EP4A对人CD34^+细胞干性因子表达的影响

Effects of Prostaglandin E2 Receptor 4 Agonist on the Expression of Stem Factors in Human CD34^+ Cells

目的:探讨前列腺素E2特异性受体激动剂4(EP4A)增强人CD34^+细胞干性因子表达的机制。方法:收集中山大学附属第一医院血液科20例健康供者经G-CSF动员后的外周血造血干细胞采集物,采用免疫磁珠法分选出人CD34^+细胞。以DMSO作对照,分别用EP4A、EP4A+EP4A拮抗剂(EP4AA)作用人CD34^+细胞72 h后,应用热图和Pathway富集分析检测与CD34^+细胞干性相关的差异基因和通路;再用qRT-PCR和Western blot检测不同组别细胞中通路蛋白和差异基因的mRNA及蛋白(β-catenin、Nanog、Oct4、Sox2、Stat3、AKT、P38)表达水平。结果:与对照组相比,EP4A组人CD34^+细胞中β-catenin、Nanog、Oct4及Sox2 mRNA及其蛋白的表达均增高,而STAT3、AKT、P38 mRNA及其蛋白的表达无变化;EP4A与EP4AA(XAV939)共同作用人CD34^+细胞时,人CD34^+细胞β-catenin表达水平明显降低,并且Nanog、Oct4及Sox2基因及蛋白表达相应减少。结论:EP4A可增加人CD34^+细胞干性因子(β-catenin、Nanog、Oct4和Sox2)表达,而不能增加STAT3、AKT、P38表达。

Objective:To investigate the effect of prostaglandin E2 recoptor 4 antagonist(EP4A)on the self-renewal ability of human CD34^+cells and its mechamism.Methods:The peripheral blood hematopoietic stem cell of 20 healthy donors received the G-CSF-mobilization were collected,then the human CD34^+cells were sorted out by MACS microbead kit.The human CD34^+cells were treated with DMSO(control group),EP4A(EP4A group)and EP4A+EP4A antagonist(EP4A+EP4A group)for 72 hours.The differential genes and pathways related with CD34^+cell stemness were detected by Thermogram and Pathway enrichment analysis.and then the expression levels of protein and gene(β-catenin,Nanog,Oct4,Sox2,Stat3,AKT,P38)were detected by qRT-PCR and Western blot respectively.Results:EP4A could elevate the mRNA and protein expression ofβ-catenin,Nanog,Oct4,Sox2,in comparison with control group,however,mRNA and protein expression of STAT3,AKT,P38 were not changed.When human CD34^+cell were cultured with EP4A+XAV939 it was found that the mRNA and protein expression ofβ-catenin was downregulated,moreover the mRNA and protein expression of Nanog,Oct4,Sox2 were reduced.Conclusion:EP4A can upregulate stemness factors-β-catenin,Nanog,Oct4 and Sox2 in human CD34^+cell in vitro,but not STAT3,AKT and P38.