Isolation and long-term serial cultivation of endothelial cells from the microvessels of the adult human dermis.
A method to isolate and maintain microvascular endothelial cells from the cutaneous vessels of adult human skin in long-term culture has been developed. Endothelial cells lining the microvessels of the papillary dermis are released from surrounding tissue during a brief trypsin incubation (0.3% trypsin, 1% EDTA). Cells are plated onto a fibronectin substrate and maintained in Leibovitz (L15) culture medium containing pooled human serum (50%) and antibiotics. Proliferation is dependent upon the presence of several additional growth factors, cholera enterotoxin (1 X 10(-9) M), isobutyl methylxanthine (3.3 X 10(-5) M), and medium conditioned by explant culture of the mouse EHS sarcoma. Using this supplemented medium, cells proliferate readily and can be cultivated serially for more than 6 passages (3 months in vitro). These cells retain their characteristic endothelial cell morphology, stain positively for Factor VIII antigen, and contain Weibel-Palade bodies.
D000941:Antigens / Q000032:analysis; D002478:Cells, Cultured; D004727:Endothelium / Q000166:cytology*; D004817:Epidermis / Q000098:blood supply*; D005169:Factor VIII / Q000032:analysis / Q000276:immunology; D005346:Fibroblast Growth Factors / Q000494:pharmacology; D006133:Growth Substances / Q000494:pharmacology; D006801:Humans; D008833:Microcirculation / Q000166:cytology*; D008854:Microscopy, Electron; D014841:von Willebrand Factor
D000941:Antigens; D006133:Growth Substances; D014841:von Willebrand Factor; D005346:Fibroblast Growth Factors; D005169:Factor VIII