Control of endothelial focal adhesions by cell shape

Endothelial cell spreading is mediated by the extracellular matrix (ECM), and it is the thought to be regulated by the binding of integrin receptors and the associated focal adhesions (FA). In the present study, we examined the relationship between FA formation and endothelial cell shape changes using microfabricated substrates patterned with islands of immobilized fibronectin (FN) surrounded by nonadhesive borders to control the degree of cell spreading. The amount of FA formation, as detected by the localized presence of vinculin and phosphotyrosine, increased in direct proportion to the degree of cell spreading. To determine whether the observed increase in FA formation was a direct result of more fibronectin in contact with spread cells, single cells were spread across multiple subcellular sized islands such that the size and spacing of the islands could vary cell spreading independently from the amount of cell-ECM contact. We found that total FA increased with cell spreading, regardless of the amount of ECM in contact with cells. The FAs formed on these substrates developed a structural directionality and quantity which correlated to the direction and magnitude of tensional stress in the actin cytoskeleton. Thus, global changes in cell shape regulate the local cytoskeletal tensions that drives FA formation.