Hyperoside Alleviates Free Fatty Acid-Induced Inflammatory Damage and Tight Junction Disruption in Mammary Epithelial Cells by Regulating Nuclear Factor Erythroid 2-Related Factor 1

Ketosis correlates with increased mastitis risk, but the underlying mechanism remains unclear. Nuclear factor erythroid 2-related factor 1 (NFE2L1) regulates inflammatory responses, and hyperoside (Hyp) exerts anti-inflammatory/antioxidant effects. However, whether NFE2L1 participates in the Hyp-mediated protection of mammary epithelial cells remains unclear. We investigated Hyp's mechanism against high-concentration free fatty acid (FFA)-induced damage via NFE2L1. Clinical ketotic cows showed reduced mammary NFE2L1, zona occludens 1 (ZO-1), occludin, claudin 1, and elevated levels of interleukin (IL)-1B and IL-6. FFAs downregulated NFE2L1, ZO-1, occludin, and activated inflammatory signaling, which Hyp (20 μM) reversed. NFE2L1 silencing abolished Hyp's protective effects. Overall, these data suggested that Hyp may serve as a promising therapeutic agent to counteract inflammation and maintain barrier integrity in the mammary tissue of clinical ketotic cows, with its efficacy dependent on NFE2L1.