Chinese Journal of Tissue Engineering Research ›› 2010, Vol. 14 ›› Issue (36): 6662-6666.doi: 10.3969/j.issn.1673-8225.2010.36.003

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Culture and identification of sheep bone marrow-derived endothelial progenitor cells

Yuan Jian-ming1, Guan Song-hui2, Dang Rui-shan1, Yang Xiang-qun1, Shen Man-ru1, Zhang Chuan-sen1   

  1. 1 Research Room of Human Anatomy, Basic Department, Second Military Medical University, Institute of Biomedical Engineering, Shanghai  200433, China; 2 Department of Laboratory, the 455 Hospital of Chinese PLA, Shanghai  200003, China
  • Online:2010-09-03 Published:2010-09-03
  • Contact: Zhang Chuan-sen, Doctor, Professor, Doctoral supervisor, Research Room of Human Anatomy, Basic Department, Second Military Medical University, Institute of Biomedical Engineering, Shanghai 200433, China chuansen@yahoo.com
  • About author:Yuan Jian-ming☆, Studying for doctorate, Technician-in-charge, Research Room of Human Anatomy, Basic Department, Second Military Medical University, Institute of Biomedical Engineering, Shanghai 200433, China yjmdragon@yahoo. com.cn
  • Supported by:

    the Natural Science Foundation of Shanghai City, No. 08441900600*

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Abstract:

BACKGROUND: Nowadays, the study of tissue-engineered venous valve has just started, the choice of seed cells is the key, endothelial progenitor cells are optimal seed cells for tissue-engineered venous valve construction in vitro.
OBJECTIVE: To study the culture method of sheep bone marrow-derived endothelial progenitor cells (EPCs) and provide experimental basis for seed cells selection of sheep tissue engineering by culture and identification of sheep bone marrow-derived endothelial progenitor cells in vitro.
METHODS: Sheep bone marrow mononuclear cells were obtained after whole bone marrow culturing selectively by using bone marrow condition medium. After primary cells were passaged, CD133-positive cells were sorted by microbeds instrument and cultured continually. Flow cytometry confirmed CD133 expression in cells before separation. The cell growth curve was drawn after separation and passage to observe cell growth capacity. Cell specific molecule CD133, D34, VWF expression were detected by immunocytochemistry. FITC labeled BS-1-lectin and DiI labeled ac-LDL were used to label cells and detect cell phagocytosis.
RESULTS AND CONCLUSION: The sheep bone marrow mononuclear cells began to adhere on day 2 in primary culture and fully integrated on day 7. Cells started to went into the logarithmic growth phase on day 2 after passage and cell morphology formed typical paving stone-like on 3-5 days after passage; cells went into platform phase on day 7 after passage. Cells microbead sorting rate was 12.6% after passage. Flow cytometry showed CD133-positive cells rate was 12.64%. Immunocytochemistry showed that cells presented CD133, CD34, VWF-positive expression. FITC-labeled BS-1-lectin and DiI-ac-LDL double positive cells rate was 85.3%. This study successfully isolated and cultured EPCs from sheep bone marrow mononuclear cells.

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