肿瘤的热休克蛋白90-肽复合物诱导产生特异性细胞毒性T淋巴细胞的体外研究

Generation of specific cytotoxicity T lymphocytes induced by tumor-derived heat shock protein 90-peptide complexes in vitro

目的 探讨肿瘤组织来源的热休克蛋白 90 肽复合物 (HSP90 PC)诱导产生特异性细胞毒性T淋巴细胞 (CTL)的作用。方法 高压液相色谱法分离纯化肾腺癌患者肿瘤组织HSP90 PC ,同时从该患者外周血扩增树突状细胞及T细胞 ,以提取的HSP90 PC冲击树突状细胞 ,将致敏的树突状细胞与T细胞混合 ,流式细胞仪检测特异性CD8+ CTL的增殖情况。结果 肿瘤组织来源的HSP90 PC致敏的树突状细胞可显著诱导T细胞增殖生成CD8+ CTL。结论 从肿瘤组织中提取的HSP90 PC具有良好的免疫原性 ,用其致敏的树突状细胞可有效诱导CTL增殖 。

Objective To investigate the specific induction of cytotoxic lymphocyte (CTL) by tumor derived heat shock protein 90 peptide complexes (HSP90 PC). Methods Heat shock protein 90 peptide complex (HSP90 PC) was isolated and purified by liquid chromatography after precipitation by 50% 70% (NH4)2SO4 saturation from 10 specimens of renal carcinoma resected from 10 patients aged 40 60 during operation. The component containing HSP90 PC was filtered and sterilized. The molecular weight and the identity of the purified HSP90 PC were confirmed by SDS PAGE and Western blotting. 10 15 ml peripheral blood was extracted from these patients. T cells were amplified. Flow cytometry was used to detect the phenotype of dendritic cells (DCs). The DCs in the experimental group were cultured for 5 days and then HSP90 PC and tumor necrosis factor (TNF) α was added into the culture. The HSP90 PC pulsed DCs were collected and co cultured with auto T cells for 72 hours. Flow cytometry was used to detect the content of CD8 +T cells. The DC of the control group were mixed directly with auto T cells and the content of CD8 +T cells was examined by flow cytometry. Results The proliferation of T cells co cultured with the HSP90 PC pulsed DCs was significantly remarkable and the content of CD8 + CTLs was significantly more in comparison with the control DC ( P <0.01). Conclusion HSP90 PC prepared from tumor tissues has strong immunogenicity and the DC sensitized thereby effectively induces the proliferation of CTL. Application of HSP90 PC provides a new approach in tumor immunotherapy.