Influence of rutin on lipopolysaccharide-induced inflammation, oxidative stress, and autophagy in bovine mammary epithelial cells.

During clinical and subclinical Escherichia coli mastitis, bovine mammary epithelial cells (BMEC) are stimulated by LPS, leading to cellular inflammatory response, oxidative stress, and autophagy. This study hypothesized that rutin might mitigate these damages in BMEC via modulation of specific signaling pathways. To simulate mammary gland inflammation, different concentrations of LPS were used to stimulate BMEC. Different concentrations of rutin were used to pretreat the BMEC. The results demonstrated that LPS stimulated the NLR family pyrin domain containing 3 (NLRP3) inflammasome and its downstream effector molecules to produce an inflammatory response. Lipopolysaccharide caused a decrease in antioxidant enzymes and the total antioxidant capacity, an increase in malondialdehyde (MDA) levels, and a decrease in silent mating type information regulation 2 homolog-1 (SIRT1) levels. Treatment with LPS altered the morphological structure of the cells, converted the autophagy marker microtubule-associated protein light chain 3 (LC3)-I into LC3-II, and the increased the level of Beclin-1. Rutin increased silent mating type information regulation 2 homolog-1 (SIRT1) activity, reduced reactive oxygen species (ROS) generation, and decreased specific oxidative stress markers MDA. Additionally, rutin enhanced antioxidant enzyme activities including total antioxidant capacity, superoxide dismutase, catalase, and glutathione peroxidase. Rutin also inhibited NLRP3 inflammasome activation, which is associated with ROS production and oxidative stress. The mechanism of the protective effect of rutin on BMEC was investigated using the SIRT1 specific inhibitor EX-527. EX-527 considerably weakened the regulating function of rutin and aggravated the LPS-induced damage to BMEC. Rutin inhibited NLRP3 inflammasome activation and the downstream effector molecule gasdermin D (GSDMD) through SIRT1, thus alleviating excessive autophagy and reducing the inflammatory damage of cells. This study confirms that rutin reduces LPS-induced inflammation, autophagy, and oxidative stress in BMEC through the SIRT1/NLRP3 pathway. These findings provide new insights into addressing mastitis in dairy cows and offer promising clinical applications for future therapeutic interventions.