| Aims: To investigate the role of micro-RNA 150 acting upon human early endothelial progenitor cells(eEPCs) and late endothelial progenitor(lEPCs) in the course of differentation and tube formation; To investigate whether miR-150 activates the differentation of EPCs via targeting c-Myb.Methods: 1. By means of Ficoll density gradient centrifugation, peripheral blood mononuclear cells(PBMCs) were isolated; Then they were cultured within special EGM-2MV medium during a period of 14 to 28 days; The characters of cultured cells were observed under the optical microscope; Flow cytometry technique and experiments of acetylated low-density lipoprotein endocytosis and ulex europaeus I lectin binding was manipulated to attain the identification. The expression level of miR-150 of eEPCs and lEPCs were examined by real time RT-PCR; 2. eEPCs and lEPCs were respectively transfected with miR-150-agomir or antagomir. The effects of miR-150 on the differentation, proliferation and tube formation were also examined; 3. After transfection, total RNAs and proteins were extracted for PCR and Western Blot accordingly; Both mRNA and proteins of the gene were testified using c-Myb.Results: 1.With the method of density gradient centrifugation, cultured cells were found to show EC specific surface markers such as CD31, CD 133, vWF and VEGFR-2 and had the ability to endocytose ac-LDL and bind UEA I lectin, displaying a high capacity of proliferation and vessel formation; 2. The up-regulation of mi R-150 in eEPCs promoted EPCs differentiation; Down-regulation of miR-150 in lEPCs inhibited the differentation, proliferation and tube formation by matrigel assay; 3.The levels of proteins were lower in which miR-150 was up-regulated.Conclusion: The expression level of miR-150 were different between eEPCs and lEPCs. The regulation of mi R-150 can influence the differentiation,proliferation and tube formation of EPCs. c-Myb was predicted to be the target gene of miR-150. |
