携带SEA和CD80双基因的重组腺病毒转染B16细胞体外诱导T淋巴细胞活化的研究

摘要

Objective:To observe whether Staphylococcus enterotoxin A( SEA)and CD80 expressing on the surface of B16 cells could induce immune response after infection with recombinant adenoviruses. Methods:Mouse spleen lymphocytes were cocultured with B16 cells after infection with empty adenoviruse vector Ad( empty)or the recombi-nant adenoviruses of Ad-MMRE-mTERT-B7,Ad-MMRE-mTERT-SEA or Ad-MMRE-mTERT-BIS. Then,lymphocyte proliferation was assayed by BrdU incorporation using a commercial kit,flow cytometric analysis was used to detect proliferation of T lymphocyte subgroups. The production of IL-2,TNF-α and IFN-γ was tested by enzyme-linked immunosorbent assay. Results:Compared with B16 infected with empty adenovirus or without infec-tion,B16 infected with recombinant adenoviruses could significantly induce proliferation of spleen lymphocytes and T lymphocyte subgroups,and increase IL-2,TNF-αand IFN-γproduction. The antitumor response was significantly stronger in B16 cells after dual-gene expression than that in B16 cells after single-gene expression. Conclusion:The results indicate that SEA and CD80 expressed on the membrane of B16 cells after infection with recombinant ade-novirus has immune activities. This study provides some experimental evidence for further immune genetherapy against malignant melanoma with the recombinant adenovirus.%目的：观察恶性黑色素瘤B16细胞经重组腺病毒感染后，表达在B16细胞膜上的SEA和CD80体外能否诱导免疫学反应。方法：B16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。结果：与感染空载体腺病毒Ad（空）和未感染B16细胞相比，经重组腺病毒感染的B16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；表达双基因的B16细胞体外诱导免疫应答的能力高于单基因过表达B16细胞。结论：B16细胞经重组腺病毒感染后，表达在细胞膜上的SEA和CD80具有免疫学活性，本研究为今后进一步采用所构建重组腺病毒进行恶性黑色素瘤的免疫基因治疗提供了实验依据。