Integrin α2β1-independent activation of platelets by simple collagen-like peptides: collagen tertiary (triple-helical) and quaternary (polymeric) structures are sufficient alone for α2β1-independent platelet reactivity

Abstract

The platelet reactivities of two simple collagen-like synthetic peptides, Gly-Lys-Hyp-(Gly-Pro-Hyp)10-Gly-Lys-Hyp-Gly and Gly-Cys-Hyp-(Gly-Pro-Hyp)10-Gly-Cys-Hyp-Gly, were investigated. Both peptides adopted a stable triple-helical conformation in solution. Following cross-linking, both peptides proved to be highly platelet-aggregatory, more active than collagen fibres, inducing aggregation at concentrations as low as 20 ng/ml. These peptides formed microaggregates in solution, and cross-linking was thought to stabilize these structures, allowing expression of their platelet reactivity at 37 degrees C. Like collagen fibres, the peptides caused platelet secretion and release of arachidonate from platelet membrane lipids as well as activation of integrin alpha IIb beta 3 culminating in aggregation. Monoclonal antibodies directed against the integrin alpha 2 beta 1 failed to prevent aggregation release of arachidonate or platelet adhesion to the peptides. Our results indicate that collagen can activate platelets by a mechanism that is independent of integrin alpha 2 beta 1 and for which collagen tertiary and quaternary structures are sufficient alone for activity without the involvement of highly specific cell-recognition sequences.

CEACAM1 negatively regulates platelet-collagen interactions and thrombus growth in vitro and in vivo

“…A "2-site, 2-step" model for plateletcollagen interactions has been proposed that requires the initial engagement of collagen to be mediated by the high-affinity receptor integrin ␣2␤1, followed by activation generated through the low-affinity collagen receptor GPVI. 37,38 In this model, integrin ␣2␤1 is now thought to have a supportive, rather than a primary role as the adhesive collagen receptor, while GPVI is a major platelet-collagen activating receptor that interacts with collagen fibrils via a triplicate GPO sequence (glycine, proline, hydroxyproline) and integrin ␣2␤1 interacts with collagen via GFOFER (glycine, phenylalanine, hydroxyproline, glutamic acid and arginine). [39][40][41] For in vitro studies, we used CRP as the GPVI-selective agonist because this peptide contains 10 GPO repeat sequences and serves as a potent activator of GPVImediated signaling and does not recognize integrin ␣2␤1 .…”

A novel interaction between FlnA and Syk regulates platelet ITAM-mediated receptor signaling and function

“…Protein G Sepharose and glutathione Sepharose beads were obtained from GE Healthcare. The CRP (amino acids GCO(GPP) 10 GCOG) was synthesized by the Tufts University Core Facility and cross-linked as described previously (Morton et al, 1995). Rhodocytin was purified from the venom of Calloselasma rhodostoma (Suzuki-Inoue et al, 2006).…”

Platelet PAR1 receptor density—Correlation to platelet activation response and changes in exposure after platelet activation

“…Chemicals used for the HEPES buffer (137 mmol/L NaCl, 2. according to their instructions [11]. The peptide solution was diluted to 55 g/mL in 0.05% acetic acid and further diluted 1:4 in HEPES buffer upon thawing.…”