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将小鼠肿瘤坏死因子(mTNF)cDNA克隆到逆转录病毒载体MNSM构建MNSM-SV40-mTNF,用小鼠白蛋白增强子/启动子Alb e/p元件置换MNSM-SV40-mTNF中SV40启动子构建成MNSM-Alb e/p-mTNF。构建物经脂质体转染法引入包装细胞PA137中制备重组病毒,在Polybrene存在条件下感染小鼠肿瘤细胞,经RNA和TNF活性分析证明Alb e/p可驱动TNF基因在合成白蛋白的小鼠肝癌细胞中特异转录和表达,转TNFα基因小鼠肝癌细胞体内致瘤性消失,体内特异抑制亲代肝癌生长。重组病毒及产病毒PA317于肝癌瘤体内显著抑制肝癌生长,特异地延长荷肝癌小鼠存活期。病理及免疫组化分析证实in situ基因治疗后,肝癌广泛坏死、出血并伴有大量Mac-11~+、CD_8~+和CD_(25)~+炎细胞浸润和纤维化。
Mice tumor necrosis factor (mTNF) cDNA was cloned into retroviral vector MNSM to construct MNSM-SV40-mTNF, and the mouse albumin enhancer/promoter Alb e/p element was used to replace SV40 promoter in MNSM-SV40-mTNF. To MNSM-Alb e/p-mTNF. The recombinants were introduced into the packaging cell PA137 by liposome transfection and the recombinant virus was infected. In the presence of Polybrene, the tumor cells were infected. The RNA and TNF activity analysis demonstrated that Alb e/p could drive the TNF gene in the synthesis of albumin. Specific transcription and expression in rat hepatoma cells, tumor necrosis of hepatoma cells transfected with TNFα gene in vivo disappeared, and the growth of parental hepatoma was specifically inhibited in vivo. The recombinant virus and virus-producing PA317 significantly inhibited the growth of hepatocellular carcinoma in hepatocellular carcinoma and specifically extended the survival period of the hepatocellular carcinoma-bearing mice. Pathological and immunohistochemical analysis confirmed extensive necrosis and hemorrhage of hepatocellular carcinoma with invasive and fibrotic Mac-11+, CD8+ and CD25+ inflammatory cells after in situ gene therapy.