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目的:探讨ClCa通道抑制剂--尼氟酸(Niflumic,NFA)对大鼠骨髓来源的晚期内皮祖细胞(endothelial progenitor cells,EPCs)生物学特性的影响。方法:密度梯度离心法分离大鼠骨髓单核细胞,应用EGM-2完全培养液进行体外培养,以第三代或第四代的晚期EPCs作为靶细胞,应用RT-PCR检测晚期EPCs上是否存在Cl Ca通道标志基因TMEM16A和Cl Ca4的表达。采用CCK-8法、Ed U标记法、划痕实验、Boyden小室实验及Matrigel法分别检测10μmol/L NFA对细胞增殖、迁移及体外血管形成能力的影响;应用荧光定量PCR及流式细胞术检测内皮分化标志v WF和CD31基因及蛋白的表达。结果:晚期EPCs表达ClCa通道标志基因TMEM16A和ClCa4;NFA抑制晚期EPCs的迁移功能(P<0.05);但对EPCs的增殖、分化及成血管能力有促进作用。NFA上调了晚期EPCs的CD31和v WF基因和蛋白表达。结论:NFA能促进EPCs的增殖、分化及成血管能力,抑制EPCs的迁移能力。NFA对EPCs生物学特性的这类影响将为心血管疾病治疗药物选择方面提供一定的参考依据。
OBJECTIVE: To investigate the effect of Niflumic (Niflumic), a ClCa channel inhibitor, on the biological characteristics of rat bone marrow-derived endothelial progenitor cells (EPCs). METHODS: Rat bone marrow mononuclear cells were isolated by density gradient centrifugation and cultured in complete EGM-2 medium. The third or fourth generation of late EPCs were used as target cells. RT-PCR was used to detect the presence or absence of advanced EPCs Expression of Cl Ca channel marker genes TMEM16A and Cl Ca4. The effects of 10 μmol / L NFA on proliferation, migration and angiogenesis were detected by CCK-8 assay, Ed U labeling assay, scratch assay, Boyden chamber assay and Matrigel assay. Fluorescent quantitative PCR and flow cytometry Endothelial differentiation markers v WF and CD31 gene and protein expression. Results: The late EPCs expressed ClEM channel marker genes TMEM16A and ClCa4. NFA inhibited the migration of advanced EPCs (P <0.05), but promoted the proliferation, differentiation and angiogenesis of EPCs. NFA up-regulates CD31 and v WF gene and protein expression in advanced EPCs. Conclusion: NFA can promote the proliferation, differentiation and angiogenesis of EPCs and inhibit the migration of EPCs. Such effects of NFA on the biological characteristics of EPCs will provide some reference for the choice of drugs for the treatment of cardiovascular diseases.