甘草总黄酮与石见穿总酚酸联用促肝星状细胞凋亡作用研究

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1、甘草总黄酮与石见穿总酚酸联用促肝星状细胞凋亡作用研究【摘要】目的研究甘草总黄酮与石见穿总酚酸联用对氧化应激的肝星状细胞(HSC)的促凋亡作用。方法MTT法检测甘草总黄酮与石见穿总酚酸联用对大鼠肝星状细胞株(HSC-T6)增殖的影响。次氮基三乙酸铁与HSC-T6体外共培养建立氧化应激模型,采用TBA法检测细胞内丙二醛(MDA)含量,运用邻苯酸酚自氧化速率法检测超氧化物歧化酶(SOD)活性。荧光探针DCFH-DA检测细胞内活性氧水平。荧光染料Hoechst33258观察细胞的形态及凋亡情况。JC-1检测细胞内线粒体膜电位变化情况。结果甘草总黄酮与石见穿总酚酸联合处理HSC-T6后,细胞的增殖受

2、到抑制,并促进HSC-T6凋亡;细胞内线粒体膜电位下降;随着药物剂量的增加细胞内活性氧含量下降。结论甘草总黄酮与石见穿总酚酸联用促进氧化应激的HSC凋亡,此作用可能与线粒体凋亡途径有关。【关键词】甘草总黄酮;石见穿总酚酸;肝星状细胞;凋亡;线粒体膜电位　Abstract：ObjectiveToinvestigatetheeffectofGlycyrrhizauralensisflavonoidsandsalvianolicacid(GS)ontheapoptosisofHSC-T6cells.MethodsHSC-T6wereincubatedwithferricnitrilotriace

3、ticacid(Fe-NTA),SODactivityandMDA15contentinHSC-T6weredetected.MTTcolorimetricmethodandtrypanbluestainingwereusedtomeasurethecellproliferentiationandsurvivalrate.Productionofreactiveoxygenspecies(ROS)ofdrug-treatedcellsweremonitoredbyafluorescentprobe2’,7’-dichlorodihydrofluorescinfluorescence(DCF

4、H-DA).MorphologicalchangesassociatedwithapoptosiswereexaminedbystainingwithHoechst33258.Moreover,changesinmitochondrialtransmembranepotentialweredetermindebythedual-emissionpotential-sensitiveprobe5,5',6,6'-tetra-chloro-1,1',3,3'-tetraethyl-imidacarbocyanineiodide(JC-1).ResultOxidativestressdecrea

5、sedtheactivityofSODinHSCs,increasedthecontentofMDAinHSC-T6.GSinhibitedthecellsproliferationanddecreasedtheiROSlevelsinadosedependentmanner,After24hourstreatmentwithGS,thetypicalapopticmorphologywasfoundinHSCs,includingthedecreaseofkaryoplasmicratioandtheincreaseofkidney-shapenuclearcells.Furthermo

6、re,GSinducedaconcentrationdecreaseinROSproductionandinmitochondrialmembranepotential.ConclusionGSplayanimportantroleinoxidativestressHSC-T6apoptosiswhichmaybecorrectedwithmitochondriaapoptosispathway.15　　Keywords：Glycyrrhizauralensis;salvianolicacid;hepaticstellatecells;apoptosis;mitochondriamembr

7、anepotential　　肝纤维化(hepaticfibrosis,HF)是慢性肝病的共同病理学基础,其中25%～40%最终发展为肝硬化甚至肝癌[1]。目前认为,肝星状细胞(hepaticstellatecells,HSC)是HF细胞外基质的主要来源细胞,HSC的活化和功能的改变是HF形成的关键[2-3],也是抗纤维化研究的核心。越来越多的实验表明,氧化应激是不同原因引起的HF发生的共同通路。实际上,铁负载、乙醇、四氯