三羟异黄酮抑制结肠癌LOVO细胞增殖作用的机制研究

Mechanism of inbibitory effect of genistein on the proliferation of colon cancer LOVO cells

【目的】探讨三羟异黄酮对以雌激素受体β表达为主的结肠癌LOVO细胞的增殖抑制作用及其机制。【方法】以不同浓度三羟异黄酮加入体外培养的LOVO细胞中,用倒置光显微镜观察细胞形态变化;通过四甲基偶氮唑盐(MTT)法检测细胞增殖抑制率;用Hoechst33342/PI双荧光染色,荧光显微镜分析细胞死亡特征;以流式细胞术检测细胞周期变化;应用半定量逆转录PCR检测caspase-3和bcl-2 mRNA的表达。【结果】10、30、60、90、120μmol/L的三羟异黄酮均可不同程度地抑制LOVO细胞生长;荧光显微镜下可见30μmol/L的三羟异黄酮既可诱导LOVO细胞凋亡,而且凋亡比例随药物浓度增高而增加,120μmol/L的三羟异黄酮可引起细胞部分坏死;30μmol/L的三羟异黄酮可明显影响LOVO的细胞周期经三羟异黄酮处理后,其中G0/G1期细胞比例逐渐减少,G2/M期细胞比例明显增高,阻断细胞生长于G2/M期。RT-RCR果结显示caspase-3 mRNA表达较对照组明显增强(P<0.01),bcl-2 mRNA表达较对照组明显减弱(P<0.01)。【结论】三羟异黄酮能够抑制LOVO细胞生长,其机制可能与影响凋亡相关基因表达有关。

[ Objective] To investigate the inhibitory effect of genistein on the proliferation of LOVO colon cancer cell merely expressing estrogen receptor- βand the mechanisms. [ Methods] The morphologic changes were detected by light microscope, the growth inhibition of cells was measured by methyl - tetrazolinm （MTT） assay and the feature of cell death was studied by Hoechst33342/PI staining. Cell cycle and apoptosis rate were detected by flow cytometry and mRNA expression of caspase-3 and bcl- 2 were measured by reverse transcription PCR （RT- PCR）. [Results] Genistein inhibited the growth of LOVO cell in a dose and time-dependent manner. Enhancement of proportion of apoptotic cell was observed by fluorescence microscope and genistein in high concentration induced cell into necrosis partially.Flow cytometry results showed apoptosis rate of LOVO was increased in a dose-dependent manner , and the fraction of G0/G1 was significantly decreased, while the fraction of G2/M was significantly increased.The result of RT-PCR showed that the mRNA expression of caspase-3 was higher and the the mRNA expression of bcl-2 was lower than the control group （ P 〈 0.01） . [Conclusions] Genistein could inhibit the cells growth of LOVO cells by affecting the gene experssion related to cell apeptesis.