Quiescence of hematopoietic stem cells and maintenance of the stem cell pool is not dependent on TGF-β signaling in vivo

Abstract In this issue’s Blood Advances Talk, Lapidot and Golan discuss how changes in daily light regulate hematopoiesis. This fascinating mechanism helps control the process of maintaining the hematopoietic stem cell pool while promoting sufficient differentiation to supply adequate numbers of functional blood cells. We hope you enjoy listening to this interesting topic.

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Hepatic Leukemia Factor Maintains Quiescence of Hematopoietic Stem Cells and Protects the Stem Cell Pool during Regeneration

Cell Reports ◽

10.1016/j.celrep.2017.11.084 ◽

2017 ◽

Vol 21 (12) ◽

pp. 3514-3523 ◽

Cited By ~ 25

Author(s):

Karolina Komorowska ◽

Alexander Doyle ◽

Martin Wahlestedt ◽

Agatheeswaran Subramaniam ◽

Shubhranshu Debnath ◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Hematopoietic Stem Cells ◽

Hematopoietic Stem ◽

Cell Pool ◽

Stem Cell Pool

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Clonal-level lineage commitment pathways of hematopoietic stem cells in vivo

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1801480116 ◽

2019 ◽

Vol 116 (4) ◽

pp. 1447-1456 ◽

Cited By ~ 29

Author(s):

Rong Lu ◽

Agnieszka Czechowicz ◽

Jun Seita ◽

Du Jiang ◽

Irving L. Weissman

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Hematopoietic Stem Cells ◽

Hematopoietic Stem Cell ◽

Lineage Commitment ◽

Self Renewal ◽

Hematopoietic Stem ◽

Hematopoietic System ◽

Different Levels

While the aggregate differentiation of the hematopoietic stem cell (HSC) population has been extensively studied, little is known about the lineage commitment process of individual HSC clones. Here, we provide lineage commitment maps of HSC clones under homeostasis and after perturbations of the endogenous hematopoietic system. Under homeostasis, all donor-derived HSC clones regenerate blood homogeneously throughout all measured stages and lineages of hematopoiesis. In contrast, after the hematopoietic system has been perturbed by irradiation or by an antagonistic anti-ckit antibody, only a small fraction of donor-derived HSC clones differentiate. Some of these clones dominantly expand and exhibit lineage bias. We identified the cellular origins of clonal dominance and lineage bias and uncovered the lineage commitment pathways that lead HSC clones to different levels of self-renewal and blood production under various transplantation conditions. This study reveals surprising alterations in HSC fate decisions directed by conditioning and identifies the key hematopoiesis stages that may be manipulated to control blood production and balance.

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Life span of multipotential hematopoietic stem cells in vivo.

Journal of Experimental Medicine ◽

10.1084/jem.171.5.1407 ◽

1990 ◽

Vol 171 (5) ◽

pp. 1407-1418 ◽

Cited By ~ 137

Author(s):

G Keller ◽

R Snodgrass

Keyword(s):

Stem Cells ◽

Gene Therapy ◽

Stem Cell ◽

Hematopoietic Stem Cells ◽

Strong Evidence ◽

Stem Cell Population ◽

Hematopoietic Stem ◽

Somatic Tissues ◽

Exogenous Factors

The findings reported in this study highlight several important features of the development of hematopoietic stem cells after transplantation into irradiated recipients. First, they demonstrate the existence of a class of primitive multipotential stem cells that can function for a significant portion of the lifetime of a mouse (15 mo). In addition, they clearly show that these primitive stem cells can be infected with recombinant retroviruses and thus would be appropriate targets for gene therapy in somatic tissues. Second, our data indicate that the progeny of some, but not all, of the primitive stem cells have fully expanded into the various hematopoietic lineages by 2 mo after reconstitution. Finally, our analysis of the secondary recipients provides strong evidence suggesting that the primitive stem cell population can actually clonally expand. Our current experiments are aimed at determining the extent to which this expansion can occur and whether or not this expansion can be influenced by exogenous factors.

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Native associations of early hematopoietic stem cells and stromal cells isolated in bone marrow cell aggregates

Blood ◽

10.1182/blood.v83.2.361.bloodjournal832361 ◽

1994 ◽

Vol 83 (2) ◽

pp. 361-369 ◽

Cited By ~ 1

Author(s):

PE Funk ◽

PW Kincade ◽

PL Witte

Keyword(s):

Stem Cells ◽

Bone Marrow ◽

Stem Cell ◽

Hematopoietic Stem Cells ◽

Stromal Cells ◽

Stromal Cell ◽

Hematopoietic Cells ◽

Hematopoietic Stem ◽

Factor C

In suspensions of murine bone marrow, many stromal cells are tightly entwined with hematopoietic cells. These cellular aggregations appear to exist normally within the marrow. Previous studies showed that lymphocytes and stem cells adhered to stromal cells via vascular cell adhesion molecule 1 (VCAM1). Injection of anti-VCAM1 antibody into mice disrupts the aggregates, showing the importance of VCAM1 in the adhesion between stromal cells and hematopoietic cells in vivo. Early hematopoietic stem cells were shown to be enriched in aggregates by using a limiting-dilution culture assay. Myeloid progenitors responsive to WEHI-3CM in combination with stem cell factor (c-kit ligand) and B220- B-cell progenitors responsive to insulin-like growth factor-1 in combination with interleukin-7 are not enriched. We propose a scheme of stromal cell-hematopoietic cell interactions based on the cell types selectively retained within the aggregates. The existence of these aggregates as native elements of bone marrow organization presents a novel means to study in vivo stem cell-stromal cell interaction.

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Hematopoietic Stem Cells Have an Intrinsic Expansion Limit

Blood ◽

10.1182/blood.v120.21.4749.4749 ◽

2012 ◽

Vol 120 (21) ◽

pp. 4749-4749

Author(s):

Shanti Rojas-Sutterlin ◽

André Haman ◽

Trang Hoang

Keyword(s):

Cell Cycle ◽

Stem Cell ◽

Chemotherapy Regimen ◽

Gene Dosage ◽

Hematopoietic Stem ◽

Cell Functions ◽

Cell Pool ◽

Stem Cell Pool

Abstract Abstract 4749 Hematopoietic stem cell (HSC) transplantation is the first successful cellular therapy and remains the only treatment providing long-term cure in acute myeloblastic leukemia. At the apex of the hematopoietic system, quiescent HSCs are spared by chemotherapeutic treatments that target proliferating cells and therefore can regenerate the entire blood system of a patient after drug exposure. Nevertheless, the consequence of repeated chemotherapy regimen on HSC function remains to be clarified. We previously showed that Scl/Tal1 gene dosage regulates HSC quiescence and functions when transplanted at limiting dilutions (Lacombe et al., 2010). In the present study, we investigate how massive expansion in vivo influences stem cell functions. To address this question, we optimized a protocol based on 5-fluorouracil (5-FU), an antimetabolite that has been used to treat colon, rectum, and head and neck cancers. In addition, we used Scl+/− mice to address the role of Scl in controlling HSCs expansion post-5-FU. We show that within 7 days following 5-FU treatment, HSCs exit quiescence and enter the cell cycle. To deplete cycling HSCs, we injected a second dose of 5-FU and showed that the stem cell pool was disseminated. Nonetheless, the remaining HSCs proliferated extensively to re-establish the HSC pool, which was twice larger than that of untreated mice. At this point, most HSCs have exited the cell cycle and were back to quiescence. Despite a near normal stem cell pool size and a quiescent status, HSCs from these 5-FU treated mice could not compete against untreated cells to regenerate the host in transplantation assays. Furthermore, we show that this extensive proliferation in vivo severely impaired the clonal expansion of individual HSC as measured by the mean activity of stem cell (MAS). Our results demonstrate that HSCs lose their competitive potential after two 5-FU treatments, suggesting that HSCs have an intrinsic expansion limit beyond which their regenerative potential is impaired. In addition, Scl is haplodeficient for cell cycle entry and cell division but Scl gene dosage does not affect this expansion limit. Therefore, our data dissociate the control of HSC expansion under extensive proliferative stress from cell cycle control during steady state. We surmise that chemotherapy regimen based on repeated administration of 5-FU or other antimetabolites are likely to severely impair long-term stem cell functions. Disclosures: No relevant conflicts of interest to declare.

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Canonical Notch Signaling Is Dispensable for the Maintenance of Adult Hematopoietic Stem Cells.

Blood ◽

10.1182/blood.v106.11.267.267 ◽

2005 ◽

Vol 106 (11) ◽

pp. 267-267 ◽

Cited By ~ 37

Author(s):

Ivan Maillard ◽

Seth E. Pross ◽

Olga Shestova ◽

Hong Sai ◽

Jon C. Aster ◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Hematopoietic Stem Cells ◽

Notch Signaling ◽

Transcriptional Activation ◽

Cell Function ◽

Fetal Life ◽

Hematopoietic Stem

Abstract Canonical Notch signaling operates through a highly conserved pathway that regulates the differentiation and homeostasis of hematopoietic cells. Ligand-receptor binding initiates proteolytic release of the Notch intracellular domain (ICN) which migrates to the nucleus, binds the transcription factor CSL/RBPJk and activates target genes through the recruitment of transcriptional coactivators of the Mastermind-like family (MAML). Notch signaling is essential for the emergence of hematopoietic stem cells (HSCs) during fetal life, but its effects on adult HSCs are controversial. In gain-of-function experiments, activation of Notch signaling in adult HSCs increased their self-renewal potential in vitro and in vivo. However, loss-of-function studies have provided conflicting results as to the role of physiological Notch signaling in HSC maintenance and homeostasis. To address this question, we expressed DNMAML1, a GFP-tagged pan-inhibitor of Notch signaling, in mouse HSCs. We have shown previously that DNMAML1 interferes with the formation of the ICN/CSL/MAML transcriptional activation complex and blocks signaling from all four Notch receptors (Notch1-4) (Maillard, Blood 2004). Transfer of DNMAML1-transduced bone marrow (BM) as compared to control GFP-transduced BM into lethally irradiated recipients gave rise to similar long-term stable expression of GFP for at least 6 months after transplant. DNMAML1 and GFP-transduced cells contributed equally to all hematopoietic lineages, except to the T cell and marginal zone B cell lineages, which are Notch-dependent. Expression of DNMAML1 did not affect the size of the BM progenitor compartment (Lin negative, Sca-1 positive, c-Kit high, or LSK cells), or the proportion of LSK cells that were negative for Flt3 and L-Selectin expression (containing long-term HSCs). The stem cell function of DNMAML1-transduced LSK cells was further assessed with in vivo competitive repopulation assays in lethally irradiated recipients. DNMAML1 and GFP-transduced LSK cells competed equally well with wild-type BM, as judged by their contribution to the myeloid lineage up to 4 months post-transplant, through two successive rounds of transplantation. Our data indicate that canonical Notch signaling is dispensable for the maintenance of stem cell function in adult HSCs.

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Mitochondria in the maintenance of hematopoietic stem cells: new perspectives and opportunities

Blood ◽

10.1182/blood-2018-10-808873 ◽

2019 ◽

Vol 133 (18) ◽

pp. 1943-1952 ◽

Cited By ~ 16

Author(s):

Marie-Dominique Filippi ◽

Saghi Ghaffari

Keyword(s):

Stem Cells ◽

Hematopoietic Stem Cells ◽

Blood Cells ◽

Self Renewal ◽

Hematopoietic Stem ◽

Hematopoietic System ◽

Cellular Energy ◽

Cell Pool ◽

Stem Cell Pool

Abstract The hematopoietic system produces new blood cells throughout life. Mature blood cells all derived from a pool of rare long-lived hematopoietic stem cells (HSCs) that are mostly quiescent but occasionally divide and self-renew to maintain the stem cell pool and to insure the continuous replenishment of blood cells. Mitochondria have recently emerged as critical not only for HSC differentiation and commitment but also for HSC homeostasis. Mitochondria are dynamic organelles that orchestrate a number of fundamental metabolic and signaling processes, producing most of the cellular energy via oxidative phosphorylation. HSCs have a relatively high amount of mitochondria that are mostly inactive. Here, we review recent advances in our understanding of the role of mitochondria in HSC homeostasis and discuss, among other topics, how mitochondrial dynamism and quality control might be implicated in HSC fate, self-renewal, and regenerative potential.

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In Vivo Fate Tracing Studies Using the SCL Stem Cell Enhancer: Embryonic Hematopoietic Stem Cells Significantly Contribute to Adult Hematopoiesis.

Blood ◽

10.1182/blood.v104.11.559.559 ◽

2004 ◽

Vol 104 (11) ◽

pp. 559-559

Author(s):

Joachim R. Gothert ◽

Sonja E. Gustin ◽

Mark A. Hall ◽

Anthony R. Green ◽

Berthold Gottgens ◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Hematopoietic Stem Cells ◽

De Novo ◽

Genetic Manipulation ◽

Fetal Liver ◽

Hematopoietic Stem ◽

Reporter Mice ◽

First Time

Abstract Evidence for the direct lineage relationship between embryonic and adult hematopoietic stem cells (HSCs) in the mouse is primarily indirect. In order to study this relationship in a direct manner we expressed the tamoxifen-inducible Cre-ERT-recombinase under the control of the SCL-stem-cell-enhancer in transgenic mice (HSC-SCL-Cre-ERT). To determine functionality, HSC-SCL-Cre-ERT transgenics were bred with the Cre-reporter-mice ROSA26R and R26R-EYFP. Flow-cytometric and transplantation studies revealed tamoxifen-dependent recombination occurring in more than 90% of adult long-term HSCs, whereas the targeted proportion within mature progenitor populations was significantly lower. Moreover, the transgene was able to irreversibly tag embryonic HSCs on days 10 and 11 of gestation. These cells contributed to bone marrow hematopoiesis five months later. In order to investigate whether the de novo HSC-generation is completed during embryogenesis, HSC-SCL-Cre-ERT marked fetal liver cells were transplanted into adult recipients. Strikingly, the proportion of marked cells within the transplanted and the in vivo-remaining HSC-compartment was not different, implying that no further HSC-generation occurred during late fetal and neonatal stages of development. These data demonstrate for the first time the direct lineage relationship between mid-gestation embryonic and adult HSCs in the mouse. Additionally, the HSC-SCL-Cre-ERT mice will provide a valuable tool to achieve temporally controlled genetic manipulation of HSCs.

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In Vitro and In Vivo Evidence That Umbilical Cord Blood (UCB)-Derived CD45-/SSEA-4+/OCT-4+/CD133+/CXCR4+/Lin- Very Small Embryonic/Epiblast Like Stem Cells (VSELs) Do Not Contain Clonogenic Hematopoietic Progenitors but Are Highly Enriched in More Primitive Stem Cells - Novel View On Hierarchy of UCB Stem Cell Compartment.

Blood ◽

10.1182/blood.v114.22.35.35 ◽

2009 ◽

Vol 114 (22) ◽

pp. 35-35 ◽

Cited By ~ 2

Author(s):

Ewa K. Zuba-Surma ◽

Izabela Klich ◽

Marcin Wysoczynski ◽

Nicholas J Greco ◽

Mary J. Laughlin ◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Hematopoietic Stem Cells ◽

Cord Blood ◽

Umbilical Cord ◽

Umbilical Cord Blood ◽

Hematopoietic Stem ◽

Hematopoietic Activity

Abstract Abstract 35 Recently, we identified in umbilical cord blood (UCB) a population of very small embryonic/epiblast-like (VSEL) stem cells (Leukemia 2007;21:297–303) that are i) smaller than erythrocytes, ii) SSEA-4+/Oct-4+/CD133+/CXCR4+/Lin−/CD45−, iii) respond to SDF-1 gradient and iv) possess large nuclei containing primitive euchromatin. We have demonstrated in vitro that UCB-derived VSELs did not reveal hematopoietic activity freshly after isolation, but grow hematopoietic colonies following co-culture/activation over OP-9 cells. To investigate the hierarchy of UCB-derived, CD45 negative VSELs, we employed staining with Aldefluor - detecting aldehyde dehydrogenase (ALDH), the enzyme expressed in primitive hematopoietic cells. Subsequently, we sorted CD45−/CD133+/ALDHhigh and CD45−/CD133+/ALDHlow sub-fractions of VSELs from UCB samples and established that freshly sorted from UCB VSELs in contrast to sorted CD45+/ CD133+/ALDHhigh and CD45+/CD133+/ALDHlow hematopoietic stem cells (HSC) did not grow colonies in vitro. However, when CD45− VSELs were activated/expanded over OP-9 stroma cells, they exhibit hematopoietic potential and grew in routine methylcellulose cultures hematopoietic colonies composed of CD45+ cells. Interestingly, while CD45−/CD133+/ALDHhigh VSELs gave raise to hematopoietic colonies after the first replating, the formation of colonies by CD45−/CD133+/ALDHlow VSELs was somehow delayed, what suggest that they needed more time to acquire hematopoietic commitment. Thus our in vitro data indicate that both populations of CD45− cells may acquire hematopoietic potential; however hematopoietic specification is delayed for CD45−/CD133+/ALDHlow cells, suggesting their more primitive nature. In parallel, real time PCR analysis confirmed that while freshly isolated CD45−/CD133+/ALDHhigh VSELs express more hematopoietic transcripts (e.g., c-myb, 80.2±27.4 fold difference), CD45−/CD133+/ALDHlow exhibit higher levels of pluripotent stem cell markers (e.g., Oct-4, 119.5±15.5 fold difference as compared to total UCB mononuclear cells) (Figure 1 panel A). Next hematopoietic potential of UCB-derived VSELs was tested in vivo after transplantation into NOD/SCID mice (Figure 1 panel B and C). We noticed that both CD45−/CD133+/ALDHhigh and CD45−/CD133+/ALDHlow VSELs, give rise to human lympho-hematopoietic chimerism in lethally irradiated NOD/SCID mice as assayed 4–6 weeks after transplantation. The level of human hematopoietic CD45+ cells in murine peripheral blood (PB), bone marrow (BM) and spleen (SP) were comparable for both transplanted UCB-VSELs fractions - 7.1±2.9% (PB), 23.2±0.2% (SP) and 25.2±1.0% (BM). In conclusion, our data suggest that freshly isolated very small CD45 negative UCB-VSELs are depleted from clonogeneic progenitors, however they are highly enriched for primitive HSC. Based on our in vitro and in vivo data we postulate following hierarchy of hematopoietic stem cells in UCB (from most primitive to more differentiated) i) CD45−/CD133+/ALDHlow, ii) CD45−/CD133+/ALDHhigh , iii) CD45+/CD133+/ALDHlow and iv) CD45−/CD133+/ALDHhigh. We also postulate that as we have already shown for murine BM-derived VSELs, human UCB-derived CD45 negative VSELs correspond to a population of most primitive long term repopulating HSC (LT-HSC). Of note, we also found that currently employed, routine UCB processing strategies may lead up to ∼50% unwanted loss of these small cells that are endowed with such remarkable hematopoietic activity! Disclosures: No relevant conflicts of interest to declare.

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