Platelets are transfused for a wide range of thrombotic deficiencies, but there areproblems. Platelet collection typically requires pooling harvests from multiple donors.Platelet transfusion risks from bacterial contamination, blood-borne pathogens, andalloimmunization are compounded because patients receive platelets from many donors.Production of autologous or compatible platelets by megakaryocytic cells (Mks) derivedfrom cultured hematopoietic stem and progenitor cells (HSPCs or CD34+ cells) wouldgreatly decrease these risks. However, generating 500 billion platelets for a singletransfusion using culture conditions that yield relatively pure (e 75%) Mk populationswould require 250 million CD34+ cells. This is equivalent to more than 50 umbilical cordblood harvests or 1-2 harvests of HSPCs from the peripheral blood of donors treated (ormobilized) with growth factors. In order for culture-derived platelet production to beeconomically feasible, it will be necessary to produce more Mk progenitors per CD34+cell, obtain a greater number of Mks per Mk progenitor, and increase Mk ploidy (platelet-producing potential). Our objective is to increase the ploidy of culture-derived Mks tolevels similar to those found in human bone marrow. We have shown that the vitaminnicotinamide (NIC) greatly increases Mk ploidy in culture. Since Mks in vivo produceseveral thousand platelets, we anticipate that Mks produced in culture with NIC couldgenerate 1000 platelets. Understanding the mechanisms responsible for NIC-mediatedincreases in Mk ploidy will facilitate regulatory approval for using NIC to produceplatelets for transplantation and is likely to lead to the discovery of even more effectiveconditions for Mk polyploidization. We propose to use RNA-interference-mediatedknockdown to test the hypothesis, based on our preliminary results, that NIC increasesMk ploidy via inhibition of the SIRT1 and SIRT2 Class III histone/protein deacetylases.We will then examine changes in the acetylation of SIRT target proteins involved in theregulation of the cell cycle and/or apoptosis. Finally, we will investigate the roles inmegakaryopoiesis of the most promising SIRT targets.
|Effective start/end date||9/15/09 → 2/29/12|
- National Heart, Lung, and Blood Institute (1R01HL093083-01A1)