Icariin alleviates the injury of Sertoli cell junction function by upregulating PKR pathway via ERα/c-fos α /c-fos signaling in aged mice

Ethnopharmacological relevenace: Sertoli cells are vital to maintain spermatogenesis and their function decline during aging. Epimedium has the effects of tonifying kidney-yang, strengthening bones and muscles, and expelling wind and dampness, and is commonly used in the treatment of kidney-yang deficiency, impotence and spermatorrhea. Icariin is the main active ingredients from Epimedium exhibiting delaying aging effects and improving male reproductive dysfunction. Whereas, it remains poorly understood how icariin alleviates age-associated decline in testicular function by protecting against the damage of junction function of Sertoli cells. Aim of the study: This study aimed to evaluate the improvement effect of icariin on Sertoli cell junction function damage and explore the underlying mechanisms. Materials and methods: Male C57BL/6 mice and mouse Sertoli cell line TM4 cells were utilized to assess the improvement effect of icariin on aging-associated Sertoli cell junction function injury. H&E &E staining, transmission electron microscopy, qPCR, Western blot, molecular docking, siRNA transfection, and immunofluorescence were performed in this study. Results: Dietary administration of icariin remarkly attenuated age-associated deterioration in spermatogenic function as evidenced by elevated testicular weight and index, sperm concentration and sperm viability. In addition, icariin protected Sertoli cell junction function from age-associated damage as proven by increased Sertoli cell numbers, improved tight junction ultrastructure, and upregulated junction-related proteins (ZO-1, Occludin and beta- Catenin). Moreover, icariin significantly upregulated ER alpha/c-fos signaling and PKR pathway in testicular Sertoli cells. Similarly, in vitro studies revealed that deletion of ER alpha, c-fos or PKR abolished the improvement effects of icariin on Sertoli cell junction function damage. Conclusions: Icariin effectively mitigates age-associated decline in testicular function by diminished Sertoli cell junction function damage through upregulating PKR pathway via ER alpha/c-fos signaling. Therefore, attenuating Sertoli cell junction function injury by the upregulation of PKR pathway via ER alpha/c-fos signaling probably indicates an effective target for the prevention and treatment of testicular spermatogenic function with aging.