p53基因修饰树突细胞诱导特异性抗肿瘤免疫应答的实验研究  

Experimental study of the specific immunoresponse induced by p53 modified dendritic cells

作　　者：王长利[1] 尹志伟[1] 任秀宝[1] 刘虹[1] 

机构地区：[1]天津医科大学附属肿瘤医院,300060

出　　处：《中华血液学杂志》2003年第12期632-635,共4页Chinese Journal of Hematology

摘　　要：目的　探讨含p5 3基因的质粒pC5 3 SN3转染的树突细胞 (DC)体外诱导特异性抗肿瘤细胞毒性T淋巴细胞 (CTL)。方法　应用脂质体介导将质粒pC5 3 SN3转染肺癌患者单个核细胞 [HLA A2 + ]衍生的DC ,然后与未经纯化的自体T细胞混合培养以诱导CTL(T pC5 3 SN3) ,并通过乳酸脱氢酶(LDH)释放实验测定其对p5 3基因突变的HLA A2 + 人肺癌细胞系Calu 6的杀伤活性。结果　用质粒pC5 3 SN3转染DC后 ,发现CD1a、CD83显著升高 ,转染前表达率分别为 (5 .4 5± 0 .89) %、(3.2 6± 0 .4 7) % ,转染后分别为 (5 2 .15± 11.5 6 ) %、(2 5 .78± 12 .35 ) %。但CD86 、CD4 0 、HLA DR等DC相关标志与转染前无明显改变。LDH释放实验显示 ,以Calu 6作为靶细胞 ,T pC5 3 SN3诱导的杀伤作用显著高于T IL 2(IL 2 10 0U ml刺激外周血单个核细胞产生的CTL) ,效靶比为 10∶1时其杀伤活性分别为 (5 6 .79±15 .6 7) %和 (39.33± 9.88) %。同时细胞表面标记检测可见T pC5 3 SN3细胞中以CD8+ 细胞为主 ,且CD6 9、CD4 5RO CD8表达均显著升高。结论　p5 3基因修饰的DC对p5 3突变的人肺癌细胞系可有效诱导HLA A2限制的特异性CTL。Objective To explore the specific cytotoxic T lymphocyte (CTL) induced by dendritic cells(DC), which were transfected by the plasmid pC53-SN3 encoding p53 gene. Methods DC derived from HLA-A2+ mononuclear cells of the 24-lung cancer patients was transfected with the plasmid pC53-SN3 by lipofectamine and then co-cultured with auto-unpurified T cells to induce potent CTL(T-pC53-SN3). The cytolysis of specific CTL against Calu-6, a HLA-A2+ human lung cancer cell line, was measured by using lactate dehydrogenase (LDH) releasing assay. Results The expression of CD 1a and CD 83, the correlative markers of DC,increased apparently after transfected with plasmid pC53-SN3, the expression rate was (5.45±0.89)% and (3.26±0 47)% versus (52.15±11.56)% and (25.78±12.35)%. CD 14 decreased apparently, but other DC correlative markers of CD 1a,CD 40, CD 86, and HLA-DR remained almost the same as that before transfection. Compared with T-IL-2, the CTL derived from PBMNC stimulated by IL-2 (100 U/ml), the cytolytic activity of T-pC53-SN3 against Calu-6 cell line showed a significant increase, but cytolytic activity was 56.79±15.67 and 39.33±9.88,respectively, when effect cells∶target cells was 10∶1. The expression of the CD 8, CD 69,and CD 45RO/CD 8 of T-pC53-SN3 cells increased significantly, but that of CD 3, CD 4, CD 86, ect, was not significantly different from those of T-pCMV-neo. Conclusions It showed that DC transfected by p53 gene could induce potent HLA-A 2 restrictive CTL to kill tumor cell efficiently.

关 键 词：P53 基因修饰 树突细胞 诱导 特异性抗肿瘤免疫应答 实验研究 肺肿瘤 

分 类 号：R73-36[医药卫生—肿瘤]