【摘要】 目的探讨自体树突状细胞(DC)与肾癌融合细胞疫苗诱导特异性抗肿瘤免疫应答。方法利用肿瘤细胞纯化技术,从手术切除的肾癌组织中分离出高纯度的肾癌细胞,用10%胎牛血清(FCS)的RPMI-1640进行原代培养。分离外周血单核细胞(PBMCs),在含重组人粒细胞-巨噬细胞集落刺激因子(rhGM-CSF)和白细胞介素4(rhIL-4)的培养条件下,诱导分化为DC。用细胞融合技术将DC与肾癌细胞融合制备DC与肾癌融合细胞疫苗。用流式细胞仪检测肾癌细胞、DC和融合细胞的表型;用噻唑盐(MTT)比色法检测融合细胞刺激自体T细胞的增殖能力,乳酸脱氢酶(LDH)释放的细胞毒实验检测融合细胞刺激的细胞毒性T淋巴细胞(CTLs)的杀瘤活性。结果DC高表达主要组织相融性复合体(MHC)Ⅰ类分子、MHCⅡ类分子和共刺激分子(CD86,CD80);原代肾癌细胞高表达一种高分子量的糖蛋白(MUC-1);自体DC与肾癌融合细胞疫苗同时表达肾癌细胞和DC细胞的表面抗原,并能有效刺激自体T细胞增殖,诱导产生的CTLs对自体肾癌细胞具有显著的杀伤活性。结论自体DC与肾癌融合细胞疫苗能诱导特异性抗自体肾癌细胞的免疫应答,为自体DC与肾癌融合细胞疫苗在治疗肾癌的临床应用中提供了实验依据。更多还原

【Abstract】 Objective To investigate whether dendritic cells fused with tumor cells could elicit in vitro antitumor responses against renal cell carcinoma (RCC) cells. Methods Renal carcinoma cells were purified from tumor tissue excised from patients with metastatic RCC through tumor cell purifying technique and cultured in RPMI-1640 medium containing 10% FCS. Monocyte-derived DCs generated from peripheral blood mononuclear cell of RCC patients were cultured in the presence of human recombinant granulocytemacrophage colony stimulating factor and interleukin-4.Tumor cells and DCs were cocultured in the presence of polyethylene glycol (PEG) to generate cell fusion. The phenotype of tumor cells, DCs and fusion cells were detected by flow cytometry. MTT was used to measure the ability of fusion cells to stimulate T cell proliferation. T cell-mediated antitumor responses were measured by lactate dehydrogenase release (LDH) assay for lysis of autologous tumor cells. Results The DCs expressed MHC classⅠ, MHC classⅡand costimulatary molecules (CD80 and CD86) , while the renal carcinoma cells expressed a high molecular glycoprotein MUC-1.The DC/tumor fusion cells coexpressed MUC-1 and the phenotype of DCs, and could stimulate T cell proliferation effectively. CTLs stimulated by the fusion vaccine showed distinct lytie activity in vitro to autologous tumor cells. Conclusion Dendritic cells fused with tumor cells can elicit distinct antitumor responses in vitro against tumor cells from patients with metastatic RCC, providing a basis for further research on the clinical application of fusion vaccine in treatment for renal cancers.更多还原