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目的观察经SEA和CD80重组腺病毒感染Hepa1-6肝癌细胞后体外能否诱导抗肿瘤免疫学反应。方法 Hepa1-6细胞分别经空载体腺病毒Ad(空)和重组腺病毒Ad-MMRE-mTERT-B7、Ad-MMRE-mTERT-SEA、Ad-MMRE-mTERT-BIS感染后,和小鼠脾淋巴细胞共培养,然后采用Brdu酶联免疫法(ELISA)检测淋巴细胞增殖;流式细胞术检测T淋巴细胞亚群增殖;ELISA法检测细胞因子IL-2、TNF-α和IFN-γ的产生;LDH释放法检测CTLs对Hepa1-6的杀伤作用。结果与感染空载体腺病毒Ad(空)和未感染Hepa1-6细胞相比,经重组腺病毒感染的Hepa1-6细胞体外能够显著诱导脾淋巴细胞的增殖,其中CD3+CD4+和CD3+CD8+T淋巴细胞增殖显著;细胞因子TNF-α、IFN-γ和IL-2的产生显著升高;CTLs对肿瘤细胞的杀伤活性显著增强;双基因过表达Hepa1-6细胞体外诱导的免疫应答高于单基因过表达Hepa-6细胞。结论 Hepa-6细胞经重组腺病毒感染后,表达在细胞膜上的SEA和CD80均具有免疫学活性,为今后采用所构建重组腺病毒进行肿瘤的免疫基因治疗提供了实验依据。
Objective To observe whether in vitro induction of antitumor immunological response to Hepa1-6 hepatoma cells transfected with SEA and CD80 recombinant adenovirus. Methods Hepa1-6 cells were infected with Ad-MMRE-mTERT-B7, Ad-MMRE-mTERT-SEA and Ad-MMRE- The proliferation of T lymphocytes was detected by flow cytometry. The production of cytokines IL-2, TNF-α and IFN-γ was detected by ELISA. The proliferation of T lymphocyte subsets was detected by Brdu enzyme-linked immunosorbent assay (ELISA) LDH release assay CTLs killing Hepa1-6. Results Hepa1-6 cells infected with recombinant adenovirus could significantly induce the proliferation of splenic lymphocytes in vitro compared with those of Ad (empty) and Hepa1-6 infected with empty vector, in which CD3 + CD4 + and CD3 + CD8 + The proliferation of T lymphocytes was significant. The production of cytokines TNF-α, IFN-γ and IL-2 were significantly increased. The killing activity of CTLs to tumor cells was significantly increased. The immune response induced by double gene Hepa1-6 cells in vitro was higher than that of Single gene overexpression Hepa-6 cells. CONCLUSIONS: Hepa-6 cells have immunological activity of SEA and CD80 expressed on the cell membrane after the recombinant adenovirus is infected, which provides an experimental basis for the future immunotherapy of tumor with the constructed recombinant adenovirus.