DOWN-REGULATION OF PLATELET ENDOTHELIAL-CELL ADHESION MOLECULE-1 RESULTS IN THROMBOSPONDIN-1 EXPRESSION AND CONCERTED REGULATION OF ENDOTHELIAL-CELL PHENOTYPE

bEND.3 cells are polyoma middle T-transformed mouse brain endothelial cells that express very little or no thrombospondin-l, a natural inhib itor of angiogenesis, but express high levels of platelet endothelial cell adhesion molecule-1 (PECAM-1) that localizes to sites of cell-cel l contact. Here, we have examined the role of PECAM-1 in regulation of bEND.3 cell proliferation, migration, morphogenesis, and hemangioma f ormation. We show that down-regulating PECAM-1 expression by antisense transfection of bEND.3 cells has a dramatic effect on their morpholog y, proliferation, and morphogenesis on Matrigel. There is an optimal l evel for PECAM-1 expression such that high levels of PECAM-1 inhibit, whereas moderate levels of PECAM-1 stimulate, endothelial cell morphog enesis. The down-regulation of PECAM-1 in bEND.3 cells resulted in ree xpression of endogenous thrombospondin-1 and its antiangiogenic recept or CD36. The expression of the vascular endothelial growth factor rece ptors flk-1 and flt-1, as well as integrins and metalloproteinases (wh ich are involved in angiogenesis), were also affected. These observati ons are consistent with the changes observed in proliferation, migrati on, and adhesion characteristics of the antisense-transfected bEND.3 c ells as well as with their lack of ability to form hemangiomas in mice . Thus, a reciprocal relationship exists between thrombospondin-l and PECAM-1 expression, such that these two molecules appear to be constit uents of a ''switch'' that regulates in concert many components of the angiogenic and differentiated phenotypes of endothelial cells.