Effects of curcumin on peroxisome proliferator-activated receptor γ expression and nuclear translocation/redistribution in culture-activated rat hepatic stellate cells
Effects of curcumin on peroxisome proliferator-activated receptor γ expression and nuclear translocation/redistribution in culture-activated rat hepatic stellate cells
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DOI：
10.1097/00029330-200705010-00011
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发表时间：
2007-05-05
影响因子：
6.1
通讯作者：
Xu Lie-ming
中科院分区：
文献类型：
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作者：
Cheng Yang;Ping Jian;Xu Lie-ming
背景 过氧化物酶体增殖物激活受体γ（PPARγ）在肝纤维发生中的功能仍然很大程度上未知。姜黄素是从姜黄中提取的天然物质，具有多种药理作用。本研究探讨姜黄素通过PPARγ信号传导对大鼠肝星状细胞(HSC)增殖、活化和凋亡的影响。方法通过Pronase E原位肝脏灌注和Nycodenz密度梯度离心从正常SD大鼠肝脏中分离HSC。用姜黄素、曲格列酮、丹酚酸 B 或 GW9662 处理细胞。通过MTT比色法测定对HSC增殖的影响。用TRIzol试剂提取总RNA，并通过半定量RT-PCR测定基因水平。总细胞和核蛋白通过10％十二烷基硫酸钠聚丙烯酰胺凝胶电泳分离和分离。通过蛋白质印迹测定蛋白质水平。 Hoechst 33258染色检测细胞凋亡。通过免疫荧光染色检测PPARγ亚细胞分布。采用明胶酶谱法测定MMP-2和9的活性。结果姜黄素以紧密依赖性方式抑制HSCs增殖。随着 HSC 随着培养时间的延长而逐渐激活，PPAR γ 核表达水平下降。姜黄素上调PPARγ表达，显着抑制α-SMA和胶原1的产生。PPARγ在细胞质和细胞核中表达，均匀分布在HSC中，但姜黄素处理后在HSC细胞核中积累，并从细胞质中消失。 Hoechst 33258 染色显示姜黄素诱导培养激活的 HSC 凋亡，并显着增加促凋亡 Bax 表达并减少抗凋亡 Bcl-2 表达。姜黄素显着下调 Cyclin D1 基因、活化的 NF kappa B p65 蛋白和 TGF beta R-1 蛋白表达。姜黄素能显着增强MMP-2和MMP-9的活性。结论姜黄素能抑制HSC的增殖和活化，诱导活化的HSCS凋亡，增强MMP-2和MMP-9的活性。姜黄素的作用是通过激活 PPAR γ 信号转导途径介导的，并与 PPAR-γ 核易位/重新分布相关。
Background The function of peroxisome proliferator-activated receptor gamma (PPAR gamma) in hepatic fibrogenesis remains largely unknown. Curcumin is a natural substance extracted form Curcuma Longa Linn and has a variety of pharmacological effects. In this study, the effects of curcumin on the proliferation, activation and apoptosis of rat hepatic stellate cells (HSCs) through PPAR gamma signaling were investigated.Methods HSCs were isolated from the normal Sprague Dawley rats th rough in situ perfusion of the liver with Pronase E and density-gradient centrifugation with Nycodenz. Cells were treated with curcumin, troglitazone, salvianolic acid B or GW9662. The effect on HSCs proliferation was determined by MTT colorimetry. Total RNA was extracted by TRizol reagent and gene levels were determined by semi-quantitative RT-PCR. Total cellular and nuclear protein were isolated and separated by 10% sodium dodecy Isulfate polyacrylamide gel electrophoresis. Protein levels were determined by Western blot. Cell apoptosis was detected by Hoechst 33258 staining. PPAR gamma subcellular distribution was detected by immunofluorescent staining. The activities of MMP-2 and 9 were measured by Gelatin zymograph assay.Results Curcumin suppressed HSCs proliferation in a close-dependent manner. As HSCs underwent gradual activation with culture prolongation the PPAR gamma nuclear expression level decreased. Curcumin up-regulated PPAR gamma expression and significantly inhibited the production of alpha-SMA and collagen 1. PPAR gamma is expressed in the cytoplasm and nucleus and is evenly distributed in HSCs, but accumulated in the nucleus of HSCs and disappeared from cytoplasm after curcumin treatment. Hoechst 33258 staining showed that curcumin induced the apoptosis of culture-activated HSCs and significantly increased pro-apoptotic Bax expression and reduced anti-apoptotic Bcl-2 expression. Cyclin D1 gene, activated NF kappa B p65 protein and TGF beta R-1 protein expression were down-regulated significantly by curcumin. The activities of MMP-2 and MMP-9 were enhanced significantly by curcumin.Conclusions Curcumin can inhibit the proliferation and activation of HSCs, induce the apoptosis of activated HSCS and enhance the activities of MMP-2 and MMP-9. The effects of curcumin are mediated through activating the PPAR gamma signal transduction pathway and associated with PPAR-gamma nuclear translocation/redistribution.