Anti-fibrogenic effect of PPAR-gamma agonists in human intestinal myofibroblasts

Abstract

BACKGROUND: Intestinal fibrosis is a serious complication of inflammatory bowel disease, including Crohn's disease and ulcerative colitis. There is no specific treatment for intestinal fibrosis. Studies have indicated that peroxisome proliferator-activated receptor- gamma (PPAR-gamma) agonists have anti-fibrogenic properties in organs besides the gut: however, their effects on human intestinal fibrosis are poorly understood. This study investigated the anti-fibrogenic properties and mechanisms of PPAR-gamma agonists on human primary intestinal myofibroblasts (HIFs). METHODS: HIFs were isolated from normal colonic tissue of patients undergoing resection due to colorectal cancer. HIFs were treated with TGF-beta1 and co-incubated with or without one of two synthetic PPAR-gamma agonists, troglitazone or rosiglitazone. mRNA and protein expression of procollagen1A1, fibronectin, and alpha-smooth muscle actin were determined by semiquantitative reverse transcription-polymerase chain reaction and Western blot. LY294002 (Akt inhibitor) was used to examine whether Akt phosphorylation was a downstream mechanism of TGF-beta1 induced expression of procollagen1A1, fibronectin, and alpha-smooth muscle actin in HIFs. The irreversible PPAR-gamma antagonist GW9662 was used to investigate whether the effect of PPAR-gamma agonists was PPAR-gamma dependent. RESULTS: Both PPAR-gamma agonists reduced the TGF-beta1-induced expression of alpha-smooth muscle actin which was integrated into stress fibers in HIFs, as determined by actin microfilaments fluorescent staining and alpha-smooth muscle actin-specific immunocytochemistry. PPAR-gamma agonists also inhibited TGF-beta1-induced mRNA and protein expressions of procollagen1A1, fibronectin, and alpha-smooth muscle actin. TGF-beta1 stimulation increased phosphorylation of downstream signaling molecules Smad2, Akt, and ERK. TGF-beta1 induced synthesis of procollagen1A1, fibronectin, and alpha-smooth muscle actin through a phosphatidylinositol 3-kinase/Akt-dependent mechanism. PPAR-gamma agonists down regulated fibrogenesis, as shown by inhibition of Akt and Smad2 phosphorylation. This anti-fibrogenic effect was PPAR-gamma independent. CONCLUSIONS: Troglitazone and rosiglitazone suppress TGF-beta1-induced synthesis of procollagen1A1, fibronectin, and alpha-smooth muscle actin in HIFs and may be useful in treating intestinal fibrosis.