Protocatechuic Acid Alleviates Intestinal Inflammation by Enriching Gut-Derived Lithocholic Acid to Suppress the TAK1/NF-ĸB/MAPK Pathway

Intestinal inflammation impairs the growth of weaned piglets. This study demonstrated that the natural anti-inflammatory compound protocatechuic acid (PCA) effectively mitigates intestinal inflammation in piglets. PCA significantly suppressed interleukin (IL) -6 and Tumor Necrosis Factor-alpha (TNF-α) (P<0.05) and enhanced intestinal barrier integrity by upregulating tight junction proteins Occludin and ZO-1 (P<0.01). Although not a direct regulator, PCA significantly inhibited the phosphorylation of TAK1 and its downstream kinases (P<0.05). 16S rDNA sequencing revealed that PCA altered the composition of the gut microbiota. Fecal microbiota transplantation (FMT) confirmed that PCA-remodeled microbiota alone could transfer the protective effect, reduced IL-6 and elevated Claudin-1 and ZO-1 in recipient mice (P<0.01). Targeted metabolomics identified a significant increase in secondary bile acids, particularly lithocholic acid (LCA), after PCA treatment (P<0.05). LCA treatment suppressed key inflammatory cytokines (IL-1β, IL-6, and TNF-α) (P<0.05), enhanced the expression of Claudin-1, improved intestinal morphology, and inhibited TAK1 phosphorylation (P<0.01). Binding assays and molecular simulations indicated that LCA directly interacts with the TAK1 catalytic site (Met104 and Lys63), disrupting its activity. In summary, PCA ameliorated intestinal inflammation by modulating the gut microbiota to enrich LCA, which directly targets and inhibits TAK1 phosphorylation.