Blood cells originate from a population of hematopoietic stem cells which are typically defined as pluripotent cells with self-renewal capacity. Although the regenerative capacity of the hematopoietic system is impressive, the replicative potential of stem cells nevertheless appears finite. Because the number of times stem cells can divide is an important consideration in the development of novel therapeutic strategies, including stem cell transplantation, ex vivo expansion and gene therapy, studies that may help define or extend the replicative potential of stem cells are of general interest. Previous studies funded by this grant with purified human candidate stem cells have shown that their functional properties change dramatically during ontogeny, that loss in their proliferative potential correlates with measurable changes in the number of telomere repeats and that they express low levels of telomerase. Here we propose to further examine the role of telomeres in the replication and differentiation of hematopoietic stem cells. For this purpose, we will use novel techniques developed in our laboratory to measure the length of telomere repeats in individual cells and on specific chromosomes in hematopoietic cells before and after transplantation and ex vivo expansion. We will use retroviral-mediated transfer of the human telomerase reverse transcriptase gene to examine the effect of artificial telomere elongation on stem cell properties that can be measured in vitro. Finally, we will specifically examine the role of l7p telomeres in the differentiation, replication and senescence of hematopoietic cells.
The specific aims of the proposal are 1) to study the telomere length in myeloid and lymphoid donor cells of short- and long-term survivors of allotransplants in comparison with cells of the transplant donor; 2) to study the effect of artificial telomere elongation on the proliferation, differentiation and replicative potential of purified """"""""candidate"""""""" stem cells; and 3) to study the role of l7p telomeres in the proliferation and replicative senescence of hematopoietic cells.
