高良姜素诱导人乳头瘤病毒阳性的宫颈癌细胞凋亡实验研究

Inductive effects of galangin on apotosis of human papilloma virus positive cervical cancer cells

目的研究高良姜素对人乳头瘤病毒(HPV)阳性人宫颈癌细胞的诱导凋亡作用及其机制。方法选择2株HPV阳性人宫颈癌细胞(Si Ha、He La)和1株HPV阴性人宫颈癌细胞(C-33-A),用不同浓度的高良姜素(20、40、80μmol/L)分别处理24、48、72 h后,采用噻唑蓝(MTT)法测定高良姜素对3株宫颈癌细胞增殖的影响;不同浓度高良姜素处理宫颈癌细胞48 h后,分别采用流式细胞仪检测细胞凋亡和细胞周期;Western blotting法检测Bcl-2家族蛋白表达情况。结果高良姜素可以剂量和时间依赖性地降低HPV阳性宫颈癌细胞Si Ha和He La的活力;而对于HPV阴性的宫颈癌细胞C-33-A活力无明显影响;细胞周期检测显示高良姜素可以使Si Ha和He La细胞的生长阻滞在G1或G2/M期;流式细胞仪分析发现40μmol/L高良姜素作用于Si Ha和He La细胞48 h后,细胞出现明显凋亡,而C-33-A细胞仅有少量凋亡;Western blotting检测发现40μmol/L高良姜素处理后,HPV阳性宫颈癌细胞中促凋亡蛋白Bad、Bid、Bax表达量升高,而抗凋亡蛋白Bcl-2、Bcl-w表达量降低。结论高良姜素能抑制HPV阳性宫颈癌细胞的增殖并诱导其凋亡,其作用机制可能与下调Bcl-2、Bcl-w基因的表达及上调Bad、Bid、Bax基因的表达有关。

Objective To explore the inductive effect of galangin on HPV-positive human cervical cancer cells and the possible mechanism. Methods Two HPV-positive human cervical cancer cell lines（Si Ha cell and He La cell） and one HPV-negative human cervical cancer cell line（C-33-A cell） were given different concentration of galangin（20, 40, and 80 μmol/L） for 24, 48, and 72 h. Three human cervical cancer cell lines and relative cell viabilities were determined by the MTT method. Apoptosis and cell cycle were analyzed by flow cytometry. Western blotting analysis was used to determine the protein expression levels of Bcl-2 family proteins. Results Cell proliferation of two HPV-positive human cervical cancer cells was significantly inhibited by galangin in a dose- and time-dependent manner, and galangin had no effect on cell proliferation of HPV-negative human cervical cancer cells. Cell cycle detection results showed that galangin could reversibly arrest the two HPV-positive cell lines, either in G1 or in G2/M phases. Flow cytometry results showed that beyond certain galangin concentration or/and over 24 h exposure, the cells underwent apoptosis. The data of Western blotting showed that 40 μmol/L galangin up-regulated the expression levels of Bad, Bid, and Bax, but down-regulated Bcl-2 and Bcl-w. Conclusion Galangin can inhibit the proliferation of HPV-positive cervical cancer cells and promote apoptosis, which may be associated with the regulation of Bcl-2 family proteins expression.