【摘要】 建立稳定表达CIITA基因的人卵巢癌细胞株HO CIITA ,分析转染CIITA基因对T细胞体外抗肿瘤免疫应答的影响。以CIITA基因逆转录病毒 (pLXSN/CIITA )转染并筛选获得HO CIITA。用FACS和RT PCR分析HLA以及抗原加工递呈基因表达水平。以磁珠法分离得到正常人外周血CD4 +/CD8+T细胞 ,分别进行混合淋巴细胞反应及细胞因子测定 (ELISA和RT PCR )。结果显示 ,转染CIITA基因后 ,使HO细胞HLAII类分子和LMP7基因表达增高 ,而Ii基因表达由阳性转为阴性 ,未检测到TAP1表达 ;HO和HO CIITA细胞刺激CD4 +T细胞分泌IL 4含量两者有显著差异。刺激 4 8h后达到顶峰 ,前者分泌量约为后者的 1/2 ,但分泌IFN γ无差异 ,RT PCR与ELISA两种测定结果一致。表明转染CIITA基因可增加肿瘤细胞表面MHCII类分子的表达 ,该作用与IFN γ具有协同效应 ;并能诱导CD4 +T细胞表达IL 4。更多还原

【Abstract】 A human ovarian cancer cell line,HO CIITA constantly expressing the CIITA gene was established and selectively cloned after transfection with pLXN/CIIYA retrovirus The expression levels of HLA molecules and antigen processing associated genes were detected with flow cytometry and RT PCR respectively The CD4 +/CD8 + T cell populations were separated from human PBL by means of magnetic beads, mixed lymphocyte reaction (MLR) between CD4 +T cells and the HO CIITA cell line was performed as usual and ELISA and RT PCR were used to analyze the expressions of IL 4 and IFN γ from the supernatants of cell cultures Experimental results showed that the expressions of HLA II molecules on the HO cells as well as that of the LMP 7 gene were elevated after transfection with the CIITA gene,but the expression of the Ii gene turned from positive to negative No expression of TAP 1 could be found On MLR,it was found that both HO and HO CIITA cells could stimulate the secretion of IL 4 with peak after stimulation for 48 hours,but the concentration of IL 4 was 1:2 between HO and HO CIITA cells There was no obvious change in the secretion of IFN γ Results of RT PCR were in accordance with those of ELISA These results suggest that the surface expression of MHC II antigens in tumor cell line could be enhanced by transfection with CIITA gene with a synergistic effect of IFN γ and an inducing effect for CD4 +T cells to secrete IL 4更多还原