熊果酸对人结肠癌HT—29细胞增殖抑制及诱导凋亡探究.docVIP

熊果酸对人结肠癌HT—29细胞增殖抑制及诱导凋亡探究 　　[摘要] 目的 探讨熊果酸（UA）诱导人结肠癌细胞HT-29凋亡的作用机制。 方法 MTT法观察UA对HT-29细胞增殖的抑制作用；荧光显微镜和流式细胞术观察HT-29细胞的凋亡情况；Western blot检测细胞色素c（cyt-c）、caspase-3及Livin蛋白的表达。 结果 UA对HT-29细胞具有明显的的增殖抑制作用，并呈一定的浓度和时间依赖性；荧光显微镜观察结果显示，50 mu;mol/L的UA作用24 h后，较多细胞呈现出凋亡的特征性改变；流式细胞术检测发现，UA可将HT-29细胞阻滞于G1期，随着UA浓度的增高，细胞凋亡率也相应增加；Western blot结果显示，UA作用24 h后，Livin蛋白和caspase-3酶原的表达逐渐降低，线粒体释放的cyt-c逐渐增多，活化的caspase-3片段逐渐增加。 结论 UA对HT-29细胞具有明显的增殖抑制作用，其机制可能与UA阻滞HT-29细胞的细胞周期、激活线粒体凋亡途径以及下调凋亡抑制蛋白Livin的表达有关。　　[关键词] 熊果酸；人结肠癌细胞HT-29；凋亡　　[中图分类号] R735.3+5 [文献标识码] A [文章编号] 1674-4721（2016）09（a）-0025-05　　[Abstract] Objective To investigate the apoptosis-inducing mechanism of ursolic aicd （UA） in human colon cancer cell HT-29. Methods The inhibitory effect of UA on the growth of HT-29 cells was evaluated by MTT assay. Apoptosis was determined by fluorescence microscopy and flow cytometry. The expressions of apoptosis-associated proteins including cytochrome c （cyt-c）， caspase-3 and Livin were assessed by Western blot analysis. Results UA inhibited HT-29 cell proliferation in a time- and dose-dependent manner. Fluorescent microscope indicated characteristics of apoptosis present in cells after UA-treated （50 mu;mol/L， 24 h）. Flow cytometry showed that UA inhibited HT-29 in G1 phase， and the apoptosis rate of HT-29 cell exposured to UA increased， corresponding to the concentration of UA. The inceasing of the release of cyt-c from the mitochondria and the activated fragment of caspase-3 were detected by Western blot， while the expression of procaspase-3 and Livin was decreased gradually. Conclusion UA can inhibit the proliferation of HT-29 cells effectively in vitro. The mechanism of above-mentioned phenomena may be related with the arrest effect of UA in the cell cycle of HT-29， activated mitochondrial apoptotic pathway， and down-regulating the expression of Livin.　　[Key words] Ursolic acid； Human colon cancer cell HT-29； Apoptosis　　熊果酸（Ursolic acid，UA），又叫乌苏酸、乌索酸，它在自然界中存在广泛，是熊果、白花蛇舌草、乌梅等多种植物的主要活性成分，具有多种生物学活性，包括