scholarly journals Distribution of cells bearing receptors for a colony-stimulating factor (CSF-1) in murine tissues.

1981 ◽  
Vol 91 (3) ◽  
pp. 848-853 ◽  
Author(s):  
P V Byrne ◽  
L J Guilbert ◽  
E R Stanley
Keyword(s):  
Bone Marrow ◽  
Binding Sites ◽  
Mononuclear Cells ◽  
Bone Marrow Cells ◽  
Mononuclear Phagocytes ◽  
Small Cells ◽  
Phagocytic Cells ◽  
Alveolar Cells ◽  
Peritoneal Cells ◽  
Blood Mononuclear Cells

CSF-1 is a subclass of the colony-stimulating factors that specifically stimulates the growth of mononuclear phagocytes. We used the binding of 125I-CSF-1 at 0 degrees C by single cell suspensions from various murine tissues, in conjunction with radioautography, to determine the frequency of binding cells, their identity, and the number of binding sites per binding cell. For all tissues examined, saturation of binding sites was achieved within 2 h at 2--3 x 10(-10) M 125I-CSF-1. The binding was irreversible and almost completely blocked by a 2 h preincubation with 5 x 10(-10) M CSF-1. 125I-CSF-1 binding was exhibited by 4.3% of bone marrow cells, 7.5% of blood mononuclear cells, 2.4% of spleen cells, 20.5% of peritoneal cells, 11.8% of pulmonary alveolar cells and 0.4% of lymph node cells. Four morphologically distinguishable cell types bound 125I-CSF-1: blast cells; mononuclear cells with a ratio of nuclear to cytoplasmic area (N/C) greater than 1; cells with indented nuclei; and mononuclear cells with N/C less than or equal to 1. No CSF-1 binding cells were detected among blood granulocytes or thymus cells. Bone marrow promyelocytes, myelocytes, neutrophilic granulocytes, eosinophilic granulocytes, nucleated erythroid cells, enucleated erythrocytes, and megakaryocytes also failed to bind. The frequency distribution of grain counts per cell for blood mononuclear cells was homogenous. In contrast, those for bone marrow, spleen, alveolar, and peritoneal cells were heterogeneous. The monocytes in blood or bone marrow (small cells, with either indented nuclei or with N/C greater than 1) were relatively uniformly labeled, possessing approximately 3,000 binding sites per cell. Larger binding cells (e.g., alveolar cells) may possess higher numbers of receptors. It is concluded that CSF-1 binding is restricted to mononuclear phagocytic cells and their precursors and that it can be used to identify both mature and immature cells of this series.

Mutant CXCR4-Triggered Apoptosis and Impaired Mobilization of Bone Marrow Cells in Myelokathexis Are Normalized upon Treatment with Specific Inhibitors.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 1631-1631
Author(s):  
Andrew G. Aprikyan ◽  
Oscar Penate ◽  
Vahagn Makaryan
Keyword(s):  
Bone Marrow ◽  
Mononuclear Cells ◽  
Bone Marrow Cells ◽  
Myeloid Cells ◽  
Specific Inhibitor ◽  
Annexin V ◽  
Whim Syndrome ◽  
Cd34 Cells ◽  
Blood Mononuclear Cells ◽  
Marrow Cells

Abstract Myelokathexis (MK) is a rare congenital disorder characterized by hypercellular marrow and severe chronic neutropenia. In some, but not all patients there is an association of Warts, Hypogammaglobulinemia, and Infections with Myelokathexis (WHIM syndrome). We and others reported that bone marrow myeloid cells in MK exhibit characteristic apoptotic features such as condensed chromatin, cytoplasmic blebbing, and cellular fragmentation. FACS analysis also demonstrated an impaired survival of myeloid cells in MK that was associated with reduced level of Bcl-X expression. Several heterozygous mutations in the CXCR4 gene have been identified in most of the families with autosomal dominant WHIM syndrome. CXCR4 is a G-protein coupled chemokine receptor and its interaction with SDF-1 ligand plays an important role in homing and mobilization of hematopoietic cells. Recently, it has been demonstrated that expression of mutant CXCR4 in human CD34+ cells results in reduced receptor internalization, enhanced calcium flux and enhanced migration of transduced CD34+ cells. However, the mechanism of mutant CXCR4-mediated neutropenia in MK remains largely unknown. We examined 6 patients representing 4 unrelated families with MK and found that all affected family members harbor heterozygous mutations in the CXCR4 gene including a previously reported R334ter mutation. We identified two novel mutations that result in frame shifts and a premature stop codon in the cytoplasmic domain of CXCR4. Expression of CXCR4 mutants in human promyelocytic HL-60 cells resulted in massive apoptosis similar to that reported for bone marrow-derived myeloid cells from MK patients. Specifically, more than 50% of HL-60 cells transfected with previously reported R334ter and novel truncation mutants were positive for annexin V, whereas 20±6% of apoptotic annexin V-positive cells were observed in control cells transfected with normal CXCR4. Impaired cell survival appeared to be due to aberrant dissipation of mitochondrial membrane potential as flow cytometry analysis revealed significantly increased DIOC6-staining in cells expressing CXCR4 mutants compared with control cells expressing normal CXCR4 (p<0.02). Moreover, the expression of CXCR4 mutants but not of the wild type form led to a significant increase in directional motility of myeloid cells towards SDF-1 (p<0.01), similar to the enhanced chemotaxis of blood mononuclear cells observed in our MK patients. These data indicate that our cellular model closely recapitulates the myelokathexis phenotype. Premature apoptosis, but not enhanced chemotaxis triggered by mutant CXCR4 was reduced to near-normal level by caspase-specific inhibitor zVAD-fmk. Interestingly, the mutant CXCR4-induced increase in directional motility to SDF-1, but not accelerated apoptosis was normalized upon treatment with protein kinase Cξ-specific inhibitor. These data suggest that accelerated apoptosis and increased chemotaxis are two independent pathways activated by mutant CXCR4. Importantly, treatment of primary blood mononuclear cells from 3 MK patients with PKCξ inhibitor restored the abnormal chemotactic properties to the levels comparable to that of treated control cells from healthy volunteers. Further studies needed for assessing the therapeutic potential of PKCξ-specific inhibitor for normalizing the impaired mobilization of bone marrow cells in MK. Our data also suggest that the PKCξ-specific inhibitor may be effective for mobilization of human hematopoietic stem cells.

scholarly journals Enrichment of Rabbit Primitive Hematopoietic Cells via MACS Depletion of CD45+ Bone Marrow Cells

2021 ◽  
Vol 7 (1) ◽  
pp. 11
Author(s):  
Jaromír Vašíček ◽  
Andrej Baláži ◽  
Miroslav Bauer ◽  
Andrea Svoradová ◽  
Mária Tirpáková ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Mononuclear Cells ◽  
Bone Marrow Cells ◽  
Quantitative Polymerase Chain Reaction ◽  
Hematopoietic Cells ◽  
Hematopoietic Stem ◽  
Stem And Progenitor Cells ◽  
Rabbit Bone ◽  
Hematopoietic Disorders ◽  
Novel Strategy

Hematopoietic stem and progenitor cells (HSC/HPCs) of human or few animal species have been studied for over 30 years. However, there is no information about rabbit HSC/HPCs, although they might be a valuable animal model for studying human hematopoietic disorders or could serve as genetic resource for the preservation of animal biodiversity. CD34 marker is commonly used to isolate HSC/HPCs. Due to unavailability of specific anti-rabbit CD34 antibodies, a novel strategy for the isolation and enrichment of rabbit HSC/HPCs was used in this study. Briefly, rabbit bone marrow mononuclear cells (BMMCs) were sorted immunomagnetically in order to remove all mature (CD45+) cells. The cells were depleted with overall purity about 60–70% and then cultured in a special medium designed for the expansion of CD34+ cells. Quantitative Polymerase Chain Reaction (qPCR) analysis confirmed the enrichment of primitive hematopoietic cells, as the expression of CD34 and CD49f increased (p < 0.05) and CD45 decreased (p < 0.001) at the end of culture in comparison to fresh BMMCs. However, cell culture still exhibited the presence of CD45+ cells, as identified by flow cytometry. After gating on CD45− cells the MHCI+MHCII−CD38+CD49f+CD90−CD117− phenotype was observed. In conclusion, rabbit HSC/HPCs might be isolated and enriched by the presented method. However, further optimization is still required.

CFU-C Populations in Blood and Bone Marrow of Dogs after Lethal Irradiation and Allogeneic Transfusion with Cryopreserved Blood Mononuclear Cells

2009 ◽  
Vol 21 (2) ◽  
pp. 115-130 ◽  
Author(s):  
Wilhelm Nothdurft ◽  
Theodor M. Fliedner ◽  
Wenceslao Calvo ◽  
Hans-Dieter Flad ◽  
Richard Huget ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Mononuclear Cells ◽  
Allogeneic Transfusion ◽  
Blood Mononuclear Cells ◽  
Lethal Irradiation

scholarly journals Investigation of Mitochondrial Transfer between Human Erythroblasts

2021 ◽  
Author(s):  
◽  
Brittany Lewer
Keyword(s):  
Bone Marrow ◽  
Mitochondrial Dna ◽  
Mononuclear Cells ◽  
Bone Marrow Cells ◽  
Qpcr Assay ◽  
Dna Polymorphisms ◽  
Mitochondrial Transfer ◽  
Hel Cells ◽  
Allele Specific

<p>The increasingly studied phenomenon of mitochondria transferring between cells contrasts the popular belief that mitochondria reside permanently within their cells of origin. Research has identified this process occurring in many tissues such as brain, lung and more recently within the bone marrow. This project aimed to investigate if mitochondria could be transferred between human erythroblasts, a context not previously studied.  Tissue microenvironments can be modelled using co-culture systems. Fluorescence activated cell sorting and a highly sensitive Allele-Specific-Blocker qPCR assay were used to leverage mitochondrial DNA polymorphisms between co-cultured populations. Firstly, HL-60ρ₀ bone marrow cells, without mitochondrial DNA, deprived of essential nutrients pyruvate and uridine were co-cultured in vitro with HEL cells, a human erythroleukemia. Secondly, HEL cells treated with deferoxamine or cisplatin, were cocultured with parental HL-60 cells in vitro. Lastly, ex vivo co-cultures between erythroblasts differentiated from mononuclear cells in peripheral blood were conducted, where one population was treated with deferoxamine.  Co-culture was able to improve recovery when HL-60ρ₀ cells were deprived of pyruvate and uridine. Improved recovery was similarly detected for HEL cells treated with deferoxamine after co-culture with HL-60 cells. Transfer of mitochondrial DNA did not occur at a detectable level in any co-culture condition tested. The high sensitivity of the allele-specific-blocker qPCR assay required completely pure populations to analyse, however this was not achieved using FACS techniques. In conclusion, results have not demonstrated but cannot exclude the possibility that erythroid cells transfer mitochondria to each other.</p>

scholarly journals Biology of human megakaryocyte factor V

Blood ◽  
1986 ◽  
Vol 67 (6) ◽  
pp. 1639-1648
Author(s):  
AM Gewirtz ◽  
M Keefer ◽  
K Doshi ◽  
AE Annamalai ◽  
HC Chiu ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Mononuclear Cells ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Small Cells ◽  
Factor V ◽  
Plasma Clot ◽  
Detection Techniques ◽  
Immunochemical Detection ◽  
Small Bone

To learn more about human megakaryocyte coagulation cofactor V (FV), we studied the expression of this protein in normal bone marrow megakaryocytes and in megakaryocytes cloned from their colony-forming unit in FV-depleted plasma clot cultures. Mouse monoclonal antibodies directed against either the light chain or an activation peptide of human FV and a rabbit polyclonal, monospecific FV antiserum were used as probes for these experiments in conjunction with a variety of immunochemical detection techniques. All morphologically recognizable megakaryocytes were shown to contain FV. The origin of this protein appeared to be both from FV bound to the cell as well as from endogenous FV in the majority of cells examined. The existence of a population of small bone marrow mononuclear cells that simultaneously expressed platelet glycoproteins and FV was also noted. Such cells represented approximately 70% of all small cells positive for platelet glycoproteins. In contrast, only about 40% of megakaryocyte colonies cloned in FV-deficient medium contained cells with immunochemically detectable FV. FV expression was most clearly demonstrated in large cells in the colonies, whereas smaller, presumably less mature cells labeled weakly or not at all. Synthesis of FV by human megakaryocytes was documented using elutriation-enriched cells incubated in 35S- methionine-containing medium. Megakaryocyte lysates and medium conditioned by these cells were subjected to immunoaffinity column purification. Column eluates analyzed by sodium dodecyl sulfate- polyacrylamide gel electrophoresis and autoradiography revealed radioactive bands comigrating with the heavy and light chains of thrombin-activated FV. These studies suggest that human megakaryocytes both bind and synthesize FV. Expression of these traits appears to be related to cell maturation, with binding ability appearing earlier than the ability to synthesize this protein. Finally, although the ability to bind FV appears to be universal among megakaryocytes, our culture data suggest that synthesis may be a restricted, or constitutively expressed property of these cells.

Sign in / Sign up

Export Citation Format

Share Document