青蒿琥酯对大鼠原代肝星状细胞产生与分泌转化生长因子β1的影响  

Impact of artesunate on the expression and secretion of transforming growth factor-β1 of primary rat hepatic stellate cells

作　　者：王媛[1] 方步武[1] 彭龙希[2] 

机构地区：[1]天津医科大学药理学教研室,300070 [2]天津医科大学总医院药剂科

出　　处：《中华肝脏病杂志》2012年第4期294-299,共6页Chinese Journal of Hepatology

基　　金：国家自然科学基金（30772856）

摘　　要：目的探讨青蒿琥酯对大鼠原代肝星状细胞（HSC）增殖的影响，从抑制HSC表达、生成和分泌转化生长因子β1（TGFβ1）这—环节探讨其抗肝纤维化的机制。方法分离大鼠HSC于培养瓶中原代培养10d，已处于培养活化状态，将HSC分为实验组和对照组，实验组以青蒿琥酯（终浓度分别为125、150、175、200、225μmol／L）作用24、48、72h。以四甲基偶氮唑盐（MTT）法检测细胞增殖率，RT-PCR法检测HSC中TGFβ1 mRNA的表达水平，Western blot法分析TGFβ1蛋白水平的变化，酶联免疫吸附法测定培养上清液中TGFβ1含量。样本均数比较采用单因素方差分析，两样本均数比较采用独立样本t检验。结果不同浓度青蒿琥酯对培养活化的HSC均有明显抑制作用，且呈剂量-效应关系和时间-效应关系，作用24h时，125、150、175、200、225μmol／L青蒿琥酯对HSC的抑制率分别为6．06%±1．44％、21．47％±5．57％、42．00％±7．36%、67．12％±4．55％、79．83％±3．67％（P值均〈0．01）。青蒿琥酯作用HSC24h能明显抑制、下调HSC表达TGFβ1 mRNA，呈剂量-效应关系（P〈0．01）；并且明显降低细胞内TGFβ1蛋白及细胞上清液中TGFβ1水平，0、150、175、200μmol／L青蒿琥酯处理组TGFμ1分别为（164．24±6.88）pg／ml、（102．68±4．45）pg／ml、（86．54±5．56）pg/ml、（56．55±5．66）pg／ml（P值均〈0．01）。结论青蒿琥酯呈剂量和时间依赖性地抑制原代分离培养活化的HSC，青蒿琥酯在体外具有抗肝纤维化的作用，与其下调TGFβ1基因及蛋白的表达、翻译与TGFβ1分泌至细胞外等环节有关。Objective To investigate the impact of the Artemisia annua plant-derived drug, artesunate, on proliferation of primary rat hepatic stellate cells （HSCs）, and to analyze the underlying molecular mechanisms of its anti-fibrogenic effects involving the inhibition of transforming growth factor-beta 1 （TGF-β1） expression and secretion in liver. Method Isolated, cultured, and activated primary rat HSCs were divided into sixteen groups, including one untreated control group and fifteen artesunate-treated experimental groups with 125, 150, 175, 200 or 225 μmol/L for 24, 48 or 72 hours. The rate of cellular proliferation was measured using the 3-（4, 5-dimethylthiazol-2-yl）-2, 5-diphenyltetrazolium bromide （MTT） assay. TGF-β1 mRNA expression was evaluated by reverse transcription-polymerase chain reaction and protein expression was evaluated by Western blotting. Enzyme-linked immunosorbent assay was used to evaluate secreted levels of TGF-β1 protein. Results Artesunate significantly inhibited proliferation of cultured HSCs in a dose- and time-dependent manner （all, P 〈 0.01）. After 24 hours of exposure, the inhibition ratios of the various artesunate concentrations were： 6.06%±1.44% （ 125 μmol/L）, 21.47% ± 5.57% （150 μmol/L）, 42.00% ± 7.36% （175 μmol/L）, 67.12% ± 4.55% （200 μmol/L）, and 79.83% ±3.67% （225 μmol/L）. Artesunate significantly inhibited the TGF-β1 mRNA expression in HSCs, and the higher the drug concentration, the higher the degree of inhibition （all, P〈 0.01）. In addition, artesunate significantly inhibited the expression of intracellular and secreted TGF-β1 protein （all, P〈 0.01）. In response to artesunate （μmol/L concentrations）, the TGF-β1 levels were （164.24±6.88） pg/ml （μmol/L）, （102.68±4.45） pg/ml （150μmol/L）, （86.54±5.56） pg/ml （175μmol/L）, and （56.55 ±5.66） pg/ml （200μmol/L）. Conclusion Artesunate exerts anti-fibrogenic effects on HSCs in vitro, possibly by reducing the expression, tran

关 键 词：青蒿 肝硬化 转化生长因子Β1 青蒿琥酯 肝星状细胞 

分 类 号：R285.5[医药卫生—中药学]