Promotion of human platelet adhesion and aggregation by a synthetic, triple-helical “mini-collagen”.

Abstract

Platelet activation and aggregation by fibrillar collagens are based on substrate primary, secondary, tertiary, and quaternary structure. Although several peptides incorporating sequences from the triple-helical domains of types I and III collagen inhibit collagen-mediated platelet aggregation, none independently promote platelet activation and aggregation. It is believed that the absence of these platelet activities is due to the lack of proper substrate tertiary and quaternary structures. We have utilized a synthetic, triple-helical "mini-collagen" that incorporates a known cell adhesion site (alpha 1(IV)1263-1277) to better understand the relationship between substrate primary, secondary, tertiary, and quaternary structure and platelet activation and aggregation. The promotion of platelet adhesion, activation, and aggregation was compared for this triple-helical polypeptide (THP), fibrillar and type IV collagens, and a single-stranded peptide (SSP) incorporating the alpha 1(IV)1263-1277 sequence. Glass-coated fibrillar and type IV collagens and the THP supported platelet adhesion at substrate concentrations of 0.33 nM, 0.20 nM, and 0.89 microM, respectively. When platelets were stirred with 10 micrograms/ml of fibrillar (0.33 nM) and type IV (0.20 nM) collagen, SSP (2.1 microM), and THP (0.89 microM), only the fibrillar collagen caused 3H-labeled arachidonic acid release, elevation of cytosolic calcium, irreversible aggregation, and secretion of granule contents. The THP (0.45-1.8 microM) effectively inhibited fibrillar collagen-mediated platelet aggregation, while the SSP did not. At a substrate concentration of 40 micrograms/ml, the THP (3.6 microM) was as effective as fibrillar collagen (1.3 nM) at inducing 3H-labeled oleic acid-labeled platelet activation and microaggregate formation, while the SSP and type IV collagen were relatively ineffective. Rotary shadowing images indicated that aggregates of the THP could form distinct quaternary structures, while the type IV collagen used here could not. These results are the first demonstrations of a synthetic peptide promoting platelet adhesion, activation, and aggregation and suggest that the combination of THP primary, secondary, tertiary, and quaternary structural features are required for platelet aggregation.