【摘要】 目的:研究T24膀胱癌细胞中Toll样受体4(Toll like receptor-4,TLR4)信号通路活化对B7-H1表达的影响,并探讨其参与膀胱癌免疫逃逸的可能机制。方法:培养膀胱肿瘤T24细胞系,以脂多糖(lipopolysaccharides,LPS)模拟TLR4在体内的激活过程,分别用1μg/mL的LPS刺激T24细胞0、2、4、6、8和12h,流式细胞技术检测T24细胞表面TLR4的表达,RT-PCR技术检测T24细胞中B7-H1mRNA的表达,蛋白质印迹法检测T24细胞中B7-H1蛋白的表达。结果:流式细胞学检测结果显示,以1μg/mL的LPS刺激不同时间T24细胞TLR4表达的阳性率,0h为(3.34±0.86)%,2h为(6.71±1.38)%,4h为(9.71±1.24)%,6h为(11.57±0.91)%,8h为(25.24±5.32)%,12h为(21.67±1.81)%,组间差异均有统计学意义,F=83.049,P<0.05。RT-PCR检测结果显示,LPS刺激0h时B7-H1mRNA的相对表达量为0.86±0.06,2h为1.30±0.11,4h为1.67±0.20,6h为1.99±0.10,8h为2.57±0.23,12h为1.51±0.04,组间差异均有统计学意义,F=84.719,P<0.05。蛋白质印迹检测结果显示,LPS刺激0h时B7-H1蛋白质相对表达量为0.66±0.08,2h为0.65±0.10,4h为0.87±0.11,6h为1.07±0.23,8h为1.45±0.19,12h为1.13±0.06,组间差异均有统计学意义,F=195.258,P<0.05。TLR4表达分别与B7-H1mRNA(r1=0.753)及蛋白(r2=0.836)表达显著正相关,P<0.05。结论:通过LPS激活TLR4信号通路可以上调B7-H1mRNA及蛋白的表达,而这有可能是TLR4信号通路参与膀胱癌免疫逃逸的机制之一。更多还原

【Abstract】 OBJECTIVE:To study the relationship between activation of TLR4-signaling pathway and B7-H1 expression in T24 bladder cancer cells,and research the role they play in bladder cancer immune evasion.METHODS:T24 bladder cancer cell lines were cultured and treated with LPS(1μg/mL) for 0,2,4,6,8,12 h.TLR4 expression was examined by flowcytometrty and B7-H1 mRNA and protein expression were examined by RT-PCR and Western-Blot respectively.One-way ANOVA and Spearman rank correlation analysis were applied in the data analysis.RESULTS:Flowcytometrty analysis showed the TLR4 expression of T24 bladder cancer cells was significantly up-regulated depending on the duration of LPS stimulation.The positive rate of TLR4 on T24 cells were 0 h(3.34±0.86)%,2 h(6.71±1.38)%,4 h(9.71±1.24)%,6 h(11.57±0.91)%,8 h(25.24±5.32)%,12 h(21.67±1.81)%,at the same time B7-H1 mRNA and protein expression showed a very similar tendency after LPS stimulation.The B7-H1 mRNA relative expression quantity were 0 h(0.86±0.06),2 h(1.30±0.11),4 h(1.67±0.20),6 h(1.99±0.10),8 h(2.57±0.23),12 h(1.51±0.04),and B7-H1 protein 0 h(0.66±0.08),2 h(0.65±0.10),4 h(0.87±0.11),6 h(1.07±0.23),8 h(1.45±0.19),12 h(1.13±0.06).One-Way ANOVA analysis showed that the differences between groups were statistically significant(F values were 83.049,84.719,195.258,P<0.05).B7-H1 mRNA and protein expression was significantly related to TLR4 expression respectively(r1=0.753,r2=0.836,P<0.05).CONCLUSION:TLR4-signaling pathway may be involved in bladder cancer immune evasion process by up-regulating B7-H1 expression.更多还原