Indole-3-acetic acid promotes differentiation while suppressing proliferation in human intestinal epithelial cells

Indole-3-acetic acid (IAA) is a tryptophan-derived microbial metabolite increasingly recognized for its role in intestinal homeostasis, immune regulation, and epithelial function. However, the IAA molecular effects on intestinal epithelial cells remain incompletely defined. Here, we investigated the effects of increasing IAA concentrations on intestinal epithelial biology using intestinal epithelial Caco-2 cell line, with a focus on cytotoxicity, epithelial repair, and differentiation. Cell viability was assessed by MTT assay; colony-forming assays were used to evaluate stemness potential; cytokine's expression was quantified by qPCR. Differentiation was analyzed through dome formation and analysis of DPPIV, SI, KLF4 and E-cadherin differentiation markers by qPCR and immunofluorescence. IAA treatment exerts concentration-dependent effects, reducing the viability and colony-forming capacity of Caco-2 cells. At higher concentrations, IAA also decreased IL-17, IL-1β, IL-6 expression. Interestingly, IAA enhanced dome formation and upregulated several differentiation markers, suggesting a shift toward a more mature epithelial phenotype. Altogether, our results indicate that IAA directly influences epithelial cell biology by modulating viability, inflammatory signaling, repair capacity, and differentiation. Our data support a potential role for this metabolite in regulating intestinal growth, repair and differentiation suggesting that IAA may contribute to intestinal homeostasis and disease pathophysiology. However, further studies are essential to understand the potential therapeutic implications related to colorectal cancer.