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目的:探讨茜草提取物蒽醌单体对人肝癌SMMC-7721细胞生长的抑制作用,诱导凋亡作用及对凋亡相关基因bcl-2表达的影响,为肝癌的基因治疗提供有效靶点。方法:采用MTT法检测生长的抑制作用;以形态学,DNA凝胶电泳,流式细胞仪单染、双染检测细胞凋亡;以RT-PCR来分析蒽醌对bcl-2 mRNA的影响。结果:蒽醌能抑制肝癌细胞的生长;蒽醌处理肝癌细胞后,倒置显微镜和光镜下可见典型的凋亡细胞;DNA琼脂糖凝胶电泳显示出典型的细胞凋亡梯形带,PI单染表明细胞周期的G1期前有异常二倍体细胞的凋亡峰,并将细胞周期阻滞在G1期,Annexin V-FITC/PI定量检测进一步证实了蒽醌诱导肝癌细胞凋亡的作用;RT-PCR检测表明蒽醌可使bcl-2 mRNA表达水平下降。结论:蒽醌能显著抑制肝癌细胞的生长,并诱导肝癌细胞凋亡,其分子机制可能与下调bcl-2基因的表达有关。
OBJECTIVE: To investigate the inhibitory effect of anthraquinone extracted from madder extract on the growth of human hepatocellular carcinoma SMMC-7721 cells, the induction of apoptosis and the expression of apoptosis-related gene bcl-2, so as to provide an effective target for the gene therapy of hepatocellular carcinoma. Methods: MTT assay was used to detect the growth inhibition. Cell apoptosis was detected by morphology, DNA gel electrophoresis, single-flow cytometry and double staining. The effect of anthraquinone on bcl-2 mRNA was analyzed by RT-PCR. Results: Anthraquinone could inhibit the growth of hepatocellular carcinoma cells. After treated with anthraquinone, typical apoptotic cells were observed under inverted microscope and light microscopy. DNA ladder agarose gel electrophoresis showed a typical apoptotic ladder, PI single staining showed Cell cycle G1 phase before abnormal apoptosis peak of diploid cells, and the cell cycle arrest in the G1 phase, Annexin V-FITC / PI quantitative detection further confirmed the anthraquinone-induced apoptosis in hepatocellular carcinoma cells; RT- PCR showed that anthraquinone decreased the expression of bcl-2 mRNA. Conclusion: Anthraquinone can significantly inhibit the growth of hepatoma cells and induce the apoptosis of hepatocellular carcinoma cells. The molecular mechanism may be related to the down-regulation of bcl-2 gene expression.