UVM Theses and Dissertations
Format:
Print
Author:
Fager, Ammon M.
Dept./Program:
Cell and Molecular Biology Program
Year:
2009
Degree:
PhD
Abstract:
The combined data from our characterization of this subpopulation demonstrated that: 1) these procoagulant platelets have a 3-7 fold increase in the expression of various adhesive receptors as compared to those platelets unable to bind FXa; 2) these platelets represent predominantly "young" platelets, as evidenced by the fact that 70% of platelets able to bind FXa are reticulated; and 3) the procoagulant subpopulation can also be defined by their expression of a non-dissociable pool of platelet-derived FVa. The critical interactions that take place between FVa and FXa in Prothrombinase assembled on an activated platelet membrane have not been investigated until now. By utilizing a chemical cross-linking approach, we have identified six regions in the FVa heavy chain (residues 317-320, 346-348, 362-364, 397-404, 502-506, & 651-655) that are involved in the interaction with FXa. Based on these data, models unique to the platelet-bound Prothrombinase were generated.
The models differ substantially from previous models in that: 1) FXa is positioned much further away from the A1 domain of FVa; 2) they contain a much higher buried surface area; and 3) the positioning of FXa allows for potential conformational changes in FVa, which may be responsible for functional differences between the plasma and platelet cofactor pools. In addition, FXa is positioned in such a way that it would not preclude FVa from being GPI anchored to the platelet membrane as has been suggested. The existence of specific receptors for FVa and FXa at the activated platelet surface has long been hypothesized, but never identified. Throughout the course of our cross-linking studies, binding partners of FXa other than FVa have repeatedly been seen.
The most prominent is a -125 kDa membrane-associated protein which "tracks" consistently with all platelet donors studied to date. Once identified, characterization of this putative receptor may lead to new targets for pharmacological intervention. Since the adherence properties of platelets and their ability to mount and sustain a procoagulant response are crucial steps in effecting both normal hemostasis and pathological thrombus formation, the identified platelet subpopulation should be significant in regulating the hemostatic response and indicates a potential role for these platelets in the initiation of thrombus formation.
The models differ substantially from previous models in that: 1) FXa is positioned much further away from the A1 domain of FVa; 2) they contain a much higher buried surface area; and 3) the positioning of FXa allows for potential conformational changes in FVa, which may be responsible for functional differences between the plasma and platelet cofactor pools. In addition, FXa is positioned in such a way that it would not preclude FVa from being GPI anchored to the platelet membrane as has been suggested. The existence of specific receptors for FVa and FXa at the activated platelet surface has long been hypothesized, but never identified. Throughout the course of our cross-linking studies, binding partners of FXa other than FVa have repeatedly been seen.
The most prominent is a -125 kDa membrane-associated protein which "tracks" consistently with all platelet donors studied to date. Once identified, characterization of this putative receptor may lead to new targets for pharmacological intervention. Since the adherence properties of platelets and their ability to mount and sustain a procoagulant response are crucial steps in effecting both normal hemostasis and pathological thrombus formation, the identified platelet subpopulation should be significant in regulating the hemostatic response and indicates a potential role for these platelets in the initiation of thrombus formation.