Dexamethasone Attenuates Oncostatin M Production via Suppressing of PI3K/Akt/NF-κB Signaling in Neutrophil-like Differentiated HL-60 Cells

Oncostatin M (OSM) plays a role in various inflammatory reactions, and neutrophils are the main source of OSM in pulmonary diseases. However, there is no evidence showing the mechanism of OSM production in neutrophils. While dexamethasone (Dex) has been known to exert anti-inflammatory activity in various fields, the precise mechanisms of OSM downregulation by Dex in neutrophils remain to be determined. Here, we examined how OSM is produced in neutrophil-like differentiated HL-60 cells. Enzyme-linked immunosorbent assay, real-time polymerase chain reaction, and Western blot analysis were utilized to assess the potential of Dex. Granulocyte-macrophage colony-stimulating factor (GM-CSF) stimulation resulted in OSM elevation in neutrophil-like dHL-60 cells. OSM elevation induced by GM-CSF is regulated by phosphatidylinositol 3-kinase (PI3K)/Akt/nuclear factor (NF)-kB signal cascades. GM-CSF stimulation upregulated phosphorylated levels of PI3K or Akt or NF-κB in neutrophil-like dHL-60 cells. Treatment with Dex decreased OSM levels as well as the phosphorylated levels of PI3K or Akt or NF-κB in neutrophil-like dHL-60 cells. Our findings show the potential of Dex in the treatment of inflammatory diseases via blocking of OSM.

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Bone marrow stromal fibroblasts secrete interleukin-6 and granulocyte- macrophage colony-stimulating factor in the absence of inflammatory stimulation: demonstration by serum-free bioassay, enzyme-linked immunosorbent assay, and reverse transcriptase polymerase chain reaction

Blood ◽

10.1182/blood.v80.5.1190.1190 ◽

1992 ◽

Vol 80 (5) ◽

pp. 1190-1198 ◽

Cited By ~ 70

Author(s):

SC Guba ◽

CI Sartor ◽

LR Gottschalk ◽

YH Jing ◽

T Mulligan ◽

...

Keyword(s):

Human Serum ◽

Enzyme Linked Immunosorbent Assay ◽

Stromal Fibroblasts ◽

Gm Csf ◽

Normal Human ◽

Macrophage Colony Stimulating Factor ◽

Polymerase Chain ◽

Macrophage Colony ◽

Stimulating Factor

Abstract Bone marrow (BM) stromal fibroblasts produce hematopoietic growth factors (HGFs) in response to inflammatory mediators such as tumor necrosis factor-alpha or interleukin-1 alpha (IL-1 alpha). In the absence of such inflammatory stimuli, production of HGFs by BM stromal cells has been problematic and controversial. In vivo, however, basal hematopoiesis maintains blood counts within a normal homeostatic range even in the absence of inflammation, and HGFs are required for progenitor cell differentiation in vitro. To better ascertain the contribution of BM stromal fibroblasts to basal hematopoiesis, we therefore studied HGF production in quiescent BM stromal fibroblasts by three sensitive assays: serum-free bioassay, enzyme-linked immunosorbent assay, and reverse transcriptase polymerase chain reaction. Stromal fibroblasts were cultured in the presence or absence of normal human serum to determine if serum factor(s) present in the noninflammatory (basal) state induce secretion of HGFs. Human serum was found to induce or enhance transcription and secretion of granulocyte- macrophage colony-stimulating factor (GM-CSF) and enhance secretion of constitutively expressed IL-6. In contrast, no secretion of either granulocyte-CSF (G-CSF) or IL-3 was found. These data indicate that factors in normal human serum are active in enhancing GM-CSF and IL-6 production by stromal fibroblasts and suggest that these growth factors contribute to the maintainance of normal, basal hematopoiesis in vivo.

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Bone marrow stromal fibroblasts secrete interleukin-6 and granulocyte- macrophage colony-stimulating factor in the absence of inflammatory stimulation: demonstration by serum-free bioassay, enzyme-linked immunosorbent assay, and reverse transcriptase polymerase chain reaction

Blood ◽

10.1182/blood.v80.5.1190.bloodjournal8051190 ◽

1992 ◽

Vol 80 (5) ◽

pp. 1190-1198 ◽

Cited By ~ 1

Author(s):

SC Guba ◽

CI Sartor ◽

LR Gottschalk ◽

YH Jing ◽

T Mulligan ◽

...

Keyword(s):

Human Serum ◽

Enzyme Linked Immunosorbent Assay ◽

Stromal Fibroblasts ◽

Gm Csf ◽

Normal Human ◽

Macrophage Colony Stimulating Factor ◽

Polymerase Chain ◽

Macrophage Colony ◽

Stimulating Factor

Bone marrow (BM) stromal fibroblasts produce hematopoietic growth factors (HGFs) in response to inflammatory mediators such as tumor necrosis factor-alpha or interleukin-1 alpha (IL-1 alpha). In the absence of such inflammatory stimuli, production of HGFs by BM stromal cells has been problematic and controversial. In vivo, however, basal hematopoiesis maintains blood counts within a normal homeostatic range even in the absence of inflammation, and HGFs are required for progenitor cell differentiation in vitro. To better ascertain the contribution of BM stromal fibroblasts to basal hematopoiesis, we therefore studied HGF production in quiescent BM stromal fibroblasts by three sensitive assays: serum-free bioassay, enzyme-linked immunosorbent assay, and reverse transcriptase polymerase chain reaction. Stromal fibroblasts were cultured in the presence or absence of normal human serum to determine if serum factor(s) present in the noninflammatory (basal) state induce secretion of HGFs. Human serum was found to induce or enhance transcription and secretion of granulocyte- macrophage colony-stimulating factor (GM-CSF) and enhance secretion of constitutively expressed IL-6. In contrast, no secretion of either granulocyte-CSF (G-CSF) or IL-3 was found. These data indicate that factors in normal human serum are active in enhancing GM-CSF and IL-6 production by stromal fibroblasts and suggest that these growth factors contribute to the maintainance of normal, basal hematopoiesis in vivo.

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Induction of anti-recombinant human granulocyte-macrophage colony- stimulating factor (Escherichia coli-derived) antibodies and clinical effects in nonimmunocompromised patients

Blood ◽

10.1182/blood.v84.12.4078.bloodjournal84124078 ◽

1994 ◽

Vol 84 (12) ◽

pp. 4078-4087 ◽

Cited By ~ 75

Author(s):

P Ragnhammar ◽

HJ Friesen ◽

JE Frodin ◽

AK Lefvert ◽

M Hassan ◽

...

Keyword(s):

Escherichia Coli ◽

Antibody Response ◽

Enzyme Linked Immunosorbent Assay ◽

Clinical Effects ◽

Immunocompromised Patients ◽

Colony Stimulating Factor ◽

Gm Csf ◽

Macrophage Colony Stimulating Factor ◽

Macrophage Colony ◽

Stimulating Factor

The pharmacokinetics of recombinant human granulocyte-macrophage colony- stimulating factor (rhGM-CSF), induction of anti-GM-CSF antibodies, and clinical effects related to the induction of the antibodies were analyzed in patients with metastatic colorectal carcinoma (CRC) who were not on chemotherapy (n = 20, nonimmunocompromised patients). rhGM- CSF (250 micrograms/m2/d; Escherichia coli-derived) was administered subcutaneously for 10 days every month for 4 months. Eight patients with multiple myeloma (MM) on intensive chemotherapy followed by rhGM- CSF treatment were also included (immunocompromised patients). After a single injection of GM-CSF at the first cycle in CRC patients, the maximum calculated concentration (Cmax) was 5.24 +/- 0.56 ng/mL; the half life (T1/2) was 2.91 +/- 0.8 hours; and the area under the concentration curve (AUC) was 30.86 +/- 6.03 hours x ng/mL (mean +/- SE). No anti-GM-CSF antibodies were detected. During the subsequent cycles, 95% of the CRC patients developed anti-GM-CSF IgG antibodies, which significantly altered the pharmacokinetics of rhGM-CSF at the third and fourth cycles with decreased Cmax (2.87 +/- 0.57 ng/mL; P < .05), T1/2 (1.57 +/- 0.2 hours; P < .05), and AUC (14.90 +/- 4.10 hours x ng/mL; P < .005). The presence of anti-GM-CSF antibodies significantly reduced the GM-CSF-induced enhancement of granulocytes, and there was a clear tendency for a decreased increment of monocytes. Antibodies diminished systemic side effects of rhGM-CSF. Only 1 of 8 MM patients showed a very low anti-GM-CSF antibody titer after GM-CSF therapy, as shown by enzyme-linked immunosorbent assay and Western blot. Therefore, in nonimmunocompromised patients, exogenous nonglycosylated GM-CSF induced an anti-GM-CSF IgG antibody response in practically all patients, which seemed to be of clinical significance. In immunocompromised patients, virtually no significant antibody response was shown.

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Identification of functionally distinct domains of human granulocyte- macrophage colony-stimulating factor using monoclonal antibodies

Blood ◽

10.1182/blood.v77.5.1033.1033 ◽

1991 ◽

Vol 77 (5) ◽

pp. 1033-1043 ◽

Cited By ~ 29

Author(s):

Y Kanakura ◽

SA Cannistra ◽

CB Brown ◽

M Nakamura ◽

GF Seelig ◽

...

Keyword(s):

Monoclonal Antibodies ◽

Neutralizing Antibodies ◽

Solid Phase ◽

Receptor Interaction ◽

Enzyme Linked Immunosorbent Assay ◽

Colony Stimulating Factor ◽

Gm Csf ◽

Macrophage Colony Stimulating Factor ◽

Macrophage Colony ◽

Stimulating Factor

Abstract Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a glycoprotein that is required for the survival, growth, and differentiation of hematopoietic progenitor cells. Although the primary structure of GM-CSF is known from cDNA cloning, the relationship between structure and function of GM-CSF is not fully understood. Fifteen different monoclonal antibodies (MoAbs) to human GM-CSF were generated to map immunologically distinct areas of the molecule. Each of the MoAbs was biotinylated and shown by enzyme-linked immunosorbent assay to bind to recombinant GM-CSF that had been affixed to a solid phase. Each of the 15 unconjugated MoAbs was then used to compete with each biotinylated MoAb for binding to GM-CSF. These cross-blocking studies identified eight distinct epitopes of native GM-CSF. Seven of these epitopes were also present in denatured GM-CSF by Western blotting, and four of the epitopes were at least partially conserved on GM-CSF that was reduced in beta-mercaptoethanol. MoAbs to four of eight epitopes neutralized both recombinant (glycosylated and nonglycosylated) and natural human GM-CSF in a GM colony-forming unit (CFU-GM) assay and blocked GM-CSF-induced activation of neutrophils. For most of the antibodies there was a good correlation between neutralizing activity and the capacity to block binding of 125I-GM-CSF to neutrophils or blasts. Non-neutralizing antibodies to one epitope partially blocked binding of 125I-GM-CSF to neutrophils. None of the MoAbs neutralized interleukin-3, G-CSF, or M-CSF. The locations of seven of the epitopes could be partially mapped with regard to the amino acid structure by determining reactivity to GM-CSF synthetic peptides or to human-mouse chimeric GM-CSFs. The neutralizing antibodies were found to map to amino acids 40–77, 78–94, or 110–127. Thus, these MoAbs are useful to identify functional domains of GM-CSF and in identifying regions that are likely to be involved in receptor interaction.

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Siglec-9 transduces apoptotic and nonapoptotic death signals into neutrophils depending on the proinflammatory cytokine environment

Blood ◽

10.1182/blood-2004-10-4112 ◽

2005 ◽

Vol 106 (4) ◽

pp. 1423-1431 ◽

Cited By ~ 139

Author(s):

Stephan von Gunten ◽

Shida Yousefi ◽

Michael Seitz ◽

Stephan M. Jakob ◽

Thomas Schaffner ◽

...

Keyword(s):

Rheumatoid Arthritis ◽

Inflammatory Diseases ◽

Interferon Α ◽

Gm Csf ◽

Independent Form ◽

Blood Neutrophils ◽

Macrophage Colony Stimulating Factor ◽

Macrophage Colony ◽

Induced Apoptosis

Abstract We report about new apoptotic and non-apoptotic death pathways in neutrophils that are initiated via the surface molecule sialic acid-binding immunoglobulin-like lectin (Siglec)-9. In normal neutrophils, Siglec-9 ligation induced apoptosis. Inflammatory neutrophils obtained from patients with acute septic shock or rheumatoid arthritis demonstrated increased Siglec-9, but normal Fas receptor-mediated cytotoxic responses when compared with normal blood neutrophils. The increased Siglec-9-mediated death was mimicked in vitro by short-term preincubation of normal neutrophils with proinflammatory cytokines, such as granulocyte/macrophage colony-stimulating factor (GM-CSF), interferon-α (IFN-α), and IFN-γ, and was demonstrated to be caspase independent. Experiments using scavengers of reactive oxygen species (ROS) or neutrophils unable to generate ROS indicated that both Siglec-9-mediated caspase-dependent and caspase-independent forms of neutrophil death depend on ROS. Interestingly, the caspase-independent form of neutrophil death was characterized by cytoplasmic vacuolization and several other nonapoptotic morphologic features, which were also seen in neutrophils present in joint fluids from rheumatoid arthritis patients. Taken together, these data suggest that apoptotic (ROS- and caspase-dependent) and nonapoptotic (ROS-dependent) death pathways are initiated in neutrophils via Siglec-9. The new insights have important implications for the pathogenesis, diagnosis, and treatment of inflammatory diseases such as sepsis and rheumatoid arthritis. (Blood. 2005;106:1423-1431)

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Cascade transactivation of growth factor receptors in early human hematopoiesis

Blood ◽

10.1182/blood.v81.6.1442.bloodjournal8161442 ◽

1993 ◽

Vol 81 (6) ◽

pp. 1442-1456 ◽

Cited By ~ 1

Author(s):

U Testa ◽

E Pelosi ◽

M Gabbianelli ◽

C Fossati ◽

S Campisi ◽

...

Keyword(s):

Growth Factor ◽

Growth Factor Receptors ◽

Cross Reactivity ◽

Gm Csf ◽

Synergistic Interactions ◽

Multipotent Progenitors ◽

Macrophage Colony Stimulating Factor ◽

Polymerase Chain ◽

Macrophage Colony ◽

Key Steps

Highly purified progenitors (including erythroid [BFU-E], granulo- monocytic [CFU-GM], multipotent [CFU-GEMM] progenitors, as well as multipotent progenitors with self-renewal capacity [CFU-B]) express high-affinity growth factor receptors (GFRs), with prevalent interleukin-3 receptors (IL-3Rs) (2,700/cell), a > or = 10-fold lower number of IL-6Rs (145/cell) and granulocyte-macrophage colony- stimulating factor receptors (GM-CSFRs) (300/cell), and a barely detectable level of erythropoietin (Ep) receptors (75/cell). Hematopoietic growth factor (HGF) dosages inducing peak clonogenetic effects are associated with partial/subtotal occupancy of the homologous HGF receptor (HGFR). Cross-reactivity between GFRs and heterologous GFs (including IL-6, IL-3, GM-CSF, Ep, and the kit ligand [KL]) was explored by competition experiments on purified progenitors with radiolabeled and excess cold HGFs at +4 degrees C. No cross- reaction was observed between IL-6R, IL-3R, EpR, and the heterologous GFs, whereas the GM-CSFR showed cross-reactivity with IL-3 and, to a lesser extent, KL. Modulation of GFRs was examined after 18 or 40 hours of incubation with GF(s) at 37 degrees C, followed by ligand-binding assay at 20 degrees C. IL-6, IL-3, GM-CSF, and Ep induce a marked down- modulation of their own receptors. Interestingly, each GF induces the transactivation of the R(s) for the “distal” GF(s): (1) IL-6 induces transactivation of IL-3R, but not of GM-CSFR/EpR; (2) IL-3 causes a rapid upmodulation of GM-CSFR/EpR (“pure” progenitors treated with IL-3 show upmodulation of GM-CSFR alpha-chain mRNA by reverse transcriptase- polymerase chain reaction); whereas (3) GM-CSF induces the transactivation of the EpR. This chain upmodulation of HGFRs may underlie the synergistic interactions between the HGFs in clonogenetic culture. It is emphasized that KL does not induce upmodulation of the other GFRs. Finally, Ep, GM-CSF, and IL-3 do not modulate the expression of the “proximal” HGFRs (ie, GM-CSFR/IL-3R/IL-6R, IL-3R/IL- 6R, and IL-6R, respectively). These results allow insight into the cellular basis of hematopoiesis, ie, the complex and coordinate interactions between HGFs and their receptors. They are compatible with a model of cascade transactivation via upmodulation of GFRs in the initial key steps of hematopoietic differentiation, whereby the action of each GF enhances the effect of the distal GF(s) by a multistep chain- potentiation mechanism.

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The genes for leukemia inhibitory factor and interleukin-6 are expressed in mouse blastocysts prior to the onset of hemopoiesis

Molecular and Cellular Biology ◽

10.1128/mcb.10.9.4953-4956.1990 ◽

1990 ◽

Vol 10 (9) ◽

pp. 4953-4956

Author(s):

R Murray ◽

F Lee ◽

C P Chiu

Keyword(s):

Stem Cells ◽

Interleukin 6 ◽

Leukemia Inhibitory Factor ◽

Embryonic Stem ◽

Polymerase Chain Reaction Analysis ◽

Inhibitory Factor ◽

Gm Csf ◽

Macrophage Colony Stimulating Factor ◽

Polymerase Chain ◽

Macrophage Colony

We have investigated the role that hemopoietic regulatory molecules may play in mouse embryogenesis prior to the appearance of hemopoietic stem cells or their microenvironments. Using polymerase chain reaction analysis, we detected mRNA transcripts for interleukin-6 (IL-6) and leukemia inhibitory factor (LIF) but not for granulocyte-macrophage colony-stimulating factor (GM-CSF) or IL-3 in mouse blastocysts at 3.5 days of gestation. Functional IL-6 protein was also detected in cultured blastocysts as a secreted product, as was an activity consistent with the presence of LIF protein. The expression of IL-6 and LIF in blastocysts prior to hemopoiesis suggests that these proteins may regulate the growth and development of trophoblasts or embryonic stem cells.

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The genes for leukemia inhibitory factor and interleukin-6 are expressed in mouse blastocysts prior to the onset of hemopoiesis.

Molecular and Cellular Biology ◽

10.1128/mcb.10.9.4953 ◽

1990 ◽

Vol 10 (9) ◽

pp. 4953-4956 ◽

Cited By ~ 70

Author(s):

R Murray ◽

F Lee ◽

C P Chiu

Keyword(s):

Stem Cells ◽

Interleukin 6 ◽

Leukemia Inhibitory Factor ◽

Embryonic Stem ◽

Polymerase Chain Reaction Analysis ◽

Inhibitory Factor ◽

Gm Csf ◽

Macrophage Colony Stimulating Factor ◽

Polymerase Chain ◽

Macrophage Colony

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Granulocyte - macrophage colony stimulating factor and interleukin-8 in the reproductive tract of ewes following oestrus and mating

Reproduction Fertility and Development ◽

10.1071/rd06137 ◽

2007 ◽

Vol 19 (4) ◽

pp. 585 ◽

Cited By ~ 6

Author(s):

Jennifer L. Scott ◽

Natkunam Ketheesan ◽

Phillip M. Summers

Keyword(s):

Reproductive Tract ◽

Staining Intensity ◽

Female Reproductive Tract ◽

Enzyme Linked Immunosorbent Assay ◽

Colony Stimulating Factor ◽

Tissue Samples ◽

Gm Csf ◽

Macrophage Colony Stimulating Factor ◽

Macrophage Colony ◽

Stimulating Factor

Cytokines produced in the female reproductive tract after mating may enhance reproductive success. The present study investigated the distribution of granulocyte–macrophage colony stimulating factor (GM-CSF) and interleukin (IL)-8 in tissues and luminal secretions from different sites in the reproductive tract of the ewe following oestrus and after natural mating. Fifteen ewes were mated with a ram for 1 h and their reproductive tracts collected 3, 6, 18, 24 or 48 h later. Another 15 ewes were used as oestrous controls. Luminal secretions and tissue samples were collected from seven sites in each reproductive tract. Secretions were analysed by enzyme-linked immunosorbent assay and tissues were stained immunohistochemically using anti-sheep GM-CSF and anti-sheep IL-8 antibodies. Both cytokines were found in luminal and glandular endometrial epithelium and, to a lesser extent, in cervical epithelium; neither was found in the vaginal epithelium. Twice as many (P < 0.05) luminal samples from mated ewes than non-mated ewes were positive for GM-CSF. The vaginal lumen contained significantly higher (P < 0.01) concentrations of IL-8 compared with other sites, irrespective of mating status. Significant differences (P < 0.05) were found in staining intensity of GM-CSF and IL-8 from different sites. Production of GM-CSF and IL-8 by reproductive tissues is likely to contribute to leucocyte infiltration into the ovine reproductive tract.

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Systematic cytokine receptor profiling reveals GM-CSF as a novel TLR-independent activator of human plasmacytoid predendritic cells

Blood ◽

10.1182/blood-2010-01-266932 ◽

2010 ◽

Vol 115 (24) ◽

pp. 5037-5040 ◽

Cited By ~ 29

Author(s):

Cristina Ghirelli ◽

Raphaël Zollinger ◽

Vassili Soumelis

Keyword(s):

Inflammatory Diseases ◽

Interferon Γ ◽

Toll Like Receptor ◽

Microbial Infection ◽

Systematic Analysis ◽

Gm Csf ◽

Protein Levels ◽

Macrophage Colony Stimulating Factor ◽

Macrophage Colony ◽

Activation Pathways

Abstract Human plasmacytoid predendritic cells (pDCs) can be activated during microbial infection through Toll-like receptor engagement. They are also involved in nonmicrobial inflammatory diseases, but their activation pathways in this context remain elusive. To identify Toll-like receptor-independent pDC activators, we performed a systematic analysis of cytokine receptors on primary human pDCs. Six receptors were expressed both at mRNA and protein levels: interleukin-3 receptor (IL-3R), IL-6R, IL-10R, IL-18R, interferon-γ receptor, and granulocyte-macrophage colony-stimulating factor (GM-CSF) receptor. Only GM-CSF and IL-3 were able to efficiently promote pDC survival and induce their differentiation into dendritic cells. Allogeneic naive CD4 T cells primed with GM-CSF–activated pDCs produced more interferon-γ and less IL-4 and IL-10 compared with IL-3–activated pDCs, indicating a shift in the Th1/Th2 balance. Our data point at a novel function of GM-CSF, which may serve as a link between a pathologic inflammatory environment, pDC activation, and the modulation of CD4 T-cell responses.

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