Hematopoietic stem cells expand during serial transplantation in vivo without apparent exhaustion

Abstract

Whether hematopoietic stem cells can proliferate without limit, or whether their regenerative capacity declines with repeated division, has been debated for decades. Prevailing opinion favours an intrinsic 'decline', a view based on the finite degree to which murine bone marrow can be serially transplanted, the diminished self-renewal of spleen colony-forming cells (CFU-s) subjected to repeated passage, and the failure of stem cells to regenerate to normal levels after even a single transplantation. However, serial transfer experiments did not specifically monitor input and output of long-lived stem cells (long-term reconstituting cells, LTRCs), leaving competing interpretations unresolved. We have re-examined the issue by quantitating 7-12 month LTRCs during sequential transplantations. Although these cells recovered to only 4% of normal levels after primary bone marrow transplantation, at each passage they increased around 10-fold relative to the amount transplanted, attaining an estimated cumulative expansion of 8400-fold over the original input after four transfers. Expansion was limited by transfer of increasing numbers of marrow cells and specifically of LRTCs, suggesting an extrinsically determined ceiling to stem cell growth. Conversely, expansion was enhanced in vivo by administration of stem cell factor (SCF, c-kit ligand) and interleukin-11. The results challenge the view that expansion of passaged stem cells is limited by exhaustion, and indicate that augmentation after transplant is limited by extrinsic mechanisms whose effects are reversible either by further transfer of the stem cells into irradiated hosts or by administration of exogenous cytokines.

Enforced expression of cyclin D2 enhances the proliferative potential of myeloid progenitors, accelerates in vivo myeloid reconstitution, and promotes rescue of mice from lethal myeloablation

“…The reason for this remains unclear, but because D-type cyclins are known to have a short half-life as a result of ubiquitin-mediated proteolysis, 31 it seems likely that mechanisms are in place in HSCs to tightly regulate cyclin D2 expression through degradation. This could be important because the size of the HSC pool is known to be tightly regulated, 32,33 and because HSCs are frequent targets for leukemic transformation. 34 These findings clearly demonstrate that the repopulating ability of hematopoietic progenitors can be improved by overexpression of positive regulators of cell cycle progression, and that this might have an effect in clinical transplantation.…”

Primitive human hematopoietic cells give rise to differentially specified daughter cells upon their initial cell division

“…Beside this, they also have the potential to expand in vivo as revealed by sequential transplantation experiments using limiting numbers of mouse HSCs to reconstitute the hematopoietic systems of primary and secondary lethally irradiated hosts. 2,3 However, although HSCs can be maintained in vitro in close contact to adequate stroma cells, [4][5][6][7][8] no evaluated in vitro condition has been reported so far, which supports the expansion of these cells over a period of several weeks. These findings demonstrate that the surrounding environment has a major influence on the cell fate of HSCs and their daughter cells.…”

Gene therapy approaches for stem cell protection

“…There is increasing evidence that the intrinsically limited competitive repopulation capacity of donor cells may be enhanced by increasing their resistance to oxidative stress. [5][6][7] Mice deficient in p21 cip/wafl or p18 INCÀ4C demonstrate altered self-renewal capacity and altered competitive repopulation capacity compared to marrow stem cells from litter mates not lacking these tumor suppressor gene or cell cycle altering gene functions. 8,9 Antioxidant approaches toward enhancing stem cell protection include transfection of the transgenes for manganese superoxide dismutase (MnSOD), catalase, glutathione, peroxidase, metallothione and others.…”

“…The concept of a limited number of niches/specific environmental sites for the multilineage or totipotential hematopoietic stem cells derives from work in experimental animals demonstrating a plateau maximum number of pluripotent stem cells which can engraft. 5,7,18 The competitive repopulation assay compares two purified (low-density gradient purified) hematopoietic stem cell populations, injected intravenously into a third irradiated murine host. [5][6][7] By comparing different ratios of the two donor stem cell populations for genotypic markers to distinguish them from each other and from the recipient, one can calculate the maximum number of stem cell sites in the host hematopoietic system.…”