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重组B7 |
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黄贵清 李成荣 赵晓东 杨锡强 蒋莉萍
【摘要】 目的 探索转B7-1基因治疗白血病的可能性。方法 通过逆转录多聚酶键反应(RT-PCR)获得人B7-1 cDNA，经酶切后克隆至逆转录病毒载体PLXSN，用磷酸钙沉淀法将其导入包装细胞PA 317，获高效表达B7-1的重组病毒，用重组病毒感染人白血病细胞株K 562和HL 60，获得稳定表达B7-1分子的肿瘤细胞，将其与人外周血单个核细胞(PBMC)混合培养，检测3HTdR掺入和细胞因子白细胞介素2(IL-2)和γ-干扰素(INF-γ)的表达。结果 B7+K 562、B7+-1HL60较载体修饰K 562(neo-K 562)、野生型K 562(Wild-K 562)和nco-HL-60、Wild-HL-60组HTdR掺入明显增强，B7+K 562组为3 680 cpm、neo-K 562组为645 cpm、Wild-K 562组为866 cpm、B7+HL60组为3 980 cpm、neo-HL 60组为486 cpm、Wild-HL-60组为742 cpm。转B7-1基因的白血病细胞能诱导PBMC产生IL-2及IFN-γ。结论 转B7-1基因的肿瘤细胞能促进T细胞增殖和分化，推测可能进一步诱导细胞毒性T淋巴细胞杀伤活性，为在临床开展转B7-1基因治疗小儿白血病打下基础。
【关键词】 逆转录病毒科 白血病 T淋巴细胞 抗原，CD80
Construction of the recombinant B7-1 retroviral vector and its effect on the function of T cells HUANG Guiqing, LI Chengrong, ZHAO Xiaodong, et al. Children′s Hospital, Chongqing Medical University, Chongqing 400014
【Abstract】 Objective To explore the possibility of transferring B7-1 gene for the treatment of leukemia. Methods B7-1 cDNA was obtained by reverse transcription polymerase chain reaction and recombined to retroviral vector PLXSN. Calcium phosphate-DNA precipitation method was used to transfer PLB 7-1 SN into packaging cells PA317. Human leukemia cells K 562 and HL-60 were infected with higher titer virus. B7-1 protein expression was confirmed by FACS technique. After PBMCs from healthy donor were cultured with B7-1+ K 562 or B7-1+ HL60, 3 HTdR incorporation and IL-2 and IFN-γ expression were determined. Results A significantly increased 3HTdR incorporation in B7-1+ leukemia cell groups was observed (3 680 cpm in K 562 and 3 980 cpm in HL60, respectively) as compared with those in B7 leukemia cell groups (645 cpm in neo-K 562, 866 cpm in Wild-K 562 and 486 cpm in neo-HL-60, 742 cpm in Wild-HL-60). Interleukin-2 and Interferon-γ expressions were found in PBMC cultured with B7-1 transfected K 562 and HL-60 cell lines, but not found in those cultured with wild or neo-ce[1] [2] [3] 下一页
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