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Additionally, histone modifications, especially H3K4me3, have been reported to be important in steroidogenesis. However, the regulation of stem Leydig cells differentiation by FGF9 and the role of histone modifications in this process remain unclear. Adult male Sprague\u2013Dawley rats were treated with ethane dimethane sulfonate to deplete Leydig cells (LCs), followed by intratesticular injection of FGF9 from post\u2013ethanedimethane sulfonate (EDS) day 14\u201328 at 0, 10, and 100\u2009ng\/testis. Quantitative real-time PCR and western blot analyses were used to detect the gene expression and protein levels of testosterone (T) synthesis. Chromatin immunoprecipitation sequencing (ChIP-seq) was employed to identify the H3K4me3-binding regions and binding levels. The results showed that FGF9 treatment led to an increase in serum T levels, LC number and the expression of LC-specific genes (Lhr, Scarb1, Stard1, Cyp11a1, Cyp17a1, Hsd17b3, and Hsd11b1). FGF9 markedly increased H3K4me3 protein levels and decreased H3K9me3 protein level. FGF9 promoted H3K4me3 on the promoter regions of Scarb1, Stard1, and Srd5a1 in vivo. In vitro studies demonstrated that FGF9 increased medium T levels and stimulated the incorporation of EdU, a marker of cell proliferation, into stem LCs after tubule culture. Moreover, Fgfr1 siRNA and WDR5-0103, the histone methyltransferases, could reverse the effect of FGF9 on promoting T production. This study supports previous ones demonstrating that FGF9 stimulates the proliferation and differentiation of stem LCs in an EDS-treated model and in vitro tubule culture model through H3K4me3 histone modifications.<\/jats:p>","DOI":"10.1093\/reprod\/xaag076","type":"journal-article","created":{"date-parts":[[2026,6,19]],"date-time":"2026-06-19T11:50:00Z","timestamp":1781869800000},"source":"Crossref","is-referenced-by-count":0,"title":["Fibroblast Growth Factor 9 promotes rat Leydig cell development via H3K4me3 histone modifications"],"prefix":"10.1093","volume":"172","author":[{"given":"Hehua","family":"Quan","sequence":"first","affiliation":[{"name":"Department of Anesthesiology and Perioperative Medicine, the Second Affiliated Hospital and Yuying Children's Hospital of Wenzhou Medical University; Key Laboratory of Pediatric Anesthesiology (Wenzhou Medical University), Ministry of Education; Key Laboratory of Precision Anesthesiology of Zhejiang Province, Wenzhou Medical University, 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Zhejiang Province, Wenzhou Medical University, Wenzhou, Zhejiang Province,","place":["China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yubin","family":"Xu","sequence":"additional","affiliation":[{"name":"Department of Anesthesiology and Perioperative Medicine, the Second Affiliated Hospital and Yuying Children's Hospital of Wenzhou Medical University; Key Laboratory of Pediatric Anesthesiology (Wenzhou Medical University), Ministry of Education; Key Laboratory of Precision Anesthesiology of Zhejiang Province, Wenzhou Medical University, Wenzhou, Zhejiang Province,","place":["China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Huiqian","family":"Zhang","sequence":"additional","affiliation":[{"name":"Department of Anesthesiology and Perioperative Medicine, the Second Affiliated Hospital and Yuying Children's Hospital of Wenzhou Medical University; Key Laboratory of Pediatric Anesthesiology (Wenzhou Medical University), Ministry of Education; Key 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