HSP70多肽刺激PBMCs体外扩增γδT细胞及其抗肿瘤效应的研究

Establishment of a novel γδT cell expansion culture system and studies on its anti tumor properties

运用分离提纯所获的肿瘤膜蛋白ＨＳＰ７０－多肽在体外刺激人外周血单个核细胞（ＰＢＭＣｓ），成功地建立了γδＴ细胞的扩增培养系统。扩增培养后γδＴ细胞从占外周血Ｔ淋巴细胞的３．５％±２．８％（健康人样本）、１１．５％±１０．８％（急性白血病样本）相应地升高至５２．８％±７．９％和６０．４％±５．９％，并以表达ＴＣＲＶγ９－Ｖδ２亚型为主。运用５１Ｃｒ细胞毒试验探讨了γδＴ细胞的杀伤效应，发现：ＨＳＰ７０－多肽激活的γδＴ细胞对一系列肿瘤细胞包括ＮＫ敏感的Ｋ５６２、ＬＡＫ敏感的Ｄａｕｄｉ以及自体或异体新鲜肿瘤细胞等都具有高效的细胞毒活性，另外，在γδＴ细胞上清中可检出高水平的ＩＦＮ－γ、ＩＬ－２、ＴＮＦ、ＩＬ－８等细胞因子。以上结果提示：γδＴ细胞作为一种细胞毒性Ｔ淋巴细胞具有高效、特异的杀伤效应，在肿瘤免疫中具有重要作用。

Despite intensive efforts,the functions of γδT cells in tumor immunity have remained undifined.We took advantage of the HSP 70 peptide,which was purified from XG 7 MM cells,to develop a novel γδT cell expansion culture system.And the cytotoxicity of HSP70 primed γδT cells was examined with 51 Cr release assay,light and electron microscope observations.The results showed:HSP70 derived from XG 7 could successfully induce proliferation of γδT cells,and HSP70 primed γδT cells exhibited a strong cytotoxicity aganist a series of tumor cells,such as NK sensitive cell K562,LAK sensitive cell Daudi,5637 and auto or allo fresh isolated tumor cells.The light and electron microscope observations showed that the target lysis could be clearly separated into three phases:1,a recognition phase,visible by random crawing of γδT cell over the target cell surface by cell to cell film interaction.2,a post recognition phase,during which firm contact between γδT cell and targets was maintained,and 3,a phase of target cells disintergration,mainly characterized by vigorous blebbing of the membrane resulting in a motionless carcass of the target cells but not in its total dissolution.Tumor cells apoptosis could be observed and a series of cytokines which showed antitumor effects were detected.Taking these results together,it can be concluded that γδT cells is a effective cytotoxicic T lymphocytes and there are potential implications in tumor immunotherapy.