Crocin Targets HSP90 to Suppress NLRP3/Caspase-1-Mediated Sertoli Cell Pyroptosis

Pyroptosis, a proinflammatory form of programmed cell death mediated by the NLRP3 inflammasome, has recently been implicated in Sertoli cell inflammatory injury. Crocin, a water-soluble carotenoid glycoside derived from Crocus sativus L. and Gardenia jasminoides Ellis, exhibits anti-inflammatory, antioxidant, and anti-apoptotic properties, yet its role in Sertoli cell pyroptosis remains unclear. Here, we investigated the pharmacological effects and mechanisms of crocin in regulating TM4 Sertoli cell viability and pyroptosis. Network pharmacology based on oligoasthenospermia (OA)-related candidate targets, together with molecular docking, identified HSP90AA1 as a candidate molecular target associated with crocin-mediated regulation of Sertoli cell pyroptosis. MST analysis further provided target-engagement evidence supporting a direct interaction between crocin and HSP90AA1. In TM4 Sertoli cells, crocin improved cell viability, enhanced Ki-67 expression, and protected against lipopolysaccharide/nigericin-induced pyroptosis, as evidenced by reduced lactate dehydrogenase release, decreased PI-positive cells, and downregulated expression of IL-1β, IL-18, NLRP3, Caspase-1, and GSDMD. Moreover, under HSP90AA1-overexpressing conditions, crocin partially reduced pyroptosis-related injury and inflammatory mediator expression, supporting the involvement of HSP90AA1 in crocin-mediated regulation of Sertoli cell pyroptosis. Collectively, these findings indicate that crocin enhances Sertoli cell proliferation and mitigates inflammasome-driven pyroptosis through an HSP90AA1-associated mechanism. These findings should be interpreted as preliminary in vitro evidence for crocin-mediated regulation of Sertoli cell proliferation and pyroptosis, and further validation in animal models and clinical samples is required.