Isolation of endothelial progenitor cells from cord blood with CD133 immunomagnetic sorting / 中国病理生理杂志

ABSTRACT

AIM: To isolate, purify and differentiate endothelial progenitor cells from cord blood in vitro and to study their biological characteristics. METHODS: CD133~+ cells were selected from fresh cord blood mononuclear cells (MNC) by magnetic activated cell-sorting system (MACS). EPC was studied by flow cytometry, immunocytochemistry and immunofluorescence staining. Isolated cells were cultured in IMDM medium supplemented with or without VEGF, bFGF, SCF. RESULTS: The percentage of CD133~+ cells of cord blood MNC was (1.41?1.14)%, and purity was 75%-85% (FACS method). CD133~+ cells were grown on fibronectin-coated chamber slides in the presence of VEGF, bFGF, SCF. Within 1-2 hours of culture cells became adherent. On day 7-10, the adherent cells displayed a typical "cobblestone" morphology. After 14 days of culture, the adherent cells revealed a heterogeneous cell population, comprising small-sized round cells, spindle-like cells and formed tube-like structure. Weibel-Palade bodies were shown on the transmission electron microscopy photomicrographs. Compared with the (original,) cell markers CD133 and CD34 decreased significantly (77.0%?3.3% to 1.6%?2.2% and 93.1%?4.7% to 37.4%?4.9%, P