非对称二甲基精氨酸对肝星状细胞的激活作用及其机制  

Effect of asymmetric dimethylarginine on the activation of hepatic stellate cells and its mechanism

作　　者：李金成[1] 昌兰[2] 吕冬[3] 姜德建[2] 谭德明[1] 

机构地区：[1]中南大学湘雅医院感染科,长沙410008 [2]中南大学药学院药理学系,长沙410008 [3]中南大学护理学院,长沙410008

出　　处：《中南大学学报（医学版）》2007年第3期427-432,共6页Journal of Central South University ：Medical Science

基　　金：湖南省自然科学基金(06jj3012)~~

摘　　要：目的:研究内源性一氧化氮合酶抑制物非对称二甲基精氨酸(asymmetric dimethylarginine,ADMA)对肝星状细胞(hepatic stellate cells,HSC)活化的影响及其机制。方法:原代分离和培养SD大鼠HSC细胞,加入不同浓度的ADMA(1,3或10μmol/L)孵育12至48h。细胞组织化学检测α-平滑肌肌动蛋白(α-smooth muscle actin,α-SMA)表达;酶联免疫吸附试验测定I型胶原的合成;逆转录PCR检测HSC转化生长因子-β1(transforming growth factor-β1,TGF-β1) mRNA水平;荧光发光法检测细胞内氧自由基(reactive oxidant species,ROS)生成;凝胶电泳迁移率试验检测核因子-κB(nuclear factor-κB,NF-κB)活性。结果:ADMA能呈浓度和时间依赖性上调HSCTGF-β1 mRNA水平,增加α-SMA阳性细胞数和促进I型胶原的合成。同时,ADMA能显著增加细胞内ROS生成和诱导NF-κB活化,而预处理抗氧化剂四氧化吡咯二硫代氨基甲酸酯能抑制ADMA(10μmol/L)诱导的NF-κB活化和TGF-β1 mRNA水平上调。结论:ADMA能诱导HSC的激活(包括收缩和分泌功能),其作用与激活细胞内ROS-NF-κB途径,上调TGF-β1表达有关。因此,ADMA可能是一种新的HSC功能调节因子,在肝纤维化的发生发展中起重要作用。Objective To investigate the effect of asymmetric dimethylarginine （ADMA） , an endogenous inhibitor of nitric oxide synthase, on the activation of hepatic stellate cells （HSCs） and its mechanism. Methods Primary HSCs isolated from SD rats were cultured arid treated with different concentrations （ 1 , 3 or 10 μmol/L） of ADMA for various periods （ 12 -48 h）. Expression of α-smooth muscle actin （α-SMA） and synthesis of type-Ⅰ collagens in HSC were determined. Mes- senger RNA levels of the transforming growth factor-β1 （ TGF-β1 ） in the HSCs were determined using RT-PCR. Intracellular reactive oxidant species （ROS） production was measured using oxidant-sensitive fluorescent indicator. Activation of nuclear factor-κB （NF-κB ） was detected by electrophoretic mobility shift assay （EMSA）. Results ADMA could increase α-SMA-positive cells ratio and Type Ⅰ collagens production of HSCs in a concentration- and time-dependent manner, concomitant with the increase of the TGF-β1 mRNA level. Treatment with ADMA （ 10 μmol/L ） significantly increased the intracellular ROS production and activated NF-κB. Such effects of ADMA on the level of TGF-β1 mRNA could be markedly attenuated by pretreatment with antioxidant pyrrolidine dithiocarbamate （25μmol/L）. Conclusion ADMA can induce the HSC activation by increasing TGF-β1 expressionthrough ROS-NF-κB-dependent pathway. Therefore, ADMA should be a novel and endogenous acti-vator of HSC, which may be involved in the development of liver fibrosis.

关 键 词：非对称二甲基精氨酸 肝星状细胞 转化生长因子-β_1 肝纤维化 

分 类 号：R575.2[医药卫生—消化系统]