β-胡萝卜素对人膀胱癌T24细胞凋亡和细胞间隙连接蛋白43表达的影响

Effects of beta-carotene on the apoptosis of human bladder cancer cell line T24 and the expression of its connexin 43

目的探讨β-胡萝卜素抑制人膀胱癌T24细胞的分子生物学机制。方法不同浓度的β-胡萝卜素作用于T24细胞,利用MTT法检测细胞增殖,流式细胞术检测其凋亡指数,半定量RT-PCR检测Cx43、Bcl-2、Bax mRNA的变化,Western blot检测Cx43、Bcl-2、Bax蛋白的表达。结果 10和20μmol/L组β-胡萝卜素均能明显抑制T24细胞的生长和诱导细胞凋亡,其抑制率分别为28.83%、63.02%,并且呈明显的剂量-效应关系(P<0.05),凋亡率分别为0.126±0.022和0.190±0.024(P≤0.01),随β-胡萝卜素浓度增加,细胞凋亡率增大,呈剂量依赖性关系(P=0.002)。与对照组相比,10和20μmol/L组β-胡萝卜素上调Cx43 mRNA及其蛋白的表达,增强细胞间的细胞间隙连接通讯(GJIC)功能(P<0.05),并且呈剂量依赖关系(P≤0.01);下调Bcl-2 mRNA及其蛋白的表达,并且呈剂量依赖关系(P≤0.01);Bax mRNA及其蛋白的表达有上调趋势,但与对照组比较,差异无显著性。结论β-胡萝卜素(10,20μmol/L)能通过有效下调Bcl-2 mRNA及其蛋白在T24细胞的表达,上调T24细胞Cx43转录水平的表达、增强细胞间隙连接所介导的GJIC功能,从而诱导膀胱肿瘤细胞凋亡,抑制其生长。

Objective To investigate the mechanism of molecular biology of beta-carotene on inhibiting the growth of human bladder cancer cell line T24.Methods The effect of different concentrations of beta-carotene（0,5,10 and 20 μmol/L） treated for 72 h on human bladder cancer T24 cells was evaluated by MTT assay and flow cytometry.Both Semi-quantitative RT-PCR and Western Blot were used to detect the expression of connexin 43（Cx43）,Bcl-2 and Bax mRNA.Results In beta-carotene treated groups at the concentration of 10 and 20 μmol/L,the proliferation of T24 cells could be inhibited and the apoptosis could be induced（28.83%,63.02%,P﹤0.05） significantly.The apoptotic index（AI） was 0.126±0.022 and 0.190±0.024 in the treated group,which was increased significantly in comparison with the control group（P≤0.01） and in a dose-response relationship（P=0.002）.In comparison with the control group,the expression of Cx43 mRNA and its protein could be up-regulated and the gap junctional intercellular communication（GJIC） of T24 cells could be enhanced in a dose-response relationship（P0.05）,but the expression of Bcl-2 mRNA and its protein was down-regulated and in a dose-response relationship significantly（P≤0.01） in the beta-carotene treated groups.There was no correlation was found between the expression of Bax mRNA and beta-carotene.Conclusion Through the effective down-regulation of the expression of Bcl-2,the up-regulation of the expression of Cx43,and the enhancement of the GJIC of T24 cells,the apoptosis of bladder tumor cells was induced by beta-carotene（10,20 μmol/L）,thereby the growth of bladder tumor cells could be inhibited.The experiment provided an evidence for the mechanism of beta-carotene on chemical prevention and treatment of bladder tumor.