title: Endothelial progenitor cells proliferated via MEK-dependent p42 MAPK signaling pathway.
abstract: Endothelial progenitor cells (EPCs) clinical applications have been well reported. However, due to low number of EPCs that could be isolated, EPCs expansion study became one of the main focuses. Some optimized mediums to culture EPCs were currently available. However, the proliferation signaling pathway is not clearly disclosed yet. Peripheral blood was collected from eight healthy subjects, followed by mononuclear cells (MNCs) isolation. MNCs were then prepared and cultured for 2 days. After that, non-adherent cells were harvested and further cultured for 3 days. Resulted colony-forming unit (CFU)-Hill colonies were documented and enumerated under an inverted light microscope. To detect membrane markers, immunofluorescence was performed to detect CD34, VEGFR-2, and CD133. Cell documentation was conducted under a fluorescence microscope. To check cell proliferation, XTT Cell Proliferation Assay Kit was used according to kit insert. To detect possible activation of p44/42 MAPK, western blot was performed to detect p44/42 MAPK and phosphorylated p44/42 MAPK. All visualized bands were captured and quantified. Our results showed that EPCs markers (CD34, CD133 and VEGFR-2) were detected in 3 days culture. From XTT cell proliferation assay and CFU enumeration results, we found that EPCs proliferated significantly (p = 0.012) with addition of supplement. Phosphorylated-p42 MAPK expression of EPCs treated with supplement was significantly higher than the one of EPCs without treatment. Significant inhibition of p42 MAPK phosphorylation by U0126 was observed (p = 0.012). By pretreatment of U0126, number of viable cells and CFUs treated with supplement was significantly decreased (p = 0.012). Our results showed that MEK-dependent p42 MAPK pathway might play an important role in EPCs proliferation.
authors.affiliation: Prodia Clinical Laboratory, Prodia Tower, Jl. Kramat Raya No. 150, Jakarta, Indonesia, ferrysandra@gmail.com.
authors.forename: Ferry
authors.initials: F
authors.lastname: Sandra
authors.forename: Yudi Her
authors.initials: YH
authors.lastname: Oktaviono
authors.forename: Mohammad Aris
authors.initials: MA
authors.lastname: Widodo
authors.forename: Yanni
authors.initials: Y
authors.lastname: Dirgantara
authors.forename: Angliana
authors.initials: A
authors.lastname: Chouw
authors.forename: Djanggan
authors.initials: D
authors.lastname: Sargowo
mesh_terms: D002448:Cell Adhesion / Q000235:genetics; D002454:Cell Differentiation / Q000235:genetics*; D002465:Cell Movement; D049109:Cell Proliferation / Q000235:genetics*; D002478:Cells, Cultured; D066026:Endothelial Progenitor Cells; D018507:Gene Expression Regulation, Developmental; D006801:Humans; D020935:MAP Kinase Signaling System / Q000235:genetics; D008297:Male; D008875:Middle Aged; D019950:Mitogen-Activated Protein Kinase 1 / Q000096:biosynthesis* / Q000235:genetics
chemical_list: D019950:Mitogen-Activated Protein Kinase 1