The circulating blood platelet plays a crucial role in blood clotting and participates in number of pathological processes. Until very recently, the physiological mechanism by which the bone marrow precursor cell, the megakaryocyte, regulated the production of its progeny platelets had been a mystery. The cytokine which regulates platelet production, thrombopoietin, and its receptor have now been purified. Although considerable effort is being directed toward developing its clinical potential, nothing is known about normal and pathological physiology. The goals of this proposal are to a number of disease processes. Using thrombocytopenic and thrombocytotic animals, the studies proposed herein will show that thrombopoietin MRNA and protein synthesis are not altered by changes in the platelet count and therefore that thrombopoietin production is not directly regulated by the circulating platelet production by studies will focus on the finding that platelets themselves bind to and clear thrombopoietin from the circulation, thereby altering circulating levels of thrombopoietin and subsequent platelet production by megakaryocytes. Beginning with its binding to platelet thrombopoietin receptors, experiments will be directed at understanding the biochemical pathway by which platelets endocytose thrombopoietin. Using cultures of megakaryocyte precursor cells efforts will also be made to determine how the thrombopoietin receptor is itself regulated. The physiological importance of platelet- dependent clearance of thrombopoietin will be confirmed in vivo by several methods. The first will measure the clearance rate of labeled thrombopoietin in normal and thrombocytopenic animals. And the second will block clearance by infusion of antibody against the count, considerable preliminary evidence suggests that its production is affected by other cytokines and small molecules. Tissue culture systems will be established to determine the mechanism by which thrombopoietin methods will be used to determine exactly which hepatic cell type is responsible for thrombopoietin production. These experiments will have great relevance to clinical applications of thrombopoietin. Moreover the studies will examine a number of clinical conditions of altered platelet production ranging from iron deficiency to essential thrombocythemia. They should show that some disorders are due to increased production of thrombopoietin while others are due to decreased clearance of thrombopoietin by the platelet.
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