Mesenchymal Stem and Progenitor Cells In the Mouse Bone Marrow Lack Expression of CD44.

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 2581-2581
Author(s):  
Hong Qian ◽  
Mikael Sigvardsson
Keyword(s):  
Bone Marrow ◽  
Progenitor Cells ◽  
Stromal Cells ◽  
Stromal Cell ◽  
Hematopoietic Cells ◽  
Hematopoietic Stem ◽  
Colony Forming Unit ◽  
Stem And Progenitor Cells ◽  
Cell Fractions

Abstract Abstract 2581 The bone marrow (BM) microenvironment consists of a heterogeneous population including mesenchymal stem cells and as well as more differentiated cells like osteoblast and adipocytes. These cells are believed to be crucial regulators of hematopoetic cell development, however, so far, their identity and specific functions has not been well defined. We have by using Ebf2 reporter transgenic Tg(Ebf2-Gfp) mice found that CD45−TER119−EBF2+ cells are selectively expressed in non-hematopoietic cells in mouse BM and highly enriched with MSCs whereas the EBF2− stromal cells are very heterogenous (Qian, et al., manuscript, 2010). In the present study, we have subfractionated the EBF2− stromal cells by fluorescent activated cell sorter (FACS) using CD44. On contrary to previous findings on cultured MSCs, we found that the freshly isolated CD45−TER119−EBF2+ MSCs were absent for CD44 whereas around 40% of the CD45−TER119−EBF2− cells express CD44. Colony forming unit-fibroblast (CFU-F) assay revealed that among the CD45−LIN−EBF2− cells, CD44− cells contained generated 20-fold more CFU-Fs (1/140) than the CD44+ cells. The EBF2−CD44− cells could be grown sustainably in vitro while the CD44+ cells could not, suggesting that Cd44− cells represents a more primitive cell population. In agreement with this, global gene expression analysis revealed that the CD44− cells, but not in the CD44+ cells expressed a set of genes including connective tissue growth factor (Ctgf), collagen type I (Col1a1), NOV and Runx2 and Necdin(Ndn) known to mark MSCs (Djouad et al., 2007) (Tanabe et al., 2008). Furthermore, microarray data and Q-PCR analysis from two independent experiments revealed a dramatic downregulation of cell cycle genes including Cdc6, Cdca7,-8 and Ki67, Cdk4-6) and up-regulation of Cdkis such as p57 and p21 in the EBF2−CD44− cells, compared to the CD44+ cells indicating a relatively quiescent state of the CD44− cells ex vivo. This was confirmed by FACS analysis of KI67 staining. Furthermore, our microarray analysis suggested high expression of a set of hematopoietic growth factors and cytokines genes including Angiopoietin like 1, Kit ligand, Cxcl12 and Jag-1 in the EBF2−CD44− stromal cells in comparison with that in the EBF2+ or EBF2−CD44+ cell fractions, indicating a potential role of the EBF2− cells in hematopoiesis. The hematopoiesis supporting activity of the different stromal cell fractions were tested by in vitro hematopoietic stem and progenitor assays- cobblestone area forming cells (CAFC) and colony forming unit in culture (CFU-C). We found an increased numbers of CAFCs and CFU-Cs from hematopoietic stem and progenitor cells (Lineage−SCA1+KIT+) in culture with feeder layer of the EBF2−CD44− cells, compared to that in culture with previously defined EBF2+ MSCs (Qian, et al., manuscript, 2010), confirming a high capacity of the EBF2−CD44− cells to support hematopoietic stem and progenitor cell activities. Since the EBF2+ cells display a much higher CFU-F cloning frequency (1/6) than the CD44−EBF2− cells, this would also indicate that MSCs might not be the most critical regulators of HSC activity. Taken together, we have identified three functionally and molecularly distinct cell populations by using CD44 and transgenic EBF2 expression and provided clear evidence of that primary mesenchymal stem and progenitor cells reside in the CD44− cell fraction in mouse BM. The findings provide new evidence for biological and molecular features of primary stromal cell subsets and important basis for future identification of stage-specific cellular and molecular interaction pathways between hematopoietic cells and their cellular niche components. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Periostin Supports Hematopoietic Stem/Progenitor Cells and Niche-Dependent Myeloblastoma Cells In Vitro

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 1494-1494 ◽  
Author(s):  
Akio Maekawa ◽  
Natsumi Hasegawa ◽  
Satowa Tanaka ◽  
Leo Matsubara ◽  
Azusa Imanishi ◽  
...  
Keyword(s):  
Stem Cells ◽  
Cell Line ◽  
Progenitor Cells ◽  
Stromal Cells ◽  
Stromal Cell ◽  
Hematopoietic Cells ◽  
Integrin Αvβ3 ◽  
Serum Starvation ◽  
Hematopoietic Stem

Abstract Periostin (POSTN), the fasciclin family extracellular matrix protein also known as osteoblast-specific factor 2 (OSF-2), was previously reported to be required for optimal B lymphopoiesis in vitro. Now, our study first demonstrates the proof that POSTN might be a bona fide niche factor for both normal and malignant myelopoiesis, indicating that it is a niche molecule for hematopoietic stem cells and diverse hematopoietic precursor cells. The Mediator, composed of about 31 subunits, is a master transcriptional coregulator complex that is essential for global transcription governed by RNA polymerase II. Among the Mediator subunits, MED1 acts as a specific coactivator for activators that include nuclear receptors and GATA1. We previously reported that Med1−/− mouse embryonic fibroblasts (MEFs) have a decreased capability to support hematopoietic stem/progenitor cells (HSPCs) relative to wild-type MEFs in vitro, and that the attenuated expression of full-length osteopontin and FGF7 in Med1−/− MEFs is responsible for the observed phenotype. The microarray analyses, showing that the expression of POSTN was also suppressed in Med1−/− MEFs, prompted us to study the role for POSTN in support of both normal and malignant HSPCs in our in vitro niche model. When bone marrow (BM) cells were cocultured with mitomycin C-treated Med1+/+ MEFs, or OP-9 or MS-5 BM stromal cells, in the presence of anti-POSTN blocking antibody, the mitogenicity and growth of BM cells were attenuated. The number of long-term culture-initiating cells (LTC-ICs), i.e., number of both granulo-monocytic and erythroid colonies, was also decreased. When BM cells were cocultured with Med1-/- MEFs in the presence of recombinant POSTN, the mitogenicity and growth of BM cells and the number of LTC-ICs were restored. These results suggest that POSTN mediates mitogenicity of BM cells and HSPCs support. The MB-1 myeloblastoma cell line, originally established from a patient with myeloid crisis chronic myeloid leukemia, is a mesenchymal stromal cell-dependent cell line. These cells are unique in that they grow by forming cobblestone areas in the presence of niche cells but die of apoptosis when detached from stromal cells, thus faithfully conforming to a stochastic model of leukemic stem cells in vitro. Intriguingly, antibody-mediated blockage of stromal cells-derived POSTN markedly reduced the mitogenicity and growth, as well as the cobblestone formation, a leukemic stem cell feature, of MB-1 myeloblastoma cells. Therefore, it appears that niche cell-derived POSTN supports niche-dependent MB-1 myeloblastoma cells. While POSTN was expressed both in BM cells and variably in different BM stromal cells, expression in the latter cells was markedly increased by tactile interaction with hematopoietic cells. Specifically, POSTN was robustly induced 6 hours after BM stromal cells were cocultured with BM cells or MB-1 myeloblastoma cells, and the induction sustained for as long as 24 hours. However, POSTN expression was not enhanced when BM cells were cocultured but physically separated from MS-5 or OP-9 cells using transwell culture wells. Therefore, the major source of POSTN in the coculture appears to be the BM stromal cells associated with hematopoietic cells. The receptor for POSTN, integrin αvβ3, was expressed abundantly in BM stromal cells. Although β3 mRNA was especially prominent in both BM cells and MB-1 cells, in accordance with a previous report that integrin β3/CD61 marks HSPCs, western blot analysis showed that αv and β3 expression levels were below the detection level on BM cells. Hence, integrin αvβ3 is scarce on BM cells compared to BM stromal cells, although it does not exclude the possibility that functional integrin αvβ3 might be enriched on HSPCs as suggested previously. When an excess amount of exogenous POSTN was added to MS-5 or OP-9 BM stromal cells after 24-h serum starvation, FAK (the immediate target of integrin αvβ3) and MAP kinases ERK1/ERK2 (the intermediate hub of various intracellular signals) were robustly phosphorylated as early as 10 min, and the phosphorylation was sustained for over 60 min. Thus, POSTN effectively activates integrin αvβ3 and subsequent intracellular signaling in BM stromal cells. These results suggest that stromal cell POSTN supports both normal HSPCs and leukemia-initiating cells in vitro, at least in part, indirectly by acting on stromal cells in an autocrine or paracrine manner. Disclosures No relevant conflicts of interest to declare.

Cell Therapy in a Mouse Model of Immune-Mediated Bone Marrow Failure.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 1689-1689
Author(s):  
Jichun Chen ◽  
Neal S. Young
Keyword(s):  
Bone Marrow ◽  
T Cells ◽  
Progenitor Cells ◽  
Stromal Cells ◽  
Stromal Cell ◽  
Adherent Cells ◽  
Hematopoietic Stem ◽  
Cell Counts ◽  
Immune Mediated ◽  
Stem And Progenitor Cells

Abstract Immune-mediated bone marrow (BM) failure has been modeled in the mouse by infusion of lymph node cells from allogeneic C57BL/6 (B6) donors into major or minor histocompatibility antigen-mismatched recipients (Chen et al., Blood 2004; Bloom et al., Exp Hematol 2004, Chen et al., J Immunol 2007). Co-infusion of limited numbers of CD4+CD25+ regulatory T lymphocytes (Tregs) can alleviate clinical manifestations by suppressing the expansion of pathogenic T cells (Chen et al., J Immunol 2007). In the current study, we investigated the effectiveness of Tregs and suppressor cells contained in BM stroma in this fatal disease. Infusion of fewer than 3 × 103 Tregs to each recipient mouse had only a minor effect in preserving BM cells and did not prevent pancytopenia. Fifteen-50 × 103 thymic Tregs was moderately protective: blood WBC, RBC, platelet and BM cell counts at three weeks after cell infusion were 197%, 116%, 155% and 158% of those of control animals that did not receive Treg infusion; 5–10 × 103 B6 splenic Tregs produced the largest effect as WBC, RBC, platelet and BM cell counts were 275%, 143%, 276%, and 198% of controls. Overall, Treg therapy was helpful but its effectiveness was limited and variable among individual recipients as no antigen-specific Tregs can be identified for the treatment of BM failure. Learned about the immunosuppressive effects of mesenchymal stem cells (MSCs), we went on to test the effectiveness of stromal cells as another therapeutic modality for BM failure, since stromal cells contain MSCs. These cells were derived from B6 BM by culture in α-modified Eagle medium at 33°C with 5% CO2 for two weeks. After separating the non-adherent cells, we detached the adherent stromal cells and infused them into TBI + B6 LN-infused C.B10 mice. Injection of 106 stromal cells at the time of LN cell infusion effectively preserved WBCs (3.09 ± 0.51 vs 0.61 ± 0.18), RBCs (8.72 ± 0.14 vs 3.52 ± 0.46), platelets (924 ± 93 vs 147 ± 25) and BM cells (186.6 ± 8.7 vs 52.7 ± 7.8) when compared to LN-cell-infused mice without stromal cell addition. Delayed stromal cell injection at day 9 after LN cell infusion had only a mild effect on the preservation of RBCs (147%), platelets (276%) and BM cells (223%) and no effect on WBCs (64%), and infusion of non-adherent cells from the same stromal cell culture had no therapeutic effect. Stromal cell-infused mice had higher proportion of FoxP3+CD4+ cells in the peripheral blood (59.7 ± 10.7% vs 29.8 ± 5.4%) and more Lin−CD117+CD34− hematopoietic stem and progenitor cells in the BM (591 ± 95 vs 60 ± 43, thousand) in comparison to LN cell infused mice without stromal cell treatment. Mitigation of pathogenic T cells, including both CD4 and CD8 T lymphocytes, is the potential mechanism for the effectiveness of Treg and stromal cell therapies that helped to protect hematopoietic stem and progenitor cells in the BM of affected animals. Figure Figure

scholarly journals Hematopoietic stem and progenitor cells regulate the regeneration of their niche by secreting Angiopoietin-1

eLife ◽  
2015 ◽  
Vol 4 ◽  
Author(s):  
Bo O Zhou ◽  
Lei Ding ◽  
Sean J Morrison
Keyword(s):  
Bone Marrow ◽  
Progenitor Cells ◽  
Stromal Cells ◽  
Leptin Receptor ◽  
Hematopoietic Cells ◽  
Perivascular Niche ◽  
Hematopoietic Stem ◽  
Stem And Progenitor Cells ◽  
And Function ◽  
Angiopoietin 1

Hematopoietic stem cells (HSCs) are maintained by a perivascular niche in bone marrow but it is unclear whether the niche is reciprocally regulated by HSCs. Here, we systematically assessed the expression and function of Angiopoietin-1 (Angpt1) in bone marrow. Angpt1 was not expressed by osteoblasts. Angpt1 was most highly expressed by HSCs, and at lower levels by c-kit+ hematopoietic progenitors, megakaryocytes, and Leptin Receptor+ (LepR+) stromal cells. Global conditional deletion of Angpt1, or deletion from osteoblasts, LepR+ cells, Nes-cre-expressing cells, megakaryocytes, endothelial cells or hematopoietic cells in normal mice did not affect hematopoiesis, HSC maintenance, or HSC quiescence. Deletion of Angpt1 from hematopoietic cells and LepR+ cells had little effect on vasculature or HSC frequency under steady-state conditions but accelerated vascular and hematopoietic recovery after irradiation while increasing vascular leakiness. Hematopoietic stem/progenitor cells and LepR+ stromal cells regulate niche regeneration by secreting Angpt1, reducing vascular leakiness but slowing niche recovery.

scholarly journals EBF1 As a Novel Regulator of Bone Marrow Mesenchymal Stromal Progenitors

Blood ◽  
2017 ◽  
Vol 130 (Suppl_1) ◽  
pp. 96-96
Author(s):  
Marta Derecka ◽  
Senthilkumar Ramamoorthy ◽  
Pierre Cauchy ◽  
Josip Herman ◽  
Dominic Grun ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Progenitor Cells ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Myeloid Cells ◽  
Bone Marrow Niche ◽  
Wild Type ◽  
Hematopoietic Stem ◽  
Stem And Progenitor Cells

Abstract Hematopoietic stem and progenitor cells (HSPC) are in daily demand worldwide because of their ability to replenish entire blood system. However, the in vitro expansion of HSPC is still a major challenge since the cues from bone marrow microenvironment remain largely elusive. Signals coming from the bone marrow niche, and specifically mesenchymal stem and progenitor cells (MSPC), orchestrate maintenance, trafficking and stage specific differentiation of HSPCs. Although, it is generally accepted that MSPCs are essential for hematopoietic homeostasis and generating multiple types of stromal cells, the exact transcriptional networks regulating MSPCs are not well established. Early B-cell factor 1 (Ebf1) has been discovered as lineage-specific transcription factor governing B lymphopoiesis. Additionally, it has been shown to play important role in differentiation of adipocytes, which are a niche component supporting hematopoietic regeneration. Thus, in this study we seek to examine if Ebf1 has an alternative function in non-hematopoietic compartment of bone marrow, specifically in mesenchymal stromal cells that maintain proper hematopoiesis. Here, we identified Ebf1 as new transcription regulator of MSPCs activity. Mesenchymal progenitors isolated from Ebf1-/- mice show diminished capacity to form fibroblasticcolonies (CFU-F) indicating reduced self-renewal. Moreover, cells expanded from these colonies display impaired in vitro differentiation towards osteoblasts, chondrocytes and adipocytes. In order to test how this defective MSPCs influence maintenance of HSPCs, we performed long-term culture-initiating cell assay (LTC-IC). After 5 weeks of co-culture of Ebf1-deficient stromal cells with wild type HSPCs we could observe significantly decreased number of cobblestone and CFU colonies formed by primitive HSPCs, in comparison to co-cultures with control stromal cells. Furthermore, in vivo adoptive transfers of wild type HSPCs to Ebf1+/- recipient mice showed a decrease in the absolute numbers of HSPCs in primary recipients and reduced donor chimerism within the HSCP compartment in competitive secondary transplant experiments. Additionally, Prx1-Cre-mediated deletion of Ebf1 specifically in MSPCs of mice leads to reduced frequency and numbers of HSPCs and myeloid cells in the bone marrow. These results confirm that mesenchymal stromal cells lacking Ebf1 render insufficient support for HSPCs to sustain proper hematopoiesis. Interestingly, we also observed a reduced ability of HSPCs sorted from Prx1CreEbf1fl/fl mice to form colonies in methylcellulose, suggesting not only impaired maintenance but also hindered function of these cells. Moreover, HSPCs exposed to Ebf1-deficient niche exhibit changes in chromatin accessibility with reduced occupancy of AP-1, ETS, Runx and IRF motifs, which is consistent with decreased myeloid output seen in Prx1CreEbf1fl/fl mice. These results support the hypothesis that defective niche can cause epigenetic reprograming of HSPCs. Finally, single cell and bulk transcriptome analysis of MSPCs lacking Ebf1 revealed differences in the niche composition and decreased expression of lineage-instructive signals for myeloid cells. Thus, our study establishes Ebf1 as a novel regulator of MSPCs playing a crucial role in the maintenance and differentiation of HSPCs. Disclosures No relevant conflicts of interest to declare.

scholarly journals Mechanism of Action of Diallyl Sulphide in Ameliorating the Hematopoietic Radiation Injury

2021 ◽  
Author(s):  
Omika Katoch ◽  
Mrinalini Tiwari ◽  
Namita Kalra ◽  
Paban K. Agrawala
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Progenitor Cells ◽  
Hematopoietic Stem ◽  
Proliferation And Differentiation ◽  
Stem And Progenitor Cells ◽  
Radiation Induced ◽  
Medicinal Properties ◽  
Marrow Cellularity

AbstractDiallyl sulphide (DAS), the pungent component of garlic, is known to have several medicinal properties and has recently been shown to have radiomitigative properties. The present study was performed to better understand its mode of action in rendering radiomitigation. Evaluation of the colonogenic ability of hematopoietic progenitor cells (HPCs) on methocult media, proliferation and differentiation of hematopoietic stem cells (HSCs), and transplantation of stem cells were performed. The supporting tissue of HSCs was also evaluated by examining the histology of bone marrow and in vitro colony-forming unit–fibroblast (CFU-F) count. Alterations in the levels of IL-5, IL-6 and COX-2 were studied as a function of radiation or DAS treatment. It was observed that an increase in proliferation and differentiation of hematopoietic stem and progenitor cells occurred by postirradiation DAS administration. It also resulted in increased circulating and bone marrow homing of transplanted stem cells. Enhancement in bone marrow cellularity, CFU-F count, and cytokine IL-5 level were also evident. All those actions of DAS that could possibly add to its radiomitigative potential and can be attributed to its HDAC inhibitory properties, as was observed by the reversal radiation induced increase in histone acetylation.

scholarly journals Primitive hematopoietic cells in murine bone marrow express the CD34 antigen

Blood ◽  
1996 ◽  
Vol 88 (10) ◽  
pp. 3774-3784 ◽  
Author(s):  
F Morel ◽  
SJ Szilvassy ◽  
M Travis ◽  
B Chen ◽  
A Galy
Keyword(s):  
Stem Cells ◽  
Monoclonal Antibody ◽  
Bone Marrow ◽  
Progenitor Cells ◽  
Significant Proportion ◽  
Hematopoietic Cells ◽  
Full Detail ◽  
Hematopoietic Stem ◽  
Cd34 Antigen

The CD34 antigen is expressed on most, if not all, human hematopoietic stem cells (HSCs) and hematopoietic progenitor cells, and its use for the enrichment of HSCs with repopulating potential is well established. However, despite homology between human and murine CD34, its expression on subsets of primitive murine hematopoietic cells has not been examined in full detail. To address this issue, we used a novel monoclonal antibody against murine CD34 (RAM34) to fractionate bone marrow (BM) cells that were then assayed in vitro and in vivo with respect to differing functional properties. A total of 4% to 17% of murine BM cells expressed CD34 at intermediate to high levels, representing a marked improvement over the resolution obtained with previously described polyclonal anti-CD34 antibodies. Sixty percent of CD34+ BM cells lacked lineage (Lin) markers expressed on mature lymphoid or myeloid cells. Eighty-five percent of Sca-1+Thy-1(10)Lin- /10 cells that are highly enriched in HSCs expressed intermediate, but not high, levels of CD34 antigen. The remainder of these phenotypically defined stem cells were CD34-. In vitro colony-forming cells, day-8 and -12 spleen colony-forming units (CFU-S), primitive progenitors able to differentiate into B lymphocytes in vitro or into T lymphocytes in SCID mice, and stem cells with radioprotective and competitive long-term repopulating activity were all markedly enriched in the CD34+ fraction after single-parameter cell sorting. In contrast, CD34-BM cells were depleted of such activities at the cell doses tested and were capable of only short-term B-cell production in vitro. The results indicate that a significant proportion of murine HSCs and multilineage progenitor cells express detectable levels of CD34, and that the RAM34 monoclonal antibody is a useful tool to subset primitive murine hematopoietic cells. These findings should facilitate more direct comparisons of the biology of CD34+ murine and human stem and progenitor cells.

scholarly journals A novel 37-Kd adhesive membrane protein from cloned murine bone marrow stromal cells and cloned murine hematopoietic progenitor cells

Blood ◽  
1993 ◽  
Vol 82 (5) ◽  
pp. 1436-1444 ◽  
Author(s):  
Y Shiota ◽  
JG Wilson ◽  
K Harjes ◽  
ED Zanjani ◽  
M Tavassoli
Keyword(s):  
Bone Marrow ◽  
Progenitor Cells ◽  
Stromal Cells ◽  
Bone Marrow Stromal Cells ◽  
Stromal Cell ◽  
Hematopoietic Cells ◽  
Marrow Stromal Cells ◽  
Hematopoietic Progenitor Cells ◽  
Single Chain ◽  
Hematopoietic Progenitor

Abstract The adhesion of hematopoietic progenitor cells to bone marrow stromal cells is critical to hematopoiesis and involves multiple effector molecules. Stromal cell molecules that participate in this interaction were sought by analyzing the detergent-soluble membrane proteins of GBI/6 stromal cells that could be adsorbed by intact FDCP-1 progenitor cells. A single-chain protein from GBI/6 cells having an apparent molecular weight of 37 Kd was selectively adsorbed by FDCP-1 cells. This protein, designated p37, could be surface-radiolabeled and thus appeared to be exposed on the cell membrane. An apparently identical 37- Kd protein was expressed by three stromal cell lines, by Swiss 3T3 fibroblastic cells, and by FDCP-1 and FDCP-2 progenitor cells. p37 was selectively adsorbed from membrane lysates by a variety of murine hematopoietic cells, including erythrocytes, but not by human erythrocytes. Binding of p37 to cells was calcium-dependent, and was not affected by inhibitors of the hematopoietic homing receptor or the cell-binding or heparin-binding functions of fibronectin. It is proposed that p37 may be a novel adhesive molecule expressed on the surface of a variety of hematopoietic cells that could participate in both homotypic and heterotypic interactions of stromal and progenitor cells.

scholarly journals In Vitro Murine Hematopoiesis Supported by Signaling from a Splenic Stromal Cell Line

2018 ◽  
Vol 2018 ◽  
pp. 1-9 ◽  
Author(s):  
Hong Kiat Lim ◽  
Pravin Periasamy ◽  
Helen C. O’Neill
Keyword(s):  
Bone Marrow ◽  
Cell Line ◽  
Stromal Cell ◽  
Hematopoietic Cells ◽  
Model Systems ◽  
Hematopoietic Stem ◽  
Cell Production ◽  
Stromal Cell Line ◽  
Reticular Cells

There are very few model systems which demonstrate hematopoiesis in vitro. Previously, we described unique splenic stromal cell lines which support the in vitro development of hematopoietic cells and particularly myeloid cells. Here, the 5G3 spleen stromal cell line has been investigated for capacity to support the differentiation of hematopoietic cells from progenitors in vitro. Initially, 5G3 was shown to express markers of mesenchymal but not endothelial or hematopoietic cells and to resemble perivascular reticular cells in the bone marrow through gene expression. In particular, 5G3 resembles CXCL12-abundant reticular cells or perivascular reticular cells, which are important niche elements for hematopoiesis in the bone marrow. To analyse the hematopoietic support function of 5G3, specific signaling pathway inhibitors were tested for the ability to regulate cell production in vitro in cocultures of stroma overlaid with bone marrow-derived hematopoietic stem/progenitor cells. These studies identified an important role for Wnt and Notch pathways as well as tyrosine kinase receptors like c-KIT and PDGFR. Cell production in stromal cocultures constitutes hematopoiesis, since signaling pathways provided by splenic stroma reflect those which support hematopoiesis in the bone marrow.

Involvement of the Integrin Alpha 6 Receptor in the Homing and Engraftment of Hematopoietic Stem and Progenitor Cells to Bone Marrow.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 1293-1293
Author(s):  
Hong Qian ◽  
Sten Eirik W. Jacobsen ◽  
Marja Ekblom
Keyword(s):  
Bone Marrow ◽  
Stem Cell ◽  
Progenitor Cells ◽  
Progenitor Cell ◽  
Bone Marrow Cells ◽  
Hematopoietic Stem ◽  
Stem And Progenitor Cells ◽  
Functional Blocking

Abstract Within the bone marrow environment, adhesive interactions between stromal cells and extracellular matrix molecules are required for stem and progenitor cell survival, proliferation and differentiation as well as their transmigration between bone marrow (BM) and the circulation. This regulation is mediated by cell surface adhesion receptors. In experimental mouse stem cell transplantation models, several classes of cell adhesion receptors have been shown to be involved in the homing and engraftment of stem and progenitor cells in BM. We have previously found that integrin a6 mediates human hematopoietic stem and progenitor cell adhesion to and migration on its specific ligands, laminin-8 and laminin-10/11 in vitro (Gu et al, Blood, 2003; 101:877). Using FACS analysis, the integrin a6 chain was now found to be ubiquitously (>95%) expressed in mouse hematopoietic stem and progenitor cells (lin−Sca-1+c-Kit+, lin−Sca-1+c-Kit+CD34+) both in adult bone marrow and in fetal liver. In vitro, about 70% of mouse BM lin−Sca-1+c-Kit+ cells adhered to laminin-10/11 and 40% adhered to laminin-8. This adhesion was mediated by integrin a6b1 receptor, as shown by functional blocking monoclonal antibodies. We also used a functional blocking monoclonal antibody (GoH3) against integrin a6 to analyse the role of the integrin a6 receptor for the in vivo homing of hematopoietic stem and progenitor cells. We found that the integrin a6 antibody inhibited the homing of bone marrow progenitors (CFU-C) into BM of lethally irradiated recipients. The number of homed CFU-C was reduced by about 40% as compared to cells incubated with an isotype matched control antibody. To study homing of long-term repopulating stem cells (LTR), antibody treated bone marrow cells were first injected intravenously into lethally irradiated primary recipients. After three hours, bone marrow cells of the primary recipients were analysed by competitive repopulation assay in secondary recipients. Blood analysis 16 weeks after transplantation revealed an 80% reduction of stem cell activity of integrin a6 antibody treated cells as compared to cells treated with control antibody. These results suggest that integrin a6 plays an important role for hematopoietic stem and progenitor cell homing in vivo.

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