Ferroptosis drives LPS-induced injury in IPEC-J2 cells and is targeted by Rutin through NFE2L2, KEAP1, TLR4 and ALOX5.

Lipopolysaccharide (LPS)-induced intestinal epithelial injury contributes to the loss of gut homeostasis. Whether ferroptosis is involved in this injury and whether rutin can attenuate ferroptosis-associated changes in porcine intestinal epithelial cells remain unclear. In this in vitro study, IPEC-J2 cells were exposed to LPS (10 μg/mL) with or without rutin pretreatment (200 μM). LPS induced a ferroptosis-associated phenotype, including cell shrinkage and death, increased malondialdehyde, reactive oxygen species (ROS), and ferrous iron (Fe2+), upregulated PTGS2 and DMT1, and decreased GPX4, SLC7A11, and FTH1 expression. Rutin pretreatment reduced TNF-α, IL-6, and IL-1β expression, limited lipid peroxidation, ROS accumulation, and LPS-induced Fe2+ overload, and partially restored GPX4 and FTH1 while reducing PTGS2. Integrated transcriptomic analysis, network pharmacology, and molecular docking identified NFE2L2, KEAP1, TLR4, and ALOX5 as candidate targets, and qRT-PCR showed that rutin reversed LPS-associated changes in these genes. These findings suggest that LPS-induced injury in IPEC-J2 cells is accompanied by ferroptosis-related changes and that rutin may attenuate this in vitro injury by coordinating antioxidant, inflammatory, iron-metabolism, and lipid-peroxidation-related gene responses. Further functional studies are needed to confirm direct target involvement and causal mechanisms.