Identification, localization, and role of fibronectin in cultured bovine endothelial cells.

Abstract

We have examined bovine aortic endothelial cell cultures for the presence of fibronectin, a high molecular weight cell-surface glycoprotein. Sparse cultures contain fibronectin only on dorsal cell surfaces at regions of cell-cell contact, as detected by immunofluorescence. In contrast, when the endothelial cells reached confluence as a highly contactinhibited monolayer, fibronectin was detected in an extracellular matrix underneath the cell monolayer but not on top of the monolayer. Sodium dodecyl sulfate/polyacrylamide gel electrophoresis of isolated extracellular matrix revealed that a predominant component of the matrix is a protein of approximately 2.3 X 105 molecular weight,.which has been identified as fibronectin.The vascular endothelium exists as a monolayer of highly flattened and contact-inhibited cells; because of their location at the interface between blood and tissue, endothelial cells are the chief elements involved in the permeability of blood vessels (1, 2). Although the side of the vascular endothelium exposed to the bloodstream is nonthrombogenic, the subendothelial matrix on which the cells rest is very thrombogenic (3,4). Thus, a disruption of the vascular endothelium can expose the underlying basement membrane, resulting.in the aggregation of platelets and thrombus formation. Therefore, factors that are involved in the attachment of endothelial cells to the basement membrane are very important for the proper functioning of the vascular endothelium.Biochemical analyses have established that basement membranes possess a highly cross-linked form of collagen (5), rendering basement membranes completely insoluble under physiological conditions. Although most biochemical studies of basement membranes have dealt with the structure and synthesis of collagen, noncollagenous matrix glycoproteins have also been found in some basement membranes (6). Recent studies indicate that one of these glycoproteins may be identical to fibronectin (7), a major cell-surface glycoprotein that is immunologically identical to cold-insoluble globulin (8), a plasma protein which is presumably the plasma form of fibronectin that is shed from cells into the blood. Fibronectin appears to be similar, if not identical, to the large external transformationsensitive (LETS) protein, which exists on the surface of certain untransformed cells, but not on the surface of transformed cells (9)(10)(11) (21), and we also show that fibronectin is a major component of the extracellular matrix produced by ABAE cells. METHODSCell Culture. The cloning, culturing, and characterization of ABAE cells have been described (21). ABAE cells are identified as endothelial cells on the basis of their morphology and ultrastructure and on their ability to synthesize Factor VIII (antihemophilic factor antigen), a marker for endothelial cells (22). These cells are routinely maintained in fibroblast growth factor (23).Electron Microscopy. Cells were prepared for electron microscopy by described methods (24), and then examined in a Hitachi S-500 ...

Autocrine fibronectin directs matrix assembly and crosstalk between cell–matrix and cell–cell adhesion in vascular endothelial cells

“…Previous studies have described vectorial secretion and basal assembly of FN in confluent BAE cell cultures (Birdwell et al, 1978;Kowalczyk et al, 1990;Unemori et al, 1990). Here, we show that autocrine FN, as opposed to exogenous soluble FN, is crucial for FN fibrillogenesis.…”

Tumour‐Induced Angiogenesis: A Review

“…The effect of haptotaxis, however, is more complicated, and not fully understood, because the EC are continually modifying the adhesive properties of their micro-environment via proteolysis (Pepper, 2001) and the synthesis of new ECM components (Birdwell et al, 1978). Perhaps the most important substance involved in cell -matrix adhesion is the ECM component, fibronectin.…”

A monoclonal antibody to human platelet glycoprotein IIIa detects a related protein in cultured human endothelial cells.

“…Both synthesize Factor-VIII-related antigen (14,15); in addition, endothelial cells have been shown to synthesize fibrinectin (16)(17)(18), thrombospondin (19,20), a mitogenic factor for smooth muscle (21), and Factor V (22), all of which are also present in platelets. Furthermore, both cell-.types have binding sites for thrombin (23)(24)(25) and Factor Xa (26,27) and provide an effective surface for prothrombin activation (26,27).…”