gp96多肽复合物介导的细胞毒性T淋巴细胞对食管腺癌细胞的免疫杀伤作用

目的:研究肿瘤细胞来源的gp96多肽复合物介导的对同一类型肿瘤的免疫治疗作用.方法:提取纯化食管腺癌细胞系SEG-1裸鼠成瘤组织的gp96蛋白多肽复合物,与外周血单个核细胞(PBMNC)诱导培养的树突状细胞(DC)结合,制备gp96-DC疫苗;台盼蓝拒染法测定淋巴细胞增殖率;ELISA方法检测细胞毒性T淋巴细胞(CTL)培养上清液的γ干扰素(IFN-γ)含量,MTT法检测CTL对靶细胞SEG-1的杀伤率.结果:55 g瘤组织提取纯化gp96蛋白120μg;单独DC、单独gp96及gp96-DC均能刺激淋巴细胞增殖,产生CTL,释放IFN-γ,对靶细胞SEG-1均显示一定的杀伤作用,以gp96-DC的作用最明显,在效靶比40:1时,杀伤率为68%.单独DC诱导的CTL对SEG-1、K562的杀伤作用与非抗原刺激的淋巴细胞比较,统计学差异无显著性.结论:肿瘤来源的热休克蛋白gp96可使DC具有更强的刺激T淋巴细胞增殖的能力,所产生的CTL对靶肿瘤细胞有明显的特异杀伤作用.

The killing effect of cytotoxic T lymhpocytes on esophageal adenocarcinoma cells mediated by gp96-peptide complexes

OBJECTIVE: To study the immunotherapeutic effect on the esophageal adenocarcinoma mediated by gp96-peptide complexes isolated from the same kind of tumor. METHODS: gp96-peptide complexes were purified from nude mice tumors burdened by subcutaneous injection of human esophageal adenocarcinoma cell line SEG-1. gp96-peptide complexes were carried by the dendritic cells(DC) induced from human peripheral blood mononuclear cells to prepare gp96-DC vaccine. The proliferation of lymphocytes was tested with trypan-blue stain. The quantity of interferon-gamma(IFN-gamma) released from cytotoxic T lymphocytes (CTL) was detected with ELISA method. The killing effect of CTL on target cell SEG-1 was measured with MTT. RESULTS: We obtained 120 microg gp96 from 55 g tumor tissue. DC, gp96, and gp96-DC all could elicit the proliferation of lymphocytes and make them becoming into CTL which released IFN-gamma and showed different degrees of killing effect on target cell SEG-1. gp96-DC has the strongest eliciting effect among them. At the ratio of E(effect) to T(target) as 40:1,the killing rate was 68%. No significant difference between the effects of CTL induced by DC alone and of lymphocytes without specific antigen on SEG-1 and K562 cells. CONCLUSION: The gp96-peptide complexes from tumors can improve the effect of eliciting lymphocyte proliferation of DC and make the lymphocyte becoming into CTL more effectively. These CTLs show prominent killing effect on the target tumor cells.