Mechanisms involved in methylmercuric chloride (MeHgCl)-induced suppression of human neutrophil apoptosis

2003 ◽  
Vol 22 (12) ◽  
pp. 629-637 ◽  
Author(s):  
éliane Moisan ◽  
édouard Kouassi ◽  
Denis Girard
Keyword(s):  
Annexin V ◽  
Surface Expression ◽  
Cytoskeletal Proteins ◽  
Cell Surface Expression ◽  
Neutrophil Apoptosis ◽  
Gm Csf ◽  
Methylmercuric Chloride ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor

We have previously demonstrated that concentrations of 1-10 μM of methylmercuric chloride (MeHgCl) that are cytotoxic to monocytes-macrophages can curiously inhibit neutrophil apoptosis by a yet unknown mechanism. In the present study, we demonstrate that, as with the cytokine granulocyte-macrophage colony-stimulating factor (GM-CSF), a classical inhibitor of neutrophil apoptosis, treatment of cells with 5 M MeHgCl inducesde novo protein synthesis and prevents the loss of expression of the antiapoptotic Mcl-1 protein. The expression of the cytoskeletal proteins gelsolin, paxillin and vinculin was similar in MeHgCl or GM-CSF-induced suppression of apoptosis. However, MeHgCl prevents the degradation of vimentin differently than GM-CSF. Apoptosis was further confirmed by flow cytometry (FITC annexin-V), and by monitoring CD16 cell surface expression. Curiously, unlike GM-CSF, MeHgCl did not prevent CD16 shedding. We conclude that, like GM-CSF, MeHgCl can delay neutrophil apoptosis by inducing de novoprotein synthesis and by preventing the loss of the antiapoptotic Mcl-1 protein. However, unlike GM-CSF, MeHgCl induces an atypical degradation of vimentin without preventing CD16 shedding.

Three conserved motifs in the extracellular domain of the human granulocyte-macrophage colony-stimulating factor receptor subunit are essential for ligand binding and surface expression

Blood ◽  
1994 ◽  
Vol 84 (8) ◽  
pp. 2539-2553 ◽  
Author(s):  
PD Doshi ◽  
JF DiPersio
Keyword(s):  
Ligand Binding ◽  
Extracellular Domain ◽  
Surface Expression ◽  
Colony Stimulating Factor ◽  
Cos Cells ◽  
Gm Csf ◽  
Alpha Subunits ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor

Abstract The receptor for the human granulocyte-macrophage colony-stimulating factor (GM-CSF) (GM-R) is a heterodimeric complex consisting of two subunits, GM-R alpha and GM-R beta. Structural analyses have shown a number of highly conserved amino acid motifs present in both GM-R alpha and GM-R beta. These motifs include QYFLY, CXW, XW, and WSXWS motifs in the extracellular domain; a conserved cysteine in the transmembrane domain; and the entire cytoplasmic domain, including the LXVLX box in the carboxy terminal region of the cytoplasmic domain. We have investigated the role of these motifs in GM-R alpha by examining the effects of specific motif mutations on ligand binding and surface expression. Transient expression of these mutant GM-R alpha subunits in COS cells shows that these extracellular motis are essential for ligand binding. Alterations of the cytoplasmic region of GM-R alpha do not alter GM-CSF binding or the reconstitution of high-affinity receptors when coexpressed with GM-R beta. Permeabilization and immunostaining of cells transfected with mutant GM-R alpha subunits yields data suggesting that each of the mutant subunits is present in the cytoplasm. Immunostaining of both intact and permeabilized COS cells transiently transfected with wild-type or mutant GM-R alpha s showed that extracellular domain mutants accumulated in the cytoplasm and were not efficiently transported to the cell surface.

scholarly journals Three conserved motifs in the extracellular domain of the human granulocyte-macrophage colony-stimulating factor receptor subunit are essential for ligand binding and surface expression

Blood ◽  
1994 ◽  
Vol 84 (8) ◽  
pp. 2539-2553 ◽  
Author(s):  
PD Doshi ◽  
JF DiPersio
Keyword(s):  
Ligand Binding ◽  
Extracellular Domain ◽  
Surface Expression ◽  
Colony Stimulating Factor ◽  
Cos Cells ◽  
Gm Csf ◽  
Alpha Subunits ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor

The receptor for the human granulocyte-macrophage colony-stimulating factor (GM-CSF) (GM-R) is a heterodimeric complex consisting of two subunits, GM-R alpha and GM-R beta. Structural analyses have shown a number of highly conserved amino acid motifs present in both GM-R alpha and GM-R beta. These motifs include QYFLY, CXW, XW, and WSXWS motifs in the extracellular domain; a conserved cysteine in the transmembrane domain; and the entire cytoplasmic domain, including the LXVLX box in the carboxy terminal region of the cytoplasmic domain. We have investigated the role of these motifs in GM-R alpha by examining the effects of specific motif mutations on ligand binding and surface expression. Transient expression of these mutant GM-R alpha subunits in COS cells shows that these extracellular motis are essential for ligand binding. Alterations of the cytoplasmic region of GM-R alpha do not alter GM-CSF binding or the reconstitution of high-affinity receptors when coexpressed with GM-R beta. Permeabilization and immunostaining of cells transfected with mutant GM-R alpha subunits yields data suggesting that each of the mutant subunits is present in the cytoplasm. Immunostaining of both intact and permeabilized COS cells transiently transfected with wild-type or mutant GM-R alpha s showed that extracellular domain mutants accumulated in the cytoplasm and were not efficiently transported to the cell surface.

scholarly journals The diadenosine polyphosphates Ap3A and Ap4A and adenosine triphosphate interact with granulocyte-macrophage colony-stimulating factor to delay neutrophil apoptosis: implications for neutrophil: platelet interactions during inflammation

Blood ◽  
1996 ◽  
Vol 87 (8) ◽  
pp. 3442-3449 ◽  
Author(s):  
L Gasmi ◽  
AG McLennan ◽  
SW Edwards
Keyword(s):  
Adenosine Triphosphate ◽  
Immune Cell ◽  
Cellular Morphology ◽  
Neutrophil Apoptosis ◽  
Colony Stimulating Factor ◽  
Diadenosine Polyphosphates ◽  
Gm Csf ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor

Incubation of neutrophils with cytokines such as granulocyte macrophage colony-stimulating factor (GM-CSF) delays their loss of function and changes in cellular morphology that are characteristic of apoptosis. Adenosine triphosphate (ATP) and the diadenosine polyphosphates Ap4A and AP3A were almost as effective as GM-CSF in delaying neutrophil apoptosis. The nucleotides could thus preserve cellular morphology, protect against chromatin fragmentation, and preserve functions such as NADPH oxidase activity and expression of CD16. Moreover, addition of ATP, AP3A and AP4A together with GM-CSF resulted in more pronounced protection from apoptosis than was observed during incubation with either the cytokine or the nucleotides alone. Because ATP, Ap3A, and AP4A may be secreted from activated platelets, these observations suggest that platelet-derived products, perhaps acting in combination with endothelial-derived or immune cell-derived cytokines, can regulate neutrophil function during certain types of inflammation.

scholarly journals Toxoplasma gondii Induces Granulocyte Colony-Stimulating Factor and Granulocyte-Macrophage Colony-Stimulating Factor Secretion by Human Fibroblasts: Implications for Neutrophil Apoptosis

2002 ◽  
Vol 70 (11) ◽  
pp. 6048-6057 ◽  
Author(s):  
Jacqueline Y. Channon ◽  
Kristin A. Miselis ◽  
Laurie A. Minns ◽  
Chaitali Dutta ◽  
Lloyd H. Kasper
Keyword(s):  
Conditioned Medium ◽  
Human Fibroblasts ◽  
Neutrophil Apoptosis ◽  
Granulocyte Colony Stimulating Factor ◽  
Colony Stimulating Factor ◽  
Gm Csf ◽  
Soluble Mediator ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor

ABSTRACT Human neutrophils are rescued from apoptosis following incubation with once-washed, fibroblast-derived Toxoplasma gondii tachyzoites. Both infected and uninfected neutrophils are rescued, implicating a soluble mediator. In this study we investigated the origin and identity of this soluble mediator. Neutrophils were incubated either with purified tachyzoites or with conditioned medium derived from T. gondii-infected human fibroblasts. Conditioned medium was found to be a potent stimulus that delayed neutrophil apoptosis up to 72 h, whereas purified and extensively washed tachyzoites had no effect. Delayed apoptosis correlated with up-regulation of the neutrophil antiapoptotic protein, Mcl-1, and the neutrophil interleukin 3 receptor α subunit (IL-3Rα), suggesting a role for granulocyte-macrophage colony-stimulating factor (GM-CSF). GM-CSF and granulocyte colony-stimulating factor (G-CSF) were measurable in conditioned medium by enzyme-linked immunosorbent assay. Neutralizing antibodies to GM-CSF and G-CSF were additive in abrogating delayed neutrophil apoptosis induced by conditioned medium. Inhibitors of Src family tyrosine kinases, Gi proteins, phosphatidylinositol 3-kinase, p44 erk1 and p42 erk2 mitogen-activated protein kinases, and Jak2 kinases partially attenuated the effect of conditioned medium, consistent with a role for G-CSF and/or GM-CSF. Hence, delayed neutrophil apoptosis is mediated by GM-CSF and G-CSF secreted by T. gondii-infected human fibroblasts. This enhanced neutrophil survival may contribute to the robust proinflammatory response elicited in the T. gondii-infected host.

scholarly journals Effects of granulocyte-macrophage colony-stimulating factor and cyclic AMP interaction on human neutrophil apoptosis

1998 ◽  
Vol 7 (6) ◽  
pp. 391-396 ◽  
Author(s):  
Cosimo Tortorella ◽  
Giuseppina Piazzolla ◽  
Felice Spaccavento ◽  
Salvatore Antonaci
Keyword(s):  
Cyclic Amp ◽  
Camp Signaling ◽  
Neutrophil Apoptosis ◽  
Colony Stimulating Factor ◽  
Dibutyryl Camp ◽  
Gm Csf ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Neutrophil Survival ◽  
Stimulating Factor

The current study was undertaken to evaluate the effects of granulocyte-macrophage colony-stimulating factor (GM-CSF) and cyclic AMP (cAMP) signaling interaction on human neutrophil apoptosis, either occurring spontaneously or induced by Fas antigen activation. Results show that GM-CSF, dibutyryl cAMP (a cAMP analog) and forskolin (an adenylate cyclase activator) are all able to suppress spontaneous neutrophil cell death. Of note however, when GM-CSF is used in combination with cAMP-elevating agents, an additive effect on neutrophil survival is observed with dibutyryl cAMP only, whereas supplementation of cell cultures with GM-CSF and forskolin results in a progressive reduction of antiapoptotic effects exerted by the single compounds. Moreover, although dibutyryl cAMP and forskolin do not affect Fas-triggered apoptotic events, they are still able to modulate the GM-CSF capacity to prolong neutrophil survival following anti-Fas IgM cell challenge, with effects similar to those respectively exerted on spontaneous neutrophil apoptosis. The data indicate that GM-CSF m ay negatively modulate the cAMP-mediated antiapoptotic pathway in human neutrophils, likely via the inhibition of adenylate cyclase activity. This would prevent an abnormal neutrophil survival as a result of cAMP signaling stimulation, which provides a novel insight into the role of GM-CSF as a physiological regulator of myeloid cell turnover.

scholarly journals Application of Western Blot for detection of neutrophil apoptosis-related proteins

2009 ◽  
Vol 57 (1) ◽  
pp. 97-104
Author(s):  
Petr Sláma
Keyword(s):  
Western Blot ◽  
Myeloid Cell ◽  
Neutrophil Apoptosis ◽  
Colony Stimulating Factor ◽  
Gm Csf ◽  
Western Blot Method ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Related Proteins ◽  
Stimulating Factor

The aim of this study was to evaluate suitability of using Western Blot for detection of neutrophil apo­pto­sis and neutrophil apoptosis-related proteins, respectively. Neutrophils were isolated from blood of healthy adult donors and incubated with G-CSF (granulocyte colony stimulating factor), GM-CSF (granulocyte-macrophage colony stimulating factor), ATP (adenosine triphosphate) and FMLP (N-formyl-methionyl-leucyl-phenylalanine). The neutrophils were incubated 4, 8 and 20 hours at 37 °C. In this assay, an expression of Mcl-1 (myeloid cell leukemia 1), XIAP (X-linked inhibitor of apoptosis) and gelsolin was analysed by Western Blot method. The results showed that Western Blot is a suitable method for detection of neutrophil apoptosis-related proteins and detection of neutrophil apoptosis, respectively.

The dual effects of TNFα on neutrophil apoptosis are mediated via differential effects on expression of Mcl-1 and Bfl-1

Blood ◽  
2008 ◽  
Vol 111 (2) ◽  
pp. 878-884 ◽  
Author(s):  
Andrew Cross ◽  
Robert J. Moots ◽  
Steven W. Edwards
Keyword(s):  
Inflammatory Diseases ◽  
Neutrophil Apoptosis ◽  
Colony Stimulating Factor ◽  
Gm Csf ◽  
Low Concentrations ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Opposing Effects ◽  
Neutrophil Survival ◽  
Stimulating Factor

Neutrophils have a very short half-life in the circulation, undergoing rapid death by apoptosis, but a number of agents can either delay or accelerate the rate at which these cells undergo death. TNFα can exert opposing, concentration-dependent effects on neutrophils to either accelerate their apoptosis or enhance their survival. We show that TNFα greatly increases the rate of turnover of Mcl-1, an antiapoptotic protein that plays a key role in neutrophil survival. In contrast to Mcl-1 turnover in control- or granulocyte-macrophage colony-stimulating factor (GM-CSF)–treated neutrophils that occurs via the proteasome, TNFα-accelerated Mcl-1 turnover occurs via activation of caspases. Mcl-1–depleted cells thus have accelerated rates of apoptosis. While TNFα had no effect on MCL-1 transcription, it induced expression of another antiapoptotic molecule, BFL-1. Low concentrations of TNFα (≤ 1 ng/mL) stimulated BFL-1 expression, whereas higher concentrations (≥ 10 ng/mL) triggered caspase-dependent acceleration of Mcl-1 turnover. These opposing effects on 2 separate antiapoptotic systems of neutrophils explain the divergent effects of TNFα on neutrophil apoptosis and have important implications for understanding how TNFα may affect immune function in inflammatory diseases.

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