VPB0903 - Matrix metalloproteinase-13: a novel player of platelet activation and in vivo thrombus formation

Background: Some MMPs are regulators of platelet function. Platelets express and release MMP-2 that enhances platelet aggregation by cleaving PAR1 at a non conventional site. Atherosclerotic plaques are rich in MMP-13 that is also known to cleave PAR1 of cardiac cells. No studies however have explored whether MMP-13 can activate platelets.

Aims: Our aim was to explore the effect of MMP-13 on platelet PAR1, in vitro platelet activation and signal transduction, and to assess whether it modulates in vivo platelet-mediated thrombosis.

Methods: The cleavage site of PAR1 by rhMMP-13 was characterized using CHO cells transfected with pcDEF3/PAR1 T7- tagged human PAR-1; platelet function was assessed by flow-cytometry; in vivo hemostasis in mice by the tail tip-transection bleeding time; in vivo arterial thrombosis by a photochemical-injury femoral artery thrombosis model.

Results: MMP-13 cleave PAR1 on CHO cells, a reaction blocked by the antibody SPAN12 directed against the PAR1 N-terminal residues 35-46. Active MMP-13 potentiated ADP-induced human platelet P-selectin expression and platelet/leukocyte aggregate formation and triggered platelet G protein activation, effects abolished by WAY170523c (MMP-13 inhibitor) and by the blockade of platelet PAR1 by SPAN12. The ex vivo formation of platelet/monocyte aggregates, and, to a lesser extent platelet/neutrophil aggregates were reduced in MMP-13-/-mice as compared with WT mice suggesting that endogenous MMP-13 plays a role in platelet activation. Femoral artery thrombosis was significantly delayed (27±1.4 vs 11.5±1.7 min, respectively, P< 0.01) and the tail tip transection bleeding time was mildly, but significantly prolonged (390±76 vs 120±11 sec, P< 0.0001) in MMP-13-/- mice as compared to WT mice.

Conclusion(s): Our data data show that active MMP-13 activates platelets by cleaving PAR1 at a non canonical site. MMP-13 plays a role in in vivo platelet activation and thrombus formation at a vessel wall-injury site. MMP-13 represents a novel modulator of platelet function and a potential target of antithrombotic therapy.