[Research research on the use of melatonin in combination with therapeutic hypothermia for the treatment of neonatal hypoxic-ischemic encephalopathy]. / è¤ªé»‘ç´ è”åˆäºšä½Žæ¸©æ²»ç–—æ–°ç”Ÿå„¿ç¼ºæ°§ç¼ºè¡€æ€§è„‘ç— çš„ç ”ç©¶è¿›å±•.

Neonatal hypoxic-ischemic encephalopathy (HIE) remains one of the leading causes of death and long-term neurodevelopmental disorders in full-term neonates, and there is currently no curative treatment. Therapeutic hypothermia is now a standard therapy for HIE in the neonatal intensive care unit, but its safety and efficacy in remote areas remains unclear. Melatonin is an indole endocrine hormone mainly produced by the pineal gland and it has the ability to easily penetrate the blood-brain barrier. Through receptor and non-receptor mechanisms, melatonin exerts anti-oxidative and anti-inflammatory effects and participates in the regulation of organelle function and the inhibition of cell death. Melatonin is considered one of the most promising drugs for the treatment of HIE based on its reliable safety profile and clinical/preclinical results. This article reviews the recent research on the use of melatonin in combination with therapeutic hypothermia for the treatment of neonatal HIE.

[Risk factors for neonatal asphyxia and establishment of a nomogram model for predicting neonatal asphyxia in Hubei Enshi Tujia and Miao Autonomous Prefecture: a multicenter study]. / æ¹–åŒ—æ©æ–½åœŸå®¶æ—è‹—æ—è‡ªæ²»å·žæ–°ç”Ÿå„¿çª’æ¯å±é™©å› ç´ åˆ†æžåŠåˆ—çº¿å›¾é¢„æµ‹æ¨¡åž‹çš„æž„å»ºï¼šä¸€é¡¹å¤šä¸­å¿ƒç ”ç©¶.

OBJECTIVES: To investigate the risk factors for neonatal asphyxia in Hubei Enshi Tujia and Miao Autonomous Prefecture and establish a nomogram model for predicting the risk of neonatal asphyxia. METHODS: A retrospective study was conducted with 613 cases of neonatal asphyxia treated in 20 cooperative hospitals in Enshi Tujia and Miao Autonomous Prefecture from January to December 2019 as the asphyxia group, and 988 randomly selected non-asphyxia neonates born and admitted to the neonatology department of these hospitals during the same period as the control group. Univariate and multivariate analyses were used to identify risk factors for neonatal asphyxia. R software (4.2.2) was used to establish a nomogram model. Receiver operator characteristic curve, calibration curve, and decision curve analysis were used to assess the discrimination, calibration, and clinical usefulness of the model for predicting the risk of neonatal asphyxia, respectively. RESULTS: Multivariate logistic regression analysis showed that minority (Tujia), male sex, premature birth, congenital malformations, abnormal fetal position, intrauterine distress, maternal occupation as a farmer, education level below high school, fewer than 9 prenatal check-ups, threatened abortion, abnormal umbilical cord, abnormal amniotic fluid, placenta previa, abruptio placentae, emergency caesarean section, and assisted delivery were independent risk factors for neonatal asphyxia (P<0.05). The area under the curve of the model for predicting the risk of neonatal asphyxia based on these risk factors was 0.748 (95%CI: 0.723-0.772). The calibration curve indicated high accuracy of the model for predicting the risk of neonatal asphyxia. The decision curve analysis showed that the model could provide a higher net benefit for neonates at risk of asphyxia. CONCLUSIONS: The risk factors for neonatal asphyxia in Hubei Enshi Tujia and Miao Autonomous Prefecture are multifactorial, and the nomogram model based on these factors has good value in predicting the risk of neonatal asphyxia, which can help clinicians identify neonates at high risk of asphyxia early, and reduce the incidence of neonatal asphyxia.

Proteasome subunit α4s is essential for formation of spermatoproteasomes and histone degradation during meiotic DNA repair in spermatocytes.

Meiosis, which produces haploid progeny, is critical to ensuring both faithful genome transmission and genetic diversity. Proteasomes play critical roles at various stages of spermatogenesis, including meiosis, but the underlying mechanisms remain unclear. The atypical proteasomes, which contain the activator PA200, catalyze the acetylation-dependent degradation of the core histones in elongated spermatids and DNA repair in somatic cells. We show here that the testis-specific proteasome subunit α4s/PSMA8 is essential for male fertility by promoting proper formation of spermatoproteasomes, which harbor both PA200 and constitutive catalytic subunits. Immunostaining of a spermatocyte marker, SYCP3, indicated that meiosis was halted at the stage of spermatocytes in the α4s-deficient testes. α4s stimulated the in vitro degradation of the acetylated core histones, instead of nonacetylated histones, by the PA200-proteasome. Deletion of α4s blocked degradation of the core histones at DNA damage loci in spermatocytes, leading to meiotic arrest at metaphase I. Thus, α4s is required for histone degradation at meiotic DNA damage loci, proper progression of meiosis, and fertility in males by promoting proper formation of spermatoproteasomes. These results are important for understanding male infertility and might provide potential targets for male contraception or treatment of male infertility.

Cyanidin-3-Ο-glucoside supplementation in cryopreservation medium improves human sperm quality.

Currently, the cryopreservation of human spermatozoa must overcome the adverse effects of excessive oxidation. In this study, we aimed to evaluate the effect of supplementation of cryopreservation medium with cyanidin-3-Ο-glucoside (C3G) on sperm quality. Semen samples were obtained from men with normozoospermia according to WHO criteria (n = 39). The sperm parameter values were compared after cryopreservation in medium supplemented with and without C3G.Compared with the control group (without additive), low doses (50 µM and 100 µM) of C3G improved sperm viability and motility and decreased the reactive oxygen species (ROS) of spermatozoa, while high doses (200 µM) of C3G did not obviously enhance sperm quality. The amount of DNA fragmentation index (DFI) and high DNA stainability (HDS) after freezing were higher in the control group than in the C3G supplementation groups. Low-concentration C3G supplementation (50 µM) was negatively correlated with sperm ROS levels (r = -0.2, p = 0.03). Collectively, our findings suggest that C3G could be an efficient semen cryoprotectant that ameliorates oxidative stress in human sperm during cryopreservation.

Extracellular HSPs: The Potential Target for Human Disease Therapy.

Heat shock proteins (HSPs) are highly conserved stress proteins known as molecular chaperones, which are considered to be cytoplasmic proteins with functions restricted to the intracellular compartment, such as the cytoplasm or cellular organelles. However, an increasing number of observations have shown that HSPs can also be released into the extracellular matrix and can play important roles in the modulation of inflammation and immune responses. Recent studies have demonstrated that extracellular HSPs (eHSPs) were involved in many human diseases, such as cancers, neurodegenerative diseases, and kidney diseases, which are all diseases that are closely linked to inflammation and immunity. In this review, we describe the types of eHSPs, discuss the mechanisms of eHSPs secretion, and then highlight their functions in the modulation of inflammation and immune responses. Finally, we take cancer as an example and discuss the possibility of targeting eHSPs for human disease therapy. A broader understanding of the function of eHSPs in development and progression of human disease is essential for developing new strategies to treat many human diseases that are critically related to inflammation and immunity.

[Correlation of the platelet count in the peripheral blood with the prostate volume].

OBJECTIVE: To investigate the relationship of the prostate volume with the count of inflammatory cells in the peripheral blood and clarify the pathogenesis of BPH. METHODS: From 2015 to 2019, we enrolled 104 men pathologically diagnosed with BPH. Using univariate and multivariate linear regression analysis models, we analyzed the correlation of the prostate volume with the neutrophil count, platelet count, neutrophil-lymphocyte ratio (NLR), platelet-WBC ratio (PWR), and lymphocyte-monocyte ratio (LMR) in the peripheral blood of the patients. RESULTS: Both the platelet count (r = 0.401, P < 0.001) and PWR (r = 0.343, P < 0.001) in the peripheral blood were positively correlated with the prostate volume and serum PSA level, but not with IPSS. No evident relationship was found between the prostate volume and the systemic inflammatory markers NLR and LMR. CONCLUSIONS: The platelet count in the peripheral blood is an important predictor of BPH and may play an important role in the development and progression of BPH.

An exploration of the role of Sertoli cells on fetal testis development using cell ablation strategy.

Sertoli cells (SCs) are presumed to be the center of testis differentiation because they provide both structural support and biological regulation for spermatogenesis. Previous studies suggest that SCs control germ cell (GC) count and Leydig cell (LC) development in mouse testes. However, the regulatory role of SCs on peritubular myoid (PTM) cell fate in fetal testis has not been clearly reported. Here, we employed Amh-Cre; diphtheria toxin fragment A (DTA) mouse model to selectively ablate SCs from embryonic day (E) 14.5. Results found that SC ablation in the fetal stage caused the disruption of testis cords and the massive loss of GCs. Furthermore, the number of α-smooth muscle actin-labeled PTM cells was gradually decreased from E14.5 and almost lost at E18.5 in SC ablation testis. Interestingly, some Ki67 and 3ß-HSD double-positive fetal LCs could be observed in Amh-Cre; DTA testes at E16.5 and E18.5. Consistent with this phenomenon, the messenger RNA levels of Hsd3b1, Cyp11a1, Lhr, Star and the protein levels of 3ß-HSD and P450Scc were significantly elevated by SC ablation. SC ablation appears to induce ectopic proliferation of fetal LCs although the total LC number appeared reduced. Together, these findings bring us a better understanding of SCs' central role in fetal testis development.

Testicular germ cell tumor: a comprehensive review.

Testicular tumors are the most common tumors in adolescent and young men and germ cell tumors (TGCTs) account for most of all testicular cancers. Increasing incidence of TGCTs among males provides strong motivation to understand its biological and genetic basis. Gains of chromosome arm 12p and aneuploidy are nearly universal in TGCTs, but TGCTs have low point mutation rate. It is thought that TGCTs develop from premalignant intratubular germ cell neoplasia that is believed to arise from the failure of normal maturation of gonocytes during fetal or postnatal development. Progression toward invasive TGCTs (seminoma and nonseminoma) then occurs after puberty. Both inherited genetic factors and environmental risk factors emerge as important contributors to TGCT susceptibility. Genome-wide association studies have so far identified more than 30 risk loci for TGCTs, suggesting that a polygenic model fits better with the genetic landscape of the disease. Despite high cure rates because of its particular sensitivity to platinum-based chemotherapy, exploration of mechanisms underlying the occurrence, progression, metastasis, recurrence, chemotherapeutic resistance, early diagnosis and optional clinical therapeutics without long-term side effects are urgently needed to reduce the cancer burden in this underserved age group. Herein, we present an up-to-date review on clinical challenges, origin and progression, risk factors, TGCT mouse models, serum diagnostic markers, resistance mechanisms, miRNA regulation, and database resources of TGCTs. We appeal that more attention should be paid to the basic research and clinical diagnosis and treatment of TGCTs.

Distinct Metabolic Features of Seminoma and Embryonal Carcinoma Revealed by Combined Transcriptome and Metabolome Analyses.

Seminoma and embryonal carcinoma (EC), two typical types of testicular germ cell tumors (TGCTs), present significant differences in growth behavior, expression characteristics, differentiation potential, clinical features, therapy, and prognosis. The purpose of this study was to compare the distinctive or preference metabolic pathways between seminoma and EC. The Cancer Genome Atlas revealed that many genes encoding metabolic enzymes could distinguish between seminoma and EC. Using well-characterized cell line models for seminoma (Tcam-2 cells) and EC (NT2 cells), we characterized their metabolite profiles using ultraperformance liquid chromatography coupled to Q-TOF mass spectrometry (UPLC/Q-TOF MS). In general, the integrated results from transcriptome and metabolite profiling revealed that seminoma and EC exhibited distinctive characteristics in the metabolisms of amino acids, glucose, fatty acids, sphingolipids, nucleotides, and drugs. Notably, an attenuation of citric acid cycle/mitochondrial oxidative phosphorylation and sphingolipid biosynthesis as well as an increase in arachidonic acid metabolism and (very) long-chain fatty acid abundance occurred in seminoma as compared with EC. Our study suggests histologic subtype-dependent metabolic reprogramming in TGCTs and will lead to a better understanding of the metabolic signatures and biology of TGCT subtypes.

In-vitro differentiation of early pig spermatogenic cells to haploid germ cells.

Spermatogonial stem cells (SSCs) self-renew and contribute genetic information to the next generation. Pig is wildly used as a model animal for understanding reproduction mechanisms of human being. Inducing directional differentiation of porcine SSCs may be an important strategy in exploring the mechanisms of spermatogenesis and developing better treatment methods for male infertility. Here, we established an in-vitro culture model for porcine small seminiferous tubule segments, to induce SSCs to differentiate into single-tail haploid spermatozoa. The culture model subsequently enabled spermatozoa to express the sperm-specific protein acrosin and oocytes to develop to blastocyst stage after round spermatid injection. The addition of retinoic acid (RA) to the differentiation media promoted the efficiency of haploid differentiation. RT-PCR analysis indicated that RA stimulated the expression of Stra8 but reduced the expression of NANOS2 in spermatogonia. Genes involved in post-meiotic development, transition protein 1 (Tnp1) and protamine 1 (Prm1) were upregulated in the presence of RA. The addition of an RA receptor (RAR) inhibitor, BMS439, showed that RA enhanced the expression of cAMP responsive-element binding protein through RAR and promoted the formation of round spermatids. We established an efficient culture system for in-vitro differentiation of pig SSCs. Our study represents a model for human testis disease and toxicology screening. Molecular regulators of SSC differentiation revealed in this study might provide a therapeutic strategy for male infertility.

Regulation of blood-testis barrier assembly in vivo by germ cells.

The assembly of the blood-testis barrier (BTB) during postnatal development is crucial to support meiosis. However, the role of germ cells in BTB assembly remains unclear. Herein, KitW/KitWV mice were used as a study model. These mice were infertile, failing to establish a functional BTB to support meiosis due to c-Kit mutation. Transplantation of undifferentiated spermatogonia derived from normal mice into the testis of KitW/KitWV mice triggered functional BTB assembly, displaying cyclic remodeling during the epithelial cycle. Also, transplanted germ cells were capable of inducing Leydig cell testosterone production, which could enhance the expression of integral membrane protein claudin 3 in Sertoli cells. Early spermatocytes were shown to play a vital role in directing BTB assembly by expressing claudin 3, which likely created a transient adhesion structure to mediate BTB and cytoskeleton assembly in adjacent Sertoli cells. In summary, the positive modulation of germ cells on somatic cell function provides useful information regarding somatic-germ cell interactions.-Li, X.-Y., Zhang, Y., Wang, X.-X., Jin, C., Wang, Y.-Q., Sun, T.-C., Li, J., Tang, J.-X., Batool, A., Deng, S.-L., Chen, S.-R., Cheng, C. Y., Liu, Y.-X. Regulation of blood-testis barrier assembly in vivo by germ cells.

Development, function and fate of fetal Leydig cells.

During fetal testis development, fetal Leydig cells (FLCs) are found to be originated from multiple progenitor cells. FLC specification and function are under tight regulation of specific genes and signaling proteins. Furthermore, Sertoli cells play a crucial role to regulate FLC differentiation during fetal testis development. FLC progenitor- and FLC-produced biomolecules are also involved in the differentiation and activity of rodent FLCs. The main function of FLCs is to produce androgens to masculinize XY embryos. However, FLCs are capable of producing androstenedione but not testosterone due to the lack of 17ß-HSD (17ß-hydroxysteroid dehydrogenase), but fetal Sertoli cells express 17ß-HSD which thus transforms androstenedione to testosterone in the fetal testis. On the other hand, FLCs produce activin A to regulate Sertoli cell proliferation, and Sertoli cells in turn modulate testis cord expansion. It is now generally accepted that adult Leydig cells (ALCs) gradually replace FLCs during postnatal development to produce testosterone to support spermatogenesis as FLCs undergo degeneration in neonatal and pre-pubertal testes. However, based on studies using genetic tracing mouse models, FLCs are found to persist in adult testes, making up ∼20% of total Leydig cells. In this review, we evaluate the latest findings regarding the development, function and fate of FLCs during fetal and adult testis development.

Signaling pathways regulating blood-tissue barriers - Lesson from the testis.

Signaling pathways that regulate blood-tissue barriers are important for studying the biology of various blood-tissue barriers. This information, if deciphered and better understood, will provide better therapeutic management of diseases particularly in organs that are sealed by the corresponding blood-tissue barriers from systemic circulation, such as the brain and the testis. These barriers block the access of antibiotics and/or chemotherapeutical agents across the corresponding barriers. Studies in the last decade using the blood-testis barrier (BTB) in rats have demonstrated the presence of several signaling pathways that are crucial to modulate BTB function. Herein, we critically evaluate these findings and provide hypothetical models regarding the underlying mechanisms by which these signaling molecules/pathways modulate BTB dynamics. This information should be carefully evaluated to examine their applicability in other tissue barriers which shall benefit future functional studies in the field. This article is part of a Special Issue entitled: Gap Junction Proteins edited by Jean Claude Herve.

CRISPR/Cas9-mediated genome editing induces gene knockdown by altering the pre-mRNA splicing in mice.

BACKGROUND: Clustered Regularly Interspaced Short Palindromic Repeats/CRISPR associated protein 9 (CRISPR/Cas9) has been wildly used to generate gene knockout models through inducing indels causing frame-shift. However, there are few studies concerning the post-transcript effects caused by CRISPR-mediated genome editing. RESULTS: In the present study, we showed that gene knockdown model also could be generated using CRISPR-mediated gene editing by disrupting the boundary of exon and intron in mice (C57BL/6 J). CRISPR induced indel at the boundary of exon and intron (5' splice site) caused alternative splicing and produced multiple different mRNAs, most of these mRNAs introduced premature termination codon causing down expression of the gene. CONCLUSIONS: These results showed that alternative splicing mutants were able to generate through CRISPR-mediated genome editing by deleting the boundary of exon and intron causing disruption of 5' splice site. Although alternative splicing was an unexpected outcome, this finding could be developed as a technology to generate gene knockdown models or to investigate pre-mRNA splicing.

GATA4 is a negative regulator of contractility in mouse testicular peritubular myoid cells

Reduced contractility of the testicular peritubular myoid (PTM) cells may contribute to human male subfertility or infertility. Transcription factor GATA4 in Sertoli and Leydig cells is essential for murine spermatogenesis, but limited attention has been paid to the potential role of GATA4 in PTM cells. In primary cultures of mouse PTM cells, siRNA knockdown of GATA4 increased the contractile activity, while GATA4 overexpression significantly attenuated the contractility of PTM cells using a collagen gel contraction assay. Using RNA sequencing and qRT-PCR, we identified a set of genes that exhibited opposite expressional alternation between Gata4 siRNA vs nontargeting siRNA-treated PTM cells and Gata4 adenovirus vs control adenovirus-treated PTM cells. Notably, ion channels, smooth muscle function, cytokines and chemokines, cytoskeleton, adhesion and extracellular matrix were the top four enriched pathways, as revealed by cluster analysis. Natriuretic peptide type B (NPPB) content was significantly upregulated by GATA4 overexpression in both PTM cells and their culture supernatant. More importantly, the addition of 100 µM NPPB could abolish the promoting effect of Gata4 silencing on PTM cell contraction. Taken together, we suggest that the inhibitory action of GATA4 on PTM cell contraction is mediated at least partly by regulating genes belonging to smooth muscle contraction pathway (e.g. Nppb).

Role of WNT signaling in epididymal sperm maturation.

PURPOSE: Spermatozoa maturation, a process required for spermatozoa to acquire progressive motility and the ability to fertilize ova, primarily occurs in the caput and corpus of the epididymis. Despite considerable efforts, the factor(s) promoting epididymal sperm maturation remains unclear. Recently, WNT signaling has been implicated in epididymal sperm maturation. METHODS: To further investigate WNT signaling function in epididymal sperm maturation, we generated Wntless conditional knockout mice (Wls cKO), Wls flox/flox ; Lcn5-Cre. RESULTS: In these mice, WNTLESS (WLS), a conserved membrane protein required for all WNT protein secretion, was specifically disrupted in the principal cells of the caput epididymidis. Immunoblot analysis showed that WLS was significantly reduced in the caput epididymidis of Wls cKO mice. In the caput epididymidis of Wls cKO mice, WNT 10A and WNT 2b, which are typically secreted by the principal cells of the caput epididymis, were not secreted. Interestingly, sperm motility analysis showed that the WLS deficiency in the caput epididymidis had no effect on sperm motility. Moreover, fertility tests showed that Wls cKO male mice had normal fertility. CONCLUSION: These results indicate that the disruption of WLS in principal cells of the caput epididymidis inhibits WNT protein secretion but has no effect on sperm motility and male fertility, suggesting that WNT signaling in the caput epididymidis may be dispensable for epididymal sperm maturation in mice.

Melatonin Regulates the Synthesis of Steroid Hormones on Male Reproduction: A Review.

Melatonin is a ubiquitous molecule and exhibits different effects in long-day and short-day breeding animals. Testosterone, the main resource of androgens in the testis, is produced by Leydig cells but regulated mainly by cytokine secreted by Sertoli cells. Melatonin acts as a local modulator of the endocrine activity in Leydig cells. In Sertoli cells, melatonin influences cellular proliferation and energy metabolism and, consequently, can regulate steroidogenesis. These suggest melatonin as a key player in the regulation of steroidogenesis. However, the melatonin-induced regulation of steroid hormones may differ among species, and the literature data indicate that melatonin has important effects on steroidogenesis and male reproduction.

Melatonin-Mediated Development of Ovine Cumulus Cells, Perhaps by Regulation of DNA Methylation.

Cumulus cells of pre-pubertal domestic animals are dysfunctional, perhaps due to age-specific epigenetic events. This study was designed to determine effects of melatonin treatment of donors on methylation modification of pre-pubertal cumulus cells. Cumulus cells from germinal vesicle stage cumulus oocyte complexes (COCs) were collected from eighteen lambs which were randomly divided into control group (C) and melatonin group given an 18 mg melatonin implant subcutaneous (M). Compared to the C group, the M group had higher concentrations of melatonin in plasma and follicular fluid (p < 0.05), greater superovulation, a higher proportion of fully expanded COCs, and a lower proportion of apoptotic cumulus cells (p < 0.05). Real-time PCR results showed that melatonin up-regulated expression of genes MT1, Bcl2, DNMT1, DNMT3a and DNMT3b, but down-regulated expression of genes p53, Caspase 3 and Bax (p < 0.05). Furthermore, melatonin increased FI of FITC (global methylation level) on cumulus cells (p < 0.05). To understand the regulation mechanism, the DNMTs promoter methylation sequence were analyzed. Compared to the C group, although there was less methylation at two CpG sites of DNMT1 (p < 0.05) and higher methylation at two CpG sites of DNMT3a (p < 0.05), there were no significant differences in methylation of the detected DNMT1 and DNMT3a promoter regions. However, there were lower methylation levels at five CpG sites of DNMT3b, which decreased methylation of detected DNMT3b promoter region on M group (p < 0.05). In conclusion, alterations of methylation regulated by melatonin may mediate development of cumulus cells in lambs.

Impaired telomere length and telomerase activity in peripheral blood leukocytes and granulosa cells in patients with biochemical primary ovarian insufficiency.

STUDY QUESTION: Are telomere length and telomerase activity associated with biochemical primary ovarian insufficiency (POI)? SUMMARY ANSWER: Shortened telomere length and diminished telomerase activity were associated with biochemical POI. WHAT IS KNOWN ALREADY: POI is a result of pathological reproductive aging and encompasses occult, biochemical and overt stages. Studies have indicated telomere length as a biomarker for biological aging. STUDY DESIGN, SIZE, DURATION: A total of 120 patients with biochemical POI and 279 control women were recruited by the Center for Reproductive Medicine of Shandong University. PARTICIPANTS/MATERIALS, SETTING, METHODS: Telomere length in peripheral blood leukocytes (LTL) and granulosa cells (GTL) was measured using a modified Quantitative Polymerase Chain Reaction technique. The relative telomerase activity (RTA) in granulosa cells was detected using a modified quantitative-telomeric repeat amplification protocol assay. MAIN RESULTS AND THE ROLE OF CHANCE: After adjusting for age, patients with biochemical POI (n = 120) exhibited significantly shorter LTLs (0.75 ± 0.09 vs 1.79 ± 0.12, P < 0.001; OR = 0.54, 95% CI = 0.43-0.68) and GTLs (0.78 ± 0.09 vs 1.90 ± 0.23, P < 0.001; OR = 0.54, 95% CI = 0.41-0.70) than the controls (n = 279 for LTLs; n = 90 for GTLs). Significantly diminished RTAs in granulosa cells were detected in patients with biochemical POI (n = 31) compared with the controls (n = 38) (1.57 ± 0.59 vs 4.63 ± 0.93, P = 0.025; OR = 0.84, 95% CI = 0.72-0.98). LARGE SCALE DATA: N/A. LIMITATIONS, REASONS FOR CAUTION: The cross-sectional nature of this study might have its limit in telomere length as well as telomerase activity along with the progressing decline in ovarian function. WIDER IMPLICATIONS OF THE FINDINGS: These findings suggest that telomere length and telomerase activity may be considered as indicators for progression of ovarian decline. STUDY FUNDING/COMPETING INTERESTS: This research was supported by the National Basic Research Program of China (973 Program) (2012CB944700), Science research foundation item of no-earnings health vocation (201402004) and the National Natural Science Foundation of China (31471352, 81270662, 81471509, 81300461, 81522018). The authors have no potential conflict of interest to declare.

EZH2 deletion promotes spermatogonial differentiation and apoptosis.

Spermatogenesis is crucial for male fertility and is therefore tightly controlled by a variety of epigenetic regulators. However, the function of enhancer of zeste homolog 2 (EZH2) in spermatogenesis and the molecular mechanisms underlying its activity remain poorly defined. Here, we demonstrate that deleting EZH2 promoted spermatogonial differentiation and apoptosis. EZH2 is expressed in spermatogonia, spermatocytes and round and elongated spermatids from stage 9 to 11 but not in leptotene and zygotene spermatocytes. Knocking down Ezh2 in vitro using a lentivirus impaired self-renewal in spermatogonial stem cells (SSCs), and the conditional knockout of Ezh2 in spermatogonial progenitors promoted precocious spermatogonial differentiation. EZH2 functions to balance self-renewal and differentiation in spermatogonia by suppressing NEUROG3 and KIT via a direct interaction that is independent of its histone methyltransferase activity. Moreover, deleting Ezh2 enhanced the activation of CASP3 in spermatids, resulting in reduced spermatozoa production. Collectively, these data demonstrate that EZH2 plays a nonclassical role in the regulation of spermatogonial differentiation and apoptosis in murine spermatogenesis.