Resveratrol inhibits NLRP3 inflammasome activation to alleviate bovine mastitis by promoting PINK1-mediated mitophagy

Objective: Negative energy balance in transition cows elevates the circulating concentrations of non-esterified fatty acids (NEFA), which can trigger mastitis and pose a severe threat to the dairy industry. Resveratrol is a polyphenolic compound with anti-inflammatory properties, yet its role in NEFA-induced inflammation in bovine mammary epithelial cells (BMECs) remains unclear. This study aimed to investigate the protective effects of resveratrol on mastitis and elucidate its underlying mechanisms. Methods: BMECs were pre-treated 100 µM RES for 24 h and then treated 0.9 mM NEFAs for 4 h, and a PINK1 inhibitor was used to assess the underlying mechanisms. Furthermore, a mouse model of mastitis was utilized to further evaluate the hepatoprotective effects of resveratrol against mastitis in vivo. Results: Resveratrol significantly attenuated the NEFA-induced inflammatory response, as evidenced by reduced levels of NLRP3 inflammasome components (NLRP3, caspase1, IL-1β) and pro-inflammatory cytokines (IL-6, IL-1β and TNF-α). Mechanistically, resveratrol promoted mitophagy by upregulating levels of LC3-II, PINK1, and Parkin, and downregulating P62 expression. Crucially, the anti-inflammatory effect of resveratrol was reversed upon inhibition of PINK1. The in vivo experiments confirmed that resveratrol alleviated mammary gland inflammation and enhanced PINK1-mediated mitophagy. Conclusion: This study demonstrates that resveratrol mitigates NLRP3-mediated inflammatory responses by activating PINK1-mediated mitophagy, suggesting its potential as a promising therapeutic candidate for mastitis in perinatal dairy cows experiencing negative energy balance.