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目的探讨二甲双胍联合柔红霉素对人结肠癌细胞(SW480)增殖和凋亡的影响。方法采用MTT法检测药物对细胞增殖抑制的影响;集落克隆形成法检测药物对细胞集落克隆形成的影响;碘化丙啶(PI)单染法检测细胞凋亡率的变化;AnnexinⅤ/PI双染法检测细胞早期凋亡的变化;蛋白质免疫印迹法检测Bax、Bcl-2蛋白的表达。试验设空白对照组、药物单用组(二甲双胍或柔红霉素)、药物合用组(二甲双胍+柔红霉素)。选择低于半数抑制浓度20 mmol·L-1的二甲双胍和0.2μmol·L-1柔红霉素单用或合用作用于SW480细胞。结果不同浓度的二甲双胍单用、两药合用24、48、72 h后的细胞存活率分别为81.23%、68.88%、57.88%、75.32%、60.98%、50.43%;与单用药组比较有著性差异(P<0.05);两药单用、合用24 h细胞凋亡率分别为24.23%、30.6%、54.83%,与对照组比较有显著性差异(P<0.05);两药合用24 h细胞早期凋亡高于药物单用,与对照组比较有显著性差异(P<0.05);两药单用或合用细胞集落克隆形成数量均减少,与对照组比较有显著性差异(P<0.05);药物合用时促进凋亡蛋白Bax的表达高于药物单用、抑制凋亡蛋白Bcl-2的表达低于药物单用。结论二甲双胍联合柔红霉素能够通过诱导细胞的凋亡,增强对SW480细胞的增殖抑制作用,其作用机制可能与增强Bax的表达,下调Bcl-2有关。
Objective To investigate the effects of metformin combined with daunorubicin on the proliferation and apoptosis of human colon cancer cell line SW480. Methods MTT assay was used to detect the effect of drugs on cell proliferation inhibition. The colony formation assay was used to detect the effect of drugs on cell colony formation. The change of apoptosis rate was detected by propidium iodide (PI) staining. AnnexinⅤ / PI double staining Method was used to detect the changes of cell apoptosis in early stage. The protein expression of Bax and Bcl-2 was detected by Western blot. The experiment set blank control group, drug group (metformin or daunorubicin), drug combination group (metformin + daunorubicin). Metformin and 0.2 μmol·L-1 daunorubicin were selected at a concentration less than 50% of the inhibitory concentration of 20 mmol·L-1 to act on SW480 cells singly or in combination. Results The cell viabilities of metformin at different concentrations were 81.23%, 68.88%, 57.88%, 75.32%, 60.98% and 50.43% at 24, 48 and 72 h, respectively. Compared with the single drug group, (P <0.05). The apoptotic rates of the two drugs alone and combined for 24 h were 24.23%, 30.6% and 54.83%, respectively, which were significantly different from those of the control group (P <0.05) (P <0.05). The numbers of single colony-forming or colony-forming colony formation of the two drugs decreased, which were significantly different from the control group (P <0.05). The combination of drugs to promote the expression of apoptosis protein Bax was higher than the drug alone, inhibiting the expression of apoptosis protein Bcl-2 was lower than the drug alone. Conclusion Metformin combined with daunorubicin can induce the apoptosis of SW480 cells and enhance its inhibitory effect on SW480 cells. The mechanism may be related to the enhancement of Bax expression and the down-regulation of Bcl-2.