Real-time analysis of the interaction of platelets with immobilized thrombospondin under flow conditions

The platelet granule protein (TS) is extracellularly secreted upon platelet activation and then binds to the platelet surface where it can interact wi th various adhesive proteins. Here, we have analyzed platelet interactions with a TS-coated surface under flow conditions, a model for platelet adhesi on onto surface-bound TS under physiological conditions. Platelets exhibite d temporary, very short-time adhesion on the TS surface, but no firm adhesi on. This adhesion was inhibited by NNKY5-5 (anti-glycoprotein (GP) Ib antib ody) and AJvW-2 (anti-von Willebrand factor (VWF)), indicating that both pl atelet GP Ib and plasma VWF contribute to this interaction. Antibodies agai nst platelet collagen receptor integrin alpha (2)beta (1) had no significan t effect. These results suggested that binding of VWF to TS is the first st ep in platelet interaction with the TS surface. By surface plasmon resonanc e spectroscopy, a dissociation constant (Kd) Of 3.97 x 10(-7) M was obtaine d for the binding reaction between immobilized TS and VWF. These results su ggest the following model for platelet interaction with the TS surface unde r flow: plasma vWF first binds to the immobilized TS and then platelets int eract with the TS-bound vWF. A low density of bound VWF would account for t he observed weak interaction between TS and platelets under flow. (C) 2001 Elsevier Science Ltd. All rights reserved.