Platelet collagen receptors and their role in modulating platelet adhesion patterns and activation on alternatively processed collagen substrates

This study examines the roles of platelet collagen receptors glycoprotein VI (GPVI), a2ß1, and the GPIb-IX-V complex in platelet activation and thrombus formation on various collagen sources from different species. Type I collagens standardly used in haematology testing, i.e. collagen type I derived from equine tendon (HORM) and rat tail collagen were evaluated. Moreover, acid soluble collagen from human umbilical cord was tested. To inhibit platelet-collagen interactions, combinations of monoclonal antibodies 6B4 and 6F1, targeting GPIba and a2ß1, respectively, were used, along with the therapeutic collagen receptor GPVI antibody glenzocimab. Our findings reveal distinct dependencies on these receptors: platelet aggregation of washed platelets to HORM collagen relied on both a2ß1 and GPVI, to acid soluble collagen mainly on GPVI, and to rat tail collagen solely on a2ß1, respectively. In whole blood perfusion assays under non-coagulating conditions, the acid soluble collagen surface triggered a more homogenous platelet adhesion when compared to the HORM collagen surface, whilst platelet adhesion on rat tail collagen varied considerably. The GPIb-IX-V complex was shown to play a key role in initial platelet adhesion and activation across all collagen surfaces at a shear rate of 1600 s-1. At 1600 s-1, inhibiting platelet a2ß1 interaction with collagen by 6F1 antibody did not affect platelet thrombus formation on acid soluble collagen, while it did reduce platelet surface coverage and P-selectin expression on HORM collagen without changing the overall thrombus morphology or contraction. Inhibiting GPVI interaction with collagen significantly reduced all thrombus parameters and abolished PS exposure and P-selectin expression on all three collagen surfaces, at both 1600 s-1 and 150 s-1. Interestingly, upon investigating combined inhibition of GPIb and a2ß1, an additive inhibitor effect of 6F1 was observed on P-selectin expression and PS-exposure on acid soluble collagen but not HORM collagen at 1600s-1, suggesting that the acid soluble collagen is well suited to study reinforcing functions of collagen receptors. Overall, this study highlights the potential advantages of using alternative collagen surfaces beyond the conventional HORM collagen to detect nuanced collagen receptor dependencies, which may prove valuable in evaluating anti-platelet medication.