Collagen-platelet interactions: evidence for a direct interaction of collagen with platelet GPIa/IIa and an indirect interaction with platelet GPIIb/IIIa mediated by adhesive proteins

Abstract

Using intact human platelets as the immunogen and a functional, collagen-coated bead agglutination assay, we have produced a murine monoclonal antibody (6F1) that blocks the interaction between platelets and collagen in the presence of Mg++. 6F1 affinity-purified the platelet glycoprotein Ia/IIa complex, and approximately 800 molecules of 6F1 bound per platelet at saturation. 6F1 nearly completely inhibited collagen-induced platelet aggregation and inhibited platelet adhesion to collagen by greater than 95% when plasma proteins were absent. Antibody 10E5, which blocks the binding of adhesive glycoproteins to GPIIb/IIIa, produced only minor inhibition (approximately 25%) of adhesion under the same circumstances. In contrast, when tested in platelet-rich plasma (PRP), 6F1 had only a minor effect on collagen-induced platelet aggregation, prolonging the lag phase but not the slope or maximum aggregation. Similarly, when collagen was precoated with plasma, 6F1 caused less inhibition of platelet adhesion (53%) than without the precoating (greater than 95%). Antibody 10E5 inhibited this adhesion by 32%, and the combination of 6F1 and 10E5 was more effective than either alone, inhibiting it by 90%. Time course studies of platelet agglutination of collagen-coated beads using PRP containing physiologic concentrations of divalent cations showed early inhibition by 6F1, indicating that the GPIa/IIa receptor operates in this environment. With more prolonged incubation, however, 6F1 was less effective; this later agglutination could be partially prevented by adding 10E5 or PGE1 to the 6F1. These data support a model wherein collagen can directly interact with GPIa/IIa and can indirectly interact with GPIIb/IIIa via intermediary adhesive proteins. The physiological significance of these interactions, and potential interactions with other receptors, remains to be established.

Allelic distribution of the glycoprotein Ia (α2‐integrin) C807T/G873A dimorphisms among Caucasian venous thrombosis patients and six racial groups

“…The GPIa/IIa complex, in whole blood, mediates the initial phase of platelet adhesion to collagen under low and high shear rates (50±1500/s) that leads to accelerated factor II conversion on the surface of the platelet and thrombus formation supported by the GPIIb/IIIa complex (Coller et al, 1989;Savage et al, 1990;Kirchhofer et al, 1995). In the presence of GPIIb/IIIa inhibitors, GPIa/IIa-mediated adhesion alone is suf®cient to induce cellular changes that catalyse factor II conversion to an extent equal to that seen in the absence of inhibitors (Savage et al, 1990).…”

Enhanced Binding of Poly(ADP-ribose)polymerase-1 and Ku80/70 to the ITGA2 Promoter via an Extended Cytosine-Adenosine Repeat

“…Murine monoclonal IgG antibody 6F1 (anti-α2β1) has been described [32] and is a gift from Dr. B. Coller (Rockefeller University, New York, NY). The murine hybridoma 12F1 producing IgG specific for α2β1 has been well characterized [33] and was generously provided by Dr V. Woods (University of California at San Diego, La Jolla, CA).…”

Distinct roles of GPVI and integrin α2β1 in platelet shape change and aggregation induced by different collagens

“…The integrin a 2 subunit (also known as GPIa) was proposed as a collagen receptor following the identi®cation of a patient with a haemorrhagic disorder whose platelets failed to respond to collagen, and which had 15 ± 25% of normal levels of GPIa on their surface (Nieuwenhuis et al, 1985). However, it has been suggested that additional abnormalities were present which were important in the aetiology of this case (Coller et al, 1989). Further patients in which dysfunctional a 2 b 1 has been associated with a selective de®cit in platelet collagen responsiveness have been identi®ed, amino acid sequences including GFOGER (Emsley et al, 2000;Knight et al, 2000).…”

“…We used reconstituted bovine collagen types I ± V, the commonly used native type I ®brils from equine tendon (Horm collagen) and the synthetic CRP, and examined their ability to induce shape change and aggregation. We investigated the role of a 2 b 1 with the anti-a 2 antibody 6F1 (Coller et al, 1989) and that of GPVI and CD36 with platelets from genetically-modi®ed mice (Febbraio et al, 1999;Park et al, 1998). In addition, we investigated the ability of the collagen agonists to activate platelets in the absence of the positive feedback pathways of ADP and thromboxane A 2 (TxA 2 ).…”