Fenofibrate suppressesinfection via autophagy-mediated cholesterol regulation in bovine mammary epithelial cells and murine mammary tissue.

Abstract

BACKGROUND: mastitis is an important disease of dairy cows; however, there are no commercialvaccines and antimicrobial resistance is increasing. Furthermore,lacks a cell wall and relies on host-derived cholesterol for survival and growth. METHODS: We evaluated effects of fenofibrate, a peroxisome proliferator-activated receptor α (PPARα) agonist, oninfection, using both bovine mammary epithelial cells and a murine mastitis model.analyses assessed autophagy, nuclear Transcription Factor EB (TFEB) and Transcription Factor E3 (TFE3) translocation, cholesterol metabolism, and bacterial localization, whereasevaluations included inflammatory responses, lysosomal/autophagy protein expression, cholesterol content, and bacterial burden. RESULTS: Fenofibrate enhanced autophagic flux via upregulation of LC3B and LAMP2, promoted nuclear translocation of TFEB and TFE3, and reduced intracellular cholesterol by repressing 3-Hydroxy-3-Methylglutaryl-Coenzyme A Reductase (HMGCR) and Sterol Regulatory Element-Binding Transcription Factor 2 (SREBF2) while increasing Acetyl-Coenzyme A Acetyltransferase 1 (ACAT1) expression. Based on confocal imaging, fenofibrate disrupted co-localization betweenand free cholesterol while enhancing its uptake by autophagosomes. In a murine mastitis model, fenofibrate alleviated inflammatory cell infiltration and cytokine release, restored lysosomal and autophagy protein expression, reduced cholesterol content, and significantly lowered bacterial burden. CONCLUSION: Fenofibrate enhanced defense capability of mammary epithelial cells againstinfection through a dual mechanism-promoting autophagy and regulating cholesterol homeostasis-thereby reducing bacterial survival and protecting tissues from damage. This discovery provides a novel strategy for prevention and treatment ofinfection, warranting further investigation in bovine models to assess pharmacokinetics, dosage, and clinical efficacy.