CPT1A Ameliorates Staphylococcus aureus-Induced Impairment of the Blood-Milk Barrier and Mitochondrial Oxidative Stress in Mastitis by Promoting NLRP3 Succinylation.

Abstract

Mastitis caused by Staphylococcus aureus is a common inflammatory disease in mammals. However, the role of carnitine palmitoyltransferase 1A (CPT1A) in this process remains poorly understood. In this study, we examined the expression and function of CPT1A using both in vitro and in vivo models. HC11 cells were transfected with a CPT1A-overexpressing plasmid and then infected with S. aureus, with or without the NOD-like receptor family pyrin domain containing 3 (NLRP3) agonist BMS-986299. In vivo, we established a mouse mastitis model by injecting S. aureus into the mammary duct and achieved local CPT1A overexpression via AAV9 delivery. The results showed that S. aureus infection significantly downregulated CPT1A expression in mammary tissue and HC11 cells. CPT1A overexpression reversed the infection-induced decrease in cell viability, restored mitochondrial membrane potential, increased antioxidant enzyme activities, reduced malondialdehyde levels, and prevented the downregulation of Occludin and Claudin-3. Mechanistically, CPT1A overexpression was associated with enhanced NLRP3 succinylation and accelerated degradation, thereby inhibiting NLRP3 expression. Administration of the NLRP3 agonist BMS-986299 partially reversed the protective effects of CPT1A overexpression both in vitro and in vivo. Collectively, these findings demonstrate that CPT1A counteracts S. aureus-induced mastitis by promoting NLRP3 succinylation, which accelerates NLRP3 degradation, thereby alleviating mitochondrial oxidative stress and preserving blood-milk barrier integrity. Targeting the CPT1A-NLRP3 axis may represent a potential therapeutic strategy for S. aureus-associated mastitis.