Pharmacological TRPM8 blockade reprograms patient-derived intestinal fibroblasts and mitigates experimental fibrosis through modulation of myofibroblast activation

Intestinal fibrosis is a major long-term complication mostly prevalent in Crohn's disease (CD) and remains without approved antifibrotic therapies. Transient receptor potential melastatin 8 (TRPM8) is a pharmacologically targetable ion channel involved in inflammatory regulation, but its contribution to intestinal fibrotic remodeling is unknown. This study investigated TRPM8 association with human stenotic CD and whether pharmacological TRPM8 modulation limits fibroblast activation and experimental intestinal fibrosis. TRPM8 expression was increased in stenotic ileocolonic tissues from CD patients and in primary patient-derived intestinal fibroblasts. In these cells, the TRPM8 antagonist AMTB reduced proliferation and migration at non-cytotoxic concentration and attenuated the expression of fibroblast activation, matrix and myofibroblast-associated markers. In mice with chronic 2,4,6-trinitrobenzenesulfonic acid (TNBS)-induced intestinal fibrosis, AMTB reduced disease activity,endoscopic severity and attenuated stromal remodeling-associated gene expression. Histological analyses further showed reduced tissue injury and collagen remodeling, supported by Masson's Trichrome staining and quantitative analysis of Picrosirius Red. In contrast, pharmacological TRPM8 activation with WS12 did not significantly modify fibroblast proliferation in vitro and/or the evaluated clinical, endoscopic and molecular readouts in vivo. AMTB also reduced the TNBS-induced T helper 17 response without reducing interleukin-23 production by colonic mononuclear phagocytes. Finally, fibroblasts isolated from AMTB-treated fibrotic mice displayed reduced expansion, lower collagenogenic gene expression, reduced contractile/activated fibroblast markers and decreased actin cytoskeletal remodeling ex vivo. Overall, these findings identify TRPM8 as a druggable regulator of fibroblast-driven intestinal remodeling and support TRPM8 antagonism as a candidate antifibrotic strategy for stricturing CD.