EPCR Limits Nitric Oxide Levels, Mediating Human and Murine Stem Cell Adhesion and Retention In The Bone Marrow, By Conjugating PAR1 and CXCR4 Signaling

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 795-795
Author(s):  
Shiri Gur Cohen ◽  
Tomer Itkin ◽  
Aya Ludin ◽  
Sagarika Chakrabarty ◽  
Orit Kollet ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Stem Cells ◽  
Bone Marrow ◽  
Stem Cell ◽  
Protein C ◽  
Cxcr4 Signaling ◽  
Cell Mobilization ◽  
Epcr Shedding

Abstract Long term repopulating hematopoietic stem cells (LTR-HSC) in the murine bone marrow (BM) highly express endothelial protein C receptor (EPCR), yet the function of EPCR in HSC is incompletely defined. EPCR is expressed primarily on endothelial cells and has anti coagulation and anti inflammatory roles. While physiological stress due to injury or bleeding is a strong inducer of HSC mobilization and leukocyte production, a role for the coagulation protease thrombin, and its major receptor PAR1 in regulation of HSC has not yet been identified. We hypothesized that thrombin plays a role in HSC mobilization in the context of injury and that, conversely, signaling involving EPCR and its ligand activated protein C (aPC) play a regulatory role in HSC maintenance. Herein, we report that murine BM EPCRhigh stem cells display enhanced CXCL12 mediated adhesion and reduced migration capacitie, while motile circulating HSC in the murine blood and spleen lack high EPCR expression. Mechanistically, we found that EPCR is a negative regulator of nitric oxide (NO) levels. EPCRhigh stem cells display low intracellular NO levels, low motility, and increased adhesion to BM stroma. Furthermore, EPCRlow transgenic mouse cells displayed reduced stem cell adhesion to BM stroma and increased motility, manifested by reduced EPCRlow HSC in the BM and their corresponding increased levels in the blood. In vitro stimulation with the EPCR ligand, aPC, which we found to be physiologically expressed adjacent to small murine BM blood vessels, augmented EPCRhigh HSC adhesion and further limited their intracellular NO content by increasing eNOS phosphorylation at Thr495 in BM HSC, causing reduced production of NO. Conversely, administration of the pro-coagulant protease thrombin to mice induced PAR1 mediated EPCR shedding from BM HSC, followed by CXCR4 upregulation on HSC, and PAR1-mediated CXCL12 secretion by BM stromal cells. Together, these events lead to loss of retention and rapid stem cell mobilization to the blood. Interestingly, shedding of EPCR was found to be mediated by elevation of intracellular NO content, leading to EPCR co-localization with Caveolin. Correspondingly, thrombin failed to induce EPCR shedding and mobilization in eNOS and PAR1 deficient mice. Additionally, we found that BM LTR-HSC functionally express the metalloproteinase TACE (ADAM17) on the cell membrane, and that in- vitro inhibition of TACE activity by a newly developed selective inhibitor, reduces thrombin- mediated EPCR shedding, suggesting the involvement of TACE in EPCR shedding and HSC mobilization. Moreover, EPCR shedding was also CXCR4 dependent, revealing a crosstalk between EPCR, PAR1 and CXCR4. HSPC mobilized by thrombin possessed increased long-term repopulation capability following transplantation into lethally irradiated recipient mice and re-synthesis of EPCR by donor HSC in the engrafted host BM. In addition, EPCR expression was re-induced on circulating stem cells following in vitro treatment with eNOS inhibitor. Interestingly, bypassing eNOS by directly injecting NO donor, induced EPCR shedding, CXCR4 upregulation and rapid HSPC mobilization in both wild type and eNOS KO mice. Importantly, we found that similar to mice, EPCR was selectively and highly expressed by primitive human BM CD34+CD38- HSC, but not in the blood circulation of clinical G-CSF mobilized stem cells or in motile cord blood stem cells. Human BM CD34+/CD38- HSC are functionally EPCRhigh cells, maintaining low levels of intracellular NO which mediates their increased adhesion, while EPCR shedding was important for their migration and mobilization. In the functional pre-clinical NOD/SCID mouse model, G-CSF mobilization induced EPCR shedding, up-regulation of PAR1 and CXCR4 on human stem and progenitor cells, while NO signaling inhibition blocked G-CSF induced mobilization and increased both murine and human EPCRhigh stem cell accumulation in the murine BM. Our results define functional roles for EPCR, on both human and murine HSC, and suggest that regulation of EPCR expression is linked to NO, PAR1 and CXCR4 signaling as a pivotal mechanism determining HSC localization and function. Our study reveals that activation of coagulation in the context of cell injury controls stem cells retention and motility, and suggests that targeting this system may be useful in improving clinical stem cell mobilization and transplantation protocols. Disclosures: No relevant conflicts of interest to declare.

scholarly journals EPCR/PAR1 Signaling Navigates Long-Term Repopulating Hematopoietic Stem Cell Bone Marrow Homing to Thrombomodulin-Enriched Blood Vessels

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 33-33 ◽  
Author(s):  
Shiri Gur Cohen ◽  
Tomer Itkin ◽  
Sagarika Chakrabarty ◽  
Claudine Graf ◽  
Orit Kollet ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Stem Cell ◽  
Stem Cell Transplantation ◽  
Protein C ◽  
No Production ◽  
Short Term ◽  
Hematopoietic Stem ◽  
Epcr Shedding

Abstract Bone marrow (BM) homing and lodgment of long-term repopulating hematopoietic stem cells (LT-HSCs) is an active and essential first step in clinical stem cell transplantation. EPCR is expressed by murine BM LT-HSCs endowed with the highest repopulation potential and its ligand, activated protein C (aPC), has anticoagulant and anti-sepsis effects in EPCR+/PAR1+ endothelial cells. We recently found that signaling cascades, traditionally viewed as coagulation and inflammation related, also independently control EPCR+ LT-HSC BM retention and recruitment to the blood via distinct PAR1 mediated pathways. EPCR/PAR1 signaling retains LT-HSCs in the BM by restricting nitric oxide (NO) production and Cdc42 activity, promoting VLA4 affinity and adhesion. Conversely, thrombin/PAR1 signaling overcome EPCR+ LT-HSC BM retention by initiating NO production, leading to TACE-‎mediated EPCR shedding, CXCR4 and PAR1 upregulation and parallel CXCL12 secretion by PAR1+ BM stromal cells, enhancing stem cell migration and mobilization. Since EPCR shedding is essential for BM LT-HSC recruitment, we tested EPCR role in LT-HSC BM homing. EPCR+ LT-HSC exhibited reduced in vitro migration towards CXCL12 and enhanced CXCL12-dependent adhesion to fibronectin. Unexpectedly, transplanted EPCR+ LT-HSCs preferentially homed ‎to the host BM, while immature progenitors were equally distributed between the BM and spleen. Specificity of BM homing was further confirmed by EPCR neutralizing antibody treatment, which blocks binding to aPC, leading to attenuated EPCR+ LT-HSC homing to the BM but not to the spleen. Importantly, short term aPC pretreatment inhibited NO production and dramatically increased EPCR+ LT-HSC BM homing. Since EPCR navigates LT-HSC to the BM, we studied the role of EPCR signaling in LT-HSC BM repopulation. Mimicking EPCR signaling by in vivo NO inhibition induced preferential expansion of blood and bone-forming stem cells and gave rise to higher donor type EPCR+ LT-HSCs in competitive repopulation assays. Similarly, repeated treatment with aPC expanded BM EPCR+ stem cells and increased competitive LT-repopulation. Importantly, loss of EPCR function reduced HSC long-term repopulation ability while maintaining their short-term repopulation activity. BM HSCs obtained from Procrlow mice, expressing markedly reduced surface EPCR, failed to compete with normal stem cells in competitive long-term repopulation assays. Consistent with inferior HSC BM repopulation, Procrlow mice exhibited reduced numbers of BM LT-HSC with reduced adhesion capacity. Additionally, these mice displayed increased HSC frequencies in the blood circulation and the spleen, which were pharmacologically corrected by inhibiting NO generation with L-NAME treatment. BM retention is essential for quiescent HSC protection from chemotherapy. Mice treated with NO donor SNAP, or with blocking EPCR antibody as well as Fr2-/-mice lacking PAR1 expression, were more susceptible to hematological failure and mortality induced by 5-FU treatment compared to control mice. Together, these results indicate a functional aPC/EPCR/PAR1 signaling pathway, regulating EPCR+ LT-HSC BM homing, adhesion and long-term repopulation potential. The thrombin-thrombomodulin (TM) complex converts protein C to its activated form aPC, facilitating high affinity binding to its receptor EPCR. To further address the preferential homing of EPCR+ LT-HSCs to the BM, we found that TM is exclusively expressed by a unique BM endothelial cell (BMEC) subpopulation, but not in the spleen. Moreover, EPCR+ LT-HSCs were found adjacent to TM+/aPC+ BMECs, imposing their adhesion and retention. Interestingly, similar to BMECs, BM EPCR+ LT-HSC also express surface TM, implying the possibility of autocrine aPC generation. Herein we define EPCR as a guidance molecule, navigating slow migrating LT-HSC in the blood flow specifically to TM+ BMEC supporting niches, maintaining NOlow stem cell retention, long-term blood production and protection from myelotoxic insult. Conversely, thrombin/PAR1 signaling oppositely increase NO generation and EPCR shedding allowing increased CXCR4-dependent LT-HSC migration and mobilization. Harnessing EPCR signaling may improve clinical stem cell transplantation, increasing LT-HSC specific BM homing and repopulation by aPC pretreatment, as well as potentially to overcome malignant stem cell chemotherapy resistance. Disclosures No relevant conflicts of interest to declare.

EPCR Guides Hematopoietic Stem Cells Homing to the Bone Marrow Independently of Niche Clearance

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 4538-4538
Author(s):  
Shiri Gur-Cohen ◽  
Francesca Avemaria ◽  
Orit Kollet ◽  
Seymen Avci ◽  
Tomer Itkin ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Stem Cell ◽  
Hematopoietic Stem Cells ◽  
Protein C ◽  
No Production ◽  
Platelet Factor ◽  
Hematopoietic Stem

Abstract Bone marrow (BM) homing and lodgment of long-term repopulating hematopoietic stem cells (LT-HSCs) are active and essential first steps during embryonic development and in clinical stem cell transplantation. Rare, BM LT-HSCs endowed with the highest self-renewal and durable repopulation potential, functionally express the anticoagulant endothelial protein C receptor (EPCR) and PAR1. In addition to coagulation and inflammation, EPCR-PAR1 signaling independently controls a BM LT-HSC retention-release switch via regulation of nitric oxide (NO) production within LT-HSCs. EPCR+ LT-HSCs are maintained in thrombomodulin+ (TM) periarterial BM microenvironments via production of activated protein C (aPC), the major ligand for EPCR. Restriction of NO production by aPC-EPCR-PAR1 signaling, activates VLA4-mediated adhesion, anchoring EPCR+ LT-HSCs to the BM and protecting them from chemotherapy insult, sparing hematological failure and premature death (Gur-Cohen S. et al, Nat Med 2015). We report that transplanted EPCR+ LT-HSCs preferentially homed ‎to and were retained in the BM, while immature progenitors were equally distributed between the BM and spleen. Specificity of BM homing was further confirmed by EPCR neutralizing treatment that block aPC binding and attenuate EPCR+ LT-HSC BM homing. Furthermore, short term aPC in vitro pretreatment dramatically augmented EPCR+ LT-HSC BM homing, lodgment and long-term repopulation. PAR1 deficient stem cells were irresponsive to treatment with aPC and displayed reduced BM homing efficiency, all pointing to the aPC-EPCR-PAR1 axis as a crucial mediator of BM LT-HSC homing. Additionally, aPC pretreated EPCR+ LT-HSCs had a striking advantage to competitively home to the BM. Consistently, BM HSCs obtained from Procrlow mice, expressing markedly reduced surface EPCR, failed to compete with wild type stem cells in competitive repopulation assays. Importantly, the competitive homing results strongly imply that the BM available niches for newly arrived EPCR+ LT-HSCs are limited. Indeed, aPC pretreated EPCR+ LT-HSCs BM homing reached a plateau, as increasing the transplanted cell dose above 5x106 BM mononuclear cells, did not yield higher donor EPCR+ LT-HSC homing. These results reveal that there is a limited BM space for newly arrived transplanted EPCR+ stem cells to non-irradiated hosts. Importantly, we found that EPCR+ LT-HSCs can engraft the BM of non-conditioned mice with high efficiency, while remaining in a dormant, non-cycling state. Furthermore, the dormant homed EPCR+ LT-HSCs were later awakened and activated solely by treating the engrafted hosts with a low dose 5-FU chemotherapy, or with NO donor SNAP, revealing that preconditioning and clearance of occupied BM HSC niches are not required. To further address the preferential homing of EPCR+ LT-HSCs to the BM, we found that TM is exclusively expressed by unique BM arterioles, and not in the spleen. BM homed EPCR+ LT-HSCs were found adjacent to TM+ arterioles, imposing their retention. Homed BM EPCR+ LT-HSCs highly express full-length TM with intact lectin-like domain, and the BM TM+ endothelium was found to be enriched with a Glycocalyx layer, in particular with Heparan Sulfate Proteoglycan-2 (HSPG-2). HSGP-2 might specifically interact with the lectin-like domain of TM-expressingLT-HSCs, providing BM specific recognition and accelerated homing. Intriguingly, stabilizing TM function by in vitro pretreatment with platelet factor-4 (PF4) bypassed BM-derived cues and increased EPCR+/TM+ LT-HSC homing also to the spleen, suggesting a supportive role for PF4, highly secreted by BM megakaryocytes, in guiding EPCR+/TM+ LT-HSCs to the BM. Herein we define EPCR as a guidance molecule, navigating LT-HSC specifically to BM TM+ aPC-secreting blood vessels, allowing stem cell retention and protection from DNA damaging agents. The BM harbors a limited number of available stem cell niches for newly arrived transplanted EPCR+/TM+ LT-HSCs, and in vitro aPC pretreatment dramatically augments EPCR+/TM+ LT-HSC BM homing. Our findings provide new mechanistic insights and identify key players concerning LT-HSC homing specifically to the BM, leading to better repopulation following transplantation. This up-to-date approach and new knowledge may potentially lead to improved BM transplantation protocols and to prevent chemotherapy resistance of EPCR-expressing cancer stem cell mediated relapse. Disclosures Ruf: Iconic Therapeutics: Consultancy.

Differential Role for VLA-5 in Mobilization of Heamotopoieic Stem Cells Toward Peripheral Blood and Homing to Bone Marrow and Spleen.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 2190-2190 ◽  
Author(s):  
Pieter K. Wierenga ◽  
Ellen Weersing ◽  
Bert Dontje ◽  
Gerald de Haan ◽  
Ronald P. van Os
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Stem Cell ◽  
Progenitor Cells ◽  
Peripheral Blood ◽  
Hematopoietic Stem ◽  
Late Antigen ◽  
Cell Mobilization

Abstract Adhesion molecules have been implicated in the interactions of hematopoietic stem and progenitor cells with the bone marrow extracellular matrix and stromal cells. In this study we examined the role of very late antigen-5 (VLA-5) in the process of stem cell mobilization and homing after stem cell transplantation. In normal bone marrow (BM) from CBA/H mice 79±3 % of the cells in the lineage negative fraction express VLA-5. After mobilization with cyclophosphamide/G-CSF, the number of VLA-5 expressing cells in mobilized peripheral blood cells (MPB) decreases to 36±4%. The lineage negative fraction of MPB cells migrating in vitro towards SDF-1α (M-MPB) demonstrated a further decrease to 3±1% of VLA-5 expressing cells. These data are suggestive for a downregulation of VLA-5 on hematopoietic cells during mobilization. Next, MPB cells were labelled with PKH67-GL and transplanted in lethally irradiated recipients. Three hours after transplantation an increase in VLA-5 expressing cells was observed which remained stable until 24 hours post-transplant. When MPB cells were used the percentage PKH-67GL+ Lin− VLA-5+ cells increased from 36% to 88±4%. In the case of M-MPB cells the number increased from 3% to 33±5%. Although the increase might implicate an upregulation of VLA-5, we could not exclude selective homing of VLA-5+ cells as a possible explanation. Moreover, we determined the percentage of VLA-5 expressing cells immediately after transplantation in the peripheral blood of the recipients and were not able to observe any increase in VLA-5+ cells in the first three hours post-tranpslant. Finally, we separated the MPB cells in VLA-5+ and VLA-5− cells and plated these cells out in clonogenic assays for progenitor (CFU-GM) and stem cells (CAFC-day35). It could be demonstared that 98.8±0.5% of the progenitor cells and 99.4±0.7% of the stem cells were present in the VLA-5+ fraction. Hence, VLA-5 is not downregulated during the process of mobilization and the observed increase in VLA-5 expressing cells after transplantation is indeed caused by selective homing of VLA-5+ cells. To shed more light on the role of VLA-5 in the process of homing, BM and MPB cells were treated with an antibody to VLA-5. After VLA-5 blocking of MPB cells an inhibition of 59±7% in the homing of progenitor cells in bone marrow could be found, whereas homing of these subsets in the spleen of the recipients was only inhibited by 11±4%. For BM cells an inhibition of 60±12% in the bone marrow was observed. Homing of BM cells in the spleen was not affected at all after VLA-5 blocking. Based on these data we conclude that mobilization of hematopoietic progenitor/stem cells does not coincide with a downregulation of VLA-5. The observed increase in VLA-5 expressing cells after transplantation is caused by preferential homing of VLA-5+ cells. Homing of progenitor/stem cells to the bone marrow after transplantation apparantly requires adhesion interactions that can be inhibited by blocking VLA-5 expression. Homing to the spleen seems to be independent of VLA-5 expression. These data are indicative for different adhesive pathways in the process of homing to bone marrow or spleen.

Beta Glucan Enhances Hematopoietic Stem Cell Mobilization.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 1348-1348
Author(s):  
Stephanie Wagner ◽  
Daniel Cramer ◽  
Richard Hansen ◽  
Ryan Reca ◽  
Mariusz Ratajczak ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Stem Cell ◽  
Mechanism Of Action ◽  
Stem Cell Mobilization ◽  
Combination Group ◽  
Beta Glucan ◽  
Hematopoietic Stem ◽  
Cell Mobilization

Abstract Background: Peripheral blood stem cell infusion is the preferred method for establishing hematopoiesis in transplantation. Use of G-CSF is now the most commonly used mobilizing agent. Despite advances in stem cell techniques and agents, studies have shown that up to 20–25% of patients exhibit poor mobilization and are not able to proceed with autotransplantation. Strategies to improve mobilization include using chemotherapy alone or in conjunction with growth factor or novel agents such as AMD3100. β-glucan PGG is a soluble yeast beta glucan with a molecular mass of 150kD comprised of a β-D-(1–3)-linked glucopyranosyl backbone with a β-D-(1–6)-linked β(1–3) side chains. In previous studies, β-glucan PGG has been shown to induce hematopoietic stem and progenitor cell (HSPC) mobilization to the periphery. In this study, we examined β-glucan PGG’s ability to mobilize HSPC alone and in conjuction with G-CSF and explored its mechanism of action. Methods: Prior to our study, dose kinetic studies were done and showed peak stem cell mobilization at 24 hours and maximum results using the 9.2 mg/kg dose with β-glucan PGG alone. In this study, four groups of wildtype (WT) C57BL/6 mice (6 mice/group) were used; control (saline injection × 4 days), G-CSF only (125ug/d × 4 days), PGG only (4.8 or 9.2mg/kg × one dose), and G-CSF/PGG combination. In the combination group, G-CSF injections were given daily for four days and one PGG injection on day three. Four hours after the last G-CSF injection, the mice were sacrificed and final white blood cell count were collected. Blood was assayed for in vitro mobilization in methycellulose culture. Peritoneal macrophage were stimulated with PGG and supernatant was harvested at times indicated and concentration of MMP-9 was determined using ELISA (R&D Systems). Results: All treated groups showed increased mobilization of all major cell lines (CFU-GM, CFU-M, and CFU-G). β-glucan PGG alone was able to mobilize peripheral stem cells at both doses (4.8mg/kg–9.3 CFU/200000 PBL and 9.2mg/kg–14 CFU/200000 PBL). The combination group (G-CSF/PGG-4.8mg/kg) showed an almost two-fold increase in CFU compared to G-CSF alone (G-CSF-30.42 CFU/200000 PBL, G-CSF/PGG(4.8)-57 CFU/200000 PBL, p=0.008). Initial in vitro chemotaxis assays revealed β-glucan PGG induces HPSC mobilization independent of SDF-1 (stromal derived factor) gradient. Our previous studies have demonstrated that β-glucan can enhance bone marrow engraftment via a CR3 dependent mechanism. However, our current study indicated that β-glucan mobilized stem cells via a CR3 independent mechanism and did not induce appreciable levels of cytokine secretion. To further explore its mechanism of action, we stimulated peritoneal macrophages with β-glucan PGG. Strikingly, β-glucan PGG stimulated macrophages to produce significant amounts of matrix metalloproteinase-9 (MMP-9). Similarly, β-glucan PGG also stimulated bone marrow-derived macrophages to secrete MMP-9. Conclusion: β-glucan PGG is an agent that enhances stem cell mobilization alone and has a synergistic effect when used in conjunction with G-CSF. The mechanism of mobilization by β-glucan PGG involves MMP-9, which results from release of pro-MMP-9 from marrow macrophages. The efficacy of β-glucan PGG and lack of proinflammatory activity make it an attractive agent to supplement mobilization with G-CSF.

An in vitro model of hematopoietic stem cell homing demonstrates rapid homing and maintenance of engraftable stem cells

Blood ◽  
2001 ◽  
Vol 98 (4) ◽  
pp. 1012-1018 ◽  
Author(s):  
Angela E. Frimberger ◽  
Allen I. Stering ◽  
Peter J. Quesenberry
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Stem Cell ◽  
Hematopoietic Stem Cell ◽  
Vitro Assay ◽  
Hematopoietic Stem ◽  
Test Cells ◽  
Stromal Layer

Hematopoietic stem cell (HSC) homing is believed to rely heavily on adhesion interactions between stem cells and stroma. An in vitro assay was developed for adhesion of engraftable HSCs in bone marrow suspensions to pre-established Dexter-type long-term bone marrow culture stromal layers. The cell numbers in the adherent layer and supernatant were examined, along with the engraftment capability of adherent layer cells to indicate the number of HSCs that homed to in vitro stroma. The cell number in the supernatant declined over the 24-hour period. The number of test cells adhering to the stromal layer increased during the first hour and then fell at 6 and 24 hours. The number of test HSCs adhering to the stromal layer was substantial at 20 minutes, increased during the first hour, and then remained constant at 1, 6, and 24 hours of adhesion. These data indicate that adhesion of engraftable HSCs occurs quickly and increases during the first hour of contact with pre-established stroma, that adhesion plateaus within 1 hour of contact, and that HSCs maintain their engraftment capability for at least 24 hours of stromal adhesion. Long-term engraftment from test cells at more than 1 hour of adhesion represents 70.7% of the predicted engraftment from equivalent numbers of unmanipulated marrow cells, indicating that 2 of 3 test engraftable HSCs adhered. These findings demonstrate the usefulness of this model system for studying stem-stromal adhesion, allowing further dissection of the mechanism of HSC homing and exploration of possible manipulations of the process.

scholarly journals Stem cell factor increases colony-forming unit-spleen number in vitro in synergy with interleukin-6, and in vivo in Sl/Sld mice as a single factor

Blood ◽  
1992 ◽  
Vol 79 (4) ◽  
pp. 913-919 ◽  
Author(s):  
DM Bodine ◽  
D Orlic ◽  
NC Birkett ◽  
NE Seidel ◽  
KM Zsebo
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Stem Cell ◽  
Growth Factors ◽  
Stem Cell Factor ◽  
Bone Marrow Cells ◽  
Cells Cultured

Abstract Hematopoiesis is thought to be modulated by interactions of progenitor cells with hematopoietic growth factors. We have shown that colony- forming units-spleen (CFU-S) and repopulating stem cells require interleukin-3 (IL-3) to survive in vitro, and that CFU-S number and long-term repopulating ability can be increased by culture in the combination of IL-3 and IL-6. In this report, we describe the effects of stem cell factor (SCF) on CFU-S and repopulating stem cells. Injection of SCF into anemic Sl/Sld mice caused a twofold and 20-fold increase in CFU-S number in the bone marrow and spleen of treated animals, respectively. After 6 days in suspension culture, CFU-S number increased threefold in cultures supplemented with SCF and IL-6, or SCF, IL-3, and IL-6 relative to the number at day 0. The long-term repopulating ability of cells cultured in SCF, IL-3, and IL-6 was approximately sevenfold better than that of cells cultured in IL-3 or SCF. Similar experiments were performed on populations of bone marrow cells enriched for, or depleted of, CFU-S by elutriation and lineage subtraction. The combination of SCF and IL-6 increased CFU-S number approximately fourfold to eightfold in the CFU-S-enriched fraction, but had no effect on the CFU-S-depleted cells. These results show that SCF alone can increase CFU-S number in vivo, and in combination with other growth factors increases CFU-S numbers in vitro.

scholarly journals Stem cell factor increases colony-forming unit-spleen number in vitro in synergy with interleukin-6, and in vivo in Sl/Sld mice as a single factor

Blood ◽  
1992 ◽  
Vol 79 (4) ◽  
pp. 913-919 ◽  
Author(s):  
DM Bodine ◽  
D Orlic ◽  
NC Birkett ◽  
NE Seidel ◽  
KM Zsebo
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Stem Cell ◽  
Growth Factors ◽  
Stem Cell Factor ◽  
Bone Marrow Cells ◽  
Cells Cultured

Hematopoiesis is thought to be modulated by interactions of progenitor cells with hematopoietic growth factors. We have shown that colony- forming units-spleen (CFU-S) and repopulating stem cells require interleukin-3 (IL-3) to survive in vitro, and that CFU-S number and long-term repopulating ability can be increased by culture in the combination of IL-3 and IL-6. In this report, we describe the effects of stem cell factor (SCF) on CFU-S and repopulating stem cells. Injection of SCF into anemic Sl/Sld mice caused a twofold and 20-fold increase in CFU-S number in the bone marrow and spleen of treated animals, respectively. After 6 days in suspension culture, CFU-S number increased threefold in cultures supplemented with SCF and IL-6, or SCF, IL-3, and IL-6 relative to the number at day 0. The long-term repopulating ability of cells cultured in SCF, IL-3, and IL-6 was approximately sevenfold better than that of cells cultured in IL-3 or SCF. Similar experiments were performed on populations of bone marrow cells enriched for, or depleted of, CFU-S by elutriation and lineage subtraction. The combination of SCF and IL-6 increased CFU-S number approximately fourfold to eightfold in the CFU-S-enriched fraction, but had no effect on the CFU-S-depleted cells. These results show that SCF alone can increase CFU-S number in vivo, and in combination with other growth factors increases CFU-S numbers in vitro.

scholarly journals Cell Source-Dependent In Vitro Chondrogenic Differentiation Potential of Mesenchymal Stem Cell Established from Bone Marrow and Synovial Fluid of Camelus dromedarius

Animals ◽  
2021 ◽  
Vol 11 (7) ◽  
pp. 1918
Author(s):  
Young-Bum Son ◽  
Yeon Ik Jeong ◽  
Yeon Woo Jeong ◽  
Mohammad Shamim Hossein ◽  
Per Olof Olsson ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Stem Cell ◽  
Synovial Fluid ◽  
Cartilage Regeneration ◽  
Cartilage Tissue ◽  
Chondrocyte Differentiation ◽  
Differentiation Potential ◽  
Proliferation Capacity

Mesenchymal stem cells (MSCs) are promising multipotent cells with applications for cartilage tissue regeneration in stem cell-based therapies. In cartilage regeneration, both bone marrow (BM-MSCs) and synovial fluid (SF-MSCs) are valuable sources. However, the cellular characteristics and chondrocyte differentiation potential were not reported in either of the camel stem cells. The in vitro chondrocyte differentiation competence of MSCs, from (BM and SF) sources of the same Camelus dromedaries (camel) donor, was determined. Both MSCs were evaluated on pluripotent markers and proliferation capacity. After passage three, both MSCs showed fibroblast-like morphology. The proliferation capacity was significantly increased in SF-MSCs compared to BM-MSCs. Furthermore, SF-MSCs showed an enhanced expression of transcription factors than BM-MSCs. SF-MSCs exhibited lower differentiation potential toward adipocytes than BM-MSCs. However, the osteoblast differentiation potential was similar in MSCs from both sources. Chondrogenic pellets obtained from SF-MSCs revealed higher levels of chondrocyte-specific markers than those from BM-MSCs. Additionally, glycosaminoglycan (GAG) content was elevated in SF-MSCs related to BM-MSCs. This is, to our knowledge, the first study to establish BM-MSCs and SF-MSCs from the same donor and to demonstrate in vitro differentiation potential into chondrocytes in camels.

Effects Of Hypoxia in Long-Term In Vitro Expansion of Human Bone Marrow Derived Mesenchymal Stem Cells

2017 ◽  
Vol 118 (10) ◽  
pp. 3072-3079 ◽  
Author(s):  
Annelise Pezzi ◽  
Bruna Amorin ◽  
Álvaro Laureano ◽  
Vanessa Valim ◽  
Alice Dahmer ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Human Bone ◽  
Human Bone Marrow ◽  
In Vitro Expansion
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