The Applicant proposes to extend ongoing mathematical and transplantation studies of stem cell dynamics and kinetics in female Safari cats, which by virtue of being heterozygous for the X-chromosome linked enzyme G-PD allow one to observe substantial variability in allelic stem cell contributions over time. In previous studies the PI has found that after transplantation with 1-2 x 107 nucleated autologous marrow cells/kg, large fluctuations in the G-6-PD phenotypes of progenitors were seen in each animal, suggesting that stem cell contributions to this compartment varied., but 1-4.5 years later and in some cats, >90% of BFU-E, CFU-GM, RBCs, polys and >80%T cells expressed a single parental G-6PD phenotype. This pattern recalled similar loss of disequilibrium observed in murine transplant studies, but with far longer kinetics, suggesting that stem cell kinetics could reflect animal size and/or longevity. Mathematical modeling of these kinetic patterns suggests that feline hematopoietic stem cells represent 1/106-1/107 nucleated marrow cell, and that these stem cells do not self-renew more frequently than once every 3 weeks.
Specific Aim 1 proposes to validate detailed computer models by performing transplants with varying numbers of cells, by retransplantation, and by experiments with retroviral marking and modification with HoxB4.
Specific Aim 2 proposes to refine simulations with additional mathematical analyses, considering hypotheses such as progressive loss of self-renewal probability with each stem cell division as well as modeling murine and human stem cell kinetics as well.
Specific Aim 3 proposes to study stem cell kinetics with aging, as well as to evaluate the contribution of genotype to stem cell behavior. Taken together, the PI believes these studies will provide insight into the kinetics and behavior of hematopoietic stem cells in large animals, to optimal strategies for gene therapy, and to disease pathogenesis.
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