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目的观察我们以前构建的端粒酶反转录酶启动子调控的葡萄球菌肠毒素A(SEA)/CD80基因重组腺病毒载体对肝癌的疗效和诱导的免疫学效应。方法重组腺病毒采用瘤体内直接注射的方式对小鼠皮下移植性肝癌进行治疗,采用RT-PCR和Western blot方法检测腺病毒注射部位的SEA和CD80 mRNA和蛋白的表达情况;采用ELISpot方法和LDH释放实验分别检测脾脏淋巴细胞中肝癌特异性IFN-γ分泌细胞的频数和细胞毒性T细胞(CTLs)对Hepa1-6细胞的特异杀伤活性;通过观察荷瘤小鼠经治疗后肿瘤体积的变化及生存时间,评价重组腺病毒对肝癌的治疗作用。结果我们构建的腺病毒能够使SEA和/或CD80mRNA和蛋白靶向地在肝癌组织中表达;与空载体组和PBS对照组相比,双基因组和单基因组分泌IFN-γ的T细胞数量均明显增多,CTL对Hepa1-6细胞的特异性杀伤作用均明显增强,荷瘤小鼠肿瘤体积明显减小,生存期明显延长;双基因组的疗效和对免疫系统的激活作用明显高于单基因组。结论我们制备的肿瘤靶向性重组腺病毒对肝癌有良好的治疗作用,联合基因治疗优于单个基因治疗。
Objective To observe the curative effect and induced immunological effect of the recombinant adenovirus vector of staphylococcal enterotoxin A (SEA) / CD80 gene regulated by telomerase reverse transcriptase promoter constructed in the past. Methods The recombinant adenovirus was directly injected into the tumor in mice to treat subcutaneous liver cancer. RT-PCR and Western blot were used to detect the expression of SEA and CD80 mRNA and protein at the site of injection. The ELISpot method and LDH The frequency of hepatocarcinoma-specific IFN-γ secreting cells in spleen lymphocytes and the specific cytotoxic T lymphocytes (CTLs) specific cytotoxic activity against Hepa 1-6 cells were detected by the release assay. The tumor volume of tumor-bearing mice was observed. Survival time, evaluate the therapeutic effect of recombinant adenovirus on liver cancer. Results The constructed adenovirus could express SEA and / or CD80 mRNA and protein in hepatocellular carcinoma. Compared with the empty vector group and the PBS control group, the numbers of T cells secreting IFN-γ in the two groups were significantly higher The specific killing effect of CTL on Hepa1-6 cells was significantly enhanced. The tumor volume of tumor-bearing mice was significantly reduced and the survival time was significantly prolonged. The efficacy and immune system activation of the double-gene group was significantly higher than that of the single genome. Conclusion We have prepared a tumor-targeting recombinant adenovirus for the treatment of liver cancer has a good effect, combined gene therapy is better than a single gene therapy.