| 1. |
- Blank Savukinas, Ulrika, et al.
(författare)
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Angptl4 maintains in vivo repopulation capacity of CD34(+) human cord blood cells.
- 2012
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Ingår i: European Journal of Haematology. - : Wiley. - 1600-0609 .- 0902-4441. ; 89:3, s. 198-205
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Tidskriftsartikel (refereegranskat)abstract
- OBJECTIVES: Methods to expand hematopoietic stem cells (HSCs) ex vivo encompass an attractive approach that would substantially broaden the clinical applicability of HSCs derived from cord blood. Recently, members of the Angiopoietin-like (Angptl) family of growth factors were shown to expand both murine and human HSCs. Specifically, Angptl5 has been implicated in the expansion of human NOD-SCID-repopulating cells (SRCs) ex vivo. Here, we sought to evaluate the potential of additional Angptls to expand human SRCs from cord blood. Additionally, the purpose of this study was to evaluate the reproducibility of Angptl-mediated expansion of SRCs across independent experiments. METHODS: Human CD34(+) cells from cord blood were cultured in vitro for eleven or eight days in the presence or absence of Angptls. The reconstitution capacity of expanded cells was subsequently measured in vivo by transplantation into NOD-SCID or NSG mice, and compared to that of uncultured cells. RESULTS: We report here that Angptl4 functions to maintain SRC-activity of CD34(+) CB-derived cells ex vivo as assayed in NOD-SCID and NSG mice. However, all Angptls tested, including Angptl1, 4, and 5, were associated with variation between experiments. CONCLUSION: Our findings indicate that Angptl4 and Angptl5 can lead to increased engraftment capacity of SRCs, but more frequently these factors are associated with maintenance of SRC-activity during ex vivo culture. Thus, Angptl-mediated expansion of SRCs ex vivo is associated with more inter-experimental variation than previously thought. We conclude that Angptls would be useful in instances where there is a need to maintain HSCs ex vivo, such as during transduction for gene therapy applications.
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| 2. |
- Blank Savukinas, Ulrika
(författare)
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Regulation of hematopoiesis by the Smad signaling pathway
- 2006
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Hematopoietic stem cells (HSCs) reside in the bone marrow (BM) of adult individuals and are ultimately responsible for the continuous production of blood cells throughout life. The regulation of HSCs in vivo is tightly regulated by both intrinsic and extrinsic factors. The Smad-signaling pathway is an evolutionary conserved signaling circuitry with critical functions during embryogenesis and throughout adult life, regulating diverse biological processes. The transforming growth factor-? (TGF-?) superfamily of ligands transduce their signals intracellularly through the Smad pathway. A large number of studies, the majority of which have been carried out in vitro, have cataloged TGF-? as a potent negative regulator of HSC proliferation. However, due to embryonic lethality of knockout mice, in vivo investigations of the role of TGF-? and the downstream Smad pathway in the context of adult hematopoiesis have been hampered. To address this, we made use of the Cre/loxP system for inducible gene deletion of two different components of the TGF-? signaling pathway, the type I TGF-? receptor and Smad4 respectively. In addition, retroviral mediated gene transfer to HSCs was used as a tool to block the entire Smad-signaling pathway, by overexpression of the inhibitory Smad7. Induced disruption of the type I TGF-? receptor in adult mice resulted in an inflammatory disorder with a lethal outcome 8-10 weeks post induction. However, all hematopoietic parameters were normal under steady state conditions as well as the regenerative- and self-renewal capacity of mutant HSCs as assessed by transplantation. Smad4 null HSCs exhibited impaired repopulative capacity in a competitive repopulation assay, a behavior that was exacerbated upon secondary transplantation. Overexpression of Smad7 in HSCs resulted in increased regenerative capacity upon secondary transplantation, with a normal lineage distribution. Taken together, our data suggests that the Smad pathway is a critical regulator of HSC self-renewal in vivo.
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| 3. |
- Blank Savukinas, Ulrika, et al.
(författare)
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Signaling pathways governing stem cell fate.
- 2008
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Ingår i: Blood. - : American Society of Hematology. - 1528-0020 .- 0006-4971. ; 111:2, s. 492-503
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Forskningsöversikt (refereegranskat)abstract
- Hematopoietic stem cells (HSCs) are historically the most thoroughly characterized type of adult stem cell, and the hematopoietic system has served as a principal model structure of stem-cell biology for several decades. However, paradoxically, although HSCs can be defined by function and even purified to near-homogeneity, the intricate molecular machinery and the signaling mechanisms regulating fate events, such as self-renewal and differentiation, have remained elusive. Recently, several developmentally conserved signaling pathways have emerged as important control devices of HSC fate, including Notch, Wingless-type (Wnt), Sonic hedgehog (Shh), and Smad pathways. HSCs reside in a complex environment in the bone marrow, providing a niche that optimally balances signals that control self-renewal and differentiation. These signaling circuits provide a valuable structure for our understanding of how HSC regulation occurs, concomitantly with providing information of how the bone marrow microenvironment couples and integrates extrinsic with intrinsic HSC fate determinants. It is the focus of this review to highlight some of the most recent developments concerning signaling pathways governing HSC fate.
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| 4. |
- Blank Savukinas, Ulrika, et al.
(författare)
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Smad7 promotes self-renewal of hematopoietic stem cells in vivo.
- 2006
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Ingår i: Blood. - : American Society of Hematology. - 1528-0020 .- 0006-4971. ; 108:13, s. 4246-4254
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Tidskriftsartikel (refereegranskat)abstract
- The Smad-signaling pathway downstream of the transforming growth factor–beta superfamily of ligands is an evolutionarily conserved signaling circuitry with critical functions in a wide variety of biologic processes. To investigate the role of this pathway in the regulation of hematopoietic stem cells (HSCs), we have blocked Smad signaling by retroviral gene transfer of the inhibitory Smad7 to murine HSCs. We report here that the self-renewal capacity of HSCs is promoted in vivo upon blocking of the entire Smad pathway, as shown by both primary and secondary bone marrow (BM) transplantations. Importantly, HSCs overexpressing Smad7 have an unperturbed differentiation capacity as evidenced by normal contribution to both lymphoid and myeloid cell lineages, suggesting that the Smad pathway regulates self-renewal independently of differentiation. Moreover, phosphorylation of Smads was inhibited in response to ligand stimulation in BM cells, thus verifying impairment of the Smad-signaling cascade in Smad7-overexpressing cells. Taken together, these data reveal an important and previously unappreciated role for the Smad-signaling pathway in the regulation of self-renewal of HSCs in vivo.
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| 5. |
- Blank Savukinas, Ulrika, et al.
(författare)
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TGF-β signaling in the control of hematopoietic stem cells.
- 2015
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Ingår i: Blood. - : American Society of Hematology. - 1528-0020 .- 0006-4971. ; 125:23, s. 3542-3550
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Tidskriftsartikel (refereegranskat)abstract
- Blood is a tissue with high cellular turnover, and its production is a tightly orchestrated process that requires constant replenishment. All mature blood cells are generated from hematopoietic stem cells (HSCs), which are the self-renewing units that sustain life-long hematopoiesis. HSC behavior, such as self-renewal and quiescence, are regulated by a wide array of factors, including external signaling cues present in the bone marrow. The Transforming Growth Factor-β (TGF-β) family of cytokines constitutes a multifunctional signaling circuitry, which regulates pivotal functions related to cell fate and behavior in virtually all tissues of the body. In the hematopoietic system, TGF-β signaling controls a wide spectrum of biological processes, from homeostasis of the immune system to quiescence and self-renewal of HSCs. Here, we review key features and emerging concepts pertaining to TGF-β and downstream signaling pathways in normal HSC biology, featuring aspects of aging, hematological disease, and how this circuitry may be exploited for clinical purposes in the future.
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| 6. |
- Blank Savukinas, Ulrika, et al.
(författare)
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The Bystander Effect : Mesenchymal Stem Cell-Mediated Lung Repair
- 2016
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Ingår i: Stem Cells. - : Oxford University Press (OUP). - 1549-4918 .- 1066-5099. ; 34:6, s. 1437-1444
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Forskningsöversikt (refereegranskat)abstract
- Mesenchymal stem or stromal cells (MSCs), a heterogeneous subset of adult stem/progenitor cells, have surfaced as potential therapeutic units with significant clinical benefit for a wide spectrum of disease conditions, including those affecting the lung. Although MSCs carry both self-renewal and multilineage differentiation abilities, current dogma holds that MSCs mainly contribute to tissue regeneration and repair by modulating the host tissue via secreted cues. Thus, the therapeutic benefit of MSCs is thought to derive from so called bystander effects. The regenerative mechanisms employed by MSCs in the lung include modulation of the immune system as well as promotion of epithelial and endothelial repair. Apart from secreted factors, a number of recent findings suggest that MSCs engage in mitochondrial transfer and shedding of membrane vesicles as a means to enhance tissue repair following injury. Furthermore, it is becoming increasingly clear that MSCs are an integral component of epithelial lung stem cell niches. As such, MSCs play an important role in coupling information from the environment to stem and progenitor populations, such that homeostasis can be ensured even in the face of injury. It is the aim of this review to outline the major mechanisms by which MSCs contribute to lung regeneration, synthesizing recent preclinical findings with data from clinical trials and potential for future therapy
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| 7. |
- Blank Savukinas, Ulrika, et al.
(författare)
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The role of Smad signaling in hematopoiesis and translational hematology.
- 2011
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Ingår i: Leukemia. - : Springer Science and Business Media LLC. - 1476-5551 .- 0887-6924. ; 25, s. 1379-1388
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Tidskriftsartikel (refereegranskat)abstract
- Hematopoietic stem cells (HSCs) reside in the bone marrow (BM) of adult individuals and function to produce and regenerate the entire blood and immune system over the course of an individual's lifetime. Historically, HSCs are among the most thoroughly characterized tissue-specific stem cells. Despite this, the regulation of fate options, such as self-renewal and differentiation, has remained elusive, partly because of the expansive plethora of factors and signaling cues that govern HSC behavior in vivo. In the BM, HSCs are housed in specialized niches that dovetail the behavior of HSCs with the need of the organism. The Smad-signaling pathway, which operates downstream of the transforming growth factor-β (TGF-β) superfamily of ligands, regulates a diverse set of biological processes, including proliferation, differentiation and apoptosis, in many different organ systems. Much of the function of Smad signaling in hematopoiesis has remained nebulous due to early embryonic lethality of most knockout mouse models. However, recently new data have been uncovered, suggesting that the Smad-signaling circuitry is intimately linked to HSC regulation. In this review, we bring the Smad-signaling pathway into focus, chronicling key concepts and recent advances with respect to TGF-β-superfamily signaling in normal and leukemic hematopoiesis.Leukemia advance online publication, 13 May 2011; doi:10.1038/leu.2011.95.
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| 8. |
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| 9. |
- Karlsson, Göran, et al.
(författare)
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Smad4 is critical for self-renewal of hematopoietic stem cells.
- 2007
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Ingår i: Journal of Experimental Medicine. - : Rockefeller University Press. - 1540-9538 .- 0022-1007. ; 204:3, s. 467-474
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Tidskriftsartikel (refereegranskat)abstract
- Members of the transforming growth factor β (TGF-β) superfamily of growth factors have been shown to regulate the in vitro proliferation and maintenance of hematopoietic stem cells (HSCs). Working at a common level of convergence for all TGF-β superfamily signals, Smad4 is key in orchestrating these effects. The role of Smad4 in HSC function has remained elusive because of the early embryonic lethality of the conventional knockout. We clarify its role by using an inducible model of Smad4 deletion coupled with transplantation experiments. Remarkably, systemic induction of Smad4 deletion through activation of MxCre was incompatible with survival 4 wk after induction because of anemia and histopathological changes in the colonic mucosa. Isolation of Smad4 deletion to the hematopoietic system via several transplantation approaches demonstrated a role for Smad4 in the maintenance of HSC self-renewal and reconstituting capacity, leaving homing potential, viability, and differentiation intact. Furthermore, the observed down-regulation of notch1 and c-myc in Smad4−/− primitive cells places Smad4 within a network of genes involved in the regulation HSC renewal.
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| 10. |
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| 11. |
- Larsson, Jonas, et al.
(författare)
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TGF-{beta} signaling-deficient hematopoietic stem cells have normal self-renewal and regenerative ability in vivo despite increased proliferative capacity in vitro.
- 2003
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Ingår i: Blood. - : American Society of Hematology. - 1528-0020 .- 0006-4971. ; 102:9, s. 3129-3135
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Tidskriftsartikel (refereegranskat)abstract
- Studies in vitro implicate transforming growth factor β (TGF-β) as a key regulator of hematopoiesis with potent inhibitory effects on progenitor and stem cell proliferation. In vivo studies have been hampered by early lethality of knock-out mice for TGF-β isoforms and the receptors. To directly assess the role of TGF-β signaling for hematopoiesis and hematopoietic stem cell (HSC) function in vivo, we generated a conditional knock-out model in which a disruption of the TGF-β type I receptor (TβRI) gene was induced in adult mice. HSCs from induced mice showed increased proliferation recruitment when cultured as single cells under low stimulatory conditions in vitro, consistent with an inhibitory role of TGF-β in HSC proliferation. However, induced TβRI null mice show normal in vivo hematopoiesis with normal numbers and differentiation ability of hematopoietic progenitor cells. Furthermore HSCs from TβRI null mice exhibit a normal cell cycle distribution and do not differ in their ability long term to repopulate primary and secondary recipient mice following bone marrow transplantation. These findings challenge the classical view that TGF-β is an essential negative regulator of hematopoietic stem cells under physiologic conditions in vivo.
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| 12. |
- Moody, Jennifer, et al.
(författare)
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Endoglin is not critical for hematopoietic stem cell engraftment and reconstitution but regulates adult erythroid development
- 2007
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Ingår i: Stem Cells. - : Oxford University Press (OUP). - 1549-4918 .- 1066-5099. ; 25:11, s. 2809-2819
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Tidskriftsartikel (refereegranskat)abstract
- Endoglin is a transforming growth factor-beta (TGF-beta) accessory receptor recently identified as being highly expressed on long-term repopulating hematopoietic stem cells (HSC) However, little is known regarding its function in these cells. We have used two complementary approaches toward understanding endoglin's role in HSC biology: one that efficiently knocks down expression via lentiviral-driven short hairpin RNA and another that uses retroviral-mediated overexpression. Altering endoglin expression had functional consequences for hematopoietic progenitors in vitro such that endoglin-suppressed myeloid progenitors (colony-forming unit-granulocyte macrophage) displayed a higher degree of sensitivity to TGF-beta-mediated growth inhibition, whereas endoglin-overexpressing cells were partially resistant. However, transplantation of transduced bone marrow enriched in primitive hematopoietic stem and progenitor cells revealed that neither endoglin suppression nor endoglin overexpression affected the ability of stem cells to short-term or long-term repopulate recipient marrow. Furthermore, transplantation of cells altered in endoglin expression yielded normal white blood cell proportions and peripheral blood platelets. Interestingly, decreasing endoglin expression increased the clonogenic capacity of early blast-forming unit-erythroid progenitors, whereas overexpression compromised erythroid differentiation at the basophilic erythroblast phase, suggesting a pivotal role for endoglin at key stages of adult erythropoietic development.
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| 13. |
- Rörby, Emma, et al.
(författare)
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Human hematopoietic stem/progenitor cells overexpressing Smad4 exhibit impaired reconstitution potential in vivo.
- 2012
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Ingår i: Blood. - : American Society of Hematology. - 1528-0020 .- 0006-4971. ; 120, s. 4343-4351
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Tidskriftsartikel (refereegranskat)abstract
- Hematopoietic stem cells (HSCs) constitute a rare population of tissue-specific cells that can self-renew and differentiate into all lineages of the blood cell system. These properties are critical for tissue regeneration and clinical applications of HSCs. Cord blood is an easily accessible source of HSCs. However, the number of HSCs from one unit is too low to effectively transplant most adult patients, and expansion of HSCs in vitro has met with limited success due to incomplete knowledge regarding mechanisms regulating self-renewal. Members of the transforming growth factor-β (TGF-β) superfamily have been shown to regulate HSCs through the Smad signaling pathway, however, its role in human HSCs has remained relatively uncharted in vivo. Therefore, we asked whether enforced expression of the common-Smad, Smad4, could reveal a role for TGF-β in human hematopoietic stem/progenitor cells (HSPCs) from cord blood. Using a lentiviral overexpression approach, we demonstrate that Smad4 overexpression sensitizes HSPCs to TGF-β, resulting in growth arrest and apoptosis in vitro. This phenotype translates in vivo into reduced HSPC reconstitution capacity yet intact lineage distribution. This suggests that the Smad pathway regulates self-renewal independently of differentiation. These findings demonstrate that the Smad signaling circuitry negatively regulates the regeneration capacity of human HSPCs in vivo.
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| 14. |
- Singbrant, Sofie, et al.
(författare)
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Smad5 is dispensable for adult murine hematopoiesis.
- 2006
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Ingår i: Blood. - : American Society of Hematology. - 1528-0020 .- 0006-4971. ; 108:12, s. 3707-3712
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Tidskriftsartikel (refereegranskat)abstract
- Smad5 is known to transduce intracellular signals from bone morphogenetic proteins (BMPs), which belong to the transforming growth factor-beta (TGF-beta) superfamily and are involved in the regulation of hematopoiesis. Recent findings suggest that BMP4 stimulates proliferation of human primitive hematopoietic progenitors in vitro, while early progenitors from mice deficient in Smad5 display increased self-renewal capacity in murine embryonic hematopoiesis. Here, we evaluate the role of Smad5 in the regulation of hematopoietic stem cell (HSC) fate decisions in adult mice by using an inducible MxCre-mediated conditional knockout model. Surprisingly, analysis of induced animals revealed unperturbed cell numbers and lineage distribution in peripheral blood (PB), bone marrow (BM), and the spleen. Furthermore, phenotypic characterization of the stem cell compartment revealed normal numbers of primitive lin(-)Sca-1(+)c-Kit(+) (LSK) cells in Smad5(-/-) BM. When transplanted in a competitive fashion into lethally irradiated primary and secondary recipients, Smad5-deficient BM cells competed normally with wild-type (wt) cells, were able to provide long-term reconstitution for the hosts, and displayed normal lineage distribution. Taken together, Smad5-deficient HSCs from adult mice show unaltered differentiation, proliferation, and repopulating capacity. Therefore, in contrast to its role in embryonic hematopoiesis, Smad5 is dispensable for hematopoiesis in the adult mouse.
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