RNA干扰沉默胃癌细胞FasL表达的抗肿瘤免疫作用

Anti-tumor effects of RNA interference silencing on the FasL expression of gastric cancer cells

目的探讨胃癌细胞FasL表达诱导肿瘤特异性CD8^+CTL凋亡介导胃癌细胞的免疫逃逸,RNA干扰沉默胃癌细胞FasL表达能通过增加肿瘤特异淋巴细胞的数量以提高机体抗肿瘤免疫作用。方法设计并筛选针对性的人胃癌细胞FasL-siRNA,应用脂质体法把siRNA双链分子(dsRNA)转入胃癌细胞,以沉默胃癌细胞中FasL的表达,设置实验组(转染目标质粒)、阴性对照组(转染空载质粒)及空白对照组(未转染)。沉默胃癌细胞FasL基因表达后,在与肿瘤特异性CTL共培养系统中观察其对CTL的凋亡诱导作用和抗肿瘤免疫功能的影响。采用流式细胞仪检测胃癌细胞Fas/FasL表达水平,采用MTT(二苯基四氮唑溴盐)法观察胃癌细胞与肿瘤特异性CTL共培养结果。结果转染FasL-siRNA的胃癌细胞经检测细胞膜的FasL表达减少,与转染前对比差异存在统计学意义(P<0.05),并且转染后实验组较阴性对照及空白对照组表达FasL量存在统计学差异(P<0.05),胃癌细胞与肿瘤特异性CTL共培养显示,实验组淋巴细胞数量(吸光度值)较阴性组和空白对照组多,差异存在统计学意义(P<0.05)。结论应用RNA干扰技术可使胃癌细胞FasL表达量下降,减少胃癌细胞通过Fas/FasL途径诱导抗肿瘤淋巴细胞凋亡的发生,增强淋巴细胞对胃癌细胞的杀伤作用。

Objective To verify the FasL expression of gastric cancer cells that induces tumor-specific CD8+CTL apoptosis mediates the immune evasion and prove RNA interference silences the FasL expres-sion of gastric cancer cells can enhance antitumor function by increasing the number of tumor-specific lymphocytes.Methods Targeted FasL shRNA was design and screened to transfect gastric cancer cells by using liposome to silence the FasL expression.The experimental group(transfect targeted plasmid),negative group and(transfect idler plasmid)blank group(without transfect)were set up,and then observing the effects of gastric cancer cells on inducing tumor-specific CTL apoptosis and antitumor function in the cell co-culture system with tumor-specific CD8+CTL.The FasL expression in gastric cancer cells was detected by flow cytometry.And the results of co-culture with gastric cancer cells and tumor-specific CD8+CTL were determined by MTT method.Results In the experimental group,those gastric cancer cells transfected by FasL-siRNA expressed a lower level of FasL which was significant in statistics compared with blank group.Compared with negative group and blank group,there existed more lymphocytes in the experimental group(differences between groups were significant in statistics,P<0.05).Conclusion In the present study,Using RNA interference reduced the FasL expression of gastric cancer cells that enhanced the function of lymphocytes killing cancer cells.