CD86对CD8^+T细胞分化的影响  

Effect of CD86 on the differentiation of CD8^+T cell

作　　者：孙斌[1] 秦成勇[1] 王延军[1] 张国全[1] 

机构地区：[1]山东大学山东省立医院消化内科,山东济南250021

出　　处：《免疫学杂志》2003年第6期443-446,共4页Immunological Journal

摘　　要：目的　探讨CD86 (B7 2 )对CD8+ T细胞分化的影响。方法　用限制性内切酶XhoⅠ酶切质粒pCDM8得到CD86基因 ,并将其插入pCDNA3,用BamHⅠ酶切鉴定。用脂质体法介导pCDNA3 CD86真核表达载体转染人肝癌细胞株HMC772 1,6 0 0 μg mLG4 18筛选 ,稳定且高表达CD86的抗性克隆用流式细胞仪进行鉴定。从健康志愿者血中分离外周血单个核细胞 (PB MC) ,使PBMC与靶细胞之比为 2 0∶1,共同培养 4 8h后 ,用流式细胞仪检测CD3+ T细胞内IL 4和IFN γ的表达率。结果　成功构建pCDNA3 CD86真核表达载体 ;CD86在HMC772 1 CD86细胞中的表达率为 30 .8% ,而在HMC772 1细胞中的表达率为 0 .98% ;健康志愿者CD3+ T细胞内IL 4和IFN γ的表达率分别为 1.92 %和 2 4 .4 % ;PBMC与靶细胞共同培养 4 8h后 ,无论是否用IFN α刺激 ,IL 4 IFN γ的阳性比值在HMC772 1 CD86转染组均大于 1,而在HMC772 1未转染组均小于 1。结论　在细胞培养中 ,CD86可诱导CD8+ T细胞活化 。Objective To investigate the effect of CD86 (B7-2) on the differentiation of CD8 +T cell in vitro. Methods CD86 gene was cut out from plasmid pCDM8 by restriction endonuclease Xho Ⅰ. It was inserted into eukaryotic expression vector pCDNA3 and identified by restriction enzyme BamH Ⅰ. Further, the recombinant plasmid vector pCDNA3-CD86 was transferred into human hepatocellular carcinoma cell line HMC7721 with cationic liposome. After selected by 600 μg/mL G418, the G418-resistant clone expressing CD86 stably and highly was isolated by flow cytometry (FCM). Finally, human peripheral blood mononuclear cells (PBMCs) were separated from peripheral blood of healthy volunteer, and co-cultured with target cells at E/T ratio of 20. 48 h later, CD3 +T cells were assayed for the secretion of the intracellular cytokines IL-4 and IFN-γ using FCM. Results The recombinant plasmid vector pCDNA3-CD86 was constructed successfully. The CD86 expression rate in HMC7731-CD86 cell was 30.8%, but the rate in HMC7721 cell was 0.98%. The expression rate of IL-4 and IFN-γ were 1.92% and 24.4% in the CD3 +T cells of healthy volunteer. After PBMCs were co-cultured with target cells for 48 h, whether target cells were stimulated by IFN-α or not, the positive rate of IL-4/IFN-γ on CD3 +T cells was more than 1 in the cell line HMC7721-CD86 groups. In contrast, the positive rate of IL-4/IFN-γ was less than 1 in the cell line HMC7721 groups. Conclusions In cell culture, CD86 can induce CD8 +T cell activation and regulates Tc2 phenotype differentiation.

关 键 词：CD86细胞 CD8^+T细胞 细胞分化 基因转染 细胞培养 

分 类 号：R392.11[医药卫生—免疫学]