CRISPR/Cas9 technology: applications in oocytes and early embryos.

CRISPR/Cas9, a highly versatile genome-editing tool, has garnered significant attention in recent years. Despite the unique characteristics of oocytes and early embryos compared to other cell types, this technology has been increasing used in mammalian reproduction. In this comprehensive review, we elucidate the fundamental principles of CRISPR/Cas9-related methodologies and explore their wide-ranging applications in deciphering molecular intricacies during oocyte and early embryo development as well as in addressing associated diseases. However, it is imperative to acknowledge the limitations inherent to these technologies, including the potential for off-target effects, as well as the ethical concerns surrounding the manipulation of human embryos. Thus, a judicious and thoughtful approach is warranted. Regardless of these challenges, CRISPR/Cas9 technology undeniably represents a formidable tool for genome and epigenome manipulation within oocytes and early embryos. Continuous refinements in this field are poised to fortify its future prospects and applications.

H3K4 Methylation Promotes Expression of Mitochondrial Dynamics Regulators to Ensure Oocyte Quality in Mice.

Significantly decreased H3K4 methylation in oocytes from aged mice indicates the important roles of H3K4 methylation in female reproduction. However, how H3K4 methylation regulates oocyte development remains largely unexplored. In this study, it is demonstrated that oocyte-specific expression of dominant negative mutant H3.3-K4M led to a decrease of the level of H3K4 methylation in mouse oocytes, resulting in reduced transcriptional activity and increased DNA methylation in oocytes, disturbed oocyte developmental potency, and fertility of female mice. The impaired expression of genes regulating mitochondrial functions in H3.3-K4M oocytes, accompanied by mitochondrial abnormalities, is further noticed. Moreover, early embryos from H3.3-K4M oocytes show developmental arrest and reduced zygotic genome activation. Collectively, these results show that H3K4 methylation in oocytes is critical to orchestrating gene expression profile, driving the oocyte developmental program, and ensuring oocyte quality. This study also improves understanding of how histone modifications regulate organelle dynamics in oocytes.

Diet-induced obesity impairs spermatogenesis: the critical role of NLRP3 in Sertoli cells.

BACKGROUND: Accumulating evidence indicates a key role of Sertoli cell (SC) malfunction in spermatogenesis impairment induced by obesity. Nucleotide-binding oligomerization domain-like receptor with a pyrin domain 3 (NLRP3) is expressed in SCs, but the role of NLRP3 in the pathological process of obesity-induced male infertility remains unclear. METHODS: NLRP3-deficient mice were fed a high-fat diet for 24 weeks to establish obesity-related spermatogenesis impairment. In another set of experiments, a lentiviral vector containing a microRNA (miR)-451 inhibitor was injected into AMP-activated protein kinase α (AMPKα)-deficient mouse seminiferous tubules. Human testis samples were obtained by testicular puncture from men with obstructive azoospermia whose samples exhibited histologically normal spermatogenesis. Isolated human SCs were treated with palmitic acid (PA) to mimic obesity model in vitro. RESULTS: Increased NLRP3 expression was observed in the testes of obese rodents. NLRP3 was also upregulated in PA-treated human SCs. NLRP3 deficiency attenuated obesity-related male infertility. SC-derived NLRP3 promoted interleukin-1ß (IL-1ß) secretion to impair testosterone synthesis and sperm performance and increased matrix metalloproteinase-8 (MMP-8) expression to degrade occludin via activation of nuclear factor-kappa B (NF-κB). Increased miR-451 caused by obesity, decreased AMPKα expression and sequentially increased NADPH oxidase activity were responsible for the activation of NLRP3. miR-451 inhibition protected against obesity-related male infertility, and these protective effects were abolished by AMPKα deficiency in mice. CONCLUSIONS: NLRP3 promoted obesity-related spermatogenesis impairment. Increased miR-451 expression, impaired AMPKα pathway and the subsequent ROS production were responsible for NLRP3 activation. Our study provides new insight into the mechanisms underlying obesity-associated male infertility.

MiR-187 regulates the proliferation, migration and invasion of human trophoblast cells by repressing BCL6-mediated activation of PI3K/AKT signaling.

INTRODUCTION: Recurrent miscarriage (RM), refers to two or more consecutive spontaneous miscarriage in a pregnant woman. RM is caused by many factors, and microRNAs play an important role in the development and pathology of RM. In the present study, we investigated the function of miR-187 in the pathogenesis of RM and its effects on human trophoblast cells. METHODS: The localization of miR-187 in the human placenta in early pregnancy was determined by in situ hybridization. QRT-PCR was used to detect the expression of miR-187 in villi of normal early pregnancy induced abortion group and recurrent spontaneous miscarriage group. Then, HTR8/SVneo cells were used to investigated the effect of miR-187 on BCL6 expression and biological activity of trophoblasts. RESULTS: We found that the expression of miR-187 in villi of RM group was higher than that of normal abortion group and miR-187 inhibited the proliferation, migration, and invasion of HTR8 cells. We also found that miR-187 promoted apoptosis, inhibited EMT, and inhibited the PI3K/AKT pathway in HTR8 cells. In addition, we also found that BCL6 is a direct target of miR-187 and is negatively regulated by miR-187. In addition, BCL6 reversed the inhibitory effects of miR-187 on HTR8/SVneo cells. These data demonstrate that miR-187-induced repression of PI3K/AKT signaling is mediated by BCL6 in HTR8 cells. DISSCUSSION: MiR-187 inhibits the proliferation, migration, and invasion of trophoblasts through a mechanism that involves regulation of BCL6.

Biological roles of l-carnitine in oocyte and early embryo development.

Poor oocyte quality is responsible for female infertility. Multiple studies have been carried out to find supplements to enhance oocyte quality and mitigate infertility problems. l-carnitine and its derivatives have diverse roles in developing oocytes and early embryos. This review focuses on the in vitro and in vivo studies that using l-carnitine alone or in combination with other supplements for oocyte quality enhancement. The key roles of l-carnitine in oocyte quality and embryo growth were summarized, and the underlying mechanism was also elucidated. l-carnitine helps in the lipid metabolism process by controlling the transfer of fatty acids to mitochondria for ß-oxidation. l-carnitine modulates glucose metabolism and enhances respiratory chain enzyme activity. Furthermore, it acts as an antioxidant to prevent oxidative damage and inhibit apoptosis, a signal in response to oxidative stress. Results show the potential of l-carnitine as a potential agent in assisted reproductive technology to improve oocyte quality and the subsequent embryonic development.

Non-Coding RNAs Regulate Placental Trophoblast Function and Participate in Recurrent Abortion.

Recurrent spontaneous abortion (RSA) is a serious pregnancy complication with an increasing clinical incidence. The various causes of recurrent abortion are complicated. Developments in genetics, immunology, and cell biology have identified important roles of non-coding RNAs (ncRNAs) in the occurrence and progress of recurrent abortion. NcRNAs can affect the growth, migration, and invasion of placental trophoblasts by regulating cell processes such as the cell cycle, apoptosis, and epithelial-mesenchymal transformation. Therefore, their abnormal expression might lead to the occurrence and development of RSA. NcRNAs include small nuclear RNA (snRNA), small nucleolar RNA (snoRNA), ribosomal RNA (rRNA), transfer, RNA (tRNA), circular RNA (cRNA), and Piwi-interacting RNA (piRNA). In this review, we discuss recent research that focused on the function and mechanism of microRNAs (miRNAs), long non-coding RNAs (lncRNAs), and circular RNA (circRNA) in regulating placental trophoblasts. The use of ncRNAs as potential diagnostic and predictive biomarkers in RSA is also discussed to provide future research insights.

Vitamin D and Polycystic Ovary Syndrome: a Narrative Review.

Polycystic ovary syndrome (PCOS) is one of the most common endocrine disorders of reproductive age women and contributes to metabolic dysfunctions including insulin resistance (IR) and dyslipidemia. Vitamin D is a steroid hormone, which is involved in calcium metabolism and bone structure and has a potential role in the prevention of many illnesses, including cancers, autoimmune disorders, hypertension, diabetes, and obesity. Recently, it has been reported that vitamin D deficiency was a common complication of PCOS and vitamin D status was associated with reproductive ability, metabolic alterations, and mental health of PCOS patients. This review summarizes the advances between vitamin D status and the pathophysiological process of PCOS. Vitamin D level was negatively associated with serum androgen level. Vitamin D treatment could reduce serum androgen and anti-MüllerianHormone (AMH) levels, and decrease endometrial thickness, which resulted in improvement of menstrual cycle and folliculogenesis of PCOS patients. Moreover, vitamin D concentrations were negatively correlated with parameters of IR and body fat mass. Vitamin D supplementation has beneficial effects on IR and lipid metabolism. In addition, a positive of vitamin D on mental health of PCOS patients was proposed. Understanding the relationship between vitamin D status and the symptoms of PCOS patients is of great clinical significance to treat and prevent the progression of PCOS.

Chromosomal polymorphisms associated with reproductive outcomes after IVF-ET.

PURPOSE: This study aimed to investigate the effect of the detail type of chromosomal polymorphisms (1/9/16qh+/-, D/G group polymorphisms, and inv(9)) on the IVF-ET outcomes. METHODS: A total of 1335 infertile couples undergoing IVF/ICSI were enrolled and comprehensively analyzed the correlation between three detail types of chromosomal polymorphisms (1/9/16qh+/-, D/G group polymorphisms, and inv(9)) and the outcome of IVF/ICSI embryo transfer. The fertilized rate, cleaved embryo rate, good-quality embryo rate, clinical pregnancy rate, implantation rate, and early stage miscarriage rate were compared between the chromosomal polymorphisms groups and the control group. RESULTS: Both the inv(9) and D/G group chromosomal polymorphisms related to female infertility significantly lead to a lower 2PN cleavage rate (86.44% vs. 97.58% and 90.67% vs. 97.58%, respectively, P < 0.05) undergoing IVF insemination, the inv(9) adversely increasing the early miscarriage rate, either undergoing IVF (21.4% vs. 3.0%, P < 0.05) or ICSI (50.0% vs. 2.0%, P < 0.05) insemination, female carriers (23.08% vs. 2.87%, P < 0.05) or male carriers (44.44% vs. 2.87%, P < 0.05). For D/G groups, ICSI insemination may increase the implantation rate (44.8% vs. 23.69%, P < 0.05) and clinical pregnancy rate (78.6% vs. 40.65%, P < 0.05). 1/9/16qh+/- had no apparent adverse effect on the patient's clinical outcomes. CONCLUSIONS: Our study suggests that chromosome karyotype analysis is necessary for IVF patients in clinical practice; we should afford individual genetic counseling suggestion according to the polymorphism types.

Epigenome editing by CRISPR/Cas9 in clinical settings: possibilities and challenges.

Epigenome editing is a promising approach for both basic research and clinical application. With the convergence of techniques from different fields, regulating gene expression artificially becomes possible. From a clinical point of view, targeted epigenome editing by CRISPR/Cas9 of disease-related genes offers novel therapeutic avenues for many diseases. In this review, we summarize the EpiEffectors used in epigenome editing by CRISPR/Cas9, current applications of epigenome editing and progress made in this field. Moreover, application challenges such as off-target effects, inefficient delivery, stability and immunogenicity are discussed. In conclusion, epigenome editing by CRISPR/Cas9 has broad prospects in the clinic, and future work will promote the application of this technology.

Gut microbiome and cancer immunotherapy.

Gut microbiome has received significant attention for its influences on a variety of host functions, especially immune modulation. With the next-generation sequencing methodologies, more knowledge is gathered about gut microbiome and its irreplaceable role in keeping the balance between human health and diseases is figured out. Immune checkpoint inhibitors (ICIs) are one of the most innovational cancer immunotherapies across cancer types and significantly expand the therapeutic options of cancer patients. However, a proportion of patients show no effective responses or develop immune-related adverse events when responses do occur. More important, it is demonstrated that the therapeutic response or treatment-limiting toxicity of cancer immunotherapy can be ameliorated or diminished by gut microbiome modulation. In this review, we first introduce the relationship between gut microbiome and cancer immunotherapy. And then, we expound the impact of gut microbiome on efficacy and toxicity of cancer immunotherapy. Further, we review approaches to manipulating gut microbiome to regulate response to ICIs. Finally, we discuss the current challenges and propose future directions to improve cancer immunotherapy via gut microbiome manipulation.

Sulforaphane ameliorates high-fat diet-induced spermatogenic deficiency in mice†.

Sulforaphane (SFN), a dietary isothiocyanate that is mainly found in cruciferous vegetables, possesses anti-oxidative and anticancer activity and modulates inflammation. However, little is known about the role of SFN in obesity-related male reproductive defects. The present study aimed to investigate the effects of SFN on high-fat diet (HFD)-induced male spermatogenic impairment and further clarify the possible underlying mechanisms. In this study, 8-week-old mice were randomly divided into four groups. Mice were fed a normal diet or an HFD with or without SFN supplementation. Sulforaphane was subcutaneously injected at a dose of 0.5 mg/kg 5 days/week for 4 weeks beginning 8 weeks after initiation of the HFD. The results demonstrated that SFN could protect against HFD-induced reproductive dysfunction in male mice. Moreover, SFN also improved reproductive ability, as demonstrated by an increased pregnancy rate and decreased embryo resorption rate in comparison to the corresponding HFD group. We also observed a decrease in apoptosis and an attenuation of endoplasmic reticulum (ER) stress after SFN treatment. In vitro studies of mouse and human sperm samples also revealed that SFN protects against the palmitic acid-induced reduction in sperm viability and motility by inhibiting ER stress in an AMP-activated protein kinase (AMPK)-dependent manner. AMPK-dependent ER stress attenuation by SFN was further confirmed using AMPK knockout mice. Taken together, these data show that SFN protects against HFD-induced male reproductive dysfunction by inhibiting ER stress and apoptosis. These findings may be helpful for identifying new therapeutic methods to treat male infertility.

Molecular detection of uterine innate lymphoid cells in the immunological mouse model of pregnancy loss.

Innate lymphoid cells (ILCs) are newly identified members of the innate lymphocyte family, which can function as adaptive T cells and act as critical modulators of inflammatory processes within different tissues and immune diseases. The role of uterine ILCs (uILCs) has recently been elucidated alongside changes associated with normal pregnancy. However, the proportions of uterine ILCs and their role in unsuccessful pregnancy remain unclear. We analyzed the characterization of uILC subsets and the expression of signature cytokines associated with ILCs in a mouse model of unsuccessful pregnancy induced by LPS, and we describe the dynamic changes they undergo during this process. We found that mice exposed to LPS display significantly higher levels of uNK cells, and uILC3s. However, a lower proportion of uILC2s and uILC1s were detected in abortion mice. In addition, we found that abortion mice display markedly higher expression of IFN-γ and IL-A17, and lower levels of IL-5. No significant differences in the expression of IL-13 and IL-22 were observed. The findings suggest that uILCs play distinct non-redundant roles during pregnancy, and uILCs may affect maternal-fetal tolerance via IL-17A, IL-5, and IFN-γ production.

Follicular dynamics of glycerophospholipid and sphingolipid metabolisms in polycystic ovary syndrome patients.

Polycystic ovary syndrome (PCOS) is a common heterogeneous disease, affecting up to 5-10% women at reproductive age. Although PCOS patients could produce morphologically normal metaphase II oocytes undergoing assisted reproductive techniques (ART), oocyte developmental competence and embryo development have been impaired in following in-vitro fertilization (IVF) steps. Follicular fluid (FF) provides a variety of information in oocyte environment when oocytes grow. In the present work, based on ultra-performance liquid chromatography coupled with mass spectrometry (UPLC-MS), the metabolic signatures of PCOS FF have been compared with healthy women using untargeted metabolomics approach. Significant abundance differences of a series of glycerolipid, glycerophospholipids, sphingolipids, and carboxylic acids have been discovered. Among them, reduced levels of phosphatidylglycerolphosphate (PGP) and a triglyceride (TG) were highly related to the lower fertilization rate in PCOS; increased abundance of lysoPE and decreased amount of PC were significantly correlated with LH/FSH (ratio of luteinizing hormone to follicle stimulating hormone). Some metabolites, including decreased sphingolipids, glycerophospholipids, and fluctuated fatty acyls, also performed close relationship with other ART and clinical results. We concluded that dysfunctions in the metabolism of glycerolipid, glycerophospholipid, sphingolipid, and glycosphingolipid biosynthesis in PCOS patients' follicles play a non-ignorable role in declining the 2 pronuclei (PN) fertilization rate during IVF procedure.

MIF May Participate in Pathogenesis of Polycystic Ovary Syndrome in Rats through MAPK Signalling Pathway.

The polycystic ovary syndrome (PCOS) model was established in rats and correlation between the expression of macrophage migration inhibitory factor (MIF) and cytokinesis with the MAPK signalling pathway in the rat ovary was measured. The PCOS model in rats was established by dehydroepiandrosterone (DHEA). Thirty sexually immature female Sprague-Dawley rats were randomly and equally assigned to three groups: control group, PCOS group, and PCOS with high-fat diet (HFD) group. Serum hormones were assayed by radioimmunoassay (RIA). The ovaries were immunohistochemically stained with MIF, and the expression of MIF, p-JNK and p-p38 was detected by Western blotting in ovaries. The serum testosterone level, LH concentration, LH/FSH ratio, fasting insulin level and HOMA IR index in the PCOS group (6.077±0.478, 13.809±1.701, 1.820±0.404, 10.83±1.123 and 1.8692±0.1096) and PCOS with HFD group (6.075±0.439, 14.075±1.927, 1.779±0.277, 10.20±1.377 and 1.7736±0.6851) were significantly higher than those in the control group (4.949±0.337, 2.458±0.509, 1.239±0.038, 9.53±0.548 and 1.5329±0.7363), but there was no significant difference between the PCOS group and PCOS with HFD group. The expression levels of MIF, p-JNK, and p-p38 in the PCOS group (0.4048±0.013, 0.6233±0.093 and 0.7987±0.061) and PCOS with HFD group (0.1929±0.012, 0.3346±0.103 and 0.3468±0.031) were obviously higher than those in control group (0.2492±0.013, 0.3271±0.093 and 0.3393±0.061), but no significant difference was observed between PCOS group and PCOS with HFD group. It was suggested that MIF may participate in the pathogenesis of PCOS through the MAPK signalling pathway in PCOS rats induced by DHEA.

CTRP3 attenuates high-fat diet-induced male reproductive dysfunction in mice.

Recent studies have suggested a role for abdominal obesity in male infertility. Previous studies have found that cell apoptosis exerts an important role in obesity-related male infertility. C1q/TNF-related protein 3 (CTRP3), a paralog of adiponectin, has been proposed to exert anti-apoptotic effects and to attenuate diabetes-related cardiac injuries. However, the role of CTRP3 in high-fat diet (HFD)-induced spermatogenic impairment remains unclear. In the present study, we fed male mice an HFD for 24 weeks to induce obesity. The expression of CTRP3 was decreased by HFD feeding. Supplementation with the recombinant human globular domain of CTRP3 (0.25 µg/g/day) for 4 weeks beginning at 20 weeks of the HFD improved spermatogenic function in the HFD-fed mice, which were characterized by improved testis morphology, increased testis weight/body weight ratio, and increased sperm count, sperm viability, and sperm motility. We also found that CTRP3 infusion resulted in the attenuation of endoplasmic reticulum (ER) stress and the activation of silence information regulator 1 (SIRT1) in the testes of obese mice. Our in vitro study also suggested that CTRP3 attenuated the palmitic acid (PA)-induced reductions in sperm viability and motility via the inhibition of ER stress. Moreover, germ cell-specific Sirtuin1 knockout abolished the protective effects of CTRP3 in vivo and in vitro. In vitro studies of human sperm showed that the protective effects of CTRP3 on sperm viability and motility were abrogated by a specific inhibitor of SIRT1. Thus, our results demonstrated that CTRP3 expression protected against HFD-induced spermatogenic deficiency through the SIRT1/ER stress pathway.

Acetyl-L-carnitine: An effective antioxidant against cryo-damage on human spermatozoa with asthenospermia.

A variety of natural and artificial cryoprotectant extenders have been explored to enhance sperm recovery following cryopreservation-thawing process. The current investigation is aimed at evaluating the effect of acetyl-L-carnitine on human spermatozoa and reactive species oxygen (ROS) level after freezing-thawing process. The spermatozoa were collected from 35 male patients diagnosed as having asthenospermia. The cryopreservation of human spermatozoa treated with acetyl-L-carnitine at different concentrations (group B: 2.5 mmol/L, group C: 7.5 mmol/L, group D: 15 mmol/L) was compared with control (group A: no acetyl-L-carnitine given). For the frozen-thawed spermatozoa, the viability, motility and DNA integrity were measured by comet assay, acrosome integrity by FITC-PNA staining and ROS level was determined in each group. The results showed that there were no significant differences in motility and viability between group A and group B, while the motility and viability of spermatozoa in group C and group D were significantly increased as compared with those in group A. As compared with group A, the values for DNA integrity parameters including comet rate (CR), tail DNA percentage (TD), tail length (TL) and Oliver tail moment (OTM) were significantly reduced in group C and group D. Group C and group D also displayed a higher proportion of intact acrosome than group A. No significant difference in ROS level was found between group A and group B, while with the increase in acetyl-L-carnitine concentration, the ROS level in groups C and D was significantly reduced as compared with that in group A. In conclusion, acetyl-L-carnitine at a concentration of 7.5 mmol/L is an effective antioxidant against cryo-damage on post-thawed human spermatozoa.

Reproductive outcomes in women with prior cesarean section undergoing in vitro fertilization: A retrospective case-control study.

The impact of prior cesarean section (CS) on the pregnancy and neonatal outcomes of in vitro fertilization and embryo transfer (IVF-ET) was investigated. A retrospective analysis was performed on 144 patients with prior CS between January 2013 and December 2015. The pregnancy, delivery, and neonatal outcomes of patients who had previous CS delivery and received IVF-ET were analyzed. The control group comprised 166 patients who had only previous vaginal delivery (VD) and received IVF-ET during the same period. The results showed that the basal follicle stimulating hormone level, estradiol level on human chorionic gonadotropin (hCG) day, gonadotrophin dosage, duration of stimulation, retrieved oocytes, fertilization rate, high-quality embryo rate, multiple birth rate, abortion rate and ectopic pregnancy rate had no significant difference between the two groups (P>0.05). The pregnancy rate (40.28% vs. 54.22%) and implantation rate (24.01% vs. 34.67%) were significantly lower (P<0.05), and the ratio of embryo difficulty transfer (9/144 vs. 0/166) was significantly higher in CS group than in VD group. The risk of pernicious placenta previa and postpartum hemorrhage in twin deliveries was significantly increased in CS group as compared with that in VD group (P<0.05), and gestational age and neonatal birth weight were significantly reduced in twin deliveries as compared with singleton deliveries in both groups (P<0.05). It was suggested that the existence of CS scar may impact embryo implantation and clinical pregnancy outcome, and increase the difficulty of ET. We should limit the number of transfer embryos to avoid multiple pregnancies and strengthen gestational supervision in patients with cesarean scar.

Follicular metabolic changes and effects on oocyte quality in polycystic ovary syndrome patients.

Polycystic ovary syndrome (PCOS) is a common complex and heterogeneous disorder, affecting up to 10% women at reproductive age. It causes three fourth of the ovulatory infertility and PCOS patients often give poor IVF quality. Although some metabolic profiles have been investigated in PCOS patient sera and urine, the follicular fluid, providing fruitful biochemical information about oocyte environment during development has been ignored. In this work, based on NMR metabolomics approach, metabolic profile of follicular fluid of PCOS patients has been explored and compared with healthy controls. Significant increases of glycoprotein, acetate, cholesterol, significant decreases of lactic acid, glutamine, pyruvate, and alanine, have been discovered in PCOS follicular fluids. Furthermore, the Pearson correlations analysis indicated significant relationship existed between ART results and NMR detected follicular metabolites. All these results indicated that PCOS may induce dyslipidemia, low-grade inflammation, and disorder of glycolysis, pyruvate and amino acid metabolism in follicular fluids.

ATP synthase ß-subunit abnormality in pancreas islets of rats with polycystic ovary syndrome and type 2 diabetes mellitus.

This study investigated the abnormal expression of ATP synthase ß-subunit (ATPsyn-ß) in pancreas islets of rat model of polycystic ovary syndrome (PCOS) with type 2 diabetes mellitus (T2DM), and the secretion function changes after up-regulation of ATP5b. Sixty female SD rats were divided into three groups randomly and equally. The rat model of PCOS with T2DM was established by free access to the high-carbohydrate/high-fat diet, subcutaneous injections of DHEA, and a single injection of streptozotocin. The pancreas was removed for the detection of the ATPsyn-ß expression by immunohistochemical staining, Western blotting and reverse transcription-PCR (RT-PCR). The pancreas islets of the rats were cultured, isolated with collagenase V and purified by gradient centrifugation, and the insulin secretion after treatment with different glucose concentrations was tested. Lentivirus ATP5b was successfully constructed with the vector of GV208 and transfected into the pancreas islets for the over-expression of ATPsyn-ß. The insulin secretion and intracellular ATP content were determined after transfection of the PCOS-T2DM pancreas islets with Lenti-ATP5b. The results showed that the expression of ATPsyn-ß protein and mRNA was significantly decreased in the pancreas of PCOS-T2DM rats. The ATP content in the pancreas islets was greatly increased and the insulin secretion was improved after the up-regulation of ATPsyn-ß in the pancreas islets transfected with lenti-ATP5b. These results indicated that for PCOS, the ATPsyn-ß might be one of the key factors for the attack of T2DM.

Diet-induced obesity impairs spermatogenesis: a potential role for autophagy.

Autophagy is an evolutionarily conserved process that plays a crucial role in maintaining a series of cellular functions. It has been found that autophagy is closely involved in the physiological process of spermatogenesis and the regulation of sperm survival and motility. However, the role of autophagy in high-fat diet (HFD)-induced impaired spermatogenesis remains unknown. This study was designed to investigate the role of autophagy in HFD-induced spermatogenesis deficiency and employed chloroquine (CQ) to inhibit autophagy and rapamycin (RAP) to induce autophagy. 3-methyladenine (3-MA) and CQ were administered via intratesticular injection in vivo. The effects of CQ and 3-MA on the parameters of spermatozoa co-cultured with palmitic acid (PA) in vitro were also investigated. Human semen samples from obese, subfertile male patients were also collected to examine the level of autophagy. The results suggested that HFD mice subjected to CQ showed improved spermatogenesis. Inhibiting autophagy with CQ improved the decreased fertility of HFD male mice. Moreover, the in vivo and in vitro results indicated that both CQ and 3-MA could suppress the pathological changes in spermatozoa caused by HFD or PA treatment. Additionally, the excessive activation of autophagy was also observed in sperm samples from obese, subfertile male patients.