目的 以CXCR4基因启动子为靶点,建立促干细胞归巢药物筛选细胞模型,并对中药单体进行筛选.方法 将人CXCR4基因启动子序列(279 bp)插入荧光素酶报告基因载体pGL4.17-Basic中,构建重组质粒pGL4.17-CXCR4,与内参质粒pRL-TK共转染293细胞,经验证CXCR4启动子转录活性后,单细胞克隆培养以获得稳转细胞株,并用于候选中药单体1～10的筛选,最后应用Western blot验证所筛选出的中药单体促进间充质干细胞(MSCs)中CXCR4表达的作用.结果 成功建立药物筛选细胞模型,筛选出候选单体6提高CXCR4启动子活性的作用最强,Western blot结果证实候选单体6可明显促进MSCs中CXCR4的表达.结论 本研究建立的药物筛选细胞模型能实现对潜在促进干细胞归巢中药有效成分的高通量筛选.%Objective To establish a cell model targeting the CXCR4 gene promoter to screen of drugs potentially promoting the stem cell homing, and to screen the monomers of traditional Chinese medicine. Methods The human CXCR4 gene promoter sequence (279 bp) was inserted into the luciferase reporter vector pGL4.17-Basic to construct the recombinant plasmid pGL4.17-CXCR4, which was co-transfected with the internal reference plasmid pRL-TK into 293 cells. After verifying transcriptional activity of CXCR4 promoter, single cell clones were cultured to obtain a stable transfectant cell line and used for screening of candidate Chinese herbal monomer (1 to 10). Finally, Western blot was used to verify the effect of the selected Chinese herbal monomer on the expression of CXCR4 in MSCs. Results The drug screening cell model was successfully established and the candidate monomer 6 was screened out because of the strongest effect on enhancing CXCR4 promoter activity. Western blot results verified that the candidate monomer 6 could promoted the expression of CXCR4 of MSCs. Conclusion The drug screening cell model established in this study was able to achieve high-throughput screening for potentially promoting the stem cell homing Chinese herbal monomer.
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