Abstract: Objective To improve the extraction method of primary endothelial cells from human saphenous vein and to provide a cell model for the study of vein graft failure after coronary artery bypass grafting. Methods Endothelial cells were extracted from abandoned human saphenous vein by enzymatic digestion and scraping(scraping the vascular intima after enzymatic digestion for further enzymatic digestion). The extracted endothelial cells were cultured in the endothelial cell culture medium containing 10% fetal bovine serum +1% endothelial cell growth supplement +100 U/mL penicillin +100 mg/mL streptomycin. The morphology and adhesion of endothelial cells were observed under an inverted microscope. The specific antigen was identified with immune fluorescence assay of von Will brand Factor(vWF). The purity was detected with flow cytometry of CD31. The activity of endothelial cells was determined with the cell growth curve plotted on different observation days. Results Under the inverted microscope, endothelial cells converged on the wall and arrayed in a typical cobblestone shape. Immune fluorescence assay confirmed the positive expression of vWF. The results of flow cytometry showed that 99.9% and 89.2% of cultured cells in the enzymatic group and scraping group were positive for CD31. Data of the cell growth curve demonstrated that there was no significant difference in endothelial cell activity between the two groups(P=0.057). Endothelial cells from both groups were able to transmit to the 7th-8th generation. Conclusion Based on the traditional enzymatic digestion method, adding the scraping and re-digestion step can extract more primary human saphenous vein endothelial cells.

Key words: Human saphenous vein, Endothelial cell, Extraction method, von Willebrand factor, Vein graft failure