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Enhanced autophagic flux contributes collagen production in hepatic stellate cells involving TGFbeta1/Smad pathway signaling
Author(s) -
Chen XingZhou,
Yao WenMin,
Zhang Guo,
Ye HuiLan,
Chen Li,
Zhang FaCan
Publication year - 2013
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.27.1_supplement.1161.3
Subject(s) - atg5 , autophagy , hepatic stellate cell , microbiology and biotechnology , chemistry , smad , intracellular , downregulation and upregulation , signal transduction , apoptosis , biology , endocrinology , biochemistry , gene
Autophagy is a conserved lysosomal degradation pathway, which has been shown to be involved in fibrogenesis in multiple organs such as kidney, heart, lung, skin and liver. However, the precise mechanism by which autophagy participates in the hepatic fibrosis remains unclear. Thus this present study was firstly designed to dynamically monitor the specific morphological change of autophagy in primary cultured hepatic stellate cells (HSCs) during the fibrogenesis process. By using fluorescence microscopy and transmission electron microscopy technique, it was found that TGFbeta1 dose‐ and time‐dependently induced increments in autophagosome numbers in HSCs. Secondly, real time RT‐PCR and western blotting was respectively performed to analyze autophagy, apoptosis and TGFbeta1/Smad signaling molecules. In the presence of rapamycin (100nM), an autophagic agonist, Atg5, beclin1, LC3‐II, Smad3 and collagen type I was found to be substantially up‐regulated in HSCs with a caspase3‐independent pattern. On the contrary, either 3‐methyladenine (3MA, 5mM) or hydroxychloroquine (HCQ, 32μg/mL), antagonists for autophagy, was found to dramatically attenuate the impact of rapamycin on the proliferation of HSCs, follow by a corresponding decline in smad3 level and cellular collagen production as well. Furthermore, small interference RNA targeting Atg5, beclin1 was added to pre‐incubate HSCs respectively. It was showed that both Atg5‐ and beclin1‐siRNA amazingly impaired TGFbeta1‐induced expression of collagen type I in these cells. Taken together, our findings indicate a critical working pattern of autophagic flux in HSCs in response to TGFbeta1 and thereby provide a novel intracellular basis for antifibrotic drug development in the future. (Supported by NSFC Grants 30630145, 881172260, 200903 to G.Z. and GWZ200958 to F.Z.)