Background: Kawasaki disease (KD) as the most commonly acquired heart disease in children worldwide is an acute systemic febrile illness with endothelial injury. The incidence of KD varies in different countries, but it is higher in Asian than in Western countries. Endothelial progenitor cells (EPCs) can differentiate into endothelial cells and serve as a potential therapeutic agent for KD. The present study aimed at exploring the simple procedures of isolation, cultivation and purification of EPCs from peripheral blood.

Methods: Five milliliters of peripheral blood was collected from the femoral artery of each Sprague-Dawley rat. Mononuclear cells were isolated by Ficoll density gradient centrifugation and cultured in a special medium, including vascular endothelial growth factor and basic fibroblast growth factor. After 6 days of continuous culture on 24-well fibronectin-coated plates, the cells expanded and differentiated into endothelial-like progenitor cells. The expression of Flk-1, von Willebrand factor (vWF), CD31, and CD34 was assessed. Surface lectin staining was performed using fluorescently labeled UEA-1 lectin at 10 µg/ml. Meanwhile, the ability for live cells to take up fluorescently labeled acetylated low-density lipoprotein (DiI-Ac-LDL) was assessed by incubation with DiI-Ac-LDL.

Results: The cells harvested by this procedure were CD31, CD34, Flk-1 positive and demonstrated double-positive fluorescence for LDL, and lectin-UEA-1.

Conclusion: Relatively purified EPCs can be obtained by certain procedures of isolation and culture. For its differentiating potential, EPCs act as an important source of mature endothelial cells (ECs).

Key words: peripheral blood; mononuclear cell; endothelial progenitor cells; cell culture