title: Expression and assembly of the angiogenic marker B-fibronectin by endothelial cells in vitro: regulation by confluency.
abstract: Recent studies have demonstrated the association of the rare fibronectin (FN) isoform B-FN with newly formed blood vessels. Although B-FN is a unique angiogenesis marker, it is not clear whether B-FN associated with the neovasculature is expressed by the endothelial cells (EC) or obtained from the microenvironment. Here we report on a study analyzing the expression and assembly of B-FN by bovine microvascular EC (BMEC) in vitro. We determined that BMEC express B-FN and assemble it into fibrils during monolayer culture. Furthermore, in addition to assembling endogenous B-FN, subconfluent BMEC can assemble exogenous B-FN provided by a non-EC source. Upon reaching confluency the assembly and expression of B-FN by BMEC is inhibited and the previously assembled B-FN is eventually eliminated from postconfluent EC monolayers. Indeed, confluent BMEC assemble neither endogenous nor exogenous B-FN, while they continue to assemble other FN isoforms. We conclude that BMEC in vitro express B-FN and assemble B-FN fibrils using endogenous as well as exogenous B-FN. The expression and assembly of B-FN are tightly regulated by confluency. Our observations in vitro are a plausible explanation for the absence of B-FN in established blood vessels in vivo, and the subsequent reappearance of B-FN in angiogenic vessels. Furthermore, since our observations of B-FN assembly in BMEC monolayers correspond to previously published observations in vivo, B-FN may be utilized as an appropriate marker for angiogenic behavior of EC in vitro.