Human Reproduction Open
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2025
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IF 11.1
Li Zhibin,Li Shuying,Xiao Yufeng,Guo Junfeng,Zhou Jianchun,Chen Yang,Yang Juan,Gong Chunli,He Bing,Wu Yuyun,Gao Nannan,Yang Huan,Gao Limin,Hu Hua,Zhang Yunfang,Yang Shiming
What are the molecular mechanisms underlying hypoxia-induced male reproductive impairment?Hypoxia compromises Septin2 (Sept2) transcription in spermatogonia, which impedes spermatogonial proliferation through protein phosphatase 2A (PP2A)-dependent AKT dephosphorylation.Hypoxia is associated with impaired spermatogenesis and poor sperm parameters in men. Spermatogonia proliferation, a crucial early step in spermatogenesis, is essential for maintaining the spermatogenic cell population and ensuring sperm quality. However, the connection between hypoxia and spermatogonial proliferation remains poorly understood, and treatment options for hypoxia-related reproductive disorders are limited.A cross-sectional study analyzed semen samples from 24 high-altitude (HA) residents, 6 pathological hypoxia (PH) patients, and 19 healthy controls to evaluate hypoxia-associated sperm parameter alterations. Complementary animal studies employing a hypobaric chamber-induced hypoxic mouse model (n = 5) confirmed reproductive impairments through assessment of birth rates, sperm quality, and testicular histopathology. Transcriptomic profiling of hypoxic versus normoxic mouse testes (n = 3/group) identified spermatogonial proliferation defects as a predominant pathological feature and pinpointed Sept2 as a candidate mediator. Subsequent mechanistic investigations employed in vitro hypoxic culture of spermatogonial cell lines under hypoxic conditions coupled with pharmacological modulation of PP2A activity in mice (n = 3-5 per intervention group) to delineate the underlying molecular pathways.Semen parameters were evaluated using computer-assisted sperm analysis (CASA; for sperm concentration, count, and motility), morphological staining (Pap staining for sperm deformity), and eosin-nigrosin staining (for sperm viability). In hypoxic mouse model, fertility outcomes were assessed through fertility assessment (mating experiments), sperm parameters (CASA), testicular histology (H&E staining), and spermatogonia proliferation (immunohistochemistry and qPCR). In hypoxic spermatogonial cell models, cell proliferation was detected using CCK-8, EdU incorporation, flow cytometry, and western blotting. Sept2 manipulation (knockdown/overexpression), followed by mechanistic analyses (dual-luciferase reporter assay, DNA pulldown/mass spectrometry, TMT-based quantitative proteomics, co-immunoprecipitation, etc), was performed to investigate the mechanism underlying hypoxia-regulated spermatogonia proliferation. The SEPT2 inhibitor forchlorfenuron (FCF), the PP2A agonists celastrol, erlotinib, and FTY720, as well as PP2A inhibitor okadaic acid (OA) were used to investigate the role of the SEPT2-PP2A-AKT axis in male fertility regulation.Both human populations (HA residents and PH patients) and mouse model consistently demonstrated hypoxia-related reproductive dysfunction. Mechanistic analyses revealed that hypoxia significantly downregulated Sept2 expression in spermatogonia, concomitant with impaired proliferative capacity. Sept2 knockdown in normoxic mice the hypoxia-induced defects in spermatogenesis. Complementary in vitro studies confirmed that Sept2 depletion impaired spermatogonial proliferation by inducing G1-S phase arrest, while its overexpression mitigated hypoxia-related proliferative defects. Further investigation revealed that hypoxia disrupts Sept2 transcription by interfering with the binding of RNA polymerase II subunit A (POLR2A) to the Sept2 promoter. The consequent reduction in Sept2 expression led to stabilization of the B56γ regulatory subunit of PP2A, resulting in enhanced AKT dephosphorylation and subsequent suppressed proliferation. Pharmacological intervention with the PP2A inhibitor OA restored reproductive competence and sperm quality in hypoxic mice, whereas PP2A agonists exacerbated these deficits.RNA-seq data are deposited in China National Center for Bioinformation (CNCB) under accession number PRJCA035733.This study focused on the effects of hypoxia on sperm parameters. Additional factors such as alterations in reproductive hormones and sexual function may contribute to hypoxia-induced infertility and warrant further research.This study identifies the SEPT2-PP2A/B56γ-AKT axis as a key regulator in hypoxia-related spermatogonia proliferation impairment. PP2A inhibitors such as OA may offer a therapeutic strategy to protect male fertility under hypoxic conditions.This work was supported by the National Natural Science Foundation of China (No. 82101688) and Natural Science Foundation of Chongqing (No. CSTB2022NSCQ-MSX0943). The authors have no conflicts of interest to declare.
研究领域:
生殖生物学
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分子生物学
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医学
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PubMed
高影响因子
Toxins
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2024
·Vol. 16
·期 3
·121
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IF 3.9
Hira Sayed,Qiongqiong Zhang,Yu Tang,Yanan Wang,Yongpeng Guo,Jianyun Zhang,Cheng Ji,Qiugang Ma,Lihong Zhao
Zearalenone (ZEN) is a non-steroidal estrogenic mycotoxin found in many agricultural products and can cause reproductive disorders, mainly affecting spermatogenesis in male animals. Rutin (RUT) is a natural flavonoid compound recognized for its significant antioxidant, anti-inflammatory and estrogenic properties. The present study aimed to determine the protective role of RUT against ZEN-induced reproductive toxicity in male mice. Twenty-four adult Kunming male mice were divided into four groups: control, RUT (500 mg/kg RUT), ZEN (10 mg/kg ZEN), ZEN + RUT (500 mg/kg RUT + 10 mg/kg ZEN), with six replicates per treatment. The results indicated that RUT mitigated ZEN-induced disruption in spermatogenic cell arrangement, decreased spermatozoa count, and increased sperm mortality in the testes. RUT significantly restored ZEN-induced reduction in T, FSH, LH, and E2 serum levels. Moreover, RUT mitigated ZEN-induced apoptosis by increasing the mRNA expression level ofbcl-2, decreasing the mRNA expression level ofkiss1-r, and decreasing the protein expression level of caspase 8 in reproductive tissues. These findings indicate the protective role of RUT against ZEN-induced reproductive toxicity in male mice by regulating gonadotropin and testosterone secretions to maintain normal spermatogenesis via the HPG axis, which may provide a new application direction for RUT as a therapeutic agent to mitigate ZEN-induced reproductive toxicity.
研究领域:
毒理学
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生殖生物学
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分子生物学
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DOI
PubMed
高影响因子
DEVELOPMENT
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2025
·Vol. 152
·期 6
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IF 3.6
Lin Zhen,Rong Bowen,Wu Meixia,Yan Junyi,Hong Tong,Hou Linjun,Tang Xinzhe,Liu Qiang,Peng Xiaozhong,Chen Yao,Lan Fei,Tong Ming-Han
ASH2L is a core component of KMT2 complexes, crucial for H3K4 trimethylation. However, its role in spermatogenesis remains elusive. Here, we demonstrate an essential role of Ash2l for meiotic prophase but dispensable for mitosis in differentiated spermatogonia. Using a germ cell-specific Ash2l knockout mouse model, we reveal that Ash2l deficiency leads to meiotic arrest and sterility in both sexes. Ash2l -deficient spermatocytes exhibit failures in chromosomal synapsis associated with persistent DMC1 foci and γH2AX, resulting in meiocyte loss due to apoptosis. Conversely, Ash2l -deficient differentiated spermatogonia show normal development. Mechanistically, Ash2l deficiency results in a global loss of H3K4me3 in promoter regions and significantly decreases expression of thousands of genes. Among these are genes involved in epigenetic silencing pathways, such as H3K9 di-methylation, DNA methylation and piRNA pathways, that are crucial for transposon repression during meiotic prophase I progression. Supporting this, we observe that Ash2l mutant spermatocytes display ectopic expression of LINE1-ORF1P. Our findings therefore reveal the previously unappreciated role of ASH2L-dependent H3K4me3 modification in spermatogenesis and provide clues to the molecular mechanisms in epigenetic disorders underlying male infertility.
研究领域:
分子生物学
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生殖生物学
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遗传学
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DOI
PubMed
高影响因子
ENVIRONMENTAL POLLUTION
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2020
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IF 6.793
Zheng Cao,Wanyue Huang,Yiran Sun,Yanfei Li
Deoxynivalenol (DON) is an unavoidable cereal crops contaminants and environmental pollutants, which seriously threated the health of human and animal. DON has been reported to exert significant toxicity effects on spermatogenesis , but the underlying mechanisms remain largely inconclusive. The blood-testis barrier (BTB) provides a specialized biochemical microenvironment for maintaining spermatogenesis. Thus, we hypothesized that DON could impair BTB and lead to spermatogenesis disorder. To confirm this hypothesis, sixty male mice were intragastrically administered with 0, 1.2, 2.4 and 4.8 mg/kg body weight DON for 28 days, and several important observations were obtained in present study. First, we found that DON induced spermatogenesis disorder, reflected by the declines of sperm concentration and quality, sperm ultrastructural damage as well as seminiferous tubular damage. Then, we proved that DON induced BTB disruption as well as decreased the expressions of BTB junction proteins, including Occludin, Connexin 43 and N-cadherin. Finally, the present study showed that DON induced inflammation and inhibited T biosynthesis in testis of mice. These results revealed that DON induced spermatogenesis disorder by BTB disruption associated with testosterone deficiency and inflammation in mice, which shed a new light on the potential mechanisms of reproductive toxicity induced by DON.
研究领域:
毒理学
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生殖生物学
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环境科学
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DOI
PubMed
高影响因子
SAUDI PHARMACEUTICAL JOURNAL
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2016
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IF 2.233
Jinpu Peng,Lianju Shen,Jinjun Chen,Xining Cao,Yue Zhou,Huali Weng,Chunlan Long,Deying Zhang,Shengfen Tu,Yan Zhang,Dawei He,Tao Lin,Guanghui Wei
This study focuses on investigation of cryptorchidism induced by flutamide (Flu) and its histopathological damage, and detects retinoic acid concentration in testicle tissue, in order to find a new method for clinical treatment to infertility caused by cryptorchidism. Twenty SD (Sprague Dawley) pregnant rats were randomly divided into Flu cryptorchidism group ( n = 10) and normal control group ( n = 10). HE stained for observing morphological difference. Transmission electron microscope (TEM) was used for observing the tight junction structure between Sertoli cells. Epididymal caudal sperms were counted and observed in morphology. The expression of stimulated by retinoic acid gene 8 (Stra8) was detected using immunohistochemistry, western blot, and Q-PCR. High performance liquid chromatography (HPLC) analysis was made on retinoic acid content. Sperm count and morphology observation confirmed cryptorchidism group was lower than normal group in sperm quantity and quality. The observation by TEM showed a loose structure of tight junctions between Sertoli cells. Immunohistochemistry, western blot, and Q-PCR showed that cryptorchidism group was significantly lower than normal group in the expression of Stra8. HPLC showed that retinoic acid content was significantly lower in cryptorchid testis than in normal testis. In the cryptorchidism model, retinoic acid content in testicular tissue has a significant reduction; testicles have significant pathological changes; damage exists in the structure of tight junctions between Sertoli cells; Stra8 expression has a significant reduction, perhaps mainly contributing to spermatogenesis disorder.
研究领域:
生殖生物学
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内分泌学
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组织病理学
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产品关联
DOI
PubMed
高影响因子
ECOTOXICOLOGY AND ENVIRONMENTAL SAFETY
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2024
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IF 6.2
Ying Wang,An-Liang Guo,Yang Xu,Xiaoyan Xu,Lin Yang,Yang Yang,Lan Chao
Background 2-ethylhexyldiphenyl phosphate (EHDPP) was used widespread in recent years and it was reported to impair reproductive behaviors and decrease fertility in male Japanese medaka. However, whether EHDPP causes spermatogenesis disturbance remains uncertain. Objectives We aimed to study the male reproductive toxicity of EHDPP and its related mechanism. Methods Human spermatocyte cell line GC-2 was treated with 10 µM, 50 µM or 100 µM EHDPP for 24 h. Male CD-1 mice aged 6 weeks were given 1, 10, or 100 mg/kg/d EHDPP daily for 42 days and then euthanized to detect sperm count and motility. Proliferation, apoptosis, oxidative stress was detected in mice and cell lines. Metabolome and transcriptome were used to detect the related mechanism. Finally, anti-oxidative reagent N-Acetylcysteine was used to detect whether it could reverse the side-effect of EHDPP both in vivo and in vitro . Results Our results showed that EHDPP inhibited proliferation and induced apoptosis in mice testes and spermatocyte cell line GC-2. Metabolome and transcriptome showed that nucleotide metabolism disturbance and DNA damage was potentially involved in EHDPP-induced reproductive toxicity. Finally, we found that excessive ROS production caused DNA damage and mitochondrial dysfunction; NAC supplement reversed the side effects of EHDPP such as DNA damage, proliferation inhibition, apoptosis and decline in sperm motility. Conclusion ROS-evoked DNA damage and nucleotide metabolism disturbance mediates EHDPP-induced germ cell proliferation inhibition and apoptosis, which finally induced decline of sperm motility.
研究领域:
毒理学
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生殖生物学
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分子生物学
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PubMed
高影响因子
Nature Communications
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2025
·Vol. 16
·期 1
·1-17
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IF 15.7
Liu Qiannan,Lu Xiaoyan,Deng Yao,Zhang Han,Wei Rumeng,Li Hongrui,Feng Ying,Wei Juan,Ma Fang,Zhang Yan,Zou Xia
Protein O-glycosylation plays critical roles in sperm formation and maturation. However, detailed knowledge on the mechanisms involved is limited due to lacking characterization of O-glycoproteome of testicular germ cells. Here, we performed a systematic analysis of site-specific O-glycosylation in mouse testis, and established an O-glycoproteome map with 349 O-glycoproteins and 799 unambiguous O-glycosites. Moreover, we comprehensively investigated the distribution properties of O-glycosylation in testis and identified a region near the N-terminal of peptidase S1 domain that is susceptible to O-glycosylation. Interestingly, we found dynamic changes with an increase Tn and a decrease T structure from early to mature developmental stages. Notably, the importance of O-glycosylation was supported by its effects on the stability, cleavage, and interaction of acrosomal proteins. Collectively, these data illustrate the global properties of O-glycosylation in testis, providing insights and resources for future functional studies targeting O-glycosylation dysregulation in male infertility.
研究领域:
生殖生物学
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蛋白质组学
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分子生物学
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DOI
PubMed
高影响因子
Oncotarget
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2017
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IF 5.168
Hao Gu,Wei Wu,Beilei Yuan,Qiuqin Tang,Dan Guo,Yiqiu Chen,Yankai Xia,Lingqing Hu,Daozhen Chen,Jiahao Sha,Xinru Wang
Genistein (GEN) is one of the isoflavones that has effect on male reproduction. However, the underlying mechanism remains unknown. miRNAs are a type of small non-coding RNAs that play important roles in spermatogenesis. We measured the GEN levels and miR-17-92 cluster expression in infertile subjects and found that miR-17-92 might be involved in GEN induced abnormal spermatogenesis. To clarify, we fed adult ICR mice with different doses of GEN (0, 0.5, 5, 50 and 250 mg/kg/day) for 35 days to study the underlying mechanism. We found that sperm average path velocity, straight-line velocity and eurvilinear velocity of the mice orally with GEN at 5mg/kg/day were significantly decreased, the expression levels of miR-17 and miR-20a in mice testis were higher in corresponding group. We also found miR-20a was the only miRNA that differentially expressed both in human and mice. By applying bioinformatics methods, Limk1 was predicted to be the target gene of miR-20a that is involved in spermatogenesis. Limk1 were significantly decreased in the corresponding group. Dual-luciferase report assay also proved that miR-20a could directly target Limk1. These results implied that Limk1 might be the target gene of miR-20a that is involved in GEN induced abnormal spermatogenesis.
研究领域:
生殖生物学
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分子生物学
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基因组学
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PubMed
高影响因子
ZOOLOGICAL RESEARCH
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2023
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IF 4.9
Cong Li,Rui Bi,Lin Wang,Yu-Hua Ma,Yong-Gang Yao,Ping Zheng
Tree shrews ( Tupaia belangeri chinensis ) share a close relationship to primates and have been widely used in biomedical research. We previously established a spermatogonial stem cell (SSC)-based gene editing platform to generate transgenic tree shrews. However, the influences of long-term expansion on tree shrew SSC spermatogenesis potential remain unclear. Here, we examined the in vivo spermatogenesis potential of tree shrew SSCs cultured across different passages. We found that SSCs lost spermatogenesis ability after long-term expansion (>50 passages), as indicated by the failure to colonize the seminiferous epithelium and generate donor spermatogonia (SPG)-derived spermatocytes or spermatids marking spermatogenesis. RNA sequencing (RNA-seq) analysis of undifferentiated SPGs across different passages revealed significant gene expression changes after sub-culturing primary SPG lines for more than 40 passages on feeder layers. Specifically, DNA damage response and repair genes (e.g., MRE11 , SMC3 , BLM , and GEN1 ) were down-regulated, whereas genes associated with mitochondrial function (e.g., NDUFA9 , NDUFA8 , NDUFA13 , and NDUFB8 ) were up-regulated after expansion. The DNA damage accumulation and mitochondrial dysfunction were experimentally validated in high-passage cells. Supplementation with nicotinamide adenine dinucleotide (NAD + ) precursor nicotinamide riboside (NR) exhibited beneficial effects by reducing DNA damage accumulation and mitochondrial dysfunction in SPG elicited by long-term culture. Our research presents a comprehensive analysis of the genetic and physiological attributes critical for the sustained expansion of undifferentiated SSCs in tree shrews and proposes an effective strategy for extended in vitro maintenance.
研究领域:
生物医学研究
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干细胞生物学
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分子遗传学
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生殖生物学
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DOI
PubMed
高影响因子
FOOD AND CHEMICAL TOXICOLOGY
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2021
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IF 6.025
Wanyue Huang,Menglin Liu,Bonan Xiao,Jian Zhang,Miao Song,Yanfei Li,Zheng Cao
Aflatoxin B 1 (AFB 1 ) is an unavoidable food and environmental contaminant, which can lead to disorders in spermatogenesis and its mechanism remains unclear. The blood-testis barrier (BTB) is responsible for ensuring normal spermatogenesis in testes. Therefore, we hypothesized that disruption of the BTB was involved in AFB 1 -induced spermatogenesis disorders. To confirm our hypothesis, male Kunming mice were orally gavaged AFB 1 (0, 0.375, 0.75, or 1.5 mg/kg) for 30 days. Primarily, we first proved that AFB 1 disrupted the BTB integrity. Then, AFB 1 decreased BTB-related junction protein expression and elevated Sertoli cell apoptosis , which were associated with oxidative stress . Additionally, AFB 1 upregulated the p-p38 MAPK/p38 MAPK ratio. These results collectively indicated that AFB 1 disrupted the BTB via reducing the expression of BTB-related junction protein and promoting apoptosis in mice testes, which were associated with the oxidative stress-mediated p38 MAPK signaling pathway.
研究领域:
毒理学
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生殖生物学
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分子生物学
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DOI
PubMed
高影响因子
MOLECULAR THERAPY
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2025
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IF 12
Mengya Zhang,Xueguang Zhang,Chen Tan,Ting Jiang,Zheng Cao,Yufei Chen,Feng Qiao,Gelin Huang,Chaoye Ma,Huixia Li,Mei Ye,Lei Wang,Liandong Zuo,Jun Yu,Xiaofeng Li,Yunfang Zhang,Yue-Qiu Tan,Shuibin Lin,Wenming Xu,Hao Chen
Recent studies have highlighted RNA modifications as integral regulators of gene expression during spermatogenesis. Ribosomal RNAs (rRNAs) are the most abundant RNA in cells, while the function and clinical relevance of rRNA modifications in spermatogenesis remain poorly understood. Here, we identified 4 pathogenic heterozygous variants of METTL5 in 1,427 patients with male infertility characterized as oligoasthenoteratozoospermia (OAT). The pathogenic variants of METTL5 led to the significantly decreased expression level of METTL5. Null mutation of 18S rRNA methyltransferase Mettl5 led to male infertility attributed to OAT during spermiogenesis, presenting defects in both the sperm head and tail. Notwithstanding the absence of notable changes in global translation after Mettl5 loss, we observed a compromised translational efficiency of mRNAs encoding proteins crucial for spermiogenesis, including Gk2 , Akap4 , Fsip2 , Odf2 , and Pgk2 . Intriguingly, therapeutic interventions via intracytoplasmic sperm injection in OAT couples with these variants resulted in successful pregnancies. These insights not only identify METTL5-mediated 18S rRNA m 6 A modification as a novel genetic determinant for OAT but also offer a new target to the genetic counseling, clinical diagnosis, and potential treatment strategies for male infertility.
研究领域:
分子生物学
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遗传学
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生殖医学
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RNA修饰
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发育生物学
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产品关联
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PubMed
高影响因子
CELL AND TISSUE RESEARCH
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2022
·Vol. 390
·期 2
·293-313
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IF 4.051
Li Zhen-Fang,Hao Shuang-Li,Wang Lan-Min,Qi Hong-Yu,Wang Jia-Ming,Tan Fu-Qing,Yang Wan-Xi
Spermatogenesis is a finely regulated process of germ cell proliferation and differentiation that leads to the production of sperm in seminiferous tubules. Although the mammalian target of rapamycin (mTOR) signaling pathway is crucial for spermatogenesis in mammals, its functions and molecular mechanisms in spermatogenesis remain largely unknown in nonmammalian species, particularly in Crustacea. In this study, we first identified es-Raptor (the core component of mTOR complex 1) and es-Rictor (the core component of mTOR complex 2) from the testis of Eriocheir sinensis . Dynamic localization of es-Raptor and es-Rictor implied that these proteins were indispensable for the spermatogenesis of E. sinensis. Furthermore, es-Raptor and es-Rictor knockdown results showed that the mature sperm failed to be released, causing almost empty lumens in the testis. We investigated the reasons for these effects and found that the actin-based cytoskeleton was disrupted in the knockdown groups. In addition, the integrity of the testis barrier (similar to the blood-testis barrier in mammals) was impaired and affected the expression of cell junction proteins. Further study revealed that es-Raptor and es-Rictor may regulate spermatogenesis via both mTORC1- and mTORC2-dependent mechanisms that involve es-rpS6 and es-Akt/es-PKC, respectively. Moreover, to explore the testis barrier in E. sinensis , we established a cadmium chloride (CdCl 2 )–induced testis barrier damage model as a positive control. Morphological and immunofluorescence results were similar to those of the es-Raptor and es-Rictor knockdown groups. Altogether, es-Raptor and es-Rictor were important for spermatogenesis through maintenance of the actin filament network and cell junctions in E. sinensis.
研究领域:
分子生物学
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生殖生物学
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甲壳动物学
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DOI
PubMed
高影响因子
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