title: Characterization of a spontaneously transformed human endothelial cell line.
abstract: BACKGROUND
  A line of cells was isolated from a focus observed in a human umbilical vein endothelial cell (HUVEC) culture, presumably the result of a spontaneous transformation event.
  
  
  METHODS
  The cell line was continuously cultured for 16 months, after which time, the proliferative rate, capacity to be cloned, and ability to be transfected was investigated. The cell line was analyzed for expression of endothelial cell markers, von Willebrand Factor, P selectin, and scavenger receptor. We also examined the tumor necrosis factor (TNF)-mediated upregulation of E-selectin, vascular cell adhesion molecule-1 and intercellular adhesion molecule-1. We evaluated the levels of expression and types of TNF-alpha receptor expressed by the cell line, and the cell lines response to interleukin-4 and interferon-gamma. CD34 expression of this cell line and the ability of transforming growth factor-beta to inhibit the TNF-alpha induction of E-selectin was examined. The ability to support neutrophil adhesion and transmigration, and generate capillary-like tubes in vitro was assessed. Finally, the karyotype and tumourigenicity of this line was established.
  
  
  RESULTS
  This cell line (C11STH) has a doubling time comparable to that of normal HUVECs and has a trisomy of chromosome 8 and 11. The cell line is capable of generating colonies at clonal density, and is transfectable with efficiencies comparable to normal HUVECs. C11STH expresses von Willebrand factor, P-selectin, and scavenger receptor to an extent similar to passaged HUVECs and can be induced to express E-selectin, VCAM, and ICAM with TNF-alpha. C11STH expresses both the p55 and p75 subunits of TNF-alpha receptor at levels similar to HUVECs. The ability of interleukin-4 to enhance the expression of VCAM and reduce the TNF-alpha-mediated expression of E-selectin is maintained in this cell line. C11STH cells are unable to induce class II major histocompatibility antigen in response to interferon-gamma. However, interferon-gamma is able to synergize with TNF to enhance the expression of E-selectin. C11STH cells do not express CD34 or show transforming growth factor-beta inhibition of TNF-alpha induced E-selectin expression, functions indicative of primary, or early passage HUVECs. The cell line retains the ability to support neutrophil adhesion and transmigration and can generate patent tubes when seeded onto complex basement membrane gels. However, the cell line no longer has the ability to generate capillary-like vessels when seeded onto collagen gels in the presence of phorbol 12-myristate 13-acetate.
  
  
  CONCLUSIONS
  C11STH has retained many of the normal characteristics of endothelial cells, is able to proliferate at clonal density, and is easily transfectable. The line will provide a useful resource for endothelial cell biology. Its ability to make capillary-like tubes on basement membrane gels, but not on collagen will provide a powerful tool with which to further analyze the processes involved in angiogenesis, and will enable us to define the role of specific proteins in angiogenesis. Since C11STH shows tube competence on Matrigel, but is not tube competent on collagen, our studies suggest that capillary-tube formation on Matrigel and collagen occur via qualitatively different mechanisms. Thus, this cell line provides the opportunity to examine the signalling mechanisms required to generate capillary tube formation. These may include the involvement of matrix molecules, the production of proteases and inhibitors, gene regulation and kinases or phosphatases.
authors.affiliation: Hanson Center for Cancer Research, IMVS, Adelaide.
authors.forename: G W
authors.initials: GW
authors.lastname: Cockerill
authors.forename: G
authors.initials: G
authors.lastname: Meyer
authors.forename: L
authors.initials: L
authors.lastname: Noack
authors.forename: M A
authors.initials: MA
authors.lastname: Vadas
authors.forename: J R
authors.initials: JR
authors.lastname: Gamble
mesh_terms: D002448:Cell Adhesion / Q000502:physiology; D015815:Cell Adhesion Molecules / Q000032:analysis; D002454:Cell Differentiation / Q000502:physiology; D002455:Cell Division / Q000502:physiology; D002461:Cell Line, Transformed; D002880:Chromosomes, Human, Pair 11; D002898:Chromosomes, Human, Pair 8; D004730:Endothelium, Vascular / Q000737:chemistry / Q000166:cytology* / Q000502:physiology; D005434:Flow Cytometry; D006801:Humans; D007150:Immunohistochemistry; D018799:Intercellular Adhesion Molecule-1 / Q000032:analysis; D007371:Interferon-gamma / Q000494:pharmacology; D015847:Interleukin-4 / Q000494:pharmacology; D007621:Karyotyping; D019007:P-Selectin; D010980:Platelet Membrane Glycoproteins / Q000032:analysis; D014162:Transfection; D014314:Trisomy; D019010:Vascular Cell Adhesion Molecule-1; D014841:von Willebrand Factor / Q000032:analysis
chemical_list: D015815:Cell Adhesion Molecules; D019007:P-Selectin; D010980:Platelet Membrane Glycoproteins; D019010:Vascular Cell Adhesion Molecule-1; D014841:von Willebrand Factor; D018799:Intercellular Adhesion Molecule-1; D015847:Interleukin-4; D007371:Interferon-gamma