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Yangjing Capsule (YC), an innovative Chinese medicine based on traditional prescription, promotes testosterone synthesis by up-regulating the expression of steroidogenic enzymes.Nur77 as a nuclear receptor is known to regulate the expression of many steroid synthetases.This study aimed to explore the potential mechanisms by which YC regulates testosterone synthesis in Leydig cells.MLTC-1 cells were treated with YC, and real-time PCR and Western blot were applied to assess the expressions of steroidogenic enzymes and Nur77.The luciferase reporter gene assay was performed to detect the activity of Nur77 gene promoter.In addition,the expressions of steroid synthases were detected after Nur77 gene was knockdown.YC significantly stimulated Nur77 production and upregulated StAR and HSD3B expression.In agreement, the activity of Nur77 gene promoter was significantly enhanced by YC.Interestingly, Nur77 knockdown blocked the above YC's effects,and consequently inhibited testosterone synthesis in MLTC-1 cells.YC promotes StAR and HSD3B expression and up-regulates testosterone synthesis in Leydig cells,which is mediated by Nur77 pathway.