Dominant role of aIIbb3 in platelet interactions with cross-linked fibrin fragment D-dimer

Abstract

Although much is known about the interaction of fibrinogen with aIIbb3, much less is known about the interaction of platelets with cross-linked fibrin. Fibrinogen residue Lys406 plays a vital role in the interaction of fibrinogen with aIIbb3, but because it participates in fibrin cross-linking, it is not available for interacting with aIIbb3. We studied the adhesion of platelets and HEK cells expressing normal and constitutively active aIIbb3 to both immobilized fibrinogen and D-dimer, a proteolytic fragment of cross-linked fibrin, as well as platelet-mediated clot retraction. Nonactivated platelets and HEK cells expressing normal aIIbb3 adhered to fibrinogen but not D-dimer, whereas activated platelets as well as HEK cells expressing activated aIIbb3 both bound to D-dimer. Small-molecule antagonists of the aIIbb3 RGD (Arg-Gly-Asp) binding pocket inhibited adhesion to D-dimer, and an Asp119Ala mutation that disrupts the b3 metal ion-dependent adhesion site inhibited aIIbb3-mediated adhesion to D-dimer. D-dimer and a polyclonal antibody against D-dimer inhibited clot retraction. The monoclonal antibody (mAb) 10E5, directed at aIIb and a potent inhibitor of platelet interactions with fibrinogen, did not inhibit the interaction of activated platelets with D-dimer or clot retraction, whereas the mAb 7E3, directed at b3, inhibited both phenomena. We conclude that activated, but not nonactivated, aIIbb3 mediates interactions between platelets and D-dimer, and by extrapolation, to cross-linked fibrin. Although the interaction of aIIbb3 with D-dimer differs from that with fibrinogen, it probably involves contributions from regions on b3 that are close to, or that are affected by, changes in the RGD binding pocket.