论文部分内容阅读
目的体外研究肾癌786-0细胞来源的exosomes介导肿瘤免疫逃逸的机制。方法采用CCK-8法检测肾癌786-0细胞来源的exosomes对Jurkat T细胞生长的影响,瑞氏-姬姆萨染色检测Jurkat T细胞形态变化,Annexin V-FITC/PI双染色流式细胞术检测Jurkat T细胞凋亡率,ELISA法检测Jurkat T细胞分泌功能,可溶性Fas阻断实验检测exosomes对Jurkat T细胞凋亡率的影响,Western blot检测exosomes中FasL、及Jurkat T细胞caspase、Bax及Bcl-2蛋白的表达。结果肾癌786-0细胞来源的exosomes可抑制Jurkat T细胞生长,10μg/mL exosomes作用于Jurkat T细胞24 h,生长抑制率为(19.64±0.92)%,72 h为(36.24±1.12)%;400μg/mL exosomes作用24 h,生长抑制率为(55.96±1.35)%,72 h为(76.51±1.37)%。Exosomes诱导Jurkat T细胞凋亡,10μg/mL exosomes作用于Jurkat T细胞8 h,凋亡率为(7.31±1.32)%,24 h为(20.19±1.47)%;400μg/mL exosomes作用8 h,凋亡率为(27.28±1.29)%,24 h为(41.72±0.88)%。Exosomes还明显抑制Jurkat T细胞IL-2、IFN-γ、IL-6、IL-10的分泌水平;exosomes高表达FasL,可溶性Fas阻断实验能逆转Jurkat T细胞的凋亡;凋亡诱导过程中caspase-3、caspase-8、caspase-9被激活,Bax/Bcl-2上调。结论肾癌786-0细胞分泌的exosomes能诱导Jurkat T细胞凋亡,介导肿瘤免疫逃逸。
OBJECTIVE: To study the mechanism of exosomes derived from renal carcinoma 786-0 cells in mediating tumor immune escape in vitro. Methods The growth of Jurkat T cells was detected by CCK-8 assay. The morphological changes of Jurkat T cells were detected by Wright-Giemsa staining. Annexin V-FITC / PI double staining flow cytometry The apoptosis of Jurkat T cells was detected by ELISA. The secretion of Jurkat T cells was detected by ELISA. The effect of exosomes on the apoptosis rate of Jurkat T cells was detected by soluble Fas-blocking assay. The expressions of FasL and Jurkat T cells caspase, Bax and Bcl- -2 protein expression. Results The exosomes derived from 786-0 cells of renal cell carcinoma could inhibit the growth of Jurkat T cells. The inhibitory rate of Jurkat T cells was (19.64 ± 0.92)% after treated with 10 μg / mL exosomes for 24 h, and (36.24 ± 1.12)% after 72 h. 400μg / mL exosomes for 24 h, the growth inhibition rate was (55.96 ± 1.35)%, 72 h (76.51 ± 1.37)%. Exosomes induced apoptosis of Jurkat T cells. The apoptotic rates of Jurkat T cells treated with 10 μg / mL exosomes for 8 h were (7.31 ± 1.32)% and (20.19 ± 1.47)% for 24 h and 400 μg / mL exosomes for 8 h, respectively The mortality rate was (27.28 ± 1.29)% and (41.72 ± 0.88)% at 24 h. Exosomes also significantly inhibited the secretion of IL-2, IFN-γ, IL-6 and IL-10 in Jurkat T cells. Exosomes overexpressed FasL. Soluble Fas blockade could reverse the apoptosis of Jurkat T cells. Caspase-3, caspase-8, caspase-9 are activated, Bax / Bcl-2 upregulation. Conclusion The exosomes secreted by renal cancer 786-0 cells induce apoptosis of Jurkat T cells and mediate tumor immune escape.