【摘要】 【目的】观察叶下珠复方Ⅱ号体外对肝星状细胞(HSC-T6)凋亡和miR-16/Bcl-2表达的影响,进一步探讨其抗肝纤维化的作用机制。【方法】体外培养HSC-T6细胞,分为空白对照组,叶下珠复方Ⅱ号高剂量组(3.6 g/L)、中剂量组(1.8 g/L)和低剂量组(0.9 g/L),采用四甲基偶氮唑盐(MTT)法检测药物对HSC-T6细胞增殖的影响,流式细胞仪检测细胞凋亡的变化,实时荧光定量PCR法检测药物处理后HSC-T6细胞miR-16和Bcl-2 mRNA表达的改变,Western Blot法测定Bcl-2蛋白的表达变化。【结果】叶下珠复方Ⅱ号处理HSC-T6细胞后,高、中、低各剂量组对HSC-T6细胞增殖均有明显的抑制作用,随着叶下珠复方Ⅱ号浓度的增加,细胞的凋亡率也逐渐增加;叶下珠复方Ⅱ号各处理组miR-16的表达显著增加,呈现明显的剂量—效应关系,且Bcl-2 mRNA和蛋白质表达显著减少,与空白对照组比较,差异均具有统计学意义(P<0.01)。【结论】叶下珠复方Ⅱ号能显著抑制HSC-T6细胞增殖和诱导其凋亡,其作用机制与提高miR-16的表达,抑制其靶基因Bcl-2的表达有关。更多还原

【Abstract】 Objective To observe the effect of compound Phyllanthus urinaria Ⅱ(CPUⅡ),a Chinese herbal recipe mainly composed of Phyllanthus urinaria Linn,on apoptosis and miR-16/Bcl-2 expression in hepatic stellate cells(HSC-T6),and to explore its anti-fibrosis mechanism.Methods HSC-T6 were cultured in vitro and were divided into 4 groups,including blank control group,and high-,middle-and low-dose CPUⅡ groups(in the dosage of 3.6,1.8,0.9 g/L,respectively).Methyl thiazolyl tetrazolium(MTT) assay was used for evaluating the effect of CPUⅡ on inhibiting HSC-T6 proliferation.Flow cytometry assay was used for studying the action of CPU Ⅱ on inducing HSC-T6 cells apoptosis.Real-time fluorescence quantitative polymerase chain reaction(PCR) was used for detecting miR-16 and Bcl-2 mRNA expression levels.The expression level of Bcl-2 protein was analyzed by Western blotting method.Results HSC-T6 proliferation was obviously inhibited in the CPUⅡ groups,and the apoptotic rate was increased gradually with the dosage.miR-16 expression level was elevated,which showed a significant dose-effect relationship.Bcl-2 mRNA and protein levels were decreased,the difference being significant compared with the blank control group(P <0.01).Conclusion CPUⅡ can effectively inhibit the proliferation of HSC-T6 and induce the apoptosis in vitro,and its therapeutic mechanism may be related with the increase of miR-16 expression and with the inhibition of the target gene Bcl-2 expression.更多还原