Toxicol Appl Pharmacol 2014 Oct 19;280(2):335-44. Epub 2014 Aug 19.
School of Pharmacy, Anhui Medical University, Mei Shan Road, Hefei, Anhui Province 230032, China; Institute for Liver Diseases of Anhui Medical University, Mei Shan Road, Hefei, Anhui Province 230032, China; Key Laboratory of Anti-inflammatory and Immune Medicine, Anhui Medical University, Ministry of Education, Mei Shan Road, Hefei, Anhui Province 230032, China. Electronic address:
Transdifferentiation of hepatic stellate cells (HSCs) into myofibroblasts plays a critical role in the development of liver fibrosis, since myofibroblasts are the key cells responsible for excessive deposition of ECM proteins. Transient receptor potential melastatin 7 (TRPM7), a non-selective cation channel with protein serine/threonine kinase activity, has been demonstrated to function in the proliferation of activated HSCs. Here, we investigated the functional role of TRPM7 in collagen deposition in activated HSC-T6 cells (a rat hepatic stellate cell line). TRPM7 mRNA and protein were measured by Real-time PCR and Western blot in TGF-β1-activated HSC-T6 cells in vitro. Results demonstrated that TRPM7 protein was dramatically increased in fibrotic human livers. Stimulation of HSC-T6 cells with TGF-β1 increased TRPM7 mRNA and protein level in a time-dependent manner. Nevertheless, TGF-β1-elicited upregulation of TRPM7 in HSC-T6 cells was abrogated by SB431542 (TGF-β1 receptor blocker) or SIS3 (inhibitor of Smad3 phosphorylation). Additionally, blockade of TRPM7 channels with non-specific TRPM7 blocker 2-APB or synthetic siRNA targeting TRPM7 attenuated TGF-β1-induced expression of myofibroblast markers, as measured by the induction of α-SMA and Col1α1. Silencing TRPM7 also increased the ratio of MMPs/TIMPs by increasing MMP-13 expression and decreasing TIMP-1 and TIMP-2 levels. Strikingly, phosphorylation of p-Smad2 and p-Smad3, associated with collagen production, was decreased in TRPM7 deficient HSC-T6 cells. These observations suggested that TGF-β1 elevates TRPM7 expression in HSCs via Smad3-dependant mechanisms, which in turn contributes Smad protein phosphorylation, and subsequently increases fibrous collagen expression. Therefore, TRPM7 may constitute a useful target for the treatment of liver fibrosis.