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目的利用逆转录病毒作为载体将小鼠白细胞介素(IL)-23基因转染小鼠乳腺癌细胞(MA-891),建立分泌IL-23的肿瘤细胞株(IL-23/MA-891),观察对其生长特点及体内成瘤情况。方法应用逆转录病毒载体(LXSN)将IL-23基因质粒经ψ2(ecotropic)和PA317(amphotropic)两种包装细胞包装后,转染MA-891细胞,经G418筛选后获得表达IL-23的IL-23/MA-891细胞。采用逆转录-聚合酶链反应(RT-PCR)鉴定转染细胞mRNA的表达;激光共聚焦显微镜(LSCM)观察其蛋白表达;用ELISA法检测IL-23/MA-891细胞培养上清诱导脾细胞分泌IFN-γ的能力;用噻唑蓝(MTT)比色法检测细胞体外增殖能力;用流式细胞仪检测细胞周期与凋亡。小鼠皮下接种转染IL-23的IL-23/MA-891细胞观察其体内致瘤性的变化,小鼠实体瘤模型研究基因转染对肿瘤的生长抑制作用。结果IL-23转染MA-891细胞后,在mRNA水平水平可获得稳定表达;LSCM可观察到其细胞内表达。IL-23基因转染MA-891细胞后,与对照组比较,其体外增殖能力不受影响,致瘤率也无改变,但IL-23基因转染组肿瘤生长明显受到抑制(P<0.05)。结论成功建立表达小鼠IL-23基因的小鼠乳腺癌细胞系(IL-23/MA-891);在体内,IL-23转染组具有明显的抗肿瘤作用。
Objective To construct murine breast cancer cell line (IL-23 / MA-891) secreting IL-23 by transfection of murine interleukin (IL) -23 gene using retrovirus as a vector. , Observe its growth characteristics and tumor formation in vivo. Methods IL-23 gene plasmids were packaged in two packaging cells of ψ2 (ecotropic) and PA317 (amphotropic) by retroviral vector (LXSN), and then transfected into MA-891 cells. After screening by G418, IL- -23 / MA-891 cells. The expression of transfected cells was identified by reverse transcription-polymerase chain reaction (RT-PCR). The expression of IL-23 / MA-891 cells was detected by laser scanning confocal microscopy (LSCM) The ability of cells to secrete IFN-γ was measured. Cell proliferation was measured by MTT colorimetric assay. Cell cycle and apoptosis were detected by flow cytometry. The in vivo tumorigenicity of IL-23 / MA-891 cells transfected with IL-23 was inoculated subcutaneously in mice, and the growth inhibition effect of gene transfection on tumor was investigated in mouse solid tumor model. Results The expression of IL-23 in MA-891 cells was stable at the mRNA level. The expression of LSCM was observed in the cells. IL-23 gene transfected MA-891 cells, compared with the control group, its in vitro proliferative capacity is not affected, the tumor-causing rate did not change, but IL-23 gene transfected tumor growth was significantly inhibited (P <0.05) . Conclusion The murine breast cancer cell line (IL-23 / MA-891) expressing mouse IL-23 gene was established successfully. In vivo, IL-23 transfection group had obvious anti-tumor effect.