1. Intestinal Myofibroblast TRPC6 Channel May Contribute to Stenotic Fibrosis in Crohnʼs Disease
- Author
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Ryuji Inoue, Kyoko Nakajima, Yaopeng Hu, Lin Hai Kurahara, Keizo Hiraishi, Midori Nakagawa, Kunihiko Aoyagi, and Miho Sumiyoshi
- Subjects
Adult ,Male ,Pathology ,medicine.medical_specialty ,Blotting, Western ,Real-Time Polymerase Chain Reaction ,TRPC4 ,TRPC6 ,Immunoenzyme Techniques ,Transforming Growth Factor beta1 ,Colonic Diseases ,Transient receptor potential channel ,Crohn Disease ,Downregulation and upregulation ,Fibrosis ,TRPC6 Cation Channel ,medicine ,Humans ,Immunoprecipitation ,Immunology and Allergy ,RNA, Messenger ,Intestinal Mucosa ,Myofibroblasts ,Cells, Cultured ,TRPC ,TRPC Cation Channels ,biology ,Reverse Transcriptase Polymerase Chain Reaction ,Gastroenterology ,Middle Aged ,medicine.disease ,Intestines ,biology.protein ,Cancer research ,Female ,ACTA2 ,Myofibroblast - Abstract
Background Intestinal fibrosis is a frequent complication of Crohn's disease (CD) and often leads to detrimental stricture formation. Myofibroblasts play active roles in mediating fibrotic changes in various tissues. We investigated whether transient receptor potential channels in intestinal myofibroblasts are involved in CD-associated intestinal fibrosis. Methods An intestinal myofibroblast cell line (InMyoFibs) was stimulated with transforming growth factor-β1 (TGF-β1) to model excessive fibrosis. Biopsy samples from nonstenotic or stenotic intestinal regions from patients with CD were used for quantitative comparisons of transient receptor potential channel and fibrosis-associated factor expression levels. Results TGF-β1 treatment transformed spindle-shaped InMyoFibs into filament-shaped cells with enhanced α-actin stress fiber formation, transient receptor potential canonical (TRPC) 4 and TRPC6 messenger RNA and protein expression, and basal- and agonist-induced Ca influxes. TGF-β1 also enhanced the formation of TRPC6/smooth muscle α-actin, TRPC6/N-cadherin, and TRPC4/N-cadherin coimmunoprecipitates. Inhibition of TRPC6 in InMyoFibs by RNA interference or dominant-negative mutations suppressed TGF-β1-induced Ca influxes, stress fiber formation, and smooth muscle α-actin expression, but increased COL1A1, interleukin (IL)-10, and IL-11 expression, as well as Smad-2, ERK, and p38-MAPK phosphorylation. Similar increases in phosphorylation levels were observed with TRPC and calcineurin inhibitors. In stenotic areas in patients with CD, TRPC6, ACTA2 (smooth muscle α-actin), CDH2 (N-cadherin), COL1A1, IL-10, and IL-11 were significantly increased. Conclusions These results suggest that augmented Ca influxes due to TRPC6 upregulation facilitate stress fiber formation and strengthen cell-cell interactions by negatively regulating the synthesis of antifibrotic factors in TGF-β1-treated myofibroblasts. Similar changes observed in stenotic areas of patients with CD suggest the therapeutic significance of targeting TRPC6.
- Published
- 2015