Hu Yi-Bing, Ye Xiao-Ting, Zhou Qing-Qing, Fu Rong-Quan
Department of Gastroenterology, Jinhua Hospital of Zhejiang University, Jinhua, China.
Department of Infectious Diseases, The Third Affiliated Hospital of Wenzhou Medical University, Rui'an, China.
Cell Physiol Biochem. 2018;51(5):2111-2122. doi: 10.1159/000495829. Epub 2018 Dec 6.
BACKGROUND/AIMS: Sestrin 2 is associated with the pathophysiology of several diseases. The aim of this study was to investigate the effects and potential mechanisms of Sestrin 2 in rat hepatic stellate cells (HSCs) during liver fibrogenesis.
In this study, Sestrin 2 protein expression was detected in rat HSC-T6 cells challenged with transforming growth factor-β (TGF-β) and in mice treated with carbon tetrachloride (CCl4), a well-known model of hepatic fibrosis. Next, HSC-T6 cells and fibrotic mice were transfected with lentivirus. The mRNA expression levels of markers of liver fibrosis [alpha-smooth muscle actin (α-SMA) and collagen 1A1 (Col1A1)] were analyzed by quantitative reverse transcription-polymerase chain reaction (RT-PCR). Cell death and proliferation were evaluated by the MTT assay, and biochemical markers of liver damage in serum [alanine transaminase (ALT) and aspartate transaminase (AST)] were also measured using a biochemical analyzer. Histopathological examination was used to evaluate the degree of liver fibrosis, and protein expression [phospho-adenosine monophosphate-activated protein kinase (p-AMPK), AMPK, phospho-mammalian target of rapamycin (p-mTOR), and mTOR] was determined by western blotting.
We found that Sestrin 2 was elevated in both the HSC-T6 cell and hepatic fibrosis models. In vitro, overexpression of Sestrin 2 attenuated the mRNA levels of α-SMA and Col1A1, suppressed α-SMA protein expression, and modulated HSC-T6 cell proliferation. In vivo, overexpression of Sestrin 2 reduced the ALT and AST levels as well as the α-SMA and Col1A1 protein expression in the CCl4 model of liver fibrosis. Moreover, the degree of liver fibrosis was ameliorated. Interestingly, overexpression of Sestrin 2 increased p-AMPK but decreased p-mTOR protein expression.
Our findings indicate that Sestrin 2 may attenuate the activation of HSCs and ameliorate liver fibrosis, most likely via upregulation of AMPK phosphorylation and suppression of the mTOR signaling pathway.
背景/目的：硒蛋白2与多种疾病的病理生理学相关。本研究旨在探讨硒蛋白2在肝纤维化过程中对大鼠肝星状细胞（HSCs）的影响及潜在机制。
在本研究中，检测了用转化生长因子-β（TGF-β）刺激的大鼠HSC-T6细胞以及用四氯化碳（CCl4）处理的小鼠（一种著名的肝纤维化模型）中硒蛋白2的蛋白表达。接下来，用慢病毒转染HSC-T6细胞和纤维化小鼠。通过定量逆转录-聚合酶链反应（RT-PCR）分析肝纤维化标志物[α-平滑肌肌动蛋白（α-SMA）和胶原蛋白1A1（Col1A1）]的mRNA表达水平。通过MTT法评估细胞死亡和增殖情况，并用生化分析仪检测血清中肝损伤的生化标志物[丙氨酸转氨酶（ALT）和天冬氨酸转氨酶（AST）]。采用组织病理学检查评估肝纤维化程度，通过蛋白质印迹法测定蛋白表达[磷酸化腺苷单磷酸活化蛋白激酶（p-AMPK）、AMPK、磷酸化雷帕霉素靶蛋白（p-mTOR）和mTOR]。
我们发现硒蛋白2在HSC-T6细胞和肝纤维化模型中均升高。在体外，硒蛋白2的过表达降低了α-SMA和Col1A1的mRNA水平，抑制了α-SMA蛋白表达，并调节了HSC-T6细胞增殖。在体内，硒蛋白2的过表达降低了肝纤维化CCl4模型中的ALT和AST水平以及α-SMA和Col1A1蛋白表达。此外，肝纤维化程度得到改善。有趣的是，硒蛋白2的过表达增加了p-AMPK但降低了p-mTOR蛋白表达。
我们的研究结果表明，硒蛋白2可能通过上调AMPK磷酸化和抑制mTOR信号通路来减轻HSCs的激活并改善肝纤维化。
