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目的:探讨不同肿瘤细胞系柯萨奇病毒-腺病毒受体(CAR)和整合素的表达水平与5型腺病毒感染效率的关系,为腺病毒基因治疗研究奠定基础。方法:利用瞬时转染CAR的真核表达质粒提高肿瘤细胞表面CAR的表达,应用抗体封闭细胞表面的CAR和整合素后,通过流式细胞仪和荧光素酶分析法测定腺病毒Ad5-CMV-EGFP和Ad5-CMV-Luc对肿瘤细胞的感染效率和基因表达水平的变化。结果:不同肿瘤细胞表面CAR和整合素的表达水平是不同的,其中SMMC-7721和A549细胞CAR的表达量最高,而K562细胞CAR的表达水平最低;荧光显微镜和流式细胞仪检测Ad5-CMV-EGFP对肿瘤细胞的感染效率,结果显示5型腺病毒对于SMMC-7721和A549细胞的感染效率最高,而对于K562细胞则很低;瞬时转染表达CAR的真核质粒可提高多种肿瘤细胞表面CAR的表达,较大幅度提高了5型腺病毒的感染效率。而抗体封闭肿瘤细胞表面的CAR或整合素后,腺病毒感染效率显著下降。结论:肿瘤细胞表面CAR和整合素的表达水平决定了腺病毒对肿瘤细胞的感染效率。
Objective: To investigate the relationship between the expression of coxsackievirus Adenovirus receptor (CAR) and integrin in different tumor cell lines and the efficiency of adenovirus type 5 infection, and to lay a foundation for the research of adenovirus gene therapy. Methods: The expression of CAR on the surface of tumor cells was enhanced by transient transfection of CAR eukaryotic expression plasmid. The CAR and integrin on the cell surface were blocked with antibodies. The expression of Ad5-CMV-1 was detected by flow cytometry and luciferase assay. EGFP and Ad5-CMV-Luc on tumor cell infection efficiency and gene expression levels. Results: The expression levels of CAR and integrin on different tumor cells were different. The expression of CAR was the highest in SMMC-7721 and A549 cells, and the lowest in CAR in K562 cells. The expression of Ad5-CMV was detected by fluorescence microscope and flow cytometry -EGFP on tumor cells. The results showed that type 5 adenovirus has the highest efficiency of infection on SMMC-7721 and A549 cells, but lower on K562 cells. Transient transfection of CAR-expressing eukaryotic plasmid can increase the expression of many tumor cells The expression of CAR on the surface greatly improves the infection efficiency of type 5 adenovirus. However, the efficiency of adenovirus infection significantly decreased after antibodies blocked the CAR or integrins on the surface of tumor cells. Conclusion: The expression level of CAR and integrin on tumor cells determines the infection efficiency of adenovirus to tumor cells.