姜黄素与5-氟尿嘧啶联用对人结肠癌HT-29细胞凋亡及环氧合酶-2表达的影响

Combined effect of curcumin and 5-FU on apoptosis and COX-2 expression in human colon carcinoma cell line HT-29

目的:探讨姜黄素是否能增强5-氟尿嘧啶(5-FU)的抗癌作用,并探讨其与细胞凋亡及COX-2蛋白表达的关系。方法:采用MTT法观察不同浓度姜黄素、5-FU单独或联合应用对HT-29细胞的生长抑制作用;流式细胞仪和荧光显微镜检测对HT-29细胞凋亡的影响;并且应用Western blotting分析联合应用后对于COX-2蛋白表达的影响。结果:姜黄素和5-FU可明显抑制HT-29细胞的生长,并呈剂量依赖性,半数生长抑制率(IC50)分别为(15.9±1.96)μmol/L和(17.3±1.85)μmol/L;当二者联合应用时,在较高的作用水平可以定量地观察到对HT-29细胞的协同抑制作用。流式细胞仪结果表明,单独应用姜黄素以及联合应用姜黄素与5-FU均能使HT-29细胞在加药后24和48 h出现凋亡峰,但联合用药所诱导的细胞凋亡比例较单独应用姜黄素低;荧光显微镜观察的结果与流式细胞仪的结果相同;免疫印迹实验结果显示两种药物联合作用可以使COX-2的表达降低约75%。结论:姜黄素与5-FU联合应用可协同抑制大肠癌细胞的增殖并协同抑制HT-29细胞内COX-2的表达。

Aim：To observe whether curcumin could enhance the anti-proliferation effect of fluorouracil （5-FU） and to investigate the possible link between this combinative effect and cell apoptosis and COX-2 expression. Methods：The inhibitory effects on HT-29 after the treatment with curcumin or 5-FU alone or in combination were investigated using MTF methods. Changes to apoptotic cell content in HT-29 cell were analyzed by the flow cytometry mad fluorescence microscope. Inhibition on COX-2 expression after the treatment of curcumin and 5-FU in combination was also evaluated by Western Blot analysis. Results： Proliferation of HT-29 cells was significantly inhibited by curcumin and/ or 5-FU in a concentration-dependent manner. The IC50 Values of curcumin and 5-FU in HT-29 cells were 15.9 ± 1.96 μmol/L and 17.3 ± 1.85 μmol/L, respectively. When HT-29 cells were treated with curcumin and 5-FU in combination, the effect of synergistic inhibition on the growth was quantitatively demonstrated at even higher levels. After the cells were treated with curcumin or curcumin and 5-FU in combination for 24 h and/or 48 h, the apoptotic peak appeared by observation with the analysis of flow cytometry. However, the ratio of apoptotie cell after being treated with the two compounds in combination was lower than that after being treated with curcumin only. The same observation was found by the means of fluorescence microscope. In addition, the level of COX-2 protein expression was reduced nearly 4-fold after the use of the two agents in combination. Conclusion： In the case of colon cancer cell line such as HT-29, combined use of curcumin and 5-FU could synergistically inhibit the cell growth and the expression of COX-2 protein.