MiR-195-5p is involved in testicular ischemia/reperfusion injury by directly targeting PELP1 and regulating spermatogonia pyroptosis

We established two models, namely a testicular torsion/ detorsion (T/D) mouse model and an oxygen-glucose deprivation/reperfusion (OGD/R)-treated germ cell model. Hematoxylin and eosin staining was performed to evaluate the testicular ischemic injury. The expression of pyroptosis-related proteins and reactive oxygen species production in testis tissues were detected using Western blotting, quantitative real-time PCR, malondialdehyde and superoxide dismutase assay kits and immunohistochemistry. Cell viability and cytotoxicity were evaluated using CCK-8 and LDH assays, whereas expression patterns of inflammatory proteins were measured using ELISA, immunofluorescence, and western blot assays. miR-195-5p interaction with PELP1 was validated by conducting the luciferase enzyme reporter test.

Ischemia/reperfusion injury (IRI), which is characterized by testicular torsion and causes permanent impairment of spermatogenic function, is linked with pyroptosis. Studies have implicated endogenous small non-coding RNAs in IRI development across various organs. In this study, we elucidated the mechanism underlying miR-195-5p's action in regulating pyroptosis in testicular IRI.

Pyroptosis-related proteins NLRP3, GSDMD, IL-1β, and IL-18 were significantly upregulated following testicular IRI. A similar pattern was observed in the OGD/R model. miR-195-5p was significantly downregulated in mouse IRI testis tissue and OGD/R-treated GC-1 cells. Notably, miR-195-5p downregulation promoted whereas its upregulation attenuated pyroptosis in OGD/R-treated GC-1 cells. Furthermore, we found that PELP1 is a miR-195-5p target. miR-195-5p attenuated pyroptosis in GC-1 cells by inhibiting PELP1 expression during OGD/R, and this protective effect was blocked upon miR-195-5p downregulation. Collectively, these results indicated that miR-195-5p inhibits testicular IRI-induced pyroptosis by targeting PELP1, suggesting that it has the potential to serve as a novel target for the future development of therapies for testicular torsion.