scholarly journals Expression and functionality of the trkA proto-oncogene product/NGF receptor in undifferentiated hematopoietic cells

Blood ◽  
1994 ◽  
Vol 83 (6) ◽  
pp. 1479-1485 ◽  
Author(s):  
S Chevalier ◽  
V Praloran ◽  
C Smith ◽  
D MacGrogan ◽  
NY Ip ◽  
...  
Keyword(s):  
Cell Line ◽  
Cell Lines ◽  
Scatchard Analysis ◽  
Colony Stimulating Factor ◽  
Ngf Receptor ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
First Time ◽  
Stimulating Factor ◽  
Hematopoietic Cell Lines

Abstract The expression of the low-affinity NGF receptor (p75) and the trkA proto-oncogene product was analyzed in a series of human hematopoietic cell lines at protein and RNA levels. We did not detect any form of NGF receptor in cell lines displaying a myelomonocytic phenotype (HL60 and U937). In contrast, cells displaying a more immature erythroleukemic phenotype (TF1 and K562) expressed TrkA in the absence of detectable p75. Scatchard analysis showed a single high-affinity site for NGF (kd = 10(-10) mol/L), with a copy number ranging from 300 to 3,000 sites per cell depending on the studied cell line. In addition, NGF induced autophosphorylation of TrkA and could substitute for granulocyte- monocyte colony-stimulating factor to trigger the proliferation of the TF1 cell line, with a half-maximal signal observed at 50 pmol/L, indicating that p75 is not required for DNA synthesis in this cell line. The physiologic relevance of NGF in early hematopoiesis was confirmed by showing that 12% to 15% of progenitor blood cells from mice treated with 5-fluorouracil expressed TrkA and that these cells could be induced to proliferate and differentiate in response to NGF in association with macrophage colony-stimulating factor. Our study demonstrates for the first time that trkA proto-oncogene expression and activation is not restricted to the nervous system, but is also an important element in early hematopoiesis.

scholarly journals Expression and functionality of the trkA proto-oncogene product/NGF receptor in undifferentiated hematopoietic cells

Blood ◽  
1994 ◽  
Vol 83 (6) ◽  
pp. 1479-1485 ◽  
Author(s):  
S Chevalier ◽  
V Praloran ◽  
C Smith ◽  
D MacGrogan ◽  
NY Ip ◽  
...  
Keyword(s):  
Cell Line ◽  
Cell Lines ◽  
Scatchard Analysis ◽  
Colony Stimulating Factor ◽  
Ngf Receptor ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
First Time ◽  
Stimulating Factor ◽  
Hematopoietic Cell Lines

The expression of the low-affinity NGF receptor (p75) and the trkA proto-oncogene product was analyzed in a series of human hematopoietic cell lines at protein and RNA levels. We did not detect any form of NGF receptor in cell lines displaying a myelomonocytic phenotype (HL60 and U937). In contrast, cells displaying a more immature erythroleukemic phenotype (TF1 and K562) expressed TrkA in the absence of detectable p75. Scatchard analysis showed a single high-affinity site for NGF (kd = 10(-10) mol/L), with a copy number ranging from 300 to 3,000 sites per cell depending on the studied cell line. In addition, NGF induced autophosphorylation of TrkA and could substitute for granulocyte- monocyte colony-stimulating factor to trigger the proliferation of the TF1 cell line, with a half-maximal signal observed at 50 pmol/L, indicating that p75 is not required for DNA synthesis in this cell line. The physiologic relevance of NGF in early hematopoiesis was confirmed by showing that 12% to 15% of progenitor blood cells from mice treated with 5-fluorouracil expressed TrkA and that these cells could be induced to proliferate and differentiate in response to NGF in association with macrophage colony-stimulating factor. Our study demonstrates for the first time that trkA proto-oncogene expression and activation is not restricted to the nervous system, but is also an important element in early hematopoiesis.

scholarly journals Growth and differentiation of a human megakaryoblastic cell line, CMK

Blood ◽  
1989 ◽  
Vol 74 (1) ◽  
pp. 42-48 ◽  
Author(s):  
N Komatsu ◽  
T Suda ◽  
M Moroi ◽  
N Tokuyama ◽  
Y Sakata ◽  
...  
Keyword(s):  
Cell Line ◽  
Colony Formation ◽  
Culture System ◽  
Dependent Manner ◽  
Colony Stimulating Factor ◽  
Gm Csf ◽  
Macrophage Colony Stimulating Factor ◽  
Hematopoietic Factors ◽  
Macrophage Colony ◽  
Stimulating Factor

Abstract Recently, a human megakaryoblastic cell line, CMK, was established from the peripheral blood of a megakaryoblastic leukemia patient with Down syndrome. Using this cell line, we studied the proliferation and differentiation of megakaryocytic cells in the presence of highly purified human hematopoietic factors and phorbol 12-myristate-13- acetate (PMA). In a methylcellulose culture system, interleukin-3 (IL- 3) and granulocyte-macrophage colony-stimulating factor (GM-CSF) facilitated colony formation by CMK cells in a dose-dependent manner. The maximum stimulating doses of these factors were 10 and 200 U/mL, respectively. These concentrations were comparable to those that stimulate activity in normal hematopoietic cells. In contrast, granulocyte-colony stimulating factor (G-CSF), macrophage-colony stimulating factor (M-CSF), and erythropoietin (EPO) had no effects on the colony formation of CMK cells. In a liquid culture system, 20% of the CMK cells expressed glycoprotein IIb/IIIa (GPIIb/IIIa) antigen without hematopoietic factors, whereas 40% of the cells expressed GPIIb/IIIa with the addition of IL-3 and GM-CSF. EPO also slightly enhanced expression of GPIIb/IIIa. On the other hand, PMA inhibited growth of CMK cells and induced most of them to express the GPIIb/IIIa antigen. Furthermore, PMA induced CMK cells to produce growth activity toward new inocula of CMK cells. This growth factor (GF) contained colony-stimulating activity (CSA) in normal bone marrow (BM) cells. The activity was believed to be attributable mainly to GM-CSF, since 64% of this activity was neutralized by anti-GM-CSF antibodies and a transcript of GM-CSF was detected in mRNA from PMA-treated CMK cells by Northern blot analysis. These observations suggest that GM-CSF, as well as IL-3, should play an important role in megakaryocytopoiesis.

scholarly journals Expression of Activated Mutants of the Human Interleukin-3/Interleukin-5/Granulocyte-Macrophage Colony-Stimulating Factor Receptor Common β Subunit in Primary Hematopoietic Cells Induces Factor-Independent Proliferation and Differentiation

Blood ◽  
1997 ◽  
Vol 90 (4) ◽  
pp. 1471-1481 ◽  
Author(s):  
Matthew P. McCormack ◽  
Thomas J. Gonda
Keyword(s):  
Amino Acid ◽  
Cell Lines ◽  
Amino Acid Substitution ◽  
Single Amino Acid ◽  
Colony Stimulating Factor ◽  
Interleukin 5 ◽  
Proliferation And Differentiation ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor

Abstract To date, several activating mutations have been discovered in the common signal-transducing subunit (hβc) of the receptors for human granulocyte-macrophage colony-stimulating factor, interleukin-3, and interleukin-5. Two of these, FIΔ and I374N, result in a 37 amino acid duplication and a single amino acid substitution in the extracellular domain of hβc, respectively. A third, V449E, results in a single amino acid substitution in the transmembrane domain. Previous studies comparing the activity of these mutants in different hematopoietic cell lines imply that the transmembrane and extracellular mutations act by different mechanisms and suggest the requirement for cell type-specific molecules in signalling. To characterize the ability of these mutant hβc subunits to mediate growth and differentiation of primary cells and hence investigate their oncogenic potential, we have expressed all three mutants in primary murine hematopoietic cells using retroviral transduction. It is shown that, whereas expression of either extracellular hβc mutant confers factor-independent proliferation and differentiation on cells of the neutrophil and monocyte lineages only, expression of the transmembrane mutant does so on these lineages as well as the eosinophil, basophil, megakaryocyte, and erythroid lineages. Factor-independent myeloid precursors expressing the transmembrane mutant display extended proliferation in liquid culture and in some cases yielded immortalized cell lines.

Perverted responses of the human granulocyte-macrophage colony-stimulating factor receptor in mouse cell lines due to cross-species β-subunit association

Blood ◽  
2001 ◽  
Vol 98 (10) ◽  
pp. 3165-3168 ◽  
Author(s):  
Barbara McClure ◽  
Frank Stomski ◽  
Angel Lopez ◽  
Joanna Woodcock
Keyword(s):  
Cell Lines ◽  
Underlying Mechanism ◽  
Mouse Cell ◽  
Colony Stimulating Factor ◽  
Murine Cell ◽  
Gm Csf ◽  
Murine Cell Line ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor

Abstract Transfected murine cell lines are commonly used to study the function of many human cytokine or receptor mutants. This study reports the inappropriate activation of the human granulocyte-macrophage colony-stimulating factor (hGM-CSF) receptor by the human GM-CSF antagonist, E21R, when the human receptor is introduced into the murine cell line BaF-B03. E21R-induced proliferation of the BaF-B03 cells is dependent on transfection with both hGM-CSF receptor α and βc subunits. Studies on the underlying mechanism revealed constitutive association between human and mouse βc and GM-CSF receptor-α, tyrosine phosphorylation of mouse and human βc, and association of phosphorylated mouse βc into an activated human GM-CSF receptor complex in response to E21R and GM-CSF. This interspecies receptor cross-talk of receptor signaling subunits may produce misleading results and emphasizes the need to use cell lines devoid of the cognate endogenous receptors for functional analysis of ligand and receptor mutants.

scholarly journals Inhibition of purinergic P2X receptor 7 (P2X7R) decreases granulocyte-macrophage colony-stimulating factor (GM-CSF) expression in U251 glioblastoma cells

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Matthew Drill ◽  
Kim L. Powell ◽  
Liyen Katrina Kan ◽  
Nigel C. Jones ◽  
Terence J. O’Brien ◽  
...  
Keyword(s):  
Cell Line ◽  
P2x Receptor ◽  
Glioblastoma Cell Line ◽  
Glioblastoma Cell ◽  
Colony Stimulating Factor ◽  
U251 Cell ◽  
Gm Csf ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor

Abstract Glioblastoma is the most aggressive form of primary brain cancer, with a median survival of 12–15 months. The P2X receptor 7 (P2X7R) is upregulated in glioblastoma and is associated with increased tumor cell proliferation. The cytokine granulocyte-macrophage colony-stimulating factor (GM-CSF) is also upregulated in glioblastoma and has been shown to have both pro- and anti-tumor functions. This study investigates the potential mechanism linking P2X7R and GM-CSF in the U251 glioblastoma cell line and the therapeutic potential of P2X7R antagonism in this setting. P2X7R protein and mRNA was demonstrated to be expressed in the U251 cell line as assessed by immunocytochemistry and qPCR. Its channel function was intact as demonstrated by live cell confocal imaging using a calcium indicator Fluo-4 AM. Inhibition of P2X7R using antagonist AZ10606120, decreased both GM-CSF mRNA (P < 0.05) and protein (P < 0.01) measured by qPCR and ELISA respectively. Neutralization of GM-CSF with an anti-GM-CSF antibody did not alter U251 cell proliferation, however, P2X7R antagonism with AZ10606120 significantly reduced U251 glioblastoma cell numbers (P < 0.01). This study describes a novel link between P2X7R activity and GM-CSF expression in a human glioblastoma cell line and highlights the potential therapeutic benefit of P2X7R inhibition with AZ10606120 in glioblastoma.

Sign in / Sign up

Export Citation Format

Share Document