SIMPLE COLLAGEN-LIKE PEPTIDES SUPPORT PLATELET-ADHESION UNDER STATIC BUT NOT UNDER FLOW CONDITIONS - INTERACTION VIA ALPHA-BETA-1 AND VON-WILLEBRAND-FACTOR WITH SPECIFIC SEQUENCES IN NATIVE COLLAGEN IS A REQUIREMENT TO RESIST SHEAR FORCES

The aim of this study was to define the need for specific collagen seq uences and the role of their conformation in platelet adhesion to coll agen under both static and flow conditions. We recently reported that simple triple-helical collagen-related peptides (CRPs), GCP(GPP*)(10) GCPG and GKP*(GPP*)(10)GKP*G (single-letter amino acid code, P* = hyd roxyproline; Morton et al, Biochem J306:337, 1995) were potent stimula tors of platelet activation and were able to support the adhesion of g el-filtered platelets examined under static conditions. The present st udy investigated whether these same peptides were able to support plat elet adhesion under more physiologic conditions by examining static ad hesion with platelet-rich plasma (PRP) and adhesion underflow conditio ns. In the static adhesion assay, we observed 20% surface coverage wit h platelet aggregates. In marked contrast, there was a total lack of a dhesion under flow conditions examined at shear rates of 50 and 300 s( -1). Thus, the interaction of platelets with the CRPs is a low-affinit y interaction unable on its own to withstand shear forces. However, th e addition of CRPs to whole blood, in the presence of 200 mu mol/LD-ar ginyl-glycyl-L-aspartyl-L-tryptophan (dRGDW) to prevent platelet aggre gation, caused an inhibition of about 50% of platelet adhesion to coll agens I and III under flow. These results suggest that the collagen tr iple helix per se, as defined by these simple collagen sequences, play s an important contributory role in the overall process of adhesion to collagen under flow. The monoclonal antibody (MoAb) 176D7, directed a gainst the alpha 2 subunit of the integrin alpha 2 beta 1, was found t o inhibit static platelet adhesion to monomeric but not fibrillar coll agens I and III. However, under flow conditions, anti-alpha 2 MoAbs (1 76D7 anf 6F1) inhibited adhesion to both monomeric and fibrillar colla gens, indicating that alpha 2 beta 1 is essential for adhesion to coll agen under flow, independent of collagen conformation, whether monomer ic or polymeric. To obtain further insight into the nature of the diff erent adhesive properties of CRPs and native collagen, we investigated the relative importance of von Willebrand factor (VWF) and the integr in alpha 2 beta 1 in platelet adhesion to collagen types I and Ill, us ing the same shear rate (300 s(-1)) as used when testing CRPs under fl ow conditions. Our results, together with recent data of others, suppo rt a two-step mechanism of platelet interaction with collagen under fl ow conditions. The first step involves adhesion via both the indirect interaction of platelet glycoprotein (GP) Ib with collagen mediated by vWF binding to specific vWF-recognition sites in collagen and the dir ect interaction between platelet alpha 2 beta 1 and specific alpha 2 b eta 1-recognition sites in collagen. This suffices to hold platelets a t the collagen surface. The second step occurs via another collagen re ceptor (thought to be GPVI) that binds to simple collagen sequences, r equired essentially to delineate the collagen triple helix. Recognitio n of the triple helix leads to strengthening of attachment and platele t activation. (C) 1998 by The American Society of Hematology.