Dietary hispidulin ameliorated chemically-induced colitis by inhibiting epithelial cell ferroptosis <i>via</i> the ACAT2-GPX4 axis and remodeling the gut microbiota

Hispidulin, a dietary flavonoid occurring naturally in brown algae, has attracted considerable interest owing to its functions in modulating inflammatory responses and neutralizing reactive oxygen species. This study examined the consequences of hispidulin in a chemically induced acute colitis model. RNA sequencing (RNA_seq), 16s rRNA_seq, and non-targeted metabolomics were conducted to determine the mechanisms underlying hispidulin-induced effects on colitis. The findings revealed that hispidulin exhibited significant therapeutic benefits against colitis. In addition, treatment with hispidulin led to a decrease in pro-inflammatory cytokine levels as well as serum FITC-dextran concentrations. RNA_seq analysis demonstrated a notable increase in colonic ACAT2 expression in mice with colitis following hispidulin treatment. Furthermore, hispidulin administration resulted in the upregulation of GPX4, a reduction in ROS levels, and inhibition of epithelial ferroptosis in mice with colitis. Both genetic inhibition of ACAT2 <i>in vitro</i> and AAV-mediated knockdown of ACAT2 <i>in vivo</i> substantially negated the effects of hispidulin on GPX4/GSH levels, ROS levels, and lipid hydroperoxide accumulation in both NCM460 cells and epithelial cells derived from DSS exposed murine models. 16s rRNA_seq results showed that hispidulin increased the abundance of <i>Lactobacillus</i>, <i>NK4A136_group</i>, <i>Oscillibacter</i>, <i>Peptococcus</i>, and <i>Adlercreutzia</i> and decreased the abundance of <i>Turicimonas</i> in colitic mice. Non-targeted metabolomics results showed that hispidulin facilitated the metabolism of galactose and promoted unsaturated fatty acid biosynthesis in colitic mice. In summary, hispidulin reduced the advancement of acute colitis in mice by reducing ACAT2-mediated ferroptosis in epithelial cells, as well as altering the composition of the gut microbiota.