论文部分内容阅读
目的:研究小鼠T淋巴细胞在转染嵌合性T细胞受体基因后的体外抗肿瘤作用。方法:应用重组DNA技术,将HER2特异性嵌合性T-细胞受体构建入逆转录病毒载体;转入包装细胞后,收集病毒上清,转染小鼠T淋巴细胞,转基因后的小鼠T淋巴细胞分别与HER2阳性(SK-OV-3)或阴性(MCF-7)的肿瘤细胞系共培养,检测其细胞因子γ干扰素释放,51Cr释放法检测CTL评价其抗肿瘤效应。结果:所构建载体经酶切鉴定符合要求,乒乓法转染包装细胞系GP+E86,检测病毒滴度为1.2×106,Retronectin结合离心法转染经抗CD3/CD28单抗活化的小鼠T淋巴细胞,转染效率可达50%以上;转染嵌合性T细胞受体基因的T淋巴细胞与HER2阳性或阴性的肿瘤细胞系共培养后可检测到HER2特异性的细胞因子γ干扰素释放,51Cr释放法测CTL可见转染嵌合性T细胞受体基因T淋巴细胞对HER2阳性的肿瘤细胞具显著杀伤效应。结论:转染嵌合性T细胞受体基因的小鼠T淋巴细胞在体外可通过细胞因子释放和CTL效应发挥显著的抗肿瘤作用。
Objective: To study the anti-tumor effect of mouse T lymphocytes in vitro after transfection of chimeric T cell receptor gene. Methods: Recombinant DNA technology was used to construct HER2-specific chimeric T-cell receptors into retroviral vectors. After transfection into packaging cells, the virus supernatants were collected and transfected into mouse T lymphocytes. Transgenic mice T lymphocytes were co-cultured with HER2-positive (SK-OV-3) or negative (MCF-7) tumor cell lines respectively to detect the release of cytokine interferon-gamma. 51Cr release assay was used to evaluate the antitumor effect of CTL. Results: The constructed vector was identified by restriction enzyme digestion. The packaging cell line GP + E86 was transfected by ping-pong method. The titer of virus was 1.2 × 106. The T cells activated by anti-CD3 / CD28 mAb were transfected by Retronectin T lymphocytes transfected with chimeric T cell receptor gene can detect HER2-specific cytokine interferon-gamma after co-cultured with HER2 positive or negative tumor cell lines Release, 51Cr release assay CTL visible transfection of chimeric T cell receptor gene T lymphocytes on HER2 positive tumor cells with a significant killing effect. CONCLUSION: Mice T lymphocytes transfected with chimeric T cell receptor gene play a significant anti-tumor effect in vitro by cytokine release and CTL effect.