Activation of the early chemiluminescence response of human neutrophils by combined treatment with tumor necrosis factor (TNF-?) and calcium ionophore A23187

1991 ◽  
Vol 111 (2) ◽  
pp. 171-173
Author(s):  
I. E. Shepetkin ◽  
E. V. Borunov ◽  
I. L. Skutina ◽  
S. A. Naumov
Keyword(s):  
Tumor Necrosis Factor ◽  
Tumor Necrosis ◽  
Combined Treatment ◽  
Calcium Ionophore ◽  
Calcium Ionophore A23187 ◽  
Human Neutrophils ◽  
Ionophore A23187 ◽  
Necrosis Factor

scholarly journals Shedding of tumor necrosis factor receptors by activated human neutrophils.

1990 ◽  
Vol 172 (2) ◽  
pp. 599-607 ◽  
Author(s):  
F Porteu ◽  
C Nathan
Keyword(s):  
Tumor Necrosis Factor ◽  
Cell Surface ◽  
Tumor Necrosis ◽  
Calcium Ionophore ◽  
Human Neutrophils ◽  
Scatchard Analysis ◽  
Ionophore A23187 ◽  
Independent Evidence ◽  
Mouse Macrophages ◽  
Necrosis Factor

The capacity of human neutrophils (PMN) to bind tumor necrosis factor (TNF) was rapidly lost when the cells were incubated in suspension with agents that can stimulate their migratory and secretory responses. Both physiological (poly)peptides (FMLP, C5a, CSF-GM) and pharmacologic agonists (PMN, calcium ionophore A23187) induced the loss of TNF receptors (TNF-R) from the cell surface. Half-maximal loss in TNF-R ensued after only approximately 2 min with 10(-7) M FMLP at 37 degrees C, and required only 10(-9) M FMLP during a 30-min exposure. However, there were no such changes even with prolonged exposure of PMN to FMLP at 4 degrees or 16 degrees C. Scatchard analysis revealed loss of TNF-binding sites without change in their affinity (Kd approximately 0.4 nM) as measured at incompletely modulating concentrations of FMLP, C5a, PMA, or A23187. The binding of anti-TNF-R mAbs to PMN decreased in parallel, providing independent evidence for the loss of TNF-R from the cell surface. At the same time, soluble TNF-R appeared in the medium of stimulated PMN. This inference was based on the PMN- and FMLP-dependent generation of a nonsedimentable activity that could inhibit the binding of TNF to fresh human PMN or to mouse macrophages, and the ability of mAbs specific for human TNF-R to abolish inhibition by PMN-conditioned medium of binding of TNF to mouse macrophages. Soluble TNF-R activity was associated with a protein of Mr approximately 28,000 by ligand blot analysis of cell-free supernatants of FMLP-treated PMN. Thus, some portion of the FMLP-induced loss of TNF-R from human PMN is due to shedding of TNF-R. Shedding was unaffected by inhibitors of serine and thiol proteases and could not be induced with phosphatidylinositol-specific phospholipase C. Loss of TNF-R from PMN first stimulated by other agents may decrease their responsiveness to TNF. TNF-R shed by PMN may be one source of the TNF-binding proteins found in body fluids, and may blunt the actions of the cytokine on other cells.

scholarly journals Tumor necrosis factor-alpha and FMLP receptors are functionally linked during FMLP-stimulated activation of adherent human neutrophils

Blood ◽  
1996 ◽  
Vol 88 (2) ◽  
pp. 690-696 ◽  
Author(s):  
KJ Balazovich ◽  
SJ Suchard ◽  
DG Remick ◽  
LA Boxer
Keyword(s):  
Tumor Necrosis Factor ◽  
Tumor Necrosis Factor Alpha ◽  
Tumor Necrosis ◽  
Actinomycin D ◽  
Tnf Alpha ◽  
Human Neutrophils ◽  
Necrosis Factor Alpha ◽  
Alpha Receptors ◽  
Factor Alpha ◽  
Necrosis Factor

Human peripheral blood neutrophils (PMN) plated onto fibrinogen and activated with FMLP release H2O2 and lactoferrin, a specific granule component, with parallel kinetics. Although tumor necrosis factor-alpha (TNF alpha) only primes PMN in suspension, it is a potent agonist of adherent PMN. Activation of adherent PMN by FMLP (10(-7) mol/L) stimulated detectable release of TNF alpha within 45 minutes of stimulation, with maximal release (45.5 pg/10(6) cells) detected by 90 minutes. TNF alpha release paralleled the release of both lactoferrin and H2O2. To determine if TNF alpha plays a role in H2O2 and lactoferrin release, we investigated the effect of anti-TNF alpha antibodies on FMLP-stimulated activation of adherent PMN. A neutralizing rabbit anti-TNF alpha antibody inhibited both H2O2 and lactoferrin release stimulated by FMLP, whereas rabbit lgG, anti-HLA- A,B,C, anti-CD 14, and anti-interleukin-8 antibodies were without effect. The simultaneous addition of TNF alpha (1,000 U/mL) with anti- TNF alpha antibody reversed the inhibition seen with anti-TNF alpha alone. Furthermore, treatment of PMN with either actinomycin D or cylcoheximide resulted in partial (33%) inhibition of H2O2 and lactoferrin release, suggesting that protein synthesis is required for FMLP-mediated activation of adherent PMN. The addition of TNF alpha to either cycloheximide or of actinomycin D-treated PMN overcame the inhibition, indicating that the effect was specific for TNF alpha. The addition of antibodies against either the 55-or 75-kD TNF alpha receptors (referred to as p55 and p75, respectively) resulted in partial (32%) inhibition of FMLP-mediated activation of H2O2 and lactoferrin release, whereas a combination of both antibodies reduced their release to control levels. These data indicate that both p55 and p75 are involved in FMLP activation of adherent PMN. Taken together, these findings indicate that the production of TNF alpha and ligation of TNF alpha receptors are central to FMLP activation of PMN adherent to fibrinogen.

scholarly journals Nitric oxide and peroxynitrite trigger and enhance release of neutrophil extracellular traps

2019 ◽  
Vol 77 (15) ◽  
pp. 3059-3075 ◽  
Author(s):  
Aneta Manda-Handzlik ◽  
Weronika Bystrzycka ◽  
Adrianna Cieloch ◽  
Eliza Glodkowska-Mrowka ◽  
Ewa Jankowska-Steifer ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Nitrosative Stress ◽  
Calcium Ionophore ◽  
Neutrophil Extracellular Traps ◽  
Calcium Ionophore A23187 ◽  
Human Neutrophils ◽  
Ionophore A23187 ◽  
Inflammatory Reactions ◽  
Extracellular Traps

Abstract Despite great interest, the mechanism of neutrophil extracellular traps (NETs) release is not fully understood and some aspects of this process, e.g. the role of reactive nitrogen species (RNS), still remain unclear. Therefore, our aim was to investigate the mechanisms underlying RNS-induced formation of NETs and contribution of RNS to NETs release triggered by various physiological and synthetic stimuli. The involvement of RNS in NETs formation was studied in primary human neutrophils and differentiated human promyelocytic leukemia cells (HL-60 cells). RNS (peroxynitrite and nitric oxide) efficiently induced NETs release and potentiated NETs-inducing properties of platelet activating factor and lipopolysaccharide. RNS-induced NETs formation was independent of autophagy and histone citrullination, but dependent on the activity of phosphoinositide 3-kinases (PI3K) and myeloperoxidase, as well as selective degradation of histones H2A and H2B by neutrophil elastase. Additionally, NADPH oxidase activity was required to release NETs upon stimulation with NO, as shown in NADPH-deficient neutrophils isolated from patients with chronic granulomatous disease. The role of RNS was further supported by increased RNS synthesis upon stimulation of NETs release with phorbol 12-myristate 13-acetate and calcium ionophore A23187. Scavenging or inhibition of RNS formation diminished NETs release triggered by these stimuli while scavenging of peroxynitrite inhibited NO-induced NETs formation. Our data suggest that RNS may act as mediators and inducers of NETs release. These processes are PI3K-dependent and ROS-dependent. Since inflammatory reactions are often accompanied by nitrosative stress and NETs formation, our studies shed a new light on possible mechanisms engaged in various immune-mediated conditions.

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