Platelet adhesion to damaged blood vessel walls is facilitated by the binding of von Willebrand's factor (VWF) to platelet membrane glycoprotein Ib (GPIb). This research proposal is a study of the biochemistry, cell physiology, and clinical significance of the GPIb receptor for VWF. Utilizing a monoclonal antibody directed at the glycocalicin fragment of GPIb, we have developed an enzyme-linked immunosorbent assay (ELISA) to detect glycocalicin in solution, and a fluorescence-activated cell sorting (FACS) assay to detect platelet surface GPIb. The FACS assay, with its ability to detect subpopulations of platelets and its requirement for only very low numbers of platelets, is central to this proposal.
The specific aims of this research program are: 1) Biochemistry: Characterize the binding site for VWF on GPIb. We have recently demonstrated that glycocalicin contains the VWF binding site. We now propose to characterize the binding site for VWF on glycocalicin by assessing digestion products of purified glycocalicin in assays developed to test glycocalicin/VWF binding. 2) Cell physiology: Test the hypothesis that GPIb is processed from the interior of the platelet to the platelet surface and thence into the plasma. New methods have been developed to quantify GPib inside the platelet (ELISA after phase partition of Triton X-114 solubilized platelets), on the platelet surface (FACS), and in the plasma (ELISA). Changes in these receptor pools will be determined after activation of washed platelets and storage of platelet concentrates. 3) Clinical significance: Investigate the GPIb defect in Wiskott-Aldrich syndrome. Our preliminary studies with FACS have suggested that patients with Wiskott-Aldrich syndrome have two populations of platelets: one GPIb-positive and the other GPIb- negative. We now propose to characterize the GPIb defect in Wiskott-Aldrich syndrome patients and carriers. The results of these experiments are likely to provide a better understanding of the pathophysiology of platelet adhesion to damaged vessel walls. Clinically applicable knowledge which may be gained from this research program includes: changes in the adhesive properties of platelet concentrates during storage; methodology for diagnosis of carriers of Wiskott-Aldrich syndrome; and new insights into the pathogenesis of atherosclerosis, in light of the possible role for VWF in this disorder.
| Michelson, A D; Adelman, B; Barnard, M R et al. (1988) Platelet storage results in a redistribution of glycoprotein Ib molecules. Evidence for a large intraplatelet pool of glycoprotein Ib. J Clin Invest 81:1734-40 |
| Michelson, A D; Barnard, M R (1988) Thrombin-induced changes in platelet membrane glycoproteins. Prog Clin Biol Res 283:159-62 |
| Michelson, A D (1988) Flow cytometric detection of distinct subpopulations of platelets in individual patients. Prog Clin Biol Res 283:131-6 |
| Michelson, A D (1987) Flow cytometric analysis of platelet surface glycoproteins: phenotypically distinct subpopulations of platelets in children with chronic myeloid leukemia. J Lab Clin Med 110:346-54 |
| Michelson, A D; Barnard, M R (1987) Thrombin-induced changes in platelet membrane glycoproteins Ib, IX, and IIb-IIIa complex. Blood 70:1673-8 |
| Michelson, A D; Marshall, P C (1987) Transient neurological disorder associated with transient erythroblastopenia of childhood. Am J Pediatr Hematol Oncol 9:161-3 |
