Blockage of TRPM7 channel induces hepatic stellate cell death through endoplasmic reticulum stress-mediated apoptosis

Aims: Proliferation is a 'multiplier' for extracellular matrix production and contraction of activated hepatic stellate cells (HSC) in fibrotic liver. Transient receptor potential melastatin-like 7 channels (TRPM7) are implicated in the survival and proliferation of several kinds of cells. This study was aimed to investigate the effect of TRPM7 blocker 2-APB on survival and proliferation of HSC and the underlying mechanisms. Main methods: Rat HSC were stimulated by 2-APB for 24 h and then collected for further use. Cell viability was detected by MIT, and apoptosis was determined by AnnexinV/PI staining and TUNEL assay. Gene expressions of TRPM7, alpha-SMA, bcl-2, bax, and endoplasmic reticulum (ER) stress key members CHOP, caspase-12, ATF4, ATF6, Xbpl, GRP78 and calnexin were evaluated with quantitative RT-PCR. Quantifications of alpha-SMA, TRPM7, CHOP and GRP78 proteins were carried out by Western blot. Transmission electron microscopy and Xbpl mRNA splicing analysis were also used for detection of ER stress. Key findings: 2-APB decreased TRPM7 and alpha-SMA expressions in primary HSC, and inhibited proliferation of activated HSC in a dose-dependent mannei. 2-APB also decreased total count of activated HSC and increased the number of apoptotic cells. 2-APB increased expressions of bax and ER stress key factors CHOP, caspase-12, ATF4, ATF6, Xbpl, GRP78 and calnexin. Meanwhile, ultra-structural ER changes and spliced Xbpl mRNA were also observed in 2-APB treated HSC. Significance: Blockage of TRPM7 could inhibit activation and proliferation of primary HSC and induce apoptotic death of activated cells, in which ER stress was identified as one of possible underlying molecular bases. (C) 2014 Elsevier Inc. All rights reserved.