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目的:研究表没食子儿茶素-3-没食子酸酯(epigallocatechin-3-gallate,EGCG)对人肺癌A549细胞增殖和凋亡影响及与CUGBP1表达的关系。方法:MTT法检测不同剂量EGCG对A549细胞增殖活性的影响;流式细胞仪检测细胞凋亡率;免疫组化和蛋白质印迹法检测各处理组CUGBP1蛋白的表达;Real-Time PCR对CUGBP1mRNA表达进行定量分析。结果:EGCG显著抑制A549细胞增殖活性,F=1 096.65,P<0.001;EGCG孵育4和24hA549细胞的凋亡率分别为(34.10±1.10)%和(51.62±1.50)%,较4和24h正常对照组的(3.90±0.97)%和(1.53±1.11)%明显增加,差异有统计学意义,F=1 254.28,P<0.001。A549细胞细胞核和细胞质内CUGBP1蛋白表达呈阳性,EGCG孵育4h可见A549细胞核CUGBP1蛋白表达呈阴性;孵育24h,部分A549细胞变圆、核固缩,CUGBP1蛋白表达又呈阳性。CUGBP1mRNA定量分析可见,EGCG明显抑制CUGBP1表达,EGCG孵育24h,A549细胞中CUGBP1mRNA相对表达量为(0.71±0.076),较4h的(0.40±0.017)高,差异有统计学意义,t=-6.782,P=0.020。结论:EGCG干扰CUGBP1基因表达抑制人肺癌细胞株A549的增殖,可能是EGCG促进A549细胞凋亡的途径之一。
Aims: To investigate the effect of epigallocatechin-3-gallate (EGCG) on the proliferation and apoptosis of human lung cancer A549 cells and its relationship with the expression of CUGBP1. Methods: MTT assay was used to detect the effect of different doses of EGCG on the proliferation of A549 cells; the apoptosis rate was detected by flow cytometry; the expression of CUGBP1 protein was detected by immunohistochemistry and Western blotting; the expression of CUGBP1 mRNA was detected by Real-Time PCR Quantitative analysis. Results: The proliferation of A549 cells was significantly inhibited by EGCG (F = 1096.65, P <0.001). The apoptotic rates of EGCG after 4 and 24h incubation were (34.10 ± 1.10)% and (51.62 ± 1.50)%, respectively, (3.90 ± 0.97)% and (1.53 ± 1.11)% in the control group were significantly increased, the difference was statistically significant, F = 1 254.28, P <0.001. The expression of CUGBP1 in nucleus and cytoplasm of A549 cells was positive. The expression of CUGBP1 protein in A549 cells was negative after incubated with EGCG for 4 h. After incubation for 24h, some A549 cells were round and nuclear pyknosis was positive and CUGBP1 protein was positive again. CUGBP1mRNA quantitative analysis showed that EGCG significantly inhibited the expression of CUGBP1, EGCG incubated 24h, A549 cells CUGBP1mRNA relative expression (0.71 ± 0.076), compared with 4h (0.40 ± 0.017), the difference was statistically significant, t = -6.782, P = 0.020. Conclusion: The interference of EGCG on CUGBP1 gene expression inhibits the proliferation of human lung cancer cell line A549 may be one of the ways of EGCG to promote the apoptosis of A549 cells.