蛇床子素诱导Hela细胞凋亡及其机制研究

Research on Apoptosis of Human Cervical Cancer Hela Cells Induced by Osthole and Its Mechanism

目的:探讨蛇床子素诱导人宫颈癌Hela细胞凋亡及其机制。方法:采用MTT法检测各浓度蛇床子素对Hela细胞增殖的抑制作用;光学显微镜观察细胞的形态学变化;凝胶电泳分析DNA的片段化改变;采用RT-PCR检测Hela细胞fas和bcl-2基因mRNA的表达水平。采用10μg/mL的蛇床子素作用于Hela细胞24h后的培养上清称为ost-La;洗去蛇床子素,用新鲜培养液进行24h再培养的Hela细胞培养上清称为ost-Lb;不经蛇床子素处理的Hela细胞同步培养上清作为对照,称为control-L;定量Elisa法测定各上清中TGF-β1含量。结果:蛇床子素可显著抑制Hela细胞的体外增殖,诱导Hela细胞的凋亡,且均呈剂量依赖性。RT-PCR:凋亡相关基因fas的表达量升高,而bcl-2的表达量下降,表达量的变化与蛇床子素的浓度有关。Elisa:TGF-β1的含量在ost-La中为16.732±3.068 ng/mL(P<0.01);在ost-Lb中为2.031±0.112 ng/mL(P<0.01);在control-L中为385.282±42.613 ng/mL。结论:子素可诱导Hela细胞的凋亡,其机制可能与促进Fas基因表达,抑制Bcl-2基因表达和TGF-β1含量改变有关。

Objective：To investigate the effects of Osthole on the apoptosis of Hela cells in vitro and to research its mechanism in detail.Methods：The inhibitory effects of Osthole on proliferation of Hela cells were detected using the MTT assay.After Hela cells were treated with different doses of Osthole,cell morphological alteration was observed by using optical microscopy,the DNA Fragmentation was examined by gel electrophoresis and RT- PCR was performed to measure the expression level of fas and bcl- 2 mRNA.The supernatant of Hela cells after treated with 10μg /mL Osthole respectively for 24 hours was called ost- La,and those followed by washing completely to remove Osthole and re- cultured with fresh medium for 24 hours was called ost- Lb.The corresponding supernatants of Hela cells treated without Osthole is control- L.The concentration of TGF- β1 in different supernatant we measured by quantitative ELISA.Results：Osthole was found to inhibit proliferation and to induce the apoptosis of Hela cells in a concentration- dependent manner.RT- PCR：The expression of apoptosis- associated gene fas was up regulated,and the expression of bcl- 2 was down regulated.The effects were dependent on concentration of Osthole.ELISA：The TGF- β1 content of ost- La was 16.732 ± 3.068 ng /mL（P 0.01）,of ost-Lb was 2.031 ±0.112ng/mL（P 0.01）,of control- L was（385.282 ±42.613） ng/mL.Conclusion：Osthole can induce apoptosis of Hela cells.Its mechanism might relate to promoting the expression of fas,inhibiting the expression of bcl- 2and changing of the TGF- β1 content.