Identification and characterization of cell-substratum adhesion receptors on cultured human endothelial cells.
A series of immunological approaches was utilized to identify the molecules involved in cell-substratum adhesion of human endothelial cells (EC) derived from adult large vessels, fat capillaries, and umbilical veins. A polyclonal antibody prepared against partially purified extracellular matrix receptors disrupted adhesion of EC to a wide variety of substrates and identified four groups of glycoproteins migrating with apparent Mr of 150, 125, 110, and 95 kD in immunoprecipitation experiments. Specific monoclonal antibodies identified these proteins as members of the Integrin family of extracellular matrix receptors and included the alpha and beta chains of the fibronectin receptor (alpha 5/beta 1), a collagen receptor (alpha 2 beta 1), a multifunctional receptor that binds to fibronectin, collagen, and laminin (alpha 3/beta 1), as well as a receptor related to platelet IIb/IIIa (alpha v/beta 3). To directly test the importance of these molecules in cell-substratum adhesion, these proteins were purified by a combination of ion exchange, lectin affinity, and immunoaffinity chromatography and used to block the biological activity of the adhesion-disrupting polyclonal antibody. Immunofluorescence experiments further supported the role of these glycoproteins in adhesion. The GPIIb/IIIa-like receptor localized to well-formed adhesion plaques on EC plated on fibrinogen, but not on fibronectin, laminin, or type IV collagen. Receptors containing the beta 1 subunit were visualized as discontinuous fibrils which colocalized with fibronectin fibrils and actin stress fibers.
Albelda
S M
SM
Department of Medicine, University of Pennsylvania School of Medicine, Philadelphia.
Daise
M
M
Levine
E M
EM
Buck
C A
CA
D000328:Adult; D000818:Animals; D000911:Antibodies, Monoclonal / Q000502:physiology; D000954:Antigens, Surface / Q000276:immunology / Q000302:isolation & purification*; D002448:Cell Adhesion; D015815:Cell Adhesion Molecules; D006224:Cricetinae; D004730:Endothelium, Vascular / Q000378:metabolism* / Q000502:physiology; D005109:Extracellular Matrix / Q000378:metabolism; D006801:Humans; D007106:Immune Sera / Q000494:pharmacology; D016023:Integrins; D008562:Membrane Glycoproteins / Q000276:immunology / Q000302:isolation & purification*; D011233:Precipitin Tests; D051381:Rats; D011971:Receptors, Immunologic / Q000302:isolation & purification*
D000911:Antibodies, Monoclonal; D000954:Antigens, Surface; D015815:Cell Adhesion Molecules; D007106:Immune Sera; D016023:Integrins; D008562:Membrane Glycoproteins; D011971:Receptors, Immunologic