小鼠Lewis肺癌细胞Foxp3的表达对T淋巴细胞增殖的影响及机制  

The expression of Foxp3 in mouse Lewis lung cancer cells and its effect and mechanism on proliferation of T lymphocytes

作　　者：历春[1] 盖晓东[2] 贾婷[1] 付海英[1] 李一[1] 

机构地区：[1]吉林大学白求恩医学院免疫学系,长春130021 [2]吉林省北华大学基础医学院病理教研室,吉林132013

出　　处：《中国免疫学杂志》2010年第11期986-991,996,共7页Chinese Journal of Immunology

基　　金：吉林大学基本科研业务费科学前沿与交叉学科创新项目(No.200903007)

摘　　要：目的:检测小鼠Lewis肺癌(LLC)细胞中Foxp3基因的表达,研究小鼠LLC细胞中Foxp3过表达对T淋巴细胞的增殖和TGF-β1和IL-10表达的影响,探讨LLC细胞中Foxp3的表达在肿瘤免疫逃逸中的作用及可能机制。方法:分别采用RT-PCR和免疫组化染色法检测LLC细胞中Foxp3的mRNA和蛋白表达。构建pcDNA3.1-Foxp3重组质粒,采用FuGENEHD瞬时转染LLC细胞,实验分3组:LLC组,pcDNA3.1-LLC组及pcDNA3.1-Foxp3-LLC组。RT-PCR和免疫组化方法分别检测转染48小时后Foxp3基因的表达,通过淋巴细胞转化试验检测Foxp3过表达对T淋巴细胞增殖的影响,RT-PCR和ELISA方法分别检测Foxp3过表达对TGF-β1和IL-10的mRNA表达及蛋白分泌的影响。结果:在LLC细胞中检测到Foxp3mRNA及蛋白的表达;瞬时转染48小时后pcDNA3.1-Foxp3-LLC组LLC细胞中Foxp3表达增高,与LLC组和pcDNA3.1-LLC组相比差异显著(P<0.01);pcDNA3.1-Foxp3-LLC组LLC细胞及培养上清对T淋巴细胞增殖的抑制率均明显高于LLC组和pcDNA3.1-LLC组(P<0.05);pcDNA3.1-Foxp3-LLC组TGF-β1、IL-10mRNA和蛋白的表达水平明显高于LLC组和pcDNA3.1-LLC组(P<0.05)。结论:LLC细胞表达Foxp3,Foxp3可能通过上调TGF-β1和IL-10的表达来抑制T淋巴细胞的增殖,进而参与肿瘤的免疫逃逸。Objective：To detect the expression of Foxp3 in Lewis lung cancer（LLC）cells and analyze the proliferation of T lymphocytes and the secretion of the inhibitory molecules TGF-β1 and IL-10 in Foxp3 overexpressing LLC cells and to study the function of Foxp3 in immune escape of the tumor and the possible mechanism.Methods：Detection of Foxp3 mRNA was performed by using RT-PCR,while protein expression was assessed by Immunohistochemistry（IHC）.The recombinant eukaryotic expressing plasmid pcDNA3.1-Foxp3 was constructed to transfect LLC cells transiently by FuGENE HD and there were three groups including LLC group,pcDNA3.1-LLC group and pcDNA3.1-Foxp3-LLC group.The expression of Foxp3 in transfected LLC cells was detected by applying RT-PCR and IHC.The proliferation of T lymphocytes was detected by lymphocyte transformation test and the expression of TGF-β1 and IL-10 was detected by using RT-PCR and ELISA kits.Results：Foxp3 mRNA and protein were detected in LLC cells.The expression of Foxp3 was significantly higher in pcDNA3.1-Foxp3-LLC group than that of in LLC group and pcDNA3.1-LLC group（P〈0.01）.The inhibitory rate of T cell proliferation for pcDNA3.1-Foxp3-LLC group was much higher than that of LLC group and pcDNA3.1-LLC group（P〈0.05）.The mRNA and protein expression of TGF-β1 and IL-10 markedly increased in pcDNA3.1-Foxp3-LLC group than that of in LLC group and pcDNA3.1-LLC group（P〈0.05）.Conclusion：LLC cells express Foxp3.Foxp3 in LLC cells could inhibit the proliferation of T lymphocytes,probably by up-regulating inhibitory molecules TGF-β1 and IL-10 and then may play an important role in tumor immune escape.

关 键 词：FOXP3 真核表达载体 基因克隆 转染 转化生长因子Β1 白介素10 

分 类 号：R730.32[医药卫生—肿瘤]