Treatment of ischemic disorders using endothelial progenitor cells derived from peripheral blood stem cells and cord blood and analysis of the signal transduction involved in neovasculalization

2004 Fiscal Year Final Research Report Summary

• Project/Area Number: 14570985
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Single-year Grants
• Section: 一般
• Research Field: Hematology
• Research Institution: Kagawa University
• Principal Investigator: KUBOTA Yoshitsugu Kagawa University, Faculty of Medicine, associated professor, 医学部附属病院, 講師 (90178054)
• Co-Investigator(Kenkyū-buntansha): KITANAKA Akira Kagawa University, Faculty of Medicine, assistant professor, 医学部, 助手 (70343308) ; OHNISHI Hiroaki Kagawa University, Faculty of Medicine, assistant professor, 医学部, 助手 (90223891) ; TSUKSMOTO Ikuko Kagawa University, Faculty of Medicine, associated professor, 医学部, 客員助教授 (10183477) ; TANAKA Terukazu Kagawa University, Faculty of Medicine, professor, 医学部, 教授 (20155146)
• Project Period (FY): 2002 – 2004
• Keywords: endothelial progenitor cells / neovascularization / peripheral blood progenitor cells / PI3-kinase / Src family / Akt / VEGF / signal transduction

Research Abstract

The aims of our project were to identify endothelial progenitor cells in the peripheral blood and harvested samples of peripheral blood progenitor cells and to analyze the signal transduction of VEGF in endothelial cells through the Src-PI3-kinase-mediated pathway.
1.Identification of the endothelial progenitor cells in the peripheral blood and harvested samples of peripheral blood progenitor cells.
After obtaining the informed consent, the peripheral blood and harvested samples of peripheral blood progenitor cells isolated from the patients with hematologic malignancies who received chemotherapy followed by G-CSF treatment were subjected to the analysis of endothelial progenitor cells. Dil-acetylated LDL- and UEA-1-positive cells were counted as endothelial progenitor cells. Number of endothelial progenitor cells in the peripheral blood from G-CSF-treated patients (n=15) was significantly increased as compared with that of healthy volunteers (n=5) (52/μL versus 2/μL). Number of endothelial progenitor cells in the harvested samples of peripheral blood progenitor cells (2〜12×10^7 cells) was significantly correlated with number of CD34-positive cells in the peripheral blood (r=0.501).
2.Analysis of the signal transduction involved in VEGF-induced neovascularization using endothelial cells.
Inhibitors of Src kinase (PP1) and PI3-kinase (LY294002) were independently suppressed VEGF-induced neovascularization in a dose-dependent manner. In addition, antisense oligonucleotides against p85 inhibited VEGF-induced neovascularization. Inhibitor of Src kinase (PP1) but not that of Jak family inhibited VEGF-induced phosphorylation of Akt (Serine 473). Taken together, the Src-PI3-kinase-mediated signaling pathway plays an important role in VEGF-induced neovascularization.