Long-term, in vitro propagation of tumor-specific endothelial cells (TEC) allows 
for functional studies and genome-wide expression profiling of clonally derived, 
well-characterized subpopulations. Using a genetically engineered mouse model of 
mammary adenocarcinoma, we have optimized an isolation procedure and defined 
growth conditions for long-term propagation of mammary TEC. The isolated TEC 
maintain their endothelial specification and phenotype in culture. Furthermore, 
gene expression profiling of multiple TEC subpopulations revealed striking, 
persistent overexpression of several candidate genes including Irx2 and Zfp503 
(transcription factors), Alcam and Cd133 (cell surface markers), Ccl4 and 
neurotensin (Nts) (angiocrine factors), and Gpr182 and Cnr2 (G protein-coupled 
receptors). Taken together, we have developed an effective method for isolating 
and culture-expanding mammary TEC, and uncovered several new TEC-selective genes 
whose overexpression persists even after long-term in vitro culture. These 
results suggest that the tumor microenvironment may induce changes in vascular 
endothelium in vivo that are stably transmittable in vitro.