腺病毒载体介导的人IL-24基因对HL-60白血病细胞体外培养的影响

Effect of Ad-IL24 on the ex vivo culture of HL-60 cells

目的构建人IL-24的腺病毒载体Ad-IL24，观察Ad-IL24对HL-60细胞体外培养的影响。方法以pcDNA3.0-IL24重组质粒为模板PCR扩增IL-24基因，酶切并连接到带有绿色荧光蛋白（GFP）标记的pAdTrack-CMV质粒上，Pme I线性化重组质粒pAdTrack-CMV-IL24，并与腺病毒质粒pAdEasy-1共转化BJ5183细菌，获得重组腺病毒载体pAdEasy-1-pTrack-CMV-IL24，经Pac I线性化后转染QBI-293A细胞，收获重组病毒Ad-IL24。将它感染HL-60细胞，通过激光扫描共聚焦显微镜（ISCM）、MTT和流式细胞术（FCM）法检测IL-24对HL-60细胞生长的影响，免疫细胞化学分析凋亡因子的改变，ELISA检测重组病毒对HL-60细胞分泌γ干扰素（IFN-γ）和肿瘤坏死因子-α（TNF-α）的影响。结果成功构建重组腺病毒载体pAdEasy-1-pTrack-CMV-IL24并获得高滴度的重组腺病毒Ad-IL24，各种检测方法表明IL-24基因能抑制HL-60细胞生长，诱导凋亡，IL-24还能下调HL-60细胞Bcl-2的表达并刺激它分泌二级细胞因子IFN-γ和TNF-α。结论IL-24能抑制HL-60细胞生长、诱导凋亡，它可通过下调HL-60细胞Bcl-2的表达并刺激其分泌具有抗肿瘤活性的二级细胞因子来发挥效应。

Objective To construct the human IL-24 recombined adenovirus （Ad-IL24） and to observe the effect of Ad-IL24 on ex vivo culture of HL-60 cells. Methods pAdTrack-CMV-IL24 was con- strutted by PCR with pcDNA3.0-IL24 recombined plasmid as a template, enzyme digestion and ligation. The pAdTrack-CMV-IL24 lineared by Pme I was co-transformed into BJ5183 with pAdEasy-1. The pAdEasy-1-pTrack-CMV-IL24 recombined adenovirus vector was lineared with Pac I and then transfected in-to QBI-293A cells. The Ad-IL24 was obtained and used to infect HL-60 cells, and the effect on HL-60 cells was tested by LSCM, MTF, FCM, ELISA and immunohistochemistry staining. Results The pAdEasy-1- pTrack-CMV-IL24 vector was constructed and the Ad-IL24 was successfully obtained. The effect of inhibiting growth of HL-60 cells and inducing cell apoptosis by IL-24 was proved by LSCM, MTT and FCM. IL-24 down-regulated the expression of anti-apoptosis factor Bcl-2 and increased the secretion of IFN-γ, TNF-α of HL-60 cells. Conclusion Ad-IL24 can inhibit the growth of HL-60 cells and induce cell apoptosis through down-regulating expression of anti-apoptosis factor and increasing the secretion of IFN-γ, TNF-α of HL-60 cells. The human Ad-IL24 may provide a basic study for the future target therapy to tumors.