the failure of acrosome biogenesis and sperm head shaping. Variants of dpy-19-like 2 (DPY19L2) are highly related to globozoospermia, but related investigations have been mainly performed in patients from Western countries. Here, we performed a screening of DPY19L2 variants in a cohort of Chinese globozoospermic patients and found that five of nine patients carried DPY19L2 deletions and the other four patients contained novel DPY19L2 point mutations, as revealed by whole-exome sequencing. Patient 3 (P3) contained a heterozygous variant (c.2126+5G>A), P6 contained a homozygous nonsense mutation (c.1720C>T, p.Arg574*), P8 contained compound heterozygous variants (c.1182-1184delATC, p.Leu394_Ser395delinsPhe; c.368A>T, p.His123Arg), and P9 contained a heterozygous variant (c.1182-1184delATCTT, frameshift). We also reported intracytoplasmic sperm injection (ICSI) outcomes in the related patients, finding that ICSI followed by assisted oocyte activation (AOA) with calcium ionophore achieved high rates of live births. In summary, the infertility of these patients results from DPY19L2 dysfunction and can be treated by ICSI together with AOA.

HMGA2 gene silencing reduces epithelial-mesenchymal transition and lymph node metastasis in cervical cancer through inhibiting the ATR/Chk1 signaling pathway.

Many cervical cancer (CC) patients suffer from cancer invasion and lymph node metastasis, resulting in poor therapeutic outcome. Evidence has indicated the involvement of misexpressed high-mobility group AT-hook 2 (HMGA2) in poor survival of cancer patients. This study hereby aims to investigate the role of HMGA2 in CC cell biological functions via the ATR/Chk1 signaling pathway. The cell line with the highest HMGA2 expression was selected to establish cell lines with wild-type and stable HMGA2 silencing. The underlying regulatory mechanisms of HMGA2 in CC cells were analyzed with the treatment of the ATR/Chk1 signaling pathway activator, inhibitor, shRNA against HMGA2 or pcDNA-HMGA2 plasmids, followed by quantification of expression levels of ATR, Chk1, Bcl-2, Bax, MMP-2, MMP-9, E-cadherin and N-cadherin. CC cell apoptosis, proliferation, migration, invasion and lymph node metastasis in nude mice were evaluated. The HeLa cell line with the highest HMGA2 expression was selected. HMGA2 inhibited the activation of the ATR/Chk1 signaling pathway. Notably, HMGA2 silencing or inhibition of the ATR/Chk1 signaling pathway inhibited epithelial mesenchymal transition (EMT), CC cell proliferation, invasion, migration, tumorigenicity and lymph node metastasis while promoting apoptosis, indicated by reduced expression of Bcl-2, MMP-2, MMP-9 and N-cadherin, with increased expression of E-cadherin and Bax. Collectively, our study provides evidence that HMGA2 gene silencing inhibits the activation of the ATR/Chk1 signaling pathway, whereby repressing EMT, proliferation, migration and invasion of CC cells and lymph node metastasis, and promoting CC cell apoptosis.

Prevalence and risk factors of monochorionic diamniotic twinning after assisted reproduction: A six-year experience base on a large cohort of pregnancies.

OBJECTIVE: To characterize the incidence and risk factors for monochorionic diamniotic(MC-DA) twinning after assisted reproductive technologies (ART). DESIGN: Retrospective population-based study. SETTING: The study was conducted in China; Department of Reproductive Medicine Center at The First Affiliated Hospital of Anhui Medical University. POPULATION: A cohort of 8860 clinical pregnancies after embryo transfer (ET) carried out in our center between 2011 and 2016 were retrospectively analyzed for the incidence of MC-DA twinning. METHODS: Logistic regression was used to model the effect on the incidence of MC-DA twinning after ART. Different clinical data (maternal age) and laboratory procedures (type of ET (fresh versus frozen), insemination (IVF or ICSI)), embryo stage at time of ET (cleavage or blastocyst)) on the incidence of MC-DA twinning were evaluated. MAIN OUTCOME MEASURES: Monochorionic-diamniotic pregnancies were identified when more than one fetal poles was visualized in one gestational sac via trans-vaginal ultrasound at early first-trimester (7 to 8 weeks). RESULTS: The overall MC-DA twinning rate was 2.55% (226/8860). Eighty one MC-DA twinnings occurred in the fresh cycles and 145 in the frozen cycles (2.67% vs. 2.49%). MC-DA twinning incidence showed no significant difference whether ICSI was performed or not (2.79% vs. 2.44%). The MZT that resulted from single embryo transfer (SET) cycles (1.99%) was slightly lower than multiple embryo transfer cycles (2.61%),but with non-significance. However, women <35 years displayed a significant higher rate (2.81%) than women ≥35 years old (1.16%). Blastocyst transfer was associated with a significantly increase in MC-DA twinning incidence than cleavage-stage embryos transfer (2.79% VS 2.02%, P = 0.008). In the results of logistic regression analysis, blastocyst transfer is a major risk factor of MZT in the fresh cycles (P = 0.044), while maternal age plays a more important role in the frozen cycles (P = 0.004). CONCLUSIONS: There is an elevated prevalence of MC-DA twinning after ART. Both maternal age and blastocyst transfer are risk factors of monozygotic pregnancy independently. Blastocyst transfer is a major risk factor of MC-DA twinning in the fresh cycles, while maternal age plays a more important role in the frozen cycles.

Efficacy of vitamin B complex as an adjuvant therapy for the treatment of complicated vulvovaginal candidiasis: An in vivo and in vitro study.

OBJECTIVE: This study aimed to explore the efficacy of vitamin B complex as an adjuvant therapy for the treatment of complicated vulvovaginal candidiasis (VVC) in vitro and in vivo. METHODS: One-hundred fifty-eight complicated VVC patients were randomly divided into group A (treated with suppository+oral antifungal agents), group B (treated with suppository+vaginal cream), and group C (treated with suppository+vaginal cream+oral vitamin B complex). A mouse model of VVC was established. Eighty VVC mice were randomly divided into 4 groups according to the dose of vitamin B complex (20 mice in each group): V1 group (injected with 150µL normal salin), V2 group (injected with 50µL vitamin B complex solution+100µL normal saline), V3 group (injected with 100µL vitamin B complex solution+50µL normal saline), and V4 group (injected with 150µL vitamin B complex solution). After 4 weeks of treatment, the vaginal secretion was obtained for microscopic smear examination. HE stainning was performed to observe histopathological changes of vaginal tissues. The expressions of inflammatory factors were detected by ELISA. Meanwhile, VVC model of vaginal epithelial cells was established. The effects of different concentrations of vitamin B complex on anti-fungal effect of fluconazole were detected in vitro. RESULTS: After the treatment, complicated patients in the group C had significantly higher effective rates than those in the group A and group B. After the intra-gastric administration, the microscopic smear examination found that obvious pseudohypha in cluster with a lot of blastospores can be seen in the vaginal secretions of mice in the V1 group under the microscope. There was significant difference between mice treated with different dosages of vitamin B complex. The inflammatory response of mice in the V1 group was significantly higher than those in other groups and the inflammation response reduced with the increase of vitamin B complex dosage. The vitamin B complex elevated the curative effects of fluconazole on VVC model of vaginal epithelial cells and significantly increased the anti-fungal effect of fluconazole. CONCLUSIONS: Our findings suggest that vitamin B complex could be an effective adjuvant therapy for complicated VVC.