目的：构建重组诱导性质粒Egr1-XPO4，探讨其与5-FU联合对肝癌细胞SK-Hep1 的协同抑制作用。方法: 将XPO4基因插入带诱导性启动子Egr1 的质粒载体中，构建Egr1-XPO4 重组质粒。将Egr1-XPO4 质粒转染肝癌细胞SK-Hep1 并经化疗药物5-FU诱导，采用Western blotting 检测转染细胞中XPO4 蛋白表达水平，CCK法检测转染诱导后SK-Hep1 细胞的增殖水平，Annexin V-FITC/PI 流式细胞仪检测SK-Hep1 细胞凋亡情况。动物实验观察Egr1-XPO4 质粒联合5-FU对裸鼠移植瘤的治疗效果。结果: 成功构建携XPO4 基因和启动子Egr1 的重组诱导性质粒Egr1-XPO4，重组质粒转染SK-Hep1 细胞并经5-FU诱导后，细胞中XPO4 蛋白表达量明显增加，且与5-FU的质量浓度明显依赖。Egr1-XPO4 转染联合5-FU诱导对SK-Hep1 细胞增殖抑制明显强于单纯转染或5-FU诱导（P<0.05），同时导致SK-Hep1 细胞的早期凋亡率显著高于单纯转染或诱导（P<0.05）。以Egr1-XPO4 质粒联合5-FU 治疗后，裸鼠肝癌移植瘤的体积明显小于Egr1-XPO4 或5-FU 单独治疗（P<0.05)。结论: 重组诱导性质粒Egr1-XPO4联合5-FU对肝癌细胞SK-Hep1 产生协同抑制作用。

Objective: To construct recombinant plasmid Egr1-XPO4 and evaluate its synergic inhibition with 5-FU against hepatocarcinoma SK-Hep1 cells. Methods: The XPO4 gene was inserted into vector carrying promoter Egr1 to construct a new recombinant vector,Egr1-XPO4, which was then transfected into human hepatocarcinoma cell line SK-Hep1 and sensitized with chemotherapeutic drug 5-FU. Western blotting was adopted to examine the protein expression of XPO4; CCK assay was used to detect SK-Hep1 cell proliferation after transfection, and Flow Cytometry with Annexin V-FITC/PI staining was used to detect the apoptosis of SK-Hep1 cells. SKHep1 cell xenograft model was constructed on nude mice, and the effect of Egr1-XPO4 in combination with 5-FU on the growth of xenograft was observed. Results: The recombinant plasmid Egr1-XPO4 was successfully constructed.With the sensitization of 5-FU, the expression of XPO4 protein in SK-Hep1 cells was significantly elevated after Egr1-XPO4 transfection, and the evlevation was in a 5-FU dose- depend manner.The combined treatment of Egr1-XPO4 and 5-FU produced a significantly stronger inhibition against SKHep1 cell proliferation and greatly promoted apoptosis of SK-Hep1cells compared with 5-FU or pEgr-XPO4 mono-treatment group (all P<0.05). And in vivo antitumor experiment showed that the tumor volume in Egr1-XPO4+5-FU treatment group was significantly smaller than that of Egr1-XPO4 or 5-FU mono-treatment group (P<0.05). Conclusion: The recombinant plasmid Egr1-XPO4 in combination with 5-FU could exertsynergic inhibitionagainst hepatocarcinomaSK-Hep1 cells.

深圳国际合作研究资助项目（No.GJHZ20170313111237888）