scholarly journals Identification and Age-dependent Increase of Platelet Biased Human Hematopoietic Stem Cells

2022 ◽  
Author(s):  
Merve Aksoz ◽  
Grigore-Aristide Gafencu ◽  
Bilyana Stoilova Stoilova ◽  
Mario Buono ◽  
Yiran Meng ◽  
...  

Hematopoietic stem cells (HSC) reconstitute multi-lineage human hematopoiesis after clinical bone marrow transplantation and are the cells-of-origin of hematological malignancies. Though HSC provide multi-lineage engraftment, individual murine HSCs are lineage-biased and contribute unequally to blood cell lineages. Now, by combining xenografting of molecularly barcoded adult human bone marrow (BM) HSCs and high-throughput single cell RNA sequencing we demonstrate that human individual BM HSCs are also functionally and transcriptionally lineage biased. Specifically, we identify platelet-biased and multi-lineage human HSCs. Quantitative comparison of transcriptomes from single HSCs from young, and aged, BM show that both the proportion of platelet-biased HSCs, and their level of transcriptional platelet priming, increases with age. Therefore, platelet-biased HSCs, as well as their increased prevalence and elevated transcriptional platelet priming during ageing, are conserved between human and murine hematopoiesis.

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 603-603 ◽  
Author(s):  
Masayuki Yamashita ◽  
Eriko Nitta ◽  
Toshio Suda

Abstract Accumulation of DNA damage in hematopoietic stem cells (HSCs) is associated with aging, bone marrow failure and development of hematological malignancies. Mutation accumulation in HSCs precedes the development of leukemia and lymphoma, and these “pre-leukemic HSCs” can survive after chemotherapy, contributing to the relapse of the disease. Thus, understanding for the DNA damage response at a HSC level is a matter of critical importance for lifelong hematopoiesis, yet the protection mechanism for HSCs from DNA damage accumulation remains to be elucidated. During our study on the response of HSCs to ionizing radiation (IR), we have detected higher responsiveness of HSCs to DNA damage compared with committed progenitor cells: higher p53 activation was observed in HSC-enriched LSK (Lin-Sca1+cKit+) cells and LT-HSCs (CD150+CD41-CD48-LSK) than in myeloid progenitor-enriched LKS- cells. Of note, when treated with 4 Gy IR, LSK cells exhibited stronger upregulation of pro-apoptotic genes Bax, Noxa and Puma compared with LKS- cells, whereas upregulation of survival-contributing p21 and Mdm2 genes was comparable between the two populations. Corresponding to such characteristic behavior, we have identified apoptosis-stimulating protein of p53 1 (Aspp1) as a novel specific regulator of HSCs that provides HSCs with high sensitivity to apoptosis. We found that mRNA and protein of Aspp1 were specifically detected in LSK cells and LT-HSCs. To uncover the roles of Aspp1 in the regulation of HSCs, we evaluated HSCs of adult Aspp1 knockout (KO) mice. These mutant mice exhibited a major increase in the absolute number of LSK cells (1.5-fold; P<0.05) and LT-HSCs (2-fold; P<0.0005). Furthermore, self-renewal capacity of Aspp1-null HSCs was significantly enhanced as measured by serial competitive bone marrow (BM) transplantation assays (P<0.01). To assess the cause of enhanced self-renewal of Aspp1-null HSCs, we examined gene expression profile of Aspp1-null LSK cells before and after BM transplantation using multiplex quantitative RT-PCR array. Aspp1-null LSK cells showed higher expression of multiple quiescence-related genes including Tek, Mpl and Ndn. In line with this, Ki67 staining revealed that Aspp1-null LSK cells showed resistance to the loss of quiescence after serial BM transplantation (P<0.01), and Aspp1 KO mice showed accelerated recovery of peripheral blood and BM when treated with a single dose of 5-FU (P<0.05). Moreover, when serially transplanted or subjected to 4 Gy IR in vivo, Aspp1-null LSK cells exhibited higher resistance to apoptosis which was detected as decreased proportion of Annexin V-positive cells (P<0.05). Gene expression analysis consistently revealed that the induction of pro-apoptotic genes Bax, Noxa and Puma was impaired in irradiated Aspp1-null LSK cells. As a result of the reduced apoptosis, Aspp1-null LSK cells exhibited the tendency to retain persistent DNA damage after genotoxic stress as assessed by γH2AX and 53BP1 foci (chi-square test, P<0.05). Importantly, by breeding Aspp1 KO mice with Mx1-Cre mice and p53flox/flox mice, we verified that Aspp1 synergized with p53 to regulate self-renewal and genomic integrity of HSCs beyond its canonical p53-dependent function. Aspp1 loss further enhanced self-renewal capacity of HSCs in a p53-null background when assayed by serial BM transplantation (P<0.05). Likewise, Aspp1 deficiency further accentuated the accumulation of DNA damage after IR exposure in the absence of p53 (P<0.05). Consequently, whereas approximately half of the recipients receiving p53-null LSK cells died of thymic lymphoma, the recipient mice transplanted with LSK cells deficient for both Aspp1 and p53 were 100% lethal within 6 months after BM transplantation (log-rank test, P<0.01). These mice succumbed to hematological malignancies, mostly T-cell acute lymphoblastic lymphoma and leukemia (ALL) (88%) but also B-cell (6%) and myeloid (6%) malignancies. Taken together, our study demonstrates that Aspp1 attenuates HSC quiescence and induces apoptosis in damaged HSCs, in both p53-dependent and -independent manners, thereby inhibiting the development of leukemia and lymphoma in conjunction with p53 in HSCs. As loss of Aspp1 expression due to aberrant methylation of its promoter has already been proven to be an independent poor prognosis factor in ALL patients, Aspp1 may be a potential target for stem cell-directed therapy of leukemia and lymphoma. Disclosures No relevant conflicts of interest to declare.


Cells ◽  
2021 ◽  
Vol 10 (8) ◽  
pp. 1849
Author(s):  
Daozheng Yang ◽  
Gerald de Haan

Hematopoietic stem cells (HSCs) sustain the lifelong production of all blood cell lineages. The functioning of aged HSCs is impaired, including a declined repopulation capacity and myeloid and platelet-restricted differentiation. Both cell-intrinsic and microenvironmental extrinsic factors contribute to HSC aging. Recent studies highlight the emerging role of inflammation in contributing to HSC aging. In this review, we summarize the recent finding of age-associated changes of HSCs and the bone marrow niche in which they lodge, and discuss how inflammation may drive HSC aging.


Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 682-682
Author(s):  
Fumi Shibata ◽  
Yuko Goto-Koshino ◽  
Miyuki Ito ◽  
Yumi Fukuchi ◽  
Yoshihiro Morikawa ◽  
...  

Abstract A variety of cell surface markers such as c-Kit, Sca-1, CD34 and Flt-3 have been utilized to prospectively isolate murine or human hematopoietic stem cells (HSCs). While murine HSCs were shown to be highly enriched in CD34−c-Kit+Sca-1+Lineage- (CD34−KSL) fraction, this population is still not homogeneous for long-term HSCs. In human, CD34+ cells are regarded as crude HSC fraction and used for clinical applications. However, quiescent human HSCs are also found in CD34− fraction, indicating that CD34 is not a bona fide marker for human HSC. Thus, novel surface markers that can be used to purify human or murine HSCs to homogeneity need to be identified. Roundabout (Robo) family proteins are immunoglobulin-type cell surface receptors that are predominantly expressed in nervous system. Slit2, a ligand for Robo, is a large leucine-rich repeat-containing secreted protein that is also expressed in brain. By binding with Robo, Slit2 acts as a repellant for axon guidance of developing neurons and they are critical for correct wiring of neuronal network. Robo family comprises four family members, Robo1 – Robo4, and Robo4 is distinct in that it is expressed specifically in endothelial cells, but not in brain. In this study, we investigated Robo4 for its possible application for HSC identification in murine and human hematopoietic system. By RT-PCR, Robo4 was specifically expressed in murine KSL fraction, and was not expressed in lineage positive cells and various progenitors such as common myeloid progenitor (CMP), granulocyte-monocyte progenitor (GMP), megakaryocyte/erythroid progenitor (MEP) and common lymphoid progenitor (CLP). Moreover, the expression of Robo4 was highest in side population of KSL cells (KSL-SP), and moderate in KSL-main population (KSL-MP) cells. Monoclonal antibody raised against Robo4 identified its high expression in KSL cells by FACS. FACS analysis of human cord blood cells revealed that Robo4 is highly expressed in CD34+ cells, and CD34+Robo4high population fell into CD38− fraction, which enriches human HSCs. Bone marrow transplantation experiments revealed that Robo4+ fraction of murine KSL cells had long-term repopulating activity, while Robo4−KSL cells not. Although both Robo4+ and Robo4− CD34−KSL cells repopulated murine hematopoietic system for long-term, Robo4+CD34−KSL cells achieved higher chimerism after repopulation compared with Robo4−CD34−KSL. To investigate the physiological role of Robo4 in HSC homeostasis, we next examined the expression of Slit2 in hematopoietic system. Interestingly, Slit2 is specifically expressed in bone marrow stromal cells, but not in hematopoietic cells. Moreover, Slit2 is induced in osteoblasts, a critical cellular component composing HSC niche, in response to myelosuppressive stress such as 5FU treatment. These results indicate that Robo4 is expressed in murine and human hematopoietic HSCs and useful for HSC purification, and Robo4 - Slit2 system may play a role in HSC physiology in niche environment under hematopoietic stress.


Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 3410-3410
Author(s):  
Takamasa Katagiri ◽  
Hiroshi Kawamoto ◽  
Takashi Nakakuki ◽  
Mariko Okada ◽  
Shigeki Ohtake ◽  
...  

Abstract Abstract 3410 Background: To sustain hematopoiesis, hematopoietic stem cells (HSCs) must, on the one hand, replenish themselves by self-renewal and on the other hand produce differentiating progenitor cells. However, it has been difficult to address the actual dynamics of hematopoiesis due to the lack of appropriate experimental systems, regardless of animal species. In the case of humans, however, we were lead to consider one “experiment of nature” that makes it possible to track the progeny of a HSC; i.e. by detecting blood cells deficient in glycosylphosphatidylinositol-anchored proteins (GPI-APs) using flow cytometry. These cells are known to be derived from HSCs with a mutation in the phosphatidylinositol N-acetylglucosaminyltransferase subunit A (PIG-A) gene, which have properties similar to normal HSCs. We recently found that GPI-APs− cells in patients with aplastic anemia (AA) and myelodysplastic syndrome (MDS) frequently show various patterns in the proportion of granulocytes and erythrocytes and that the individual patterns of the two lineage combinations can persist for many years. GPI-APs− cells were rather frequently found in patients with less severe bone marrow (BM) failure. We thus reasoned that the dynamics of HSCs visualized in these cases might reflect those occurring during normal hematopoiesis. Objectives/Methods: We determined the proportion of GPI-APs− cells in six major lineages, namely granulocytes (G), monocytes (M), erythrocytes (E), T cells (T), NK cells (NK) and B cells (B), in peripheral blood cells from 601 BM failure patients using a highly sensitivity flow cytometric analysis and classified the lineage combinations of GPI-APs− cells in patients possessing GPI-APs− cells. We have attempted to test our idea of hematopoiesis by modeling and simulation. Results: Of 601 patients with BM failure, GPI-APs− cells cells were detected in at least one lineage of cells of 250 patients (42%) and the lineage combinations of GPI-APs− cells were classified into 16 different patterns such as GEM, GEMTBNK and GE. Of special interest was our finding that in all 250 cases, the same combinations of lineages were detected regardless of the interval between the first and second analysis and there was a clear trend toward the pattern of the higher the percentage of GPI-APs− granulocytes in patients, the greater the number of GPI-APs− cell lineages. It was clear from our studies that most of the small clones contain only limited lineage cells and even in the case of larger clones, e.g. clones of 1–3% that might be maintained with more than two HSCs, the majority (81%) were non-full-lineage clones. Based on these results indicating that most individual human HSCs only give rise to a limited range of hematopoietic progeny, we proposed a new idea that the observed patterns could be explained by assuming the presence of mosaic-like hematopoietic environments that could simultaneously support the “commitment” of early multipotent progenitors to a certain lineage. We have attempted to test this idea by modeling and simulation and assumed that a self-renewing HSC is located in a particular location in BM and that uncommitted progenitors derived from this HSC can reach to a certain defined area (Figure), which we termed the “commitment sphere”. If the BM microenvironment is mosaic in terms of function in the commitment of progenitors towards a certain lineage, and the size of such mosaic is as large as commitment sphere, then production of progenitors of limited lineages can occur. Indeed, by simulation of many virtual HSCs in a certain type of mosaic environment, formation of similar clone types was recapitulated. We also performed simulations of many virtual HSCs in the same mosaic environment while changing clone size (i.e. changing the size of the commitment sphere). The relationship between clone size and number of lineages in each clone in vivo was quite similar to that of the simulation model. Thus, these simulations provide a reasonable explanation for the observed in vivo findings. Conclusions: Individual HSCs in humans produce only restricted lineages of blood cells. The paradoxical phenomenon can be explained by a model in which the BM microenvironment is mosaic in supporting commitment of progenitors towards distinct lineages. Disclosures: No relevant conflicts of interest to declare.


Blood ◽  
2003 ◽  
Vol 102 (3) ◽  
pp. 881-886 ◽  
Author(s):  
Ewa Sitnicka ◽  
Natalija Buza-Vidas ◽  
Staffan Larsson ◽  
Jens M. Nygren ◽  
Karina Liuba ◽  
...  

Abstract The cytokine tyrosine kinase receptors c-kit and flt3 are expressed and function in early mouse and human hematopoiesis. Through its ability to promote ex vivo expansion and oncoretroviral transduction of primitive human hematopoietic progenitors, the flt3 ligand (FL) has emerged as a key stimulator of candidate human hematopoietic stem cells (HSCs). However, recent studies in the mouse suggest that though it is present on short-term repopulating cells, flt3 is not expressed on bone marrow long-term reconstituting HSCs, the ultimate target for the development of cell replacement and gene therapy. Herein we demonstrate that though only a fraction of human adult bone marrow and cord blood CD34+long-term culture-initiating cells (LTC-ICs) express flt3, most cord blood lymphomyeloid HSCs capable of in vivo reconstituting nonobese diabetic/severe combined immunodeficiency (NOD/SCID) mice are flt3+. The striking difference in flt3 and c-kit expression on mouse and candidate human HSCs translated into a corresponding difference in flt3 and c-kit function because FL was more efficient than SCF at supporting the survival of candidate human HSCs. In contrast, SCF is far superior to FL as a viability factor for mouse HSCs. Thus, the present data provide compelling evidence for a contrasting expression and response pattern of flt3 and c-kit on mouse and human HSCs.


Stem Cells ◽  
2013 ◽  
Vol 31 (3) ◽  
pp. 536-546 ◽  
Author(s):  
Takamasa Katagiri ◽  
Hiroshi Kawamoto ◽  
Takashi Nakakuki ◽  
Ken Ishiyama ◽  
Mariko Okada-Hatakeyama ◽  
...  

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