title: Establishment and characterization of a line of adipose-derived human microvascular endothelial cells (HADMEC-5) transformed by simian virus 40 large T antigen expression: application to endotoxin research.
abstract: This paper reports the establishment and initial characterization of an immortalized line of human, adipose tissue-derived microvascular endothelial cells. Transfection of primary endothelial cell cultures was accomplished by the introduction of a plasmid, which contained simian virus 40 large T antigen DNA as well as a Rous sarcoma viral promoter region. One emergent colony, termed HADMEC-5, was isolated and has been passaged 45 times to date. The cells express simian virus 40 large T antigen protein, are immunohistochemically positive for factor VIII-related antigen, bind Ulex europaeus lectin, and accumulate Dil-labeled acetylated low density lipoprotein. The HADMEC-5 line demonstrates a highly proliferative growth rate in the absence of supplemental growth factors, when compared with primary cultures of nontransformed endothelial cells. HADMEC-5 growth remains serum dependent, but exhibits a lower serum requirement than nontransformed cells. The transformed cells grow well upon a variety of matrix compounds and in a variety of growth media. When grown upon Matrigel, the HADMEC-5 cells form three dimensional tube-like structures. The HADMEC-5 line was also tested for its ability to produce eicosanoids in response to bacterial lipopolysaccharide. The transformed cells, tested at passages 7 and 45, displayed a dose-dependent production of prostaglandin E2 in response to lipopolysaccharide in a manner similar to that seen in primary cell cultures. Threshold sensitivity to lipopolysaccharide was 10 pg/mL of media. The HADMEC-5 cell line represents a unique model in which to investigate lipopolysaccharide interactions with microvessel-derived endothelial cells and is of potential value in the study of other aspects of endothelial cell physiology.
authors.affiliation: Department of Physiology, Jefferson Medical College, Thomas Jefferson University, Philadelphia, Pennsylvania 19107, USA.
authors.forename: J T
authors.initials: JT
authors.lastname: Flynn
authors.forename: M
authors.initials: M
authors.lastname: Westbrooks
authors.forename: K A
authors.initials: KA
authors.lastname: Lucas
mesh_terms: D000273:Adipose Tissue / Q000098:blood supply*; D000952:Antigens, Polyomavirus Transforming / Q000096:biosynthesis*; D002455:Cell Division / Q000502:physiology; D002461:Cell Line, Transformed; D002472:Cell Transformation, Viral; D015232:Dinoprostone / Q000096:biosynthesis; D004730:Endothelium, Vascular / Q000166:cytology* / Q000187:drug effects; D006801:Humans; D007150:Immunohistochemistry; D008070:Lipopolysaccharides / Q000494:pharmacology*; D008833:Microcirculation / Q000187:drug effects / Q000502:physiology
chemical_list: D000952:Antigens, Polyomavirus Transforming; D008070:Lipopolysaccharides; D015232:Dinoprostone