Hydroxysafflor yellow a attenuates oxygen-glucose deprivation/ reoxygenation induced endothelial pyroptosis via PARP-1/NLRP3 pathway

Introduction Dysfunction of brain microvascular endothelial cells (BMECs) induced by oxidative stress represents a critical event in the pathogenesis of ischemia/reperfusion (I/R) injury. Although previous investigations have demonstrated the protective effects of Hydroxysafflor yellow A (HSYA) against I/R injury, the precise underlying mechanisms remain incompletely understood. Methods An oxygen-glucose deprivation/reoxygenation (OGD/R) model was established in rat BMECs (rBMECs). We analyzed cell viability, proliferation, oxidative stress markers (SOD, JC-1), and pyroptosis-related protein expression (NLRP3, Caspase-1, GSDMD, IL-1β). The NLRP3 inhibitor MCC950 was utilized to elucidate HSYA’s regulatory role in OGD/R-induced pyroptosis. The network pharmacology approach was used to identify potential targets of HSYA against ischemia/reperfusion (I/R) injury. Molecular docking, molecular dynamics (MD) simulation, CETSA, DARTS assays, along with PARP-1 overexpression/inhibition experiments were performed to elucidate the underlying mechanism of HSYA in ameliorating I/R injury. Results The results indicated that HSYA enhanced cell viability and proliferation of rBMECs exposed to OGD/R injury, accompanied by increased SOD activity and preserved MMP. HSYA suppressed the expression of pyroptosis-related proteins (NLRP3, Caspase-1, GSDMD, and IL-1β). The protective effects of HSYA were comparable to those observed with the NLRP3 inhibitor MCC950. Cotreatment afforded superior protection and more pronounced inhibition of NLRP3-mediated pyroptosis in OGD/R-induced rBMECs. Network pharmacology identified PARP-1 as a key target of HSYA against I/R injury. This interaction was validated through molecular docking and MD simulation, which revealed stable binding with high-affinity. Further experimental validation using CETSA and DARTS assays confirmed the direct binding of HSYA to PARP-1. Modulation of PARP-1 activity resulted in altered NLRP3 expression; Notably, both the PARP-1 inhibitor Olaparib and HSYA suppressed NLRP3, suggesting that the protective effects of HSYA may be attributed to direct targeting of PARP-1/NLRP3 pathway. Discussion This study demonstrates that HSYA protects rBMECs against OGD/R injury by directly targeting PARP-1, thereby inhibiting the NLRP3-mediated pyroptosis pathway. These findings reveal a novel mechanism of HSYA in mitigating I/R injury and implicate PARP-1 as a promising therapeutic target. Nevertheless, several limitations should be considered. The precise molecular details between PARP-1 and the NLRP3 inflammasome pathway require further elucidation, and our findings remain to be validated using animal models of cerebral I/R injury.