A HUMAN VASCULAR ENDOTHELIAL-CELL MODEL TO STUDY ANGIOGENESIS AND TUMORIGENESIS

Endothelial cell biology has recently been the subject of considerable interest in thrombosis and cancer research. However, the successful e stablishment of immortalized human endothelial cells which retain diff erentiated cell characteristics has been rare. We have successfully es tablished immortalized human umbilical vein endothelial cells (HUVECs) by human papilloma virus (HPV)-16 E6-E7, HPV-16 E6, E7 and E6-E7 were successfully introduced into HUVEC cells. Both E6 and E7 cultures had an extended lifespan but eventually underwent senescence. E6-E7 cultu res 4-5-2G, however, acquired an indefinite lifespan in culture but di d not undergo malignant conversion. Telomerase activity was not detect ed in either E6 or E7 cultures; however, telomerase was detected in E6 -E7 4-5-2G cells. The cells exhibited a 'cobblestone' morphology and d eveloped a capillary-like tube structure upon reaching confluence. The 4-5-2G line expressed Factor VIII related antigen and took up DiI-Ac- LDL as markers of endothelial origin. The line expressed integrin subu nits (alpha v beta 3, alpha v beta 5,beta 1, alpha 2, alpha 3, beta 4 and alpha 6) consistent with an endothelial origin. The higher passage of 4-5-2G line showed a similar intensity of integrin immunostaining to that of primary HUVECS. Subsequent infection of these immortal cell s with the Kirsten murine sarcoma virus which contains an activated K- ras oncogene induced morphological transformation that led to the acqu isition of invasion capability and neoplastic properties. Telomerase w as also detected in the tumorigenic v-Ki-ras transformed cell line. Th ese cell lines should be useful for studies of the molecular mechanism s underlying normal and neoplastic endothelial cell proliferation and migration, and might also provide an in vitro model for development of pharmacologic and gene therapy for cardiovascular thrombosis and canc er.