Basic Aspects of Hematopoietic Stem Cells and Aging

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Project Summary: Changes in hematopoiesis and hematopoietic stem cells are an important component of aging. Many critical studies on the roles in hematopoietic aging of telomere dysfunction, cellular senescence and induction of the senescence-associated secretory phenotype, sitruin family effects, mitochondrial metabolism, the mechanistic target of rapamycin (mTOR), decline in the immune system and changes induced by a variety of toxic entities many of which involve Reactive Oxygen Species (ROS) were carried out employing highly purified hematopoietic stem cells. Studies on purified murine hematopoietic stem cells as exemplified by lineage negative c- kit+Sca-1+ (LSK) cells with or without CD150 marking, have indicated that with aging the number of phenotypically defined stem cells increases while the function of these cells decreases and myeloid skewing occurs. Much of the work on hematopoiesis in the aged has relied on studies of purified stem cells and is based on the assumption that these cells are non cycling or dormant. However, our work has shown that the long-term multi-lineage repopulating cell in lethally irradiated hosts is actively cycling and always changing. The key to these findings was the study of whole un-separated marrow stem cells with long-term multi-lineage engraftment as the assay. We feel that most studies to date on hematopoietic stem cells in aging have used purified stem cells, which are not representative of the true marrow stem cell populations. We propose here to study the content, cell cycle status and differentiation characteristics of un-separated marrow stem cells in young and aged mice and in male and female mice. We will separate cells into G0, G1 and S/G2/M fractions and evaluate engraftment. We will also utilize tritiated thymidine suicide to assess cycle status of stem cells in un- separated marrow and in vivo bromodeoxyuridine (BrdU) labeling to assess the flux of stem cells through cell cycle. These studies should give an important new base for further studies on the effects of aging on hematopoietic stem cells.