ABSTRACT Triptolide is a diterpene triepoxide isolated from the traditional Chinese medicinal vine Trypterygium wilfordii hook f. (T. wilfordii). It possesses multiple biological activities, such as antitumor, immunosuppression, and antifertility. Previous studies suggested that triptolide might be a potential candidate for post-testicular male contraceptive agent. Nevertheless, the mechanisms of triptolide-induced reproductive toxicity remain unclear. In the present study, the results of reverse-transcription PCR and Western blotting revealed that triptolide inhibited the expression of Cx43 compared with the different effect of estradiol in cultured SCs from male rats. Further computational study revealed that triptolide can bind to the active site of human estrogenic 17beta-hydroxysteroid dehydrogenase and human estrogen receptorbeta. Therefore, our results indicated that male reproductive toxicity induced by triptolide was associated with the effects on intratesticular estrogen levels and estrogen receptors rather than its cytotoxicity.