scholarly journals Identification of Constituents of Human Neutrophil Azurophil Granules That Mediate Fungistasis againstHistoplasma capsulatum

2000 ◽  
Vol 68 (10) ◽  
pp. 5668-5672 ◽  
Author(s):  
Simon L. Newman ◽  
Lisa Gootee ◽  
Joelle E. Gabay ◽  
Michael E. Selsted
Keyword(s):  
Human Neutrophil ◽  
Histoplasma Capsulatum ◽  
Cathepsin G ◽  
Human Neutrophils ◽  
Dependent Manner ◽  
Subsequent Growth ◽  
Concentration Dependent Manner ◽  
Fungistatic Activity ◽  
Maximum Inhibition ◽  
Azurophil Granule

ABSTRACT Previously we demonstrated that human neutrophils mediate potent and long-lasting fungistasis against Histoplasma capsulatumyeasts and that all of the fungistatic activity resides in the azurophil granules. In the present study, specific azurophil granule constituents with fungistatic activity were identified by incubation with H. capsulatum yeasts for 24 h and by quantifying the subsequent growth of yeasts via the incorporation of [3H]leucine. Human neutrophil defensins HNP-1, HNP-2, and HNP-3 inhibited the growth of H. capsulatum yeasts in a concentration-dependent manner with maximum inhibition at 8 μg/ml. At a concentration of 4 μg/ml, all possible paired combinations of defensins exhibited additive fungistatic activity against H. capsulatum yeasts. Cathepsin G and bactericidal-permeability-increasing protein (BPI) also mediated fungistasis against H. capsulatum in a concentration-dependent manner. The fungistatic activities of combinations of cathepsin G and BPI were additive, as were those of combinations of cathepsin G or BPI with HNP-1, HNP-2, and HNP-3. Lysozyme and elastase exhibited modest antifungal activity, and azurocidin and proteinase 3 exhibited no significant fungistasis against H. capsulatum yeasts. Thus, defensins, cathepsin G, and BPI are the major anti-H. capsulatum effector molecules in the azurophil granules of human neutrophils.

Neutrophil Elastase Potentiates Cathepsin G-lnduced Platelet Activation

1992 ◽  
Vol 68 (05) ◽  
pp. 570-576 ◽  
Author(s):  
Mary A Selak
Keyword(s):  
Neutrophil Elastase ◽  
Cell Activation ◽  
Thromboxane A2 ◽  
Creatine Phosphate ◽  
Dense Granule ◽  
Cathepsin G ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Low Concentrations ◽  
High Concentrations

SummaryWe have previously demonstrated that human neutrophil cathepsin G is a strong platelet agonist that binds to a specific receptor. This work describes the effect of neutrophil elastase on cathepsin G-induced platelet responses. While platelets were not activated by high concentrations of neutrophil elastase by itself, elastase enhanced aggregation, secretion and calcium mobilization induced by low concentrations of cathepsin G. Platelet aggregation and secretion were potentiated in a concentration-dependent manner by neutrophil elastase with maximal responses observable at 200 nM. Enhancement was observed when elastase was preincubated with platelets for time intervals of 10–60 s prior to addition of a low concentration of cathepsin G and required catalytically-active elastase since phenylmethanesulphonyl fluoride-inhibited enzyme failed to potentiate cell activation. Neutrophil elastase potentiation of platelet responses induced by low concentrations of cathepsin G was markedly inhibited by creatine phosphate/creatine phosphokinase and/or indomethacin, indicating that the synergism between elastase and cathepsin G required the participation of ADP and thromboxane A2. On the other hand, platelet responses were not attenuated by the PAF antagonist BN 52021, signifying that PAF-acether did not play a role in elastase potentiation. At higher concentrations porcine pancreatic elastase exhibits similar effects to neutrophil elastase, demonstrating that the effect of elastase was not unique to the neutrophil protease. While neutrophil elastase failed to alter the ability of cathepsin G to hydrolyze a synthetic chromogenic substrate, preincubation of platelets with elastase increased the apparent affinity of cathepsin G binding to platelets. In contrast to their effect on cathepsin G-induced platelet responses, neither neutrophil nor pancreatic elasatse potentiated aggregation or dense granule release initiated by ADP, PAF-acether, arachidonic acid or U46619, a thromboxane A2 mimetic. Moreover, unlike its effect on cathepsin G, neutrophil elastase inhibited thrombin-induced responses. The current observations demonstrate that elastase can potentiate platelet responses mediated by low concentrations of cathepsin G, suggesting that both enzymes may function synergistically to activate platelets under conditions where neutrophil degranulation occurs.

scholarly journals Inter-α inhibitor proteins maintain neutrophils in a resting state by regulating shape and reducing ROS production

2018 ◽  
Vol 2 (15) ◽  
pp. 1923-1934 ◽  
Author(s):  
Soe Soe Htwe ◽  
Hidenori Wake ◽  
Keyue Liu ◽  
Kiyoshi Teshigawara ◽  
Barbara S. Stonestreet ◽  
...  
Keyword(s):  
Resting State ◽  
Smooth Surface ◽  
Spherical Shape ◽  
Human Neutrophils ◽  
Dependent Manner ◽  
Ros Production ◽  
Concentration Dependent Manner ◽  
Key Points

Key Points IAIP, but not bikunin, maintains spherical shape, small size, and smooth surface of human neutrophils and supports capillary passage. IAIP reduced ROS production from neutrophils in a concentration-dependent manner probably through the p47phox phosphorylation on Ser328.

scholarly journals Modulatory Effect of Chelidonium majus Extract and Its Alkaloids on LPS-Stimulated Cytokine Secretion in Human Neutrophils

Molecules ◽  
2020 ◽  
Vol 25 (4) ◽  
pp. 842 ◽  
Author(s):  
Sylwia Zielińska ◽  
Monika Ewa Czerwińska ◽  
Magdalena Dziągwa-Becker ◽  
Andrzej Dryś ◽  
Mariusz Kucharski ◽  
...  
Keyword(s):  
Biological Activities ◽  
Cytokine Secretion ◽  
Human Neutrophils ◽  
Root Extract ◽  
Dependent Manner ◽  
Chelidonium Majus ◽  
Concentration Dependent Manner ◽  
Cytotoxic Effects ◽  
Tnf Α ◽  
High Cytotoxicity

Due to certain differences in terms of molecular structure, isoquinoline alkaloids from Chelidonium majus engage in various biological activities. Apart from their well-documented antimicrobial potential, some phenanthridine and protoberberine derivatives as well as C. majus extract present with anti-inflammatory and cytotoxic effects. In this study, the LC–MS/MS method was used to determine alkaloids, phenolic acids, carboxylic acids, and hydroxybenzoic acids. We investigated five individually tested alkaloids (coptisine, berberine, chelidonine, chelerythrine, and sanguinarine) as well as C. majus root extract for their effect on the secretion of IL-1β, IL-8, and TNF-α in human polymorphonuclear leukocytes (neutrophils). Berberine, chelidonine, and chelerythrine significantly decreased the secretion of TNF-α in a concentration-dependent manner. Sanguinarine was found to be the most potent inhibitor of IL-1β secretion. However, the overproduction of IL-8 and TNF-α and a high cytotoxicity for these compounds were observed. Coptisine was highly cytotoxic and slightly decreased the secretion of the studied cytokines. The extract (1.25–12.5 μg/mL) increased cytokine secretion in a concentration-dependent manner, but an increase in cytotoxicity was also noted. The alkaloids were active at very low concentrations (0.625–2.5 μM), but their potential cytotoxic effects, except for chelidonine and chelerythrine, should not be ignored.

Functional properties of noradrenergic nervous system in rat colonic mucosa: uptake of [3H]norepinephrine

1981 ◽  
Vol 241 (2) ◽  
pp. G137-G142
Author(s):  
Z. C. Wu ◽  
T. S. Gaginella
Keyword(s):  
Colonic Mucosa ◽  
Close Association ◽  
Physiological Role ◽  
Dependent Manner ◽  
Noradrenergic Neurons ◽  
Concentration Dependent Manner ◽  
Colonic Epithelial Cells ◽  
High Affinity ◽  
Maximum Inhibition ◽  
6 Hydroxydopamine

The accumulation of exogenous [3H]norepinephrine ([3H]NE) by rat colonic mucosa was studied. Uptake was linear for 10 min and reached a maximum after 90 min. The process was concentration dependent and saturable, having a Km of 1.67 X 10(-6) M and Vmax of 0.57 nmol.g-1.min-1. The inhibitor of specific norepinephrine uptake, desmethylimipramine (DMI), inhibited uptake in a concentration-dependent manner; the maximum inhibition was 81% at 10 microM. Normetanephrine also inhibited uptake at 100 microM. Reserpine, at concentrations ranging from 10(-7) to 10(-5) M, prevented the accumulation of [3H]NE, with maximum inhibition being 47% of control. Accumulation by mucosa obtained from rats sympathectomized with 6-hydroxydopamine was only 33% of control; DMI did not further reduce this uptake. Colonic epithelial cells were isolated and were found to also accumulate [3H]NE, but this accumulation was not affected by DMI. It is concluded that rat colonic mucosa contains noradrenergic neurons capable of accumulating exogenously administered norepinephrine by a specific and high-affinity process. The presence of a functional noradrenergic neural network in close association with the epithelium suggests that this system may play a physiological role in modulating colonic mucosal function.

scholarly journals Cathepsin G is a strong platelet agonist released by neutrophils

1988 ◽  
Vol 251 (1) ◽  
pp. 293-299 ◽  
Author(s):  
M A Selak ◽  
M Chignard ◽  
J B Smith
Keyword(s):  
Biological Activity ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Enzymic Activity ◽  
Calcium Mobilization ◽  
Cathepsin G ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Alpha 1 Antitrypsin ◽  
Acid Gel

The present studies were undertaken to characterize a serine protease released by N-formyl-L-Met-L-Leu-L-Phe (fMet-Leu-Phe)-stimulated neutrophils that rapidly induces platelet calcium mobilization, secretion and aggregation. The biological activity associated with this protease was unaffected by leupeptin, was only weakly diminished by N-p-tosyl-L-Lys-chloromethane, but was strongly inhibited by alpha 1-antitrypsin, soyabean trypsin inhibitor, N-tosyl-L-Phe-chloromethane and benzoyloxycarbonyl-Gly-Leu-Phe-chloromethane (Z-Gly-Leu-PheCH2Cl). These observations indicated that the biological activity of neutrophil supernatants could be attributed to a chymotrypsin-like enzyme such as cathepsin G. Furthermore, platelet aggregation and 5-hydroxytryptamine release induced by cell-free supernatants from fMet-Leu-Phe-stimulated neutrophils were found to be blocked by antiserum to cathepsin G in a concentration-dependent manner but were unaffected by antiserum to elastase. The biological activity present in neutrophil supernatants co-purified with enzymic activity for cathepsin G during sequential Aprotinin-Sepharose affinity chromatography and carboxymethyl-Sephadex chromatography. SDS/polyacrylamide-gel electrophoresis of the reduced, purified protein, demonstrated three polypeptides with apparent Mr values of 31,500, 29,000 and 28,000 and four polypeptides were resolved on acid-gel electrophoresis. Purified cathepsin G from neutrophils cross-reacted with anti-(cathepsin G) serum in a double immunodiffusion assay and elicited platelet calcium mobilization, 5-hydroxytryptamine secretion and aggregation. Calcium mobilization and secretion induced by low concentrations of cathepsin G were partially dependent on arachidonic acid metabolites and ADP, while stimulation by higher enzyme concentrations was independent of amplification pathways, indicating that cathepsin G is a strong platelet agonist. These results suggest that pathological processes which stimulate neutrophils and release cathepsin G can in turn result in the recruitment and activation of platelets.

Fungistatic Activity of Human Neutrophils against Histoplasma capsulatum: Correlation with Phagocytosis

1991 ◽  
Vol 164 (1) ◽  
pp. 158-162 ◽  
Author(s):  
E. Brummer ◽  
N. Kurita ◽  
S. Yosihida ◽  
K. Nishimura ◽  
M. Miyaji
Keyword(s):  
Histoplasma Capsulatum ◽  
Human Neutrophils ◽  
Fungistatic Activity

Inhibition of ADAMTS13 Activity By Human Neutrophil Peptide 1 (HNP-1): Potential Link Between Inflammation, Onset of Thrombotic Thrombocytopenic Purpura, and Other Thrombosis

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 599-599 ◽  
Author(s):  
Jialing Bao ◽  
Khalil Bdeir ◽  
Don L. Siegel ◽  
Douglas B. Cines ◽  
X. Long Zheng
Keyword(s):  
Thrombotic Thrombocytopenic Purpura ◽  
Human Neutrophil ◽  
Conflicts Of Interest ◽  
Human Monoclonal Antibody ◽  
Amino Acid Sequences ◽  
Dependent Manner ◽  
Adamts13 Activity ◽  
Single Chain ◽  
Concentration Dependent Manner ◽  
Thrombocytopenic Purpura

Abstract Thrombotic thrombocytopenic purpura (TTP) is a potentially fatal syndrome associated with severe deficiency of plasma ADAMTS13 activity resulting from either mutations or autoantibodies. However, patients with severe ADAMTS13 deficiency do not always develop TTP. Rather, a trigger, such as infection or inflammation, often precedes the onset of the TTP syndrome. We hypothesized that antimicrobial human neutrophil peptides 1-3 (HNPα1-3) or α-defensins, the most abundant proteins in the granules of neutrophils, which are released at site of inflammation, activate platelets, and inhibit fibrinolysis, might help to initiate TTP. This question arose because we noted that the amino acid sequences of HNPα1-3, which are nearly identical except for one residue at the N-terminus, all contain a motif (RRY) similar to exosite 3 (659RRYGEEY665) in the spacer domain of ADAMTS13 (Fig. 1) that was shown to be critical for recognition of von Willebrand factor (VWF). Here, we found that both purified and synthetic HNPα1 bind to FRETS-VWF73 and plasma-derived VWF and inhibit proteolytic cleavage of these substrates in a concentration-dependent manner. At the final concentrations of 10 micro mol/L and 150 micro mol/L, HNPα1 completely abolished the cleavage of FRETS-VWF73 (IC50=3.5 micro mol/L) (Fig. 2) and VWF (IC50=75 micro mol/L) (not shown), respectively. Such concentrations are readily attained locally after systemic infection. Deletion or alanine substitution within the RRY motif of HNPα1 completely abolished its ability to inhibit ADAMTS13 activity assessed by FRETS-VWF73 and VWF multimer analysis. This suggests that an interaction of the RRY motif in HNPα1 with the central A2 domain of VWF is required to mediate its inhibition. In support of this hypothesis, HNPα1 interacts with a human monoclonal antibody against ADAMTS13 scFv (the single chain fragment of variable region) designated 4-20, but not scFv3-1, both isolated by phage display from patients with acquired autoimmune TTP. Hydrogen-deuterium exchange mass spectrometry has shown that the binding site for scFv4-20, but not scFv3-1, contains the RRY sequence. These results suggest that HNPα1-3 released from neutrophils following infection or inflammation may inhibit residual plasma ADAMTS13 activity in vivo similar to anti-ADAMTS13 autoantibodies by interfering with its interaction with VWF, thereby triggering the onset of hereditary and acquired autoimmune TTP. Our findings suggest a potential novel link between systemic inflammation and the pathogenesis of TTP and possibility other thrombotic sequelae. Figure 2 Figure 2. Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.

scholarly journals Responsiveness of human neutrophils to interleukin-4: induction of cytoskeletal rearrangements, de novo protein synthesis and delay of apoptosis

1997 ◽  
Vol 325 (1) ◽  
pp. 147-153 ◽  
Author(s):  
Denis GIRARD ◽  
Robert PAQUIN ◽  
André D. BEAULIEUL
Keyword(s):  
Protein Synthesis ◽  
De Novo ◽  
Biological Activities ◽  
Interleukin 4 ◽  
Human Neutrophils ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Cytoskeletal Rearrangements ◽  
First Time ◽  
De Novo Protein Synthesis

Interleukin-4 (IL-4) and IL-13 are cytokines that share many biological activities. We have previously demonstrated that IL-13 affects a number of neutrophil responses, and here we extend our observations to IL-4. We present, for the first time, direct evidence for the presence of functional IL-4 receptors on human neutrophils. We report that IL-4 induces RNA synthesis in a concentration-dependent manner and, based on observations of the induction of morphological cell shape changes and spreading onto glass, we demonstrate that IL-4 activates neutrophil cytoskeletal rearrangements. We further show that IL-4 is a potent activator of de novo protein synthesis in neutrophils, and we identify by microsequencing one of these proteins as the cytoskeletal protein actin. We were also able to demonstrate for the first time that actin is cleaved into at least two fragments of ∼ 30 kDa (pI 5.4) and ∼ 25 kDa (pI 5.0) in neutrophils. Finally, we report that IL-4 delays neutrophil apoptosis, as assessed by morphological observations from cytocentrifuge preparations, as well as by measurement of differences in staining by flow cytometry with both propidium iodide and Hoechst reagent. Taken together, we conclude that IL-4 is a more potent neutrophil agonist than previously believed. We discuss the possibility that the induction of the de novo synthesis of actin by IL-4 is related to the mechanism by which this cytokine delays apoptosis; in addition, the cleavage of this protein is likely to contribute to the apoptotic process.

scholarly journals Neutrophil Extracellular Traps Activate Proinflammatory Functions of Human Neutrophils

2021 ◽  
Vol 12 ◽  
Author(s):  
Daniel Dömer ◽  
Tabea Walther ◽  
Sonja Möller ◽  
Martina Behnen ◽  
Tamás Laskay
Keyword(s):  
B Cell ◽  
Adaptive Immune System ◽  
Neutrophil Extracellular Traps ◽  
Microbial Pathogens ◽  
Human Neutrophils ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Extracellular Traps ◽  
Proinflammatory Role

Neutrophil extracellular traps (NETs) consist of decondensed nuclear chromatin that is associated with proteins and are released by neutrophils during an inflammatory response. Released NETs are able to capture pathogens, prevent their dissemination and potentially kill them via antimicrobial peptides and proteins that are associated with the decondensed chromatin. In addition to their antimicrobial functions, NETs have also been shown to exert immunomodulatory effects by activation and differentiation of macrophages, dendritic cells and T cells. However, the effect of NETs on neutrophil functions is poorly understood. Here we report the first comprehensive study regarding the effects of NETs on human primary neutrophils in vitro. NETs were isolated from cultures of PMA-exposed neutrophils. Exposure of neutrophils to isolated NETs resulted in the activation of several neutrophil functions in a concentration-dependent manner. NETs induced exocytosis of granules, the production of reactive oxygen species (ROS) by the NADPH oxidase NOX2, NOX2-dependent NET formation, increased the phagocytosis and killing of microbial pathogens. Furthermore, NETs induced the secretion of the proinflammatory chemokine IL-8 and the B-cell-activating cytokine BAFF. We could show that the NET-induced activation of neutrophils occurs by pathways that involve the phosphorylation of Akt, ERK1/2 and p38. Taken together our results provide further insights into the proinflammatory role of NETs by activating neutrophil effector function and further supports the view that NETs can amplify inflammatory events. On the one hand the amplified functions enhance the antimicrobial defense. On the other hand, NET-amplified neutrophil functions can be involved in the pathophysiology of NET-associated diseases. In addition, NETs can connect the innate and adaptive immune system by inducing the secretion of the B-cell-activating cytokine BAFF.

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