Andreas Trumpp

Hematopoietic Stem Cells Reversibly Switch from Dormancy to Self-Renewal during Homeostasis and Repair

Bone marrow hematopoietic stem cells (HSCs) are crucial to maintain lifelong production of all blood cells. Although HSCs divide infrequently, it is thought that the entire HSC pool turns over every few weeks, suggesting that HSCs regularly enter and exit cell cycle. Here, we combine flow cytometry with label-retaining assays (BrdU and histone H2B-GFP) to identify a population of dormant mouse HSCs (d-HSCs) within the lin(-)Sca1+cKit+CD150+CD48(-)CD34(-) population. Computational modeling suggests that d-HSCs divide about every 145 days, or five times per lifetime. d-HSCs harbor the vast majority of multilineage long-term self-renewal activity. While they form a silent reservoir of the most potent HSCs during homeostasis, they are efficiently activated to self-renew in response to bone marrow injury or G-CSF stimulation. After re-establishment of homeostasis, activated HSCs return to dormancy, suggesting that HSCs are not stochastically entering the cell cycle but reversibly switch from dormancy to self-renewal under conditions of hematopoietic stress.

Bone-marrow haematopoietic-stem-cell niches.

Adult stem cells hold many promises for future clinical applications and regenerative medicine. The haematopoietic stem cell (HSC) is the best-characterized somatic stem cell so far, but in vitro expansion has been unsuccessful, limiting the future therapeutic potential of these cells. Here we review recent progress in characterizing the composition of the HSC bone-marrow microenvironment, known as the HSC niche. During homeostasis, HSCs, and therefore putative bone-marrow HSC niches, are located near bone surfaces or are associated with the sinusoidal endothelium. The molecular crosstalk between HSCs and the cellular constituents of these niches is thought to control the balance between HSC self-renewal and differentiation, indicating that future successful expansion of HSCs for therapeutic use will require three-dimensional reconstruction of a stem-cell–niche unit.

IFNα activates dormant haematopoietic stem cells in vivo.

Maintenance of the blood system is dependent on dormant haematopoietic stem cells (HSCs) with long-term self-renewal capacity. After injury these cells are induced to proliferate to quickly re-establish homeostasis. The signalling molecules promoting the exit of HSCs out of the dormant stage remain largely unknown. Here we show that in response to treatment of mice with interferon-α (IFNα), HSCs efficiently exit G0 and enter an active cell cycle. HSCs respond to IFNα treatment by the increased phosphorylation of STAT1 and PKB/Akt (also known as AKT1), the expression of IFNα target genes, and the upregulation of stem cell antigen-1 (Sca-1, also known as LY6A). HSCs lacking the IFNα/β receptor (IFNAR), STAT1 (ref. 3) or Sca-1 (ref. 4) are insensitive to IFNα stimulation, demonstrating that STAT1 and Sca-1 mediate IFNα-induced HSC proliferation. Although dormant HSCs are resistant to the anti-proliferative chemotherapeutic agent 5-fluoro-uracil, HSCs pre-treated (primed) with IFNα and thus induced to proliferate are efficiently eliminated by 5-fluoro-uracil exposure in vivo. Conversely, HSCs chronically activated by IFNα are functionally compromised and are rapidly out-competed by non-activatable Ifnar-/- cells in competitive repopulation assays. Whereas chronic activation of the IFNα pathway in HSCs impairs their function, acute IFNα treatment promotes the proliferation of dormant HSCs in vivo. These data may help to clarify the so far unexplained clinical effects of IFNα on leukaemic cells, and raise the possibility for new applications of type I interferons to target cancer stem cells.

Identification of a population of blood circulating tumor cells from breast cancer patients that initiates metastasis in a xenograft assay.

It has been hypothesized that carcinoma metastasis is initiated by a subpopulation of circulating tumor cells (CTCs) found in the blood of patients. However, although the presence of CTCs is an indicator of poor prognosis in several carcinoma entities, the existence and phenotype of metastasis-initiating cells (MICs) among CTCs has not been experimentally demonstrated. Here we developed a xenograft assay and used it to show that primary human luminal breast cancer CTCs contain MICs that give rise to bone, lung and liver metastases in mice. These MIC-containing CTC populations expressed EPCAM, CD44, CD47 and MET. In a small cohort of patients with metastases, the number of EPCAM+CD44+CD47+MET+ CTCs, but not of bulk EPCAM+ CTCs, correlated with lower overall survival and increased number of metastasic sites. These data describe functional circulating MICs and associated markers, which may aid the design of better tools to diagnose and treat metastatic breast cancer.

The bone marrow stem cell niche grows up: mesenchymal stem cells and macrophages move in

Ehninger and Trumpp discuss the role of monocytes/macrophages and other niche cells in the regulation of HSC mobilization and retention.

Transcriptional Heterogeneity and Lineage Commitment in Myeloid Progenitors

Within the bone marrow, stem cells differentiate and give rise to diverse blood cell types and functions. Currently, hematopoietic progenitors are defined using surface markers combined with functional assays that are not directly linked with in vivo differentiation potential or gene regulatory mechanisms. Here, we comprehensively map myeloid progenitor subpopulations by transcriptional sorting of single cells from the bone marrow. We describe multiple progenitor subgroups, showing unexpected transcriptional priming toward seven differentiation fates but no progenitors with a mixed state. Transcriptional differentiation is correlated with combinations of known and previously undefined transcription factors, suggesting that the process is tightly regulated. Histone maps and knockout assays are consistent with early transcriptional priming, while traditional transplantation experiments suggest that in vivo priming may still allow for plasticity given strong perturbations. These data establish a reference model and general framework for studying hematopoiesis at single-cell resolution.

Awakening dormant haematopoietic stem cells

Haematopoietic stem cells (HSCs) in mouse bone marrow are located in specialized niches as single cells. During homeostasis, signals from this environment keep some HSCs dormant, which preserves long-term self-renewal potential, while other HSCs actively self renew to maintain haematopoiesis. In response to haematopoietic stress, dormant HSCs become activated and rapidly replenish the haematopoietic system. Interestingly, three factors — granulocyte colony-stimulating factor, interferon-α and arsenic trioxide — have been shown to efficiently activate dormant stem cells and thereby could break their resistance to anti-proliferative chemotherapeutics. Thus, we propose that two-step strategies could target resistant leukaemic stem cells by priming tumours with activators of dormancy followed by chemotherapy or targeted therapies.

THE EVOLVING CONCEPT OF CANCER AND METASTASIS STEM CELLS

The cancer stem cell (CSC) concept, which arose more than a decade ago, proposed that tumor growth is sustained by a subpopulation of highly malignant cancerous cells. These cells, termed CSCs, comprise the top of the tumor cell hierarchy and have been isolated from many leukemias and solid tumors. Recent work has discovered that this hierarchy is embedded within a genetically heterogeneous tumor, in which various related but distinct subclones compete within the tumor mass. Thus, genetically distinct CSCs exist on top of each subclone, revealing a highly complex cellular composition of tumors. The CSC concept has therefore evolved to better model the complex and highly dynamic processes of tumorigenesis, tumor relapse, and metastasis.

Telomere dysfunction induces environmental alterations limiting hematopoietic stem cell function and engraftment

Cell-intrinsic checkpoints limit the proliferative capacity of primary cells in response to telomere dysfunction. It is not known, however, whether telomere dysfunction contributes to cell-extrinsic alterations that impair stem cell function and organ homeostasis. Here we show that telomere dysfunction provokes defects of the hematopoietic environment that impair B lymphopoiesis but increase myeloid proliferation in aging telomerase knockout (Terc−/−) mice. Moreover, the dysfunctional environment limited the engraftment of transplanted wild-type hematopoietic stem cells (HSCs). Dysfunction of the hematopoietic environment was age dependent and correlated with progressive telomere shortening in bone marrow stromal cells. Telomere dysfunction impaired mesenchymal progenitor cell function, reduced the capacity of bone marrow stromal cells to maintain functional HSCs, and increased the expression of various cytokines, including granulocyte colony-stimulating factor (G-CSF), in the plasma of aging mice. Administration of G-CSF to wild-type mice mimicked some of the defects seen in aging Terc−/− mice, including impairment of B lymphopoiesis and HSC engraftment. Conversely, inhibition of G-CSF improved HSC engraftment in aged Terc−/− mice. Taken together, these results show that telomere dysfunction induces alterations of the environment that can have implications for organismal aging and cell transplantation therapies.

Exit from dormancy provokes DNA-damage-induced attrition in haematopoietic stem cells

Haematopoietic stem cells (HSCs) are responsible for the lifelong production of blood cells. The accumulation of DNA damage in HSCs is a hallmark of ageing and is probably a major contributing factor in age-related tissue degeneration and malignant transformation. A number of accelerated ageing syndromes are associated with defective DNA repair and genomic instability, including the most common inherited bone marrow failure syndrome, Fanconi anaemia. However, the physiological source of DNA damage in HSCs from both normal and diseased individuals remains unclear. Here we show in mice that DNA damage is a direct consequence of inducing HSCs to exit their homeostatic quiescent state in response to conditions that model physiological stress, such as infection or chronic blood loss. Repeated activation of HSCs out of their dormant state provoked the attrition of normal HSCs and, in the case of mice with a non-functional Fanconi anaemia DNA repair pathway, led to a complete collapse of the haematopoietic system, which phenocopied the highly penetrant bone marrow failure seen in Fanconi anaemia patients. Our findings establish a novel link between physiological stress and DNA damage in normal HSCs and provide a mechanistic explanation for the universal accumulation of DNA damage in HSCs during ageing and the accelerated failure of the haematopoietic system in Fanconi anaemia patients.