Erschienen in:
01.05.2006 | Basic Science Research
Expansion of Canine Bone Marrow-Derived Endothelial Progenitor Cells and Dynamic Observation
verfasst von:
YingFeng Wu, MD, Jian Zhang, MD, YongQuan Gu, MD, JianXin Li, PhD, Bing Chen, MD, LianRui Guo, MD, Tao Luo, MD, ZhongGao Wang, MD
Erschienen in:
Annals of Vascular Surgery
|
Ausgabe 3/2006
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Abstract
We cultured bone marrow-derived endothelial progenitor cells by a simple ex vivo expansion method and observed the expansion efficacy. Bone marrow mononuclear cells from five mongrel adult dogs were cultured with EGM-2MV medium in culture flasks coated with fibronectin. Morphology was observed with phase contrast microscopy, and a growth curve was constructed to evaluate the efficacy of expansion. Incorporation of Dil-ac-LDL was tested to evaluate the function. At different time points, immunocytochemical staining for flk-1, CD133, and factor VIII-related antigens was done and compared to staining of endothelial cells and mesenchymal stem cells and percentages of CD133, vascular endothelial growth factor receptor 2 (VEGFR-2), and the double-positive cells measured with flow cytometry to identify the quality and efficacy of expansion. Cluster-like attached cells grew to confluence at an average time of 10 days, and the mean number of cells harvested from 1 mL of bone marrow was (1.3 ± 0.3) × 106. The cells presented a cobblestone-like appearance and took up Dil-ac-LDL. Immunocytochemistry showed that flk-1, CD133, and factor VIII-related antigens were positive. Flow cytometry showed that VEGFR-2 and CD133 double-positive cells augmented 242-fold at the tenth day. Ex vivo expansion can effectively proliferate endothelial progenitor cells from bone marrow; the expansion efficacy could meet the requirements for tissue engineering of blood vessels.