RNA m6A modification regulates L1 retrotransposons in human spermatogonial stem cell differentiation in vitro and in vivo.

The maintenance of genome integrity in the germline is crucial for mammalian development. Long interspersed element type 1 (LINE-1, L1) is a mobile genetic element that makes up about 17% of the human genome and poses a threat to genome integrity. N6-methyl-adenosine (m6A) plays an essential role in regulating various biological processes. However, the function of m6A modification in L1 retrotransposons and human germline development remains largely unknown. Here we knocked out the m6A methyltransferase METTL3 or the m6A reader YTHDF2 in human embryonic stem cells (hESCs) and discovered that METTL3 and YTHDF2 are crucial for inducing human spermatogonial stem cells (hSSCs) from hESCs in vitro. The removal of METTL3 or YTHDF2 resulted in increased L1 retrotransposition and reduced the efficiency of SSC differentiation in vitro. Further analysis showed that YTHDF2 recognizes the METTL3-catalyzed m6A modification of L1 retrotransposons and degrades L1 mRNA through autophagy, thereby blocking L1 retrotransposition. Moreover, the study confirmed that m6A modification in human fetal germ cells promotes the degradation of L1 retrotransposon RNA, preventing the insertion of new L1 retrotransposons into the genome. Interestingly, L1 retrotransposon RNA was highly expressed while METTL3 was significantly downregulated in the seminal plasma of azoospermic patients with meiotic arrest compared to males with normal fertility. Additionally, we identified some potentially pathogenic variants in m6A-related genes in azoospermic men with meiotic arrest. In summary, our study suggests that m6A modification serves as a guardian of genome stability during human germline development and provides novel insights into the function and regulatory mechanisms of m6A modification in restricting L1 retrotransposition.

A comprehensive meta-analysis to identify susceptibility genetic variants for precocious puberty.

PURPOSE: Currently, several genetic variants in ERα gene (rs2234693 and rs9340799), ERß gene (rs1256049 and rs4986938), KISS1 gene (rs4889, rs1132506 and rs5780218), LIN28B gene (rs314263, rs314276 and rs314280), and MKRN3 gene (rs2239669) have been repeatedly explored for their contribution to precocious puberty (PP) susceptibility. However, the results remain conflicting rather than conclusive. We here performed a meta-analysis to identify the real susceptibility genetic variants for PP. METHODS: After screening by inclusion criteria, 20 related studies were finally included in this meta-analysis. The odds ratios and 95% confidence intervals were calculated to assess the strength of association. Sensitive analysis, publication bias, and trial sequential analysis (TSA) were performed to evaluate the stability and reliability of results. RESULTS: Rs2234693, rs9340799, and rs1256049 were significantly associated with PP susceptibility (p < 0.0084). Stratified analysis according to ethnicity showed that rs2234693 and rs9340799 were significantly associated with PP susceptibility in Asian and Chinese populations. Stratified analysis according to PP subtype showed that rs2234693 and rs9340799 were significantly associated with idiopathic central PP susceptibility in Asian and Chinese populations (p < 0.0084). The results of publication bias, sensitivity analysis, and TSA provided solid evidence for the association between these three variants and PP susceptibility. CONCLUSIONS: Rs2234693 and rs9340799 in ERα gene and rs1256049 in ERß gene may serve as susceptive factors for PP development. The present finding should be confirmed in replication studies and reinforced in functional studies, which will ultimately improve the feasibility of the application of these three PP-susceptible loci in clinical practice.

Palmitoylation of vacuole membrane protein 1 promotes small extracellular vesicle secretion via interaction with ALIX and influences intercellular communication.

BACKGROUND: Small extracellular vesicles (EVs), exemplified by exosomes, mediate intercellular communication by transporting proteins, mRNAs, and miRNAs. Post-translational modifications are involved in controlling small EV secretion process. However, whether palmitoylation regulates small EV secretion, remains largely unexplored. METHODS: Vacuole Membrane Protein 1 (VMP1) was testified to be S-palmitoylated by Palmitoylation assays. VMP1 mutant plasmids were constructed to screen out the exact palmitoylation sites. Small EVs were isolated, identified and compared between wild-type VMP1 or mutant VMP1 transfected cells. Electron microscope and immunofluorescence were used to detect multivesicular body (MVB) number and morphology change when VMP1 was mutated. Immunoprecipitation and Mass spectrum were adopted to identify the protein that interacted with palmitoylated VMP1, while knock down experiment was used to explore the function of targeted protein ALIX. Taking human Sertoli cells (SCs) and human spermatogonial stem cell like cells (SSCLCs) as a model of intercellular communication, SSCLC maintenance was detected by flow cytometry and qPCR at 12 days of differentiation. In vivo, mouse model was established by intraperitoneal injection with palmitoylation inhibitor, 2-bromopalmitate (2BP) for 3 months. RESULTS: VMP1 was identified to be palmitoylated at cysteine 263,278 by ZDHHC3. Specifically, palmitoylation of VMP1 regulated its subcellular location and enhanced the amount of small EV secretion. Mutation of VMP1 palmitoylation sites interfered with the morphology and biogenesis of MVBs through suppressing intraluminal vesicle formation. Furthermore, inhibition of VMP1 palmitoylation impeded small EV secretion by affecting the interaction of VMP1 with ALIX, an accessory protein of the ESCRT machinery. Taking SCs and SSCLCs as a model of intercellular communication, we discovered VMP1 palmitoylation in SCs was vital to the growth status of SSCLCs in a co-culture system. Inhibition of VMP1 palmitoylation caused low self-maintenance, increased apoptosis, and decreased proliferation rate of SSCLCs. In vivo, intraperitoneal injection of 2BP inhibited VMP1 palmitoylation and exosomal marker expression in mouse testes, which were closely associated with the level of spermatogenic cell apoptosis and proliferation. CONCLUSIONS: Our study revealed a novel mechanism for small EV secretion regulated by VMP1 palmitoylation in Sertoli cells, and demonstrated its pivotal role in intercellular communication and SSC niche.

Sertoli cell-only syndrome: advances, challenges, and perspectives in genetics and mechanisms.

Male infertility can be caused by quantitative and/or qualitative abnormalities in spermatogenesis, which affects men's physical and mental health. Sertoli cell-only syndrome (SCOS) is the most severe histological phenotype of male infertility characterized by the depletion of germ cells with only Sertoli cells remaining in the seminiferous tubules. Most SCOS cases cannot be explained by the already known genetic causes including karyotype abnormalities and microdeletions of the Y chromosome. With the development of sequencing technology, studies on screening new genetic causes for SCOS are growing in recent years. Directly sequencing of target genes in sporadic cases and whole-exome sequencing applied in familial cases have identified several genes associated with SCOS. Analyses of the testicular transcriptome, proteome, and epigenetics in SCOS patients provide explanations regarding the molecular mechanisms of SCOS. In this review, we discuss the possible relationship between defective germline development and SCOS based on mouse models with SCO phenotype. We also summarize the advances and challenges in the exploration of genetic causes and mechanisms of SCOS. Knowing the genetic factors of SCOS offers a better understanding of SCO and human spermatogenesis, and it also has practical significance for improving diagnosis, making appropriate medical decisions, and genetic counseling. For therapeutic implications, SCOS research, along with the achievements in stem cell technologies and gene therapy, build the foundation to develop novel therapies for SCOS patients to produce functional spermatozoa, giving them hope to father children.

Analysis of different plant- and animal-based dietary patterns and their relationship with serum uric acid levels in Chinese adults.

BACKGROUND: Dietary patterns play an important role in regulating serum uric acid levels in the body, but evidence for the association between different kinds of plant-based and animal-based dietary patterns and individual serum uric acid levels is scarce and inconsistent. METHODS: We analyzed data from the sixth wave of the China Health and Nutrition Survey. The plant-based diet of 7,806 participants was determined using three consecutive 24-hour dietary recalls, and latent profile analysis was used to identify dietary patterns among participants. Serum uric acid levels were analyzed using the enzymatic colorimetric method. The association between intakes of different types of dietary pattern and individual serum uric acid levels was analyzed using linear regression analysis, after adjusting for confounding variables. RESULTS: We identified three types of plant-based dietary patterns, namely, low tuber starches and vegetable plant-based diet (LTVP), high cereal, tuber starches and vegetable plant-based diet (HCTVP), and high legume and fruit plant-based diet (HLFP). We also identified three types of animal-based dietary patterns, namely, high milk and egg animal-based diet (HMiEA), low egg and fish animal-based diet, and high meat and fish animal-based diet (HMeFA). Significant coefficients for participant serum uric acid levels were observed for the HCTVP diet (ß = -0.022, P = 0.031) and HMeFA diet (ß = 0.061, P < 0.001). The median intake of foods in the HCTVP diet was as follows: cereals and cereal products, 444.83 g/d; tubers and starch products, 166.67 g/d; dried legumes and legume products, 8.33 g/d; vegetables and vegetable products, 333.33 g/d; and fruits and fruit products, 0 g/d. The median intake of foods in the HMeFA diet was as follows: meat and meat products, 73.33 g/d; poultry and poultry products, 0 g/d; milk and milk products, 0 g/d; eggs and egg products, 26.67 g/d; and fish, shellfish, and mollusks, 180.00 g/d. CONCLUSION: We showed that individual serum uric acid levels (1) might decrease under the plant-based HCTVP diet, (2) might increase under the animal-based HMeFA diet, (3) might not decrease under the plant-based HLFP diet, and (4) might not increase under the animal-based HMiEA diet. Further studies are needed to confirm these associations.

Hardness and superconductivity in tetragonal LiB4 and NaB4.

Boron-based compounds have triggered substantial attention due to their multifunctional properties, incorporating excellent hardness and superconductivity. While tetragonal metal borides LiB4 and NaB4 with BaAl4-type structure and striking clathrate boron motif have been induced under compression, there is still a lack of deep understanding of their potential properties at ambient pressure. We herein conduct a comprehensive study on I4/mmm-structured LiB4 and NaB4 under ambient pressure via first-principles calculations. Remarkably, both LiB4 and NaB4 are found to possess high Vickers hardness of 39 GPa, which is ascribed to the robust boron framework with strong covalency. Furthermore, their high hardness values together with distinguished stability make them highly potential superhard materials. Meanwhile, electron-phonon coupling analysis reveals that both LiB4 and NaB4 are conventional phonon-mediated superconductors, with critical temperatures of 6 and 8 K at 1 atmosphere pressure (atm), respectively, mainly arising from the coupling of B 2p electronic states and the low-frequency phonon modes associated with Li-, Na-, and B-derived vibrations. This work provides valuable insights into the mechanical and superconducting behaviors of metal borides and will boost further studies of emergent borides with multiple functionalities.

[Delivery of epididymis-specific mRNAs from mouse epididymosomes into N2a and TM4 cells].

OBJECTIVE: To investigate whether mouse epididymis-specific mRNAs Adam7 and Crisp1 can be delivered into N2a and TM4 cells, and to provide an experimental basis for exploring the function of epididymal mRNAs. METHODS: Using RT-PCR, we detected the presence of epididymis-specific genes (Adam7, Crisp1, Defb22, Wfdc2, and Wfdc9) in the testis, epididymis, epididymosome and sperm of adult male BALB/c mice as well as in the human testis, seminal vesicles and sperm. We isolated epididymosomes of BALB/c mice by low-speed centrifugation, filtration and ultracentrifugation, fluorescently labeled them by PKH26, co-incubated them for 1 hour with the N2a and TM4 cells after 24 hours of starvation culture, and observed whether they were fused with the N2a and TM4 cells and ingested using the epididymosomes without PKH26 labeling, PKH26 dye without epididymosomes, and non- epididymosome or -PKH26 dye as controls. Then we detected the epididymis-specific genes in the N2a and TM4 cells after 1-hour co-incubation by RT-PCR. RESULTS: Adam7 and Crisp1 were present in the mouse epididymis, epididymosomes and sperm, and in the human seminal vesicles and sperm as well, but not in the testes of either the mice or men. PKH26 and Hoechst33258 fluorescence double-labeling showed that the mouse epididymosomes were fused with the N2a and TM4 cells and ingested; RT-PCR revealed the mRNAs of Adam7 and Crisp1 in the N2a and TM4 cells after 1-hour co-incubation; and Western blot exhibited the CRISP1 protein in the N2a and TM4 cells incubated with epididymosomes. CONCLUSION: Epididymosomes can deliver epididymis-specific mRNAs Adam7 and Crisp1 into N2a and TM4 cells, where Crisp1 may be translated into proteins, though their function and significance need to be further studied.

[Molecular markers for predicting the success of micro-TESE in non-obstructive azoospermia].

Non-obstract azoospermia (NOA) is a serious male infertility disease. At present, testicular sperm extraction (micro-TESE) is performed in combination with intracytoplasmic sperm injection (ICSI) technology, NOA patients can have their own consanguine offspring. However, due to the invasiveness and uncertainty of micro-TESE surgery, it is difficult for patients to accept it. Therefore, finding an accurate method to predict the possibility of micro-TESE successful sperm retrival would be beneficial to azoospermia patients. Many genes are transcribed and expressed during spermatogenesis, and molecular assays have irreplaceable sensitivity and specificity in predicting the success sperm retrivel of micro-TESE. This article reviews the methods to predict the success sperm retrivel of micro-TESE including mRNA, non-coding RNA (piRNA, microRNA, cirRNA, tFRNAs) and some protein so far, to provide certain reference value for clinical and subsequent research.

Diurnal rhythm of human semen quality: analysis of large-scale human sperm bank data and timing-controlled laboratory study.

STUDY QUESTION: Can we identify diurnal oscillations in human semen parameters as well as peak times of semen quality? SUMMARY ANSWER: Human semen parameters show substantial diurnal oscillation, with most parameters reaching a peak between 1100 and 1500 h. WHAT IS KNOWN ALREADY: A circadian clock appears to regulate different physiological functions in various organs, but it remains controversial whether diurnal rhythms occur in human semen parameters. STUDY DESIGN, SIZE, DURATION: The medical record of a provincial human sperm bank (HSB) with 33 430 semen samples collected between 0800 and 1700 h from 1 March 2010 to 8 July 2015 was used to analyze variation in semen parameters among time points. A laboratory study was conducted to collect semen samples (n = 36) from six volunteers at six time points with identical time intervals (2 days plus 4 h) between 6 June and 8 July in 2019, in order to investigate the diurnal oscillation of semen parameters in vivo, with a strictly controlled abstinence period. Therefore, the sperm bank study with a large sample size and the in vivo study with a strictly controlled abstinence period in a 24-h time window could be compared to describe the diurnal rhythms in human semen parameters. PARTICIPANTS/MATERIALS, SETTING, METHODS: Samples were obtained from potential HSB donors and from participants in the laboratory study who were volunteers, recruited by flyers distributed in the community. Total sperm count, sperm concentration, semen volume, progressive motility and total motility were assessed using computer-aided sperm analysis. In addition, sperm chromatin integrity parameters (DNA fragmentation index and high DNA stainability) were assessed by the sperm chromatin structure assay, and sperm viability was measured with flow cytometry in the laboratory study. MAIN RESULTS AND THE ROLE OF CHANCE: The 33 430 samples from the HSB showed a temporal variation in total sperm count, sperm concentration, semen volume, progressive motility and total motility (all P < 0.001) between 0800 and 1700 h. Consequently, the eligibility of semen samples for use in ART, based on bank standards, fluctuated with time point. Each hour earlier/later than 1100 h was associated with 1.14-fold risk of ineligibility. Similarly, the 36 samples taken during the 24-h time window showed diurnal oscillation. With the pre-collection abstinence period strictly controlled, most semen parameters reached the most favorable level between 1100 and 1500 h. LIMITATIONS, REASONS FOR CAUTION: Some of the possible confounding factors, such as energy intake, which might influence semen quality or diurnal rhythms, were not adjusted for in the analyses. In addition, the findings should be considered with caution because the study was conducted in a specific population, time and place, while the timing of oscillations could differ with changing conditions. WIDER IMPLICATIONS OF THE FINDINGS: The findings could help us to estimate semen quality more precisely and to obtain higher quality sperm for use in ART and in natural conception. STUDY FUNDING/COMPETING INTEREST(S): This study was supported by the National Natural Science Foundation of China (81871208) and National Key R&D Program of China (2017YFC1002001). There are no conflicts of interest to declare. TRIAL REGISTRATION NUMBER: N/A.

The developmental miR-17-92 cluster and the Sfmbt2 miRNA cluster cannot rescue the abnormal embryonic development generated using obstructive epididymal environment-producing sperm in C57BL/6 J mice.

BACKGROUND: Sperm, during epididymal transit, acquires microRNAs(miRNAs), which are crucial for embryonic development. However, whether sperm miRNAs influenced by an obstructive epididymal environment affect embryonic development remains unknown. METHOD: The sham operation and vasectomy were performed in C57BL/6 J mice to create the control group (CON) and the obstructive epididymal environment group(OEE) group, respectively. The morphology of the testis and epididymis was observed using hematoxylin and eosin staining (HE staining) to establish the OEE mice model. The sperm quality test, intracytoplasmic sperm injection (ICSI), and epididymosomes fusion were employed to observe the effect of the obstructive epididymal environment on sperm and resultant embryonic development. The alteration of the sperm small RNA (sRNA) profile was analyzed by sRNA sequencing. RT-qPCR and DNA methylation were applied to observe the effect of obstructive epididymis on the expression of sperm miRNAs. The miRNAs microinjection was used to explore the impacts of sperm miRNAs on embryonic development. RESULTS: We confirmed postoperative 8-week mice as the OEE mice model by examining the morphology of the testis and epididymis. In the OEE group, we observed that sperm quality degraded and the development potential of embryos was reduced, which can be saved by the normal epididymal environment. The sperm sRNA sequencing revealed that the expression of the developmental miR-17-92 cluster and the Sfmbt2 miRNA cluster was downregulated in the OEE group. The expression of these two miRNA clusters in epididymis was also downregulated and regulated by DNA methylation. However, the downregulation of either the miR-17-92 cluster or the Sfmbt2 miRNA cluster in normal zygotes did not impair embryonic development. CONCLUSION: The obstructive epididymal environment influences sperm quality and resultant embryonic development, as well as the abundance of the developmental miR-17-92 cluster and the Sfmbt2 miRNA cluster in sperm, but these miRNA clusters are not the cause of abnormal embryonic development. It implies that epididymis is important in early embryonic development and may play a potential role in sperm epigenome.

The dose-effect relationships of cigarette and alcohol consumption with depressive symptoms: a multiple-center, cross-sectional study in 5965 Chinese middle-aged and elderly men.

BACKGROUND: Although association of depressive symptoms with cigarette or alcohol is well documented, the dose-response relationship between them is rarely studied. This study aims to evaluate dose-response relationships of cigarette and alcohol consumption with depressive symptoms in Chinese middle-aged and elderly men, providing evidence to guide cigarette and alcohol control. METHODS: This multiple-center, cross-sectional study including 5965 Chinese men aged 40-79 years was conducted in 2013-2016 in China. Depressive symptoms were evaluated by Beck Depression Inventory-Short Form. History of cigarette smoking and alcohol drinking were collected with a structured questionnaire. Prevalence of depressive symptoms was compared depending on cigarette and alcohol consumption. Adjusted odds ratios (OR) and 95% confidence interval (CI) were estimated by binary logistic regression. Interpolation analysis was applied to test dose-effect relationships. RESULTS: A parabolic-shaped relationship was observed between cigarette consumption and depressive symptoms. Compared to never smokers, 59.0% (OR = 1.59, 95% CI 1.30-1.94) and 29.0% (OR = 1.29, 95% CI 1.08-1.54) higher odds of depressive symptoms were observed in men smoking < 10 cigarettes/day and 10-20 cigarettes/day, whereas, similar odds of depressive symptoms among men smoking > 20 cigarettes/day (P = 0.092). An inverted J-shaped relationship was observed between alcohol consumption and depressive symptoms. Compared to never drinkers, a tendency of higher prevalence of depressive symptoms (OR = 1.16, 95% CI 0.99-1.36) was observed in men drinking < 140 g/week, and similar prevalence was observed in those drinking 140-280 g/week (P = 0.920), whereas, 29.4% (OR = 0.71, 95% CI 0.57-0.88) lower odds in men drinking > 280 g/week. CONCLUSIONS: Associations of cigarette smoking and alcohol drinking with depressive symptoms differ with consumption in middle-aged and elderly men. Health-care providers should exercise great caution on depressive symptoms in conducting cigarette and alcohol control.

Competitive CatSper Activators of Progesterone from Rhynchosia volubilis.

The root Rhynchosia volubilis was widely used for contraception in folk medicine, although its molecular mechanism on antifertility has not yet been revealed. In human sperm, it was reported that the cation channel of sperm, an indispensable cation channel for the fertilization process, could be regulated by various steroid-like compounds in plants. Interestingly, these nonphysiological ligands would also disturb the activation of the cation channel of sperm induced by progesterone. Therefore, this study aimed to explore whether the compounds in R. volubilis affect the physiological regulation of the cation channel of sperm. The bioguided isolation of the whole herb of R. volubilis has resulted in the novel discovery of five new prenylated isoflavonoids, rhynchones A - E (1:  - 5: ), a new natural product, 5'-O-methylphaseolinisoflavan (6: ) (1H and 13C NMR data, Supporting Information), together with twelve known compounds (7:  - 18: ). Their structures were established by extensive spectroscopic analyses and drawing a comparison with literature data, while their absolute configurations were determined by electronic circular dichroism calculations. The experiments of intracellular Ca2+ signals and patch clamping recordings showed that rhynchone A (1: ) significantly reduced cation channel of sperm activation by competing with progesterone. In conclusion, our findings indicat that rhynchone A might act as a contraceptive compound by impairing the activation of the cation channel of sperm and thus prevent fertilization.

Multiple circRNAs regulated by QKI5 conjointly spongemiR-214-3p to antagonize bisphenol A-inducedspermatocyte toxicity.

Although circular RNAs (circRNAs) are found to play important roles in many pathophysiological processes, the canonical theory that they act as microRNA sponges is now more and more challenged, given that most circRNAs only have few binding sites in a particular microRNA. Our previous study revealed that some up-regulated circRNAs play protective roles in bisphenol A (BPA)-induced toxicity in GC-2 germ cells. Here by CCK-8 assay, apoptosis assay, qRT-PCR and western blot analysis, we further discover that circRNAs (represented by circDcbld2, circMapk1 and circTbcld20) can cooperatively sponge miR-214-3p and then up-regulate AKT1 in ameliorating BPA-induced reproductive toxicity. They share binding sites with miR-214-3p and collectively reinforce the sponging effects. In addition, the upstream regulation mechanism, proven by bioinformatics analysis and in vitro gain- and loss-of-function study, shows that down-regulation of RNA binding protein QKI5 after BPA exposure can increase the expressions of these protective circRNAs, and thus activate the cell protective process. The QKI5-circDcbld2/circMapk1/circTblcd20-miR-214-3p-AKT1 axis ameliorates the toxic effect of BPA on GC-2 cells. Many other circRNAs up-regulated upon BPA treatment and QKI5 down-regulation also show binding sites with miR-214-3p. Thus the above axis may also be extrapolated to other circRNAs. Our results enrich the context of circRNA sponge mode and may provide new ideas in future multiple nucleic acid therapy.

sncRNAs in Epididymosomes: The Contribution to Embryonic Development and Offspring Health.

Much progress has been made in determining that paternal environmental exposures can remodel their spermatozoa small noncoding RNAs (sncRANs) and, in turn, affect the phenotypes of their offspring. Studies have shown that changes in the spermatozoa sncRNAs profile occur during passing through the epididymis. Due to the absence of transcription and translation in the epididymis, spermatozoa remodel their sncRNAs profile through communication with the epididymal microenvironment. Since epididymosomes contribute to the process of spermatozoa maturation by mediating the crosstalk between the epididymis and the passing spermatozoa, they are considered to be the leading candidate to mediate these changes. Previous studies and reviews on the role of epididymal transfer proteins in sperm maturation and function are myriad. This review focuses on the role and mechanisms of epididymosome-mediated transfer of sncRNAs cargoes onembryonic development and offspring health.

Androgen-dependent miR-125a-5p targets LYPLA1 and regulates global protein palmitoylation level in late-onset hypogonadism males.

Late-onset hypogonadism (LOH) is defined as a clinical and biochemical syndrome with multiple symptoms caused by testosterone deficiency in aging males. An in-depth exploration of the molecular mechanism underlying LOH development is insufficient. We previously identified miR-125a-5p as a dysregulated microRNA in LOH patients and potential diagnostic biomarker for LOH. The present study demonstrated that plasma miR-125a-5p was upregulated after testosterone supplementation in both LOH patients and castrated mice, and positively associated with the testosterone concentrations, suggesting direct regulation of miR-125a-5p expression by testosterone. Androgen response element in the promoter of miR-125a-5p was subsequently identified. Target gene screening and confirmation verified that LYPLA1, encoding acyl-protein thioesterase 1 which catalyzed protein depalmitoylation process, was a target gene of miR-125a-5p. Furthermore, in cells cultured with testosterone deprivation and organs from castrated mice, testosterone deficiency led to decreased global protein palmitoylation level. In aging males, global protein palmitoylation in peripheral blood showed a notable decline in LOH patients contrast to the normal elderly males. And the palmitoylation level was positively correlative with serum testosterone concentrations. Our results suggested that testosterone could regulate global palmitoylation level through miR-125a-5p/LYPLA1 signaling pathway. Given that protein palmitoylation is pivotal for protein function and constitutes the pathogenesis of various diseases, testosterone/miR-125a-5p/LYPLA1 may contribute to the molecular mechanism underlying multiple symptoms caused by testosterone deficiency in LOH patients, and aberrant global palmitoylation could be a potential biomarker for LOH.

Effect of mitophagy in oocytes and granulosa cells on oocyte quality†.

Mitophagy is the process by which cells selectively remove supernumerary or damaged mitochondria through autophagy, and is crucial for mitochondrial homeostasis and cell survival. Mitochondria play vital roles in determining the developmental competence of oocytes. During the early stages of oogenesis, aberrant mitochondria can be removed by mitophagy. After oocyte formation, mitophagy is not actively initiated to clear damaged mitochondria despite the presence of mitophagy regulators in oocytes, which leads to the transmission of dysfunctional mitochondria from the oocyte to the embryo. However, granulosa cells around oocytes can improve mitochondrial function through mitophagy, thereby improving oocyte developmental capacity. Furthermore, this review discusses recent work on the substances and environmental conditions that affect mitophagy in oocytes and granulosa cells, thus providing new directions for improving oocyte quality during assisted reproductive technology treatment.

Transfer- or 'transmission'-RNA fragments? The roles of tsRNAs in the reproductive system.

Transfer-RNAs (tRNAs) help ribosomes decode mRNAs and synthesize proteins; however, tRNA fragments produced under certain conditions, known as tRNA-derived small RNAs (tsRNAs), have been found to play important roles in pathophysiological processes. In the reproductive system, tsRNAs are abundant in gametes and embryos and at the maternal-fetal interface, as well as in microvesicles like epididymosomes, seminal plasma exosomes, and syncytiotrophoblast-derived extracellular vesicles. tsRNAs can affect gamete cell maturation, zygote activation, and early embryonic development. tsRNAs can transmit epigenetic information to later generations. In particular, exposure to environmental factors such as nutrition, isoproterenol, and poly(I:C) may allow tsRNAs to transfer information to the gametes or placenta to alter offspring phenotype. The underlying mechanisms of tsRNAs action include transposon silencing, translation regulation, and target mRNA degradation. Herein, we review the currently reported tsRNAs in the reproductive system, their validated functions, and potential roles. A better understanding of this field may help to provide useful recommendations or develop strategies to increase fertility and conception of healthy babies.

Clinical application value of Inhibin B alone or in combination with other hormone indicators in subfertile men with different spermatogenesis status: A study of 324 Chinese men.

BACKGROUND: In this study, we investigated the clinical value of serum Inhibin B alone or in combination with other hormone indicators in subfertile men. METHODS: This is a multicenter study involving 324 men from different cities in China. Testicular volume, routine semen analysis, serum Inhibin B, anti-Müllerian hormone (AMH), follicle-stimulating hormone (FSH), luteinizing hormone (LH), testosterone, estradiol, and prolactin were measured. Testicular tissue samples were also analyzed in 78 of 129 patients with azoospermia to distinguish impaired spermatogenesis from obstructive azoospermia. RESULTS: The concentration of Inhibin B, FSH, and AMH is related to spermatogenesis. For men with impaired spermatogenesis, including mild-to-moderate oligozoospermia (IMO) and severe oligozoospermia (ISO), serum levels of Inhibin B and FSH are highly correlated with sperm counting. However, in patients with idiopathic moderate oligozoospermia or severe oligozoospermia, there was no significant correlation between Inhibin B (or FSH) and sperm concentration. The upper cutoff value of Inhibin B to diagnose ISO is 58.25 pg/ml with a predictive accuracy of 80.65%. To distinguish between nonobstructive azoospermia (NOA) and obstructive azoospermia (OA), the area under the curve (AUC) for AMH + Inhibin B + FSH is very similar to Inhibin B (0.943 vs. 0.941). The cutoff level of Inhibin B to diagnose nonobstructive azoospermia is 45.9 pg/ml with a positive and negative prediction accuracy of 97.70% and 85.71%, respectively. CONCLUSION: In summary, Inhibin B is a promising biomarker alone or in combination with other hormone indicators for the diagnosis of testicular spermatogenesis status, helping clinical doctors to distinguish NOA from OA.

[Functional polymorphism sites in non-coding regions of folate metabolism-related genes in the reproductive-aged population of Hubei Province].

Objective: To investigate the distribution of the functional polymorphisms in the non-coding regions of folate metabolism-related genes in the reproductive-aged population of Hubei Province. METHODS: Using Sanger sequencing, we examined the polymorphisms of the genes MTR (rs28372871 and rs1131450), MTRR (rs326119) and CBS (rs2850144) in 790 subjects before and during pregnancy from April 2020 to March 2021. We compared the distributions of the four loci between different populations. RESULTS: The distributions of the four genotypes of rs28372871, rs1131450, rs326119 and rs2850144 all conformed to the Hardy-Weinberg equilibrium (HWE). Statistically significant differences were observed in the polymorphism distribution of rs28372871 between Hubei and Jiangsu (P < 0.05), in that of rs1131450 between Hubei and Shanghai (P < 0.05), and in that of rs2850144 between Hubei and Yazd, Iran (P < 0.01). CONCLUSIONS: This was the first investigation on the distribution of MTR, MTRR and CBS gene polymorphisms in the reproductive-aged population of Hubei Province. The effects of the functional loci in both encoding and non-coding regions of folate metabolism-related genes have to be comprehensively considered so as to formulate an appropriate folate-supplementary protocol.

Frequent and mild scrotal heat stress in mice epigenetically alters glucose metabolism in the male offspring.

Several studies have reported that health problems occur in assisted reproductive technology (ART)-conceived offspring. Recently, investigations have demonstrated that paternal environmental conditions influence offspring health. However, it is unclear whether the factors that cause male infertility per se affect offspring health and contribute to health problems in ART-born children. Scrotal heat stress represents a common cause for oligoasthenozoospermia, and in these cases, in vitro fertilization-embryo transfer (IVF-ET) is typically recommended for those individuals trying to conceive. We exposed C57BL/6J male mice to frequent and mild scrotal heat stress (fmSHS) (39°C for 30 min once weekly for 5 consecutive wk). Sperm was subjected to IVF-ET with oocytes of untreated C57BL/6J females to produce offspring mice. Glucose intolerance and insulin resistance was observed in the male offspring mice derived from fmSHS-exposed fathers. Islets, after evaluation, remained unchanged. Genes involved in glucose metabolism, especially, those in insulin signaling pathways, showed dysregulation in the liver of the fmSHS-derived male offspring. Differentially methylated regions were found in the sperm of fmSHS-exposed mice by whole genome bisulfite sequencing. Interestingly, abnormal methylation of some genes with altered expression in offspring was observed in both the sperm of fmSHS fathers and the liver of their male offspring. Our results suggest that the factors that cause male infertility can affect male offspring health by an epigenetic mechanism.