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目的 用逆转录病毒载体将CD80 基因导入人卵巢癌细胞系TYK ,观察表达CD80 基因的TYK(TYK hCD80 )细胞体外诱导细胞毒T淋巴细胞 (CTL)的增殖及其杀瘤活性。方法 用逆转录病毒载体将CD80 基因导入TYK细胞 ,流式细胞仪测定其CD80 基因的表达。用TYK hCD80 细胞在抗CD3 单克隆抗体 (单抗 )存在下刺激外周血单个核细胞 (PBMC) ,诱导CTL ,3 H 胸腺嘧啶核苷掺入法测定其增殖活性 ,四甲基偶氮唑蓝法测定CTL的杀瘤活性。结果 转染后经G418(geneticin ,是一种氨基糖甙类抗生素 )筛选 ,流式细胞仪检测 ,CD80 基因表达率最高为 84 9%。TYK hCD80 、TYK在抗CD3 单抗存在下刺激PBMC增殖时 ,3 H 胸腺嘧啶核苷掺入量分别为 (4 0 6 0 4± 842 )、(80 0 0± 5 94)cpm ,两者比较 ,差异有显著性 (P <0 0 5 )。TYK hCD80 体外诱导的CTL较TYK诱导者对TYK细胞的杀伤率显著增高(P <0 0 5 )。结论 表达CD80 基因的卵巢癌细胞在抗CD3 单抗存在下能诱导产生CTL ,增殖快且有较高的杀伤活性 ,可能为卵巢癌的免疫基因治疗提供实验依据
Objective To introduce CD80 gene into human ovarian cancer cell line TYK by retroviral vector and observe the proliferation and cytotoxicity of cytotoxic T lymphocyte (CTL) induced by TYK (TYK hCD80) cells expressing CD80 gene in vitro. Methods CD80 gene was transduced into TYK cells by retroviral vector and the expression of CD80 gene was determined by flow cytometry. Peripheral blood mononuclear cells (PBMCs) were stimulated with TYK hCD80 cells in the presence of anti-CD3 monoclonal antibody (mAb), and CTL and 3 H thymidine incorporation assay were used to determine the proliferative activity. Determination of cytotoxic activity of CTL. Results After transfected with G418 (geneticin, an aminoglycoside antibiotic), the highest expression rate of CD80 gene was 84.9% by flow cytometry. When TYK hCD80 and TYK stimulated the proliferation of PBMC in the presence of anti-CD3 mAb, the incorporation of 3 H thymidine was (40 06 ± 842) and (80 0 0 ± 5 94) cpm, respectively , The difference was significant (P <0 05). The killing rate of TYK hCD80 induced by CTK in TYK cells was significantly higher than that induced by TYK (P <0.05). Conclusion The ovarian cancer cells expressing CD80 gene can induce the production of CTL in the presence of anti-CD3 mAb, with rapid proliferation and high killing activity, which may provide experimental evidence for immunological gene therapy of ovarian cancer