肿瘤特异性T细胞受体基因转染诱导记忆性T细胞体外分化的研究

目的 探讨肝癌特异性T细胞受体基因转染对记忆性T细胞体外诱导分化的影响.方法 以肝癌特异性T细胞受体V β7.1转染PBMC,流式细胞术检测目的基因表达.肝癌细胞BEL-7402刺激诱导记忆性T细胞;以流式细胞术检测记忆性T细胞标志性表面分子表达验证记忆性T细胞表型及分化亚群:以Annexin V-PI双染检测肿瘤细胞的凋亡,ELISA检测细胞因子IFN-γ分泌探讨记忆性T细胞免疫功能.结果 TCRVβ7.1基因成功转染到T细胞并得到有效表达.与PBMC对照组相比.TCRVβ37.1转染组CD45RO表达显著上升.以CD45RA及CCR7表达分析记忆性T细胞亚群显示其表型主要为效应型记忆性T细胞.分化后的记忆性T细胞诱导肿瘤细胞凋亡率及IFN-γ分泌显著上升.结论 肿瘤特异性TCR基因转染可促进以效应型记忆性T细胞(TEM)为主的记忆性T细胞分化.TEM将通过诱导凋亡与分泌细胞因子作用发挥其免疫功能.

Tumor-specific T cell recptor gene transfection promotes memory T cell differentiation in vitro

OBJECTIVE: To investigate effect of tumor-specific T cell receptor gene transfection on memory T cell differentiation in vitro. METHODS: TCRVbeta7.1 gene was transferred into peripheral blood mononuclear cells (PBMCs) obtained from healthy adults, and the expression of Vbeta7.1 was detected by flow cytometry before and after the transfection. Memory T cell differentiation was induced by stimulation with the hepatocarcinoma cell line BEL-7402 in vitro. The expression of surface molecules CD45RO, CD45RA and CCR7 was analyzed by flow cytometry to identify the phenotype and subsets of the memory T cells. Fluorescence-activated cell sorting was performed to detect the apoptosis of the tumor cells, and enzyme-linked immunoabsorbent assay was used to determine the production of interferon-gamma (IFN-gamma) for assessing the immune function of the memory T cells. RESULTS: Flow cytometry showed that TCRVbeta7.1 gene was efficiently expressed after transfection. After stimulation by the tumor cells in vitro, the expression of CD45RO in TCRVbeta7.1 gene-modified T cells increased gradually, and analysis of the coexpression of CD45RA and CCR7 revealed that the effector memory T cells constituted the majority of the differentiated memory T cells. The apoptotic rate of the tumor cells induced by the T cells increased significantly with also obviously increased INF-gamma secretion in the memory T cells. CONCLUSION: Tumor-specific TCRVbeta7.1 gene transfection can promote the differentiation of the memory T cells, the majority of which belongs to effector memory T cells that perform immune functions by inducing apoptosis and cytokine secretion.