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 Identification and characterization of a low-affinity granulocyte- macrophage colony-stimulating factor receptor on primary and cultured human melanoma cells

Blood ◽  
1991 ◽  
Vol 78 (3) ◽  
pp. 609-615 ◽  
Author(s):  
GC Baldwin ◽  
DW Golde ◽  
GF Widhopf ◽  
J Economou ◽  
JC Gasson
Keyword(s):  
Cell Lines ◽  
Melanoma Cell ◽  
Melanoma Cells ◽  
Human Melanoma ◽  
Colony Stimulating Factor ◽  
Gm Csf ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor ◽  
Melanoma Cell Lines

Abstract Hematopoietic growth factor receptors are present on cells of normal nonhematopoietic tissues such as endothelium and placenta. We previously demonstrated functional human granulocyte-macrophage colony- stimulating factor (GM-CSF) receptors on small cell carcinoma of the lung cell lines, and others have reported that certain solid tumor cell lines respond to GM-CSF in clonogenic assays. In the current study, we examine human melanoma cell lines and fresh specimens of melanoma to determine whether they have functional GM-CSF receptors. Scatchard analyses of 125I-GM-CSF equilibrium binding to melanoma cell lines showed a mean of 542 +/- 67 sites per cell with a kd of 0.72 +/- 0.14 nmol/L. Cross-linking studies in the melanoma cell line, M14, showed a major GM-CSF receptor species of 84,000 daltons. Under the conditions tested, the M14 cells did not have a proliferative response to GM-CSF in vitro, nor was any induction of primary response genes detected by Northern analysis in response to GM-CSF. Studies to determine internal translocation of the receptor-ligand complex indicated less than 10% of the 125I-GM-CSF internalized was specifically bound to receptors. Primary melanoma cells from five surgical specimens had GM-CSF receptors; Scatchard analysis was performed on one sample, showing 555 sites/cell with a kd of 0.23 nmol/L. These results indicate that human tumor cells may express a low-affinity GM-CSF receptor protein that localizes to the cell surface and binds ligand, but lacks functional components or accessory factors needed to transduce a signal.

scholarly journals Production of granulocyte-macrophage colony-stimulating factor by Abelson virus-induced tumorigenic mast cell lines

Blood ◽  
1986 ◽  
Vol 68 (5) ◽  
pp. 1074-1081
Author(s):  
SW Chung ◽  
PM Wong ◽  
G Shen-Ong ◽  
S Ruscetti ◽  
T Ishizaka ◽  
...  
Keyword(s):  
Mast Cells ◽  
Cell Lines ◽  
Colony Formation ◽  
Northern Analysis ◽  
Pokeweed Mitogen ◽  
Colony Stimulating Factor ◽  
Gm Csf ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor

We have recently described a system that supports the development of continuously growing and tumorigenic cell lines after infection of individual multilineage hematopoietic colonies with Abelson murine leukemia virus (A-MuLV). We now provide definitive evidence that these transformed lines express features characteristic of mast cells. Although these lines have been maintained in some cases for more than a year in the absence of exogenous growth factors other than those present in fetal calf serum, colony formation could consistently after 2 months, and variably after 5 months, be shown to be increased several fold when pokeweed mitogen-stimulated spleen cell conditioned medium (CM) was added to the cultures. CM from the A-MuLV-transformed lines was then tested for its ability to stimulate hematopoietic colony formation by cells from both fetal and adult tissues. Four of four randomly selected cell lines produced factors that were active on erythropoietic, granulopoietic, and in some cases pluripotent progenitors. Removal of viral particles from the CM from one of the lines (27d1) by either heat inactivation or high-speed centrifugation did not alter the colony-stimulating activity detected. When CM from 27d1 cells was tested for its ability to stimulate the proliferation of interleukin 3 (IL3) granulocyte-macrophage colony-stimulating factor (GM-CSF)-dependent FDC-P1 cells, a positive result was obtained. This stimulatory activity was not reduced in the presence of neutralizing anti-IL 3 immunoglobulin (Ig), suggesting that the activity detected was GM-CSF and not IL 3. This was confirmed by the lack of expression of the IL 3 gene in 27d1 cells as determined by Northern analysis of 27d1 cell RNA. Furthermore, S1 analysis of mRNA from 27d1 cells as well as two other lines indicated that the GM-CSF gene in all three was transcriptionally active. Taken together, these data suggest that A- MuLV transformation of normal mast cells or their precursors under certain conditions commonly activates the production of GM-CSF.

Characterization of a cell-type-restricted negative regulatory activity of the human granulocyte-macrophage colony-stimulating factor gene

1994 ◽  
Vol 14 (3) ◽  
pp. 2213-2221
Author(s):  
J K Fraser ◽  
J J Guerra ◽  
C Y Nguyen ◽  
J E Indes ◽  
J C Gasson ◽  
...  
Keyword(s):  
Cell Lines ◽  
Regulatory Element ◽  
Myelogenous Leukemia ◽  
Resting Cells ◽  
Colony Stimulating Factor ◽  
Mobility Shift ◽  
Gm Csf ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor

Human granulocyte-macrophage colony-stimulating factor (GM-CSF) stimulates the proliferation and maturation of normal myeloid progenitor cells and can also stimulate the growth of acute myelogenous leukemia (AML) blasts. GM-CSF is not normally produced by resting cells but is expressed by a variety of activated cells including T lymphocytes, macrophages, and certain cytokine-stimulated fibroblasts and endothelial cells. Production of GM-CSF by cultured AML cells has been demonstrated, and GM-CSF expression by normal myeloid progenitors has been postulated to play a role in myelopoiesis. We have investigated the regulation of expression of GM-CSF in AML cell lines, and our results demonstrate the presence of a strong constitutive promoter element contained within 53 bp upstream of the cap site. We have also identified a negative regulatory element located immediately upstream of the positive regulatory element (within 69 bp of the cap site) that is active in AML cell lines but not T cells or K562 CML cells. Competition transfection and mobility shift studies demonstrate that this activity correlates with binding of a 45-kDa protein.

Characterization of cytokine LD78 gene promoters: positive and negative transcriptional factors bind to a negative regulatory element common to LD78, interleukin-3, and granulocyte-macrophage colony-stimulating factor gene promoters

1993 ◽  
Vol 13 (5) ◽  
pp. 2787-2801
Author(s):  
H Nomiyama ◽  
K Hieshima ◽  
K Hirokawa ◽  
T Hattori ◽  
K Takatsuki ◽  
...  
Keyword(s):  
Cell Lines ◽  
Regulatory Elements ◽  
Gene Promoters ◽  
Colony Stimulating Factor ◽  
Interleukin 3 ◽  
Gm Csf ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor

Cytokine LD78 is a human counterpart of the mouse macrophage inflammatory protein 1 alpha/hematopoietic stem cell inhibitor. Promoters of the LD78 alpha and LD78 beta genes showed similar inducible activities in two leukemic cell lines, K562 and Jurkat, but the induction mechanisms differed between the two cell lines. Further characterization of the LD78 alpha promoter indicated that multiple positive and negative regulatory elements are present, some of which are differentially required for induction and repression of the promoter activity in different cells. One of the negative regulatory elements, ICK-1, functioned in both cell lines in the absence and presence of stimulation and was shown to be a recognition site for positive and negative transcriptional factors. This ICK-1 element contained a direct repeat, and similar repeats were also found in the negative regulatory elements of hematopoietic growth factor interleukin-3 (IL-3) and granulocyte-macrophage colony-stimulating factor (GM-CSF) gene promoters. Nuclear extracts from K562 and Jurkat cells formed several protein-DNA complexes with the LD78 alpha ICK-1 element, one of which was also observed with the IL-3 and GM-CSF ICK-1 elements. Results from in vivo and in vitro analyses suggested that the protein forming this complex functions as a negative factor. The binding affinity of this protein, ICK-1A, to the LD78 alpha ICK-1 element was low and was significantly affected by the incubation temperature and the salt concentration in the binding buffer. ICK-1B, another protein bound specifically by the LD78 alpha ICK-1 element, was shown to be a positive factor important for induction of the promoter. These results suggested that ICK-1A plays an important role in balanced expression of LD78, IL-3, and GM-CSF during hematopoietic cell growth and differentiation.

Effects of Constitutive Production of Colony-Stimulating Factors by Human Lung Cancer Cells on Invasive Capacity.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 4211-4211
Author(s):  
Yoshiki Uemura ◽  
Makoto Kobayashi ◽  
Hideshi Nakata ◽  
Tetsuya Kubota ◽  
Hirokuni Taguchi
Keyword(s):  
Lung Cancer ◽  
Cancer Cells ◽  
Tumor Cells ◽  
Cell Lines ◽  
Colony Stimulating Factor ◽  
Gelatinase Activity ◽  
Gm Csf ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor

Abstract In advanced clinical stage, many cases of lung cancer produce colony-stimulating factors (CSFs), including granulocyte-colony stimulating factor (G-CSF), granulocyte macrophage-colony stimulating factor (GM-CSF), and macrophage-colony stimulating factor (M-CSF). The biological properties of the overproduction of CSFs by tumor cells are not well understood. We previously found that CSFs produced by two lung cancer cell lines, OKa-C-1 and MI-4, which constitutively produce abundant amounts of G-CSF, GM-CSF, and M-CSF, stimulate the tumor’s own growth through an autoctine mechanism. In this study, we examined whether CSFs produced by tumor cells contribute to the invasion of the tumor itself. Invasive behaviors of the cancer cells were stimulated by exogenous CSFs but were suppressed by neutralizing antibodies against the CSFs. Matrix-degrading proteinases also are essential for successful tumor cell metastasis. Gelatin zymographs of conditioned media revealed two major bands of gelatinase activity at 68 and 92 kDa. The enzyme activities at 68 and 92 kDa were significantly enhanced in the presence of each CSF in the cell lines but were suppressed by the antibodies against the CSFs. The two gelatinase bands were characterized as matrix metalloproteinase (MMP)-2 and MMP-9, respectively. Our findings suggest that CSFs produced by tumor cells might be closely associated with the tumor invasion through gelatinase activation in CSF-producing lung cancer cells. In addition, an increase of invasion capacity and gelatinase activity by the CSFs might be mediated through the p44/42 mitogen-activated protein kinase (MAPK) signaling pathway. Figure Figure

scholarly journals Production of granulocyte-macrophage colony-stimulating factor by Abelson virus-induced tumorigenic mast cell lines

Blood ◽  
1986 ◽  
Vol 68 (5) ◽  
pp. 1074-1081 ◽  
Author(s):  
SW Chung ◽  
PM Wong ◽  
G Shen-Ong ◽  
S Ruscetti ◽  
T Ishizaka ◽  
...  
Keyword(s):  
Mast Cells ◽  
Cell Lines ◽  
Colony Formation ◽  
Northern Analysis ◽  
Pokeweed Mitogen ◽  
Colony Stimulating Factor ◽  
Gm Csf ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor

Abstract We have recently described a system that supports the development of continuously growing and tumorigenic cell lines after infection of individual multilineage hematopoietic colonies with Abelson murine leukemia virus (A-MuLV). We now provide definitive evidence that these transformed lines express features characteristic of mast cells. Although these lines have been maintained in some cases for more than a year in the absence of exogenous growth factors other than those present in fetal calf serum, colony formation could consistently after 2 months, and variably after 5 months, be shown to be increased several fold when pokeweed mitogen-stimulated spleen cell conditioned medium (CM) was added to the cultures. CM from the A-MuLV-transformed lines was then tested for its ability to stimulate hematopoietic colony formation by cells from both fetal and adult tissues. Four of four randomly selected cell lines produced factors that were active on erythropoietic, granulopoietic, and in some cases pluripotent progenitors. Removal of viral particles from the CM from one of the lines (27d1) by either heat inactivation or high-speed centrifugation did not alter the colony-stimulating activity detected. When CM from 27d1 cells was tested for its ability to stimulate the proliferation of interleukin 3 (IL3) granulocyte-macrophage colony-stimulating factor (GM-CSF)-dependent FDC-P1 cells, a positive result was obtained. This stimulatory activity was not reduced in the presence of neutralizing anti-IL 3 immunoglobulin (Ig), suggesting that the activity detected was GM-CSF and not IL 3. This was confirmed by the lack of expression of the IL 3 gene in 27d1 cells as determined by Northern analysis of 27d1 cell RNA. Furthermore, S1 analysis of mRNA from 27d1 cells as well as two other lines indicated that the GM-CSF gene in all three was transcriptionally active. Taken together, these data suggest that A- MuLV transformation of normal mast cells or their precursors under certain conditions commonly activates the production of GM-CSF.

scholarly journals Identification and characterization of a low-affinity granulocyte- macrophage colony-stimulating factor receptor on primary and cultured human melanoma cells

Blood ◽  
1991 ◽  
Vol 78 (3) ◽  
pp. 609-615 ◽  
Author(s):  
GC Baldwin ◽  
DW Golde ◽  
GF Widhopf ◽  
J Economou ◽  
JC Gasson
Keyword(s):  
Cell Lines ◽  
Melanoma Cell ◽  
Melanoma Cells ◽  
Human Melanoma ◽  
Colony Stimulating Factor ◽  
Gm Csf ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor ◽  
Melanoma Cell Lines

Hematopoietic growth factor receptors are present on cells of normal nonhematopoietic tissues such as endothelium and placenta. We previously demonstrated functional human granulocyte-macrophage colony- stimulating factor (GM-CSF) receptors on small cell carcinoma of the lung cell lines, and others have reported that certain solid tumor cell lines respond to GM-CSF in clonogenic assays. In the current study, we examine human melanoma cell lines and fresh specimens of melanoma to determine whether they have functional GM-CSF receptors. Scatchard analyses of 125I-GM-CSF equilibrium binding to melanoma cell lines showed a mean of 542 +/- 67 sites per cell with a kd of 0.72 +/- 0.14 nmol/L. Cross-linking studies in the melanoma cell line, M14, showed a major GM-CSF receptor species of 84,000 daltons. Under the conditions tested, the M14 cells did not have a proliferative response to GM-CSF in vitro, nor was any induction of primary response genes detected by Northern analysis in response to GM-CSF. Studies to determine internal translocation of the receptor-ligand complex indicated less than 10% of the 125I-GM-CSF internalized was specifically bound to receptors. Primary melanoma cells from five surgical specimens had GM-CSF receptors; Scatchard analysis was performed on one sample, showing 555 sites/cell with a kd of 0.23 nmol/L. These results indicate that human tumor cells may express a low-affinity GM-CSF receptor protein that localizes to the cell surface and binds ligand, but lacks functional components or accessory factors needed to transduce a signal.

scholarly journals Expression of the interleukin-3 and granulocyte-macrophage colony- stimulating factor genes in Friend spleen focus-forming virus-induced erythroleukemia

Blood ◽  
1992 ◽  
Vol 79 (9) ◽  
pp. 2423-2431 ◽  
Author(s):  
Y Shimada ◽  
G Migliaccio ◽  
S Ruscetti ◽  
JW Adamson ◽  
AR Migliaccio
Keyword(s):  
Cell Lines ◽  
Disease Process ◽  
Colony Stimulating Factor ◽  
Interleukin 3 ◽  
Gm Csf ◽  
Erythroid Hyperplasia ◽  
Immortalized Cells ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor

Abstract Friend spleen focus-forming virus (F-SFFV) is a replication-defective retrovirus that induces a multistage erythroleukemia in mice. In the first stage, expression of the SFFV envelope glycoprotein results in erythroid hyperplasia. Subsequently, the F-SFFV integrates near the Spi- 1 gene and activates its expression, resulting in immortalized cells that represent a second stage in the disease process. We report here that media conditioned by erythroleukemia cell lines or leukemic spleen cells induced by the polycythemia-inducing strain of F-SFFV (F-SFFVp), but not medium conditioned by SFFVp-induced hyperplastic spleens, promote the proliferation of normal granulocyte-macrophage progenitor cells and of granulocyte-macrophage colony-stimulating factor (GM-CSF)- and/or interleukin-3 (IL-3)-dependent cell lines. The colony- stimulating activity of the conditioned media from four of five of the lines studied was neutralized by antibodies specific for IL-3 and/or GM- CSF, and IL-3 and GM-CSF-specific mRNA could be detected in the cells after amplification by the polymerase chain reaction. No rearrangements of the IL-3 or GM-CSF genes were observed by Southern blot analysis. However, as previously shown for SFFV-induced cell lines, the Spi-1 gene was expressed in all of these cells. Because the Spi-1 gene encodes a transcription factor whose cognate sequences are present in the promoter region of many hematopoietic growth factor genes, including IL-3 and GM-CSF, Spi-1 activation may be inducing the expression of these genes.

scholarly journals Recombinant Toxins Containing Human Granulocyte-Macrophage Colony-Stimulating Factor and Either Pseudomonas Exotoxin or Diphtheria Toxin Kill Gastrointestinal Cancer and Leukemia Cells

Blood ◽  
1997 ◽  
Vol 90 (1) ◽  
pp. 252-259 ◽  
Author(s):  
Robert J. Kreitman ◽  
Ira Pastan
Keyword(s):  
Cell Lines ◽  
Diphtheria Toxin ◽  
Leukemia Cells ◽  
Colony Stimulating Factor ◽  
Binding Studies ◽  
Pseudomonas Exotoxin ◽  
Gm Csf ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor

Abstract The granulocyte-macrophage colony-stimulating factor receptor (GM-CSFR) is a potential target for toxin-directed therapy, because it is overexpressed on many leukemias and solid tumors and apparently not on stem cells. To investigate the potential therapeutic use of GM-CSF toxins, we fused human GM-CSF to truncated forms of either Pseudomonas exotoxin (PE) or diphtheria toxin (DT) and tested the cytotoxicity of the resulting GM-CSF–PE38KDEL and DT388–GM-CSF on human gastrointestinal (GI) carcinomas and leukemias. Toward gastric and colon cancer cell lines, GM-CSF–PE38KDEL was much more cytotoxic than DT388–GM-CSF, with IC50s (concentration resulting in 50% inhibition of protein synthesis) of 0.5 to 10 ng/mL compared with 4 to 400 ng/mL, respectively. In contrast, toward leukemia lines and fresh bone marrow cells DT388–GM-CSF was more cytotoxic than GM-CSF–PE38KDEL. The cytotoxicity of both GM-CSF–PE38KDEL and DT388–GM-CSF toward the human cells was specific, because it could be competed by an excess of GM-CSF. Binding studies indicated that human GM-CSF receptors were present on all of the human GI and leukemic cell lines tested, at levels of 540 to 3,700 sites per cell (kd = 0.2 to 2 nmol/L), and the number of sites per cell did not correlate with the cell type. A similar pattern of cytotoxicity was found with recombinant immunotoxins binding to the transferrin receptor, in that anti-TFR(Fv)–PE38KDEL was much more cytotoxic than DT388–anti-TFR(Fv) toward GI cells, but both were similar in their cytotoxic activity toward leukemia cells. The fact that PE is more effective than DT in killing GI but not leukemic tumor cells targeted by GM-CSF indicates a fundamental difference in the way PE or DT gains access to the cytosol in these cells. GM-CSF–PE38KDEL and DT388–GM-CSF deserve further evaluation as possible treatments for selected tumors.

scholarly journals Hematopoietic Effects of Paeoniflorin and Albiflorin on Radiotherapy-Induced Myelosuppression Mice

2016 ◽  
Vol 2016 ◽  
pp. 1-8 ◽  
Author(s):  
Yingli Zhu ◽  
Linyuan Wang ◽  
Zhihui Yang ◽  
Jingxia Wang ◽  
Wei Li ◽  
...  
Keyword(s):  
Bone Marrow ◽  
White Blood Cells ◽  
Underlying Mechanism ◽  
Mrna Levels ◽  
Colony Stimulating Factor ◽  
Gm Csf ◽  
Protein Levels ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor

Paeonia lactifloraroot (baishao in Chinese) is a commonly used herb in traditional Chinese medicine (TCM). Paeoniflorin (PF) and albiflorin (AF) are two major active constituents ofP. lactiflora. In this paper, we aimed to investigate the hematopoietic effects of PF and AF on myelosuppression mice induced by radiotherapy and to explore the underlying mechanism. The finding indicated that PF and AF significantly increased the numbers of white blood cells (WBC) and reversed the atrophy of thymus. Furthermore, PF and AF increased the levels of granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin-3 (IL-3) and reduced the levels of tumor necrosis factor-α(TNF-α) in serum and increased the level of colony-stimulating factor (G-CSF) in plasma. Lastly, PF and AF not only enhanced the mRNA levels of GM-CSF and G-CSF in the spleens, but also increased the protein levels of G-CSF and GM-CSF in bone marrow. Our results suggest that PF and AF may promote the recovery of bone marrow hemopoietic function in a myelosuppressed mouse model.

Sign in / Sign up

Export Citation Format

Share Document