scholarly journals Hematopoietic Progenitor Cells (HPC) from Mobilized Peripheral Blood Display Enhanced Migration and Marrow Homing Compared to Steady-State Bone Marrow HPC

2007 ◽  
Vol 35 (2) ◽  
pp. 326-334 ◽  
Author(s):  
Halvard Bonig ◽  
Gregory V. Priestley ◽  
Vivian Oehler ◽  
Thalia Papayannopoulou
Keyword(s):  
Bone Marrow ◽  
Steady State ◽  
Progenitor Cells ◽  
Peripheral Blood ◽  
Hematopoietic Progenitor Cells ◽  
Hematopoietic Progenitor ◽  
Mobilized Peripheral Blood

AMD3100 Signals Via the Nervous System, Inducing Release to the Circulation of Bone Marrow SDF-1, Which Is Crucial for Progenitor Cell Mobilization.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 1315-1315 ◽  
Author(s):  
Ayelet Dar ◽  
Alexander Kalinkovich ◽  
Neta Netzer ◽  
Raanan Margalit ◽  
Amir Schajnovitz ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Nervous System ◽  
Steady State ◽  
Progenitor Cells ◽  
Peripheral Blood ◽  
Neutralizing Antibodies ◽  
Progenitor Cell ◽  
Hematopoietic Progenitor Cells ◽  
Hematopoietic Progenitor

Abstract AMD3100, a bicyclam antagonist of the chemokine receptor CXCR4 in vitro, has been shown to induce rapid mobilization of human and murine maturing leukocytes and immature hematopoietic stem and progenitor cells in vivo. In addition, AMD3100 combined with G-CSF, synergistically augments mobilization of human progenitor cells (Broxmeyer & Srour et al, JEM, 2005). However, the mechanism of AMD3100-induced mobilization is currently poorly understood. We report that AMD3100-induced mobilization in mice was accompanied with rapid increase in functional SDF-1 concentrations in the circulation and their parallel decrease in the bone marrow within 1 hour. Biotinylated SDF-1 (bSDF-1) directly injected into the femur was detected in the peripheral blood, adjacent bones and spleen as early as 10 minutes post administration. Interestingly, AMD3100 induced significant elevations in bone marrow-derived bSDF-1 concentrations in the peripheral blood. Similarly, G-CSF induced mobilization was initiated (24 hours post a single injection of G-CSF), by SDF-1 release to the circulation. Administration of neutralizing antibodies against CXCR4 to either untreated or AMD3100 treated mice markedly reduced SDF-1 levels in the peripheral blood, coinciding with increased retention levels of this ligand in the bone marrow. In vitro, AMD3100 directly induced SDF-1 release from the human osteoblast cell line MG-63 in a bell shaped dose response. Inhibition of CXCR4-dependent release of SDF-1 during homeostasis or upon treatment with AMD3100, correlated with selective reduction in recruitment of hematopoietic progenitor cells but not mature leukocytes to the circulation. Importantly, injection of neutralizing antibodies against SDF-1 (but not matched control antibodies) resulted in decreased steady state egress and AMD3100-induced mobilization of hematopoietic progenitor cells. Rapid recruitment (within 1 hour) of hematopoietic progenitor cells and maturing leukocytes out of the bone marrow as well as SDF-1 release were dependent on signals from the nervous system. Administration of the b2 adrenergic agonist (clenbuterol) inhibited endogenous SDF-1 and exogenous bSDF-1 release to the circulation and reduced progenitor cell egress, both during steady state and AMD3100-induced mobilization, while administration of the b2 adrenergic antagonist (propranolol) resulted in opposite effects. Based on our results we propose a model in which egress and mobilization of immature progenitor cells differs from that of maturing leukocytes and is more dependent on SDF-1/CXCR4 interactions. In addition to hematopoietic progenitor cells, also bone marrow stromal cells induce homeostatic secretion of SDF-1, which is increased during mobilization and stress induced recruitment. Secretion of this ligand is also CXCR4-dependent, revealing orchestrated mutual and reciprocal SDF1/CXCR4 interactions and a cross-talk with the nervous system, which regulates progenitor cell egress and recruitment.

Murine Hematopoietic Progenitor Cells With Colony-Forming or Radioprotective Capacity Lack Expression of the β2-Integrin LFA-1

Blood ◽  
1999 ◽  
Vol 93 (1) ◽  
pp. 107-112 ◽  
Author(s):  
Johannes F.M. Pruijt ◽  
Yvette van Kooyk ◽  
Carl G. Figdor ◽  
Roel Willemze ◽  
Willem E. Fibbe
Keyword(s):  
Bone Marrow ◽  
Steady State ◽  
Progenitor Cells ◽  
Mononuclear Cells ◽  
Hematopoietic Progenitor Cells ◽  
Colony Stimulating Factor ◽  
Hematopoietic Progenitor ◽  
Β2 Integrin ◽  
Stimulating Factor

Recently, we have demonstrated that antibodies that block the function of the β2-integrin leukocyte function-associated antigen-1 (LFA-1) completely abrogate the rapid mobilization of hematopoietic progenitor cells (HPC) with colony-forming and radioprotective capacity induced by interleukin-8 (IL-8) in mice. These findings suggested a direct inhibitory effect of these antibodies on LFA-1–mediated transmigration of stem cells through the bone marrow endothelium. Therefore, we studied the expression and functional role of LFA-1 on murine HPC in vitro and in vivo. In steady state bone marrow ± 50% of the mononuclear cells (MNC) were LFA-1neg. Cultures of sorted cells, supplemented with granulocyte colony-stimulating factor (G-CSF)/granulocyte-macrophage colony-stimulating factor (GM-CSF)/IL-1/IL-3/IL-6/stem cell factor (SCF) and erythropoietin (EPO) indicated that the LFA-1neg fraction contained the majority of the colony-forming cells (CFCs) (LFA-1neg 183 ± 62/7,500 cells v LFA-1pos 29 ± 17/7,500 cells,P < .001). We found that the radioprotective capacity resided almost exclusively in the LFA-1neg cell fraction, the radioprotection rate after transplantation of 103, 3 × 103, 104, and 3 × 104 cells being 63%, 90%, 100%, and 100% respectively. Hardly any radioprotection was obtained from LFA-1pos cells. Similarly, in cytokine (IL-8 and G-CSF)–mobilized blood, the LFA-1neg fraction, which comprised 5% to 10% of the MNC, contained the majority of the colony-forming cells, as well as almost all cells with radioprotective capacity. Subsequently, primitive bone marrow-derived HPC, represented by Wheat-germ-agglutinin (WGA)+/Lineage (Lin)−/Rhodamine (Rho)− sorted cells, were examined. More than 95% of the Rho− cells were LFA-1neg. Cultures of sorted cells showed that the LFA-1neg fraction contained all CFU. Transplantation of 150 Rho− LFA-1neg or up to 600 Rho−LFA-1pos cells protected 100% and 0% of lethally irradiated recipient mice, respectively. These results show that primitive murine HPC in steady-state bone marrow and of cytokine-mobilized blood do not express LFA-1.

The RNA Editing Enzyme ADAR1 Is Required for Survival of Differentiating Hematopoietic Progenitor Cells in Adult Mice.

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 1395-1395
Author(s):  
Feng Xu ◽  
Qingde Wang ◽  
Hongmei Shen ◽  
Hui Yu ◽  
Yanxin Li ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Progenitor Cells ◽  
Rna Editing ◽  
Cell Population ◽  
Peripheral Blood ◽  
Hematopoietic Progenitor Cells ◽  
Hematopoietic Progenitor ◽  
Hematopoietic Stem ◽  
Low Level ◽  
Adult Mice

Abstract Adenosine Deaminases Acting on RNA (ADAR) are RNA-editing enzymes converting adenosine residues into inosine (A-to-I) in many double-stranded RNA substrates including coding and non-coding sequences as well as microRNAs. Disruption of the ADAR1 gene in mice results in fetal liver, but not yolk sac, defective erythropoiesis and death at E11.5 (Wang Q et al, Science 2000). Subsequently, a conditional knockout mouse model confirmed these findings and showed massively increased cell death in the affected organs (Wang Q et al, JBC 2004). However, the actual impact of ADAR1 absence on definitive or adult hematopoiesis has not been examined. To define the role of ADAR1 in adult hematopoiesis, we first examined the expression of ADAR1 in different hematopoietic stem/progenitor cell subsets isolated from bone marrow by real-time RT-PCR. ARAR1 was present in hematopoietic stem cells (HSCs) at relatively low level and increased in hematopoietic progenitor cells (HPCs). A series of functional hematopoietic assays were then undertaken. A conditional deletion of ADAR1 was achieved by transducing Lin− or Lin−cKit+ bone marrow cells from ADAR1-lox/lox mice with a MSCV retroviral vector co-expressing Cre and GFP. PCR analysis confirmed the complete deletion of ADAR1 in the transduced cells within 72 hours after the transduction. This system allowed us to evaluate the acute effect of ADAR1 deletion in a specific hematopoietic cell population. Following 4 days of in vitro culture after transduction, the absolute number of Lin− Sca1+ cells in the Cre transduced group was similar to the input number; however the differentiating Lin+ cells significantly decreased whereas both the Lin−Sca1+ and Lin+ cells in the vector (MSCV carrying GFP alone) transduced group increased during culture. Moreover, the colony forming cell (CFC) assay showed much fewer and smaller colonies that contained dead cells from the gene deleted group as compared to those from the control group (p&lt;0.001). The TUNEL assay showed a dramatic increase of apoptosis in the Lin+ population but not in the Lin− cells. Given the mixed genetic background of the ADAR1-lox/lox mice, repopulation of the transduced hematopoietic cells in vivo was examined in immunodeficient mice. Sublethally irradiated (3.5 Gy) NOD/SCID-γcnull recipient were transplanted with either 1.5 × 105 Cre or vector transduced Lin− ADAR1-lox/lox cells. Multi-lineage engraftment in peripheral blood was monitored monthly. While the vector transduced cells were able to constitute more than 90% in multiple lineages of the peripheral blood at 1 to 3 months, Cre-transduced cells were virtually undetectable at all the time points (n=9 to 13, p&lt;0.001). A similar result was found in the hematopoietic organs, including the bone marrow, spleen and thymus. Interestingly, however, the Lin−Sca1+cKit+ cell population was preserved in the Cre transduced group despite the very low level of total donor-derived cells in the bone marrow (n=6 to 7, p&lt;0.01). Consistently, the single cell culture experiment demonstrated that there was no significant difference between ADAR−/− and wild-type HSCs in terms of survival and division during the first 3 days of culture. Taken together, our current study demonstrates nearly absolute requirement of ADAR1 for hematopoietic repopulation in adult mice and it is also suggested that ADAR1 has a preferential effect on the survival of differentiating progenitor cells over more primitive cells.

Angiogenic Factors eNOS and VEGF Have Increased Expression in Granulocytes of JAK2V617F Homozygous Forms of Polycythemia Vera,

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 3847-3847
Author(s):  
Vladan P Cokic ◽  
Dragana Markovic ◽  
Olivera Mitrovic ◽  
Sanja Vignjevic ◽  
Dragoslava Djikic ◽  
...  
Keyword(s):  
Gene Expression ◽  
Bone Marrow ◽  
Transcription Factor ◽  
Progenitor Cells ◽  
Peripheral Blood ◽  
Protein Level ◽  
Angiogenic Factors ◽  
Hematopoietic Progenitor Cells ◽  
Hematopoietic Progenitor ◽  
Vegf Protein

Abstract Abstract 3847 The microvessel density of bone marrow is increased in myeloproliferative neoplasms (MPN) parallel with vascular endothelial growth factor (VEGF). VEGF-mediated angiogenesis requires nitric oxide (NO) production from activated endothelial NO synthase (eNOS). NO as well as hypoxia stimulate the VEGF gene expression and angiogenesis by enhancing hypoxia inducible factor (HIF)-1 activity. We studied 126 newly diagnosed patients with BCR-ABL− MPN: 64 polycythemia vera (PV), 36 essential thrombocythemia (ET), 26 primary myelofibrosis (PMF) and 12 healthy individuals. We performed a combined analysis of hematopoietic CD34+ progenitor cells and granulocytes in peripheral blood of these individuals. The eNOS protein level is more than three-fold elevated in granulocytes of JAK2V617F homozygous PV patients. The essential inducer of angiogenesis VEGF-A has also about three-fold elevation at the protein level in granulocytes of PV patients, with major increases in JAK2V617F homozygous forms. Immunohistochemical analysis reveal that the percentage of VEGF-A-positive cells is increased in bone marrow of PV (5.58±0.7%) compared to normal controls (2.78±0.7%) and VEGF-A mRNA levels are increased in hematopoietic progenitor cells of PV origin. Transcription factor HIF-1α gene expression is decreased in hematopoietic progenitor cells and increased in granulocytes of PV patients. Negative regulator of HIF-1α activity, a transcription factor HIF-3α, has decreased expression in hematopoietic progenitor cells and not changed in granulocytes. In contrast to PV patients, PMF and ET disorders with a minor JAK2 mutation burden demonstrate reduced eNOS and VEGF protein levels and decreased HIF-1a gene expression in peripheral blood granulocytes, although the increase in percentage of VEGF-A-positive cells in bone marrow observed in PV patients is also evident. The present results expand the significance of JAK2V617F mutation in induction of angiogenic factors eNOS and VEGF in granulocytes of PV patients with enhanced HIF-1α presence. Moreover, the stromal and hematopoietic cells also show increased VEGF protein expression in bone marrow of PV patients. Therefore, we find that variations in angiogenic factors expression among MPN patients appear to be related to JAK2V617F mutation allele burden. Disclosures: No relevant conflicts of interest to declare.

Murine Hematopoietic Progenitor Cells With Colony-Forming or Radioprotective Capacity Lack Expression of the β2-Integrin LFA-1

Blood ◽  
1999 ◽  
Vol 93 (1) ◽  
pp. 107-112 ◽  
Author(s):  
Johannes F.M. Pruijt ◽  
Yvette van Kooyk ◽  
Carl G. Figdor ◽  
Roel Willemze ◽  
Willem E. Fibbe
Keyword(s):  
Bone Marrow ◽  
Steady State ◽  
Progenitor Cells ◽  
Mononuclear Cells ◽  
Hematopoietic Progenitor Cells ◽  
Colony Stimulating Factor ◽  
Hematopoietic Progenitor ◽  
Β2 Integrin ◽  
Stimulating Factor

Abstract Recently, we have demonstrated that antibodies that block the function of the β2-integrin leukocyte function-associated antigen-1 (LFA-1) completely abrogate the rapid mobilization of hematopoietic progenitor cells (HPC) with colony-forming and radioprotective capacity induced by interleukin-8 (IL-8) in mice. These findings suggested a direct inhibitory effect of these antibodies on LFA-1–mediated transmigration of stem cells through the bone marrow endothelium. Therefore, we studied the expression and functional role of LFA-1 on murine HPC in vitro and in vivo. In steady state bone marrow ± 50% of the mononuclear cells (MNC) were LFA-1neg. Cultures of sorted cells, supplemented with granulocyte colony-stimulating factor (G-CSF)/granulocyte-macrophage colony-stimulating factor (GM-CSF)/IL-1/IL-3/IL-6/stem cell factor (SCF) and erythropoietin (EPO) indicated that the LFA-1neg fraction contained the majority of the colony-forming cells (CFCs) (LFA-1neg 183 ± 62/7,500 cells v LFA-1pos 29 ± 17/7,500 cells,P &lt; .001). We found that the radioprotective capacity resided almost exclusively in the LFA-1neg cell fraction, the radioprotection rate after transplantation of 103, 3 × 103, 104, and 3 × 104 cells being 63%, 90%, 100%, and 100% respectively. Hardly any radioprotection was obtained from LFA-1pos cells. Similarly, in cytokine (IL-8 and G-CSF)–mobilized blood, the LFA-1neg fraction, which comprised 5% to 10% of the MNC, contained the majority of the colony-forming cells, as well as almost all cells with radioprotective capacity. Subsequently, primitive bone marrow-derived HPC, represented by Wheat-germ-agglutinin (WGA)+/Lineage (Lin)−/Rhodamine (Rho)− sorted cells, were examined. More than 95% of the Rho− cells were LFA-1neg. Cultures of sorted cells showed that the LFA-1neg fraction contained all CFU. Transplantation of 150 Rho− LFA-1neg or up to 600 Rho−LFA-1pos cells protected 100% and 0% of lethally irradiated recipient mice, respectively. These results show that primitive murine HPC in steady-state bone marrow and of cytokine-mobilized blood do not express LFA-1.

scholarly journals Arhgap21 Expression in Bone Marrow Niche Is Crucial for Hematopoietic Progenitor Homing and Short Term Reconstitution after Transplantation

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 1591-1591
Author(s):  
Juliana M. Xavier ◽  
Lauremilia Ricon ◽  
Karla Priscila Vieira ◽  
Longhini Ana Leda ◽  
Carolina Bigarella ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Stem Cell ◽  
Progenitor Cells ◽  
Peripheral Blood ◽  
Hematopoietic Progenitor Cells ◽  
Bone Marrow Niche ◽  
Wild Type ◽  
Hematopoietic Progenitor ◽  
Short Term ◽  
Hematopoietic Stem

Abstract The microenvironment of the bone marrow (BM) is essential for retention and migration of hematopoietic progenitor cells. ARHGAP21 is a negative regulator of RhoGTPAses, involved in cellular migration and adhesion, however the role of ARHGAP21 in hematopoiesis is unknown. In order to investigate whether downregulation of Arhgap21 in microenvironment modulates bone marrow homing and reconstitution, we generated Arhgap21+/-mice using Embryonic Stem cell containing a vector insertion in Arhgap21 gene obtained from GeneTrap consortium and we then performed homing and bone marrow reconstitution assays. Subletally irradiated (9.5Gy) Arhgap21+/- and wild type (WT) mice received 1 x 106 BM GFP+cells by IV injection. For homing assay, 19 hours after the transplant, Lin-GFP+ cells were analyzed by flow cytometry. In reconstitution and self-renew assays, the GFP+ cell percentage in peripheral blood were analyzed 4, 8, 12 and 16 weeks after transplantation. Hematopoietic stem cells [GFP+Lin-Sca+c-Kit+ (LSK)] were counted after 8 and 16 weeks in bone marrow after primary transplant and 16 weeks after secondary transplant. The percentage of Lin-GFP+ hematopoietic progenitor cells that homed to Arhgap21+/-recipient (mean± SD) (2.07 ± 0.85) bone marrow was lower than those that homed to the WT recipient (4.76 ± 2.60); p=0.03. In addition, we observed a reduction (WT: 4.22 ±1.39; Arhgap21+/-: 2.17 ± 0.69; p=0.001) of Lin- GFP+ cells in Arhgap21+/-receptor spleen together with an increase of Lin- GFP+ population in Arhgap21+/-receptor peripheral blood (WT: 8.07 ± 3.85; Arhgap21+/-: 14.07 ±5.20; p=0.01), suggesting that hematopoietic progenitor cells which inefficiently homed to Arhgap21+/-bone marrow and spleen were retained in the blood stream. In bone marrow reconstitution assay, Arhgap21+/-receptor presented reduced LSK GFP+ cells after 8 weeks (WT: 0.19 ±0.03; Arhgap21+/-0.12±0.05; p=0.02) though not after 16 weeks from primary and secondary transplantation. The reduced LSK percentage after short term reconstitution was reflected in the lower GFP+ cells in peripheral blood 12 weeks after transplantation (WT: 96.2 ±1.1; Arhgap21+/-94.3±1.6; p=0.008). No difference was observed in secondary transplantation, indicating that Arhgap21reduction in microenvironment does not affect normal hematopoietic stem cell self-renewal. The knowledge of the niche process in regulation of hematopoiesis and their components helps to better understand the disordered niche function and gives rise to the prospect of improving regeneration after injury or hematopoietic stem and progenitor cell transplantation. In previous studies, the majority of vascular niche cells were affected after sublethal irradiation, however osteoblasts and mesenchymal stem cells were maintained (Massimo Dominici et al.; Blood; 2009.). RhoGTPase RhoA, which is inactivated by ARHGAP21 (Lazarini et al.; Biochim Biophys acta; 2013), has been described to be crucial for osteoblasts and mesenchymal stem cell support of hematopoiesis (Raman et al.; Leukemia; 2013). Taken together, these results suggest that Arhgap21 expression in bone marrow niche is essential for homing and short term reconstitution support. Moreover, this is the first study to investigate the role of Arhgap21 in bone marrow niche. Figure 1 Reduced homing and short term reconstitution in Arhgap21 +/- recipients. Bone marrow cells from GFP+ mice were injected into wild-type and Arhgap21+/- sublethally irradiated mice. 19 hours after the transplant, a decreased homing was observed to both bone marrow (a) and spleen (b) together with an increase of retained peripheral blood (c) Lin-GFP+ cells. In serial bone marrow transplantation, Arhgap21+/- presented reduced bone marrow LSK GFP+ cells 8 weeks (d) and peripheral blood GFP+ cells 12 weeks (e) after primary transplantation, though not 16 weeks after primary (f) and 16 weeks after secondary (g) transplantations. The result is expressed by means ±SD of 2 independent experiments. Figure 1. Reduced homing and short term reconstitution in Arhgap21+/- recipients. Bone marrow cells from GFP+ mice were injected into wild-type and Arhgap21+/- sublethally irradiated mice. 19 hours after the transplant, a decreased homing was observed to both bone marrow (a) and spleen (b) together with an increase of retained peripheral blood (c) Lin-GFP+ cells. In serial bone marrow transplantation, Arhgap21+/- presented reduced bone marrow LSK GFP+ cells 8 weeks (d) and peripheral blood GFP+ cells 12 weeks (e) after primary transplantation, though not 16 weeks after primary (f) and 16 weeks after secondary (g) transplantations. The result is expressed by means ±SD of 2 independent experiments. Disclosures No relevant conflicts of interest to declare.

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