Influence of botulinum C2 toxin on F-actin and N-formyl peptide receptor dynamics in human neutrophils.

Stimulation of human neutrophils with the chemotactic N-formyl peptide causes production of oxygen radicals and conversion of monomeric actin (G-actin) to polymeric actin (F-actin). The effects of the binary botulinum C2 toxin on the amount of F-actin and on neutrophil cell responses were studied. Two different methods for analyzing the actin response were used in formyl peptide-stimulated cells: staining of F-actin with rhodamine-phalloidin and a transient right angle light scatter. Preincubation of neutrophils with 400 ng/ml component I and 1,600 ng/ml component II of botulinum C2 toxin for 30 min almost completely inhibited the formyl peptide-stimulated polymerization of G-actin and at the same time decreased the amount of F-actin in unstimulated neutrophils by an average of approximately 30%. Botulinum C2 toxin preincubation for 60 min destroyed approximately 75% of the F-actin in unstimulated neutrophils. Right angle light scatter analysis showed that control neutrophils exhibited the transient response characteristic of actin polymerization; however, after botulinum C2 toxin treatment, degranulation was detected. Single components of the binary botulinum C2 toxin were without effect on the actin polymerization response. Fluorescence flow cytometry and fluorospectrometric binding studies showed little alteration in N-formyl peptide binding or dissociation dynamics in the toxin-treated cells. However, endocytosis of the fluorescent N-formyl peptide ligand-receptor complex was slower but still possible in degranulating neutrophils treated with botulinum C2 toxin for 60 min. The half-time of endocytosis, estimated from initial rates, was 4 and 8 min in control and botulinum C2 toxin-treated neutrophils, respectively.

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Actin Polymerization, Calcium-Transients, and Phospholipid Metabolism in Human Neutrophils After Stimulation with Interleukin-8 and N-formyl Peptide

Journal of Investigative Dermatology ◽

10.1111/1523-1747.ep12371788 ◽

1994 ◽

Vol 102 (3) ◽

pp. 310-314 ◽

Cited By ~ 33

Author(s):

Johannes Norgauer ◽

Jean Krutmann ◽

Gustav J. Dobos ◽

Alexis E. Traynor-Kaplan ◽

Zenaida G. Oades ◽

...

Keyword(s):

Actin Polymerization ◽

Phospholipid Metabolism ◽

Interleukin 8 ◽

Calcium Transients ◽

Human Neutrophils ◽

Formyl Peptide

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Chemical Composition and Immunomodulatory Activity of Essential Oils from Rhododendron albiflorum

Molecules ◽

10.3390/molecules26123652 ◽

2021 ◽

Vol 26 (12) ◽

pp. 3652

Author(s):

Igor A. Schepetkin ◽

Gulmira Özek ◽

Temel Özek ◽

Liliya N. Kirpotina ◽

Andrei I. Khlebnikov ◽

...

Keyword(s):

Chemical Composition ◽

Essential Oils ◽

Neutrophil Migration ◽

Compositional Analysis ◽

Microglial Cells ◽

Formyl Peptide Receptor ◽

Human Neutrophils ◽

Immunomodulatory Activity ◽

Hl60 Cells ◽

Formyl Peptide

Rhododendron (Ericaceae) extracts contain flavonoids, chromones, terpenoids, steroids, and essential oils and are used in traditional ethnobotanical medicine. However, little is known about the immunomodulatory activity of essential oils isolated from these plants. Thus, we isolated essential oils from the flowers and leaves of R. albiflorum (cascade azalea) and analyzed their chemical composition and innate immunomodulatory activity. Compositional analysis of flower (REOFl) versus leaf (REOLv) essential oils revealed significant differences. REOFl was comprised mainly of monoterpenes (92%), whereas sesquiterpenes were found in relatively low amounts. In contrast, REOLv was primarily composed of sesquiterpenes (90.9%), with a small number of monoterpenes. REOLv and its primary sesquiterpenes (viridiflorol, spathulenol, curzerene, and germacrone) induced intracellular Ca2+ mobilization in human neutrophils, C20 microglial cells, and HL60 cells transfected with N-formyl peptide receptor 1 (FPR1) or FPR2. On the other hand, pretreatment with these essential oils or component compounds inhibited agonist-induced Ca2+ mobilization and chemotaxis in human neutrophils and agonist-induced Ca2+ mobilization in microglial cells and FPR-transfected HL60 cells, indicating that the direct effect of these compounds on [Ca2+]i desensitized the cells to subsequent agonist activation. Reverse pharmacophore mapping suggested several potential kinase targets for these compounds; however, these targets were not supported by kinase binding assays. Our results provide a cellular and molecular basis to explain at least part of the beneficial immunotherapeutic properties of the R. albiflorum essential oils and suggest that essential oils from leaves of this plant may be effective in modulating some innate immune responses, possibly by inhibition of neutrophil migration.

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Salmonella typhimurium attachment to human intestinal epithelial monolayers: transcellular signalling to subepithelial neutrophils.

The Journal of Cell Biology ◽

10.1083/jcb.123.4.895 ◽

1993 ◽

Vol 123 (4) ◽

pp. 895-907 ◽

Cited By ~ 304

Author(s):

B A McCormick ◽

S P Colgan ◽

C Delp-Archer ◽

S I Miller ◽

J L Madara

Keyword(s):

Epithelial Cells ◽

Salmonella Typhimurium ◽

Apical Membrane ◽

Coli Strain ◽

Formyl Peptide Receptor ◽

Human Neutrophils ◽

Classical Pathway ◽

Intestinal Epithelial ◽

Formyl Peptide

In human intestinal disease induced by Salmonella typhimurium, transepithelial migration of neutrophils (PMN) rapidly follows attachment of the bacteria to the epithelial apical membrane. In this report, we model those interactions in vitro, using polarized monolayers of the human intestinal epithelial cell, T84, isolated human PMN, and S. typhimurium. We show that Salmonella attachment to T84 cell apical membranes did not alter monolayer integrity as assessed by transepithelial resistance and measurements of ion transport. However, when human neutrophils were subsequently placed on the basolateral surface of monolayers apically colonized by Salmonella, physiologically directed transepithelial PMN migration ensued. In contrast, attachment of a non-pathogenic Escherichia coli strain to the apical membrane of epithelial cells at comparable densities failed to stimulate a directed PMN transepithelial migration. Use of the n-formyl-peptide receptor antagonist N-t-BOC-1-methionyl-1-leucyl-1- phenylalanine (tBOC-MLP) indicated that the Salmonella-induced PMN transepithelial migration response was not attributable to the classical pathway by which bacteria induce directed migration of PMN. Moreover, the PMN transmigration response required Salmonella adhesion to the epithelial apical membrane and subsequent reciprocal protein synthesis in both bacteria and epithelial cells. Among the events stimulated by this interaction was the epithelial synthesis and polarized release of the potent PMN chemotactic peptide interleukin-8 (IL-8). However, IL-8 neutralization, transfer, and induction experiments indicated that this cytokine was not responsible for the elicited PMN transmigration. These data indicate that a novel transcellular pathway exists in which subepithelial PMN respond to lumenal pathogens across a functionally intact epithelium. Based on the known unique characteristics of the intestinal mucosa, we speculate that IL-8 may act in concert with an as yet unidentified transcellular chemotactic factor(s) (TCF) which directs PMN migration across the intestinal epithelium.

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Primary granule exocytosis in human neutrophils is regulated by Rac-dependent actin remodeling

AJP Cell Physiology ◽

10.1152/ajpcell.00239.2008 ◽

2008 ◽

Vol 295 (5) ◽

pp. C1354-C1365 ◽

Cited By ~ 59

Author(s):

Troy Mitchell ◽

Andrea Lo ◽

Michael R. Logan ◽

Paige Lacy ◽

Gary Eitzen

Keyword(s):

Actin Polymerization ◽

Microscopic Analysis ◽

Formyl Peptide Receptor ◽

Human Neutrophils ◽

Cell Cortex ◽

Secretory Cells ◽

Actin Remodeling ◽

Granule Exocytosis ◽

Primary Granule

The actin cytoskeleton regulates exocytosis in all secretory cells. In neutrophils, Rac2 GTPase has been shown to control primary (azurophilic) granule exocytosis. In this report, we propose that Rac2 is required for actin cytoskeletal remodeling to promote primary granule exocytosis. Treatment of neutrophils with low doses (≤10 μM) of the actin-depolymerizing drugs latrunculin B (Lat B) or cytochalasin B (CB) enhanced both formyl peptide receptor- and Ca2+ionophore-stimulated exocytosis. Higher concentrations of CB or Lat B, or stabilization of F-actin with jasplakinolide (JP), inhibited primary granule exocytosis measured as myeloperoxidase release but did not affect secondary granule exocytosis determined by lactoferrin release. These results suggest an obligatory role for F-actin disassembly before primary granule exocytosis. However, lysates from secretagogue-stimulated neutrophils showed enhanced actin polymerization activity in vitro. Microscopic analysis showed that resting neutrophils contain significant cortical F-actin, which was redistributed to sites of primary granule translocation when stimulated. Exocytosis and actin remodeling was highly polarized when cells were primed with CB; however, polarization was reduced by Lat B preincubation, and both polarization and exocytosis were blocked when F-actin was stabilized with JP. Treatment of cells with the small molecule Rac inhibitor NSC23766 also inhibited actin remodeling and primary granule exocytosis induced by Lat B/fMLF or CB/fMLF, but not by Ca2+ionophore. Therefore, we propose a role for F-actin depolymerization at the cell cortex coupled with Rac-dependent F-actin polymerization in the cell cytoplasm to promote primary granule exocytosis.

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Cloning of a cDNA encoding a receptor related to the formyl peptide receptor of human neutrophils

Gene ◽

10.1016/0378-1119(92)90208-7 ◽

1992 ◽

Vol 118 (2) ◽

pp. 303-304 ◽

Cited By ~ 35

Author(s):

H.Daniel Perez ◽

Richard Holmes ◽

Edward Kelly ◽

John McClary ◽

William H. Andrews

Keyword(s):

Formyl Peptide Receptor ◽

Human Neutrophils ◽

Peptide Receptor ◽

Formyl Peptide

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Calcium mobilization, actin polymerization and right-angle light scatter responses to leukotriene B4, 12(R)- and 12(S)-hydroxyeicosatetraenoic acid in human neutrophils

Life Sciences ◽

10.1016/0024-3205(88)90465-1 ◽

1988 ◽

Vol 42 (6) ◽

pp. 727-733 ◽

Cited By ~ 17

Author(s):

P.H. Naccache ◽

N. Faucher ◽

S. Therrien ◽

P. Borgeat

Keyword(s):

Actin Polymerization ◽

Leukotriene B4 ◽

Calcium Mobilization ◽

Human Neutrophils ◽

Light Scatter ◽

Hydroxyeicosatetraenoic Acid

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Bioactive Secondary Metabolites of a Marine Bacillus sp. Inhibit Superoxide Generation and Elastase Release in Human Neutrophils by Blocking Formyl Peptide Receptor 1

Molecules ◽

10.3390/molecules18066455 ◽

2013 ◽

Vol 18 (6) ◽

pp. 6455-6468 ◽

Cited By ~ 9

Author(s):

Shun-Chin Yang ◽

Chwan-Fwu Lin ◽

Wen-Yi Chang ◽

Jimmy Kuo ◽

Yin-Ting Huang ◽

...

Keyword(s):

Secondary Metabolites ◽

Formyl Peptide Receptor ◽

Human Neutrophils ◽

Bacillus Sp ◽

Superoxide Generation ◽

Peptide Receptor ◽

Bioactive Secondary Metabolites ◽

Formyl Peptide

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N-Formyl peptide receptor subtypes in human neutrophils activate l-plastin phosphorylation through different signal transduction intermediates

Biochemical Journal ◽

10.1042/bj20031114 ◽

2004 ◽

Vol 377 (2) ◽

pp. 469-477 ◽

Cited By ~ 25

Author(s):

Marie-Hélène PACLET ◽

Clare DAVIS ◽

Peter KOTSONIS ◽

Jasminka GODOVAC-ZIMMERMANN ◽

Anthony W. SEGAL ◽

...

Keyword(s):

Signal Transduction ◽

Formyl Peptide Receptor ◽

Actin Binding ◽

Human Neutrophils ◽

Receptor Subtypes ◽

Peptide Receptor ◽

High Affinity ◽

Formyl Peptide ◽

Fmlp Receptor ◽

Phosphoinositide 3 Kinase

We investigated the coupling of the fMLP (N-formyl-l-methionyl-l-leucyl-l-phenylalanine; ‘chemotactic peptide’) receptor with phosphorylation of the actin-binding protein l-plastin in neutrophils. Using two-dimensional IEF (isoelectric focusing)/PAGE and MALDI–TOF (matrix-assisted laser desorption ionization–time-of-flight)-MS, l-plastin was identified as a major phosphoprotein in fMLP-stimulated neutrophils whose phosphorylation was dependent on phosphoinositide 3-kinase, PLD (phospholipase D) and PKC (protein kinase C) activity. Two fMLP receptor subtypes were identified in neutrophils, characterized by a distinct sensitivity to fMLP and antagonistic peptides. Both receptor subtypes induced the phosphorylation of l-plastin. l-plastin phosphorylation induced by low-affinity fMLP receptors involves an action of phosphoinositide 3-kinase, PLD and PKC isotypes. In contrast, none of these intermediates are utilized by high-affinity fMLP receptors in the phosphorylation of l-plastin. However, the PKC inhibitor Ro-31-8220 inhibits l-plastin phosphorylation induced by the high-affinity fMLP receptor. Thus, an as yet unknown Ro-31-8220-sensitive kinase regulates l-plastin phosphorylation in response to the high-affinity fMLP receptor. The results suggest a model in which receptor subtypes induce a similar endpoint event through different signal-transduction intermediates. This may be relevant in the context of cell migration in which one receptor subpopulation may become desensitized in a concentration gradient of chemoattractant.

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