Differential effects of tyrosine kinase inhibitors and an inhibitor of the mitogen-activated protein kinase cascade on degranulation and superoxide production of human neutrophil granulocytes

Mócsai, Attila; Bánfi, Botond; Kapùs, András; Farkas, Gyöngyi; Geiszt, Miklós; Buday, László; Farago, Anna F.; Ligeti, Erzsébet

doi:10.1016/s0006-2952(97)00245-1

Cited by 45 publications

(25 citation statements)

References 33 publications

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“…An additional factor is that azurophil degranulation requires relatively high concentrations of agonist, and is highly dependent on Ca 2ϩ signaling pathways. Our findings are in accord with the report that inhibition of the MAPK pathway does not block superoxide generation and azurophil granule release (86).…”

Section: Discussionsupporting

confidence: 93%

L-Selectin Signaling of Neutrophil Adhesion and Degranulation Involves p38 Mitogen-activated Protein Kinase

Smolen¹,

Petersen²,

Koch³

et al. 2000

Journal of Biological Chemistry

135

104

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The adhesion molecules known as selectins mediate the capture of neutrophils from the bloodstream. We have previously reported that ligation and cross-linking of L-selectin on the neutrophil surface enhances the adhesive function of ␤ 2 -integrins in a synergistic manner with chemotactic agonists. In this work, we examined degranulation and adhesion of neutrophils in response to cross-linking of L-selectin and addition of interleukin-8. Cross-linking of L-selectin induced priming of degranulation that was similar to that observed with the alkaloid cytochalasin B. Activation mediated by L-selectin of neutrophil shape change and adhesion through CD11b/CD18 were strongly blocked by Merck C, an imidazole-based inhibitor of p38 mitogen-activated protein kinase (MAPK), but not by a structurally similar non-binding regioisomer. Priming by L-selectin of the release of secondary, tertiary, and secretory, but not primary, granules was blocked by inhibition of p38 MAPK. Peak phosphorylation of p38 MAPK was observed within 1 min of cross-linking L-selectin, whereas phosphorylation of ERK1/2 was highest at 10 min. Phosphorylation of p38 MAPK, but not ERK1/2, was inhibited by Merck C. These data suggest that signal transduction as a result of clustering L-selectin utilizes p38 MAPK to effect neutrophil shape change, integrin activation, and the release of secondary, tertiary, and secretory granules.Neutrophils circulate in the vasculature in a passive state and become more adhesive upon stimulation at sites of inflammation. Margination to the vessel wall and subsequent transmigration and phagocytosis (1) requires a number of surface proteins, including the ␤ 2 -integrins and the selectins, as mediators of adherence to the endothelium (2-5). A sequence of molecular and biophysical events has been identified that facilitates neutrophil activation and increased adherence during the acute inflammatory response in vivo. Neutrophils entering post-capillary venules adjacent to inflammatory foci develop transient rolling adhesive interactions with endothelium via selectins (6). Following exposure to inflammatory cytokines such as tumor necrosis factor and interleukin-1, endothelial cells are induced to express E-selectin and P-selectin (6). Several surface glycoproteins on neutrophils, including L-selectin and P-selectin glycoprotein ligand 1, present oligosaccharide moieties that serve as counter receptors for E-selectin and P-selectin. In conjunction with neutrophil membrane L-selectin, which recognizes oligosaccharides on endothelial cells,

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Section: Discussionsupporting

confidence: 93%

L-Selectin Signaling of Neutrophil Adhesion and Degranulation Involves p38 Mitogen-activated Protein Kinase

Smolen¹,

Petersen²,

Koch³

et al. 2000

Journal of Biological Chemistry

135

104

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“…HL-60 and PLB-985 cells were differentiated for 5 days with 1.2% Me 2 SO and 0.5% dimethylformamide, respectively. Human neutrophils were purified as described previously (40). For transfection, the coding regions of human NOX5␤ cDNA and mouse NOX4 cDNA (both with an inserted Kozak sequence) were subcloned into pcDNA3.1 (Invitrogen, Groningen, Netherlands).…”

Section: Methodsmentioning

confidence: 99%

A Ca2+-activated NADPH Oxidase in Testis, Spleen, and Lymph Nodes

Bánfi¹,

Molnár²,

Maturana³

et al. 2001

Journal of Biological Chemistry

Self Cite

539

528

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Superoxide and its derivatives are increasingly implicated in the regulation of physiological functions from oxygen sensing and blood pressure regulation to lymphocyte activation and sperm-oocyte fusion. Here we describe a novel superoxide-generating NADPH oxidase referred to as NADPH oxidase 5 (NOX5). NOX5 is distantly related to the gp91 phox subunit of the phagocyte NADPH oxidase with conserved regions crucial for the electron transport (NADPH, FAD and heme binding sites). However, NOX5 has a unique N-terminal extension that contains three EF hand motifs. The mRNA of NOX5 is expressed in pachytene spermatocytes of testis and in B-and T-lymphocyte-rich areas of spleen and lymph nodes. When heterologously expressed, NOX5 was quiescent in unstimulated cells. However, in response to elevations of the cytosolic Ca 2؉ concentration it generated large amounts of superoxide. Upon Ca 2؉ activation, NOX5 also displayed a second function: it became a proton channel, presumably to compensate charge and pH alterations due to electron export. In summary, we have identified a novel NADPH oxidase that generates superoxide and functions as a H ؉ channel in a Ca 2؉ -dependent manner. NOX5 is likely to be involved in Ca 2؉ -activated, redox-dependent processes of spermatozoa and lymphocytes such as sperm-oocyte fusion, cell proliferation, and cytokine secretion.

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“…The GST-Syk-(SH2) 2 fusion protein was coupled to glutathione-Sepharose beads (GE Healthcare) and the result was incubated with cell lysates prepared with Triton X-100 (ref. 36) or radioimmunoprecipitation assay buffer (20 mM Tris, pH 7.5, 150 mM NaCl, 1% (volume/volume) Triton X-100 buffer, 1% (weight/volume) sodium deoxycholate, 0.1% (weight/volume) SDS, 1 mM Na 3 VO 4 , 50 mM NaF, 2 mM EDTA, 1 mM Pefabloc, 10 µg/ml of leupeptin, 2 µg/ml of aprotinin, 1 mM dithiothreitol, 1 µg/ml of pepstatin and 1 mM di-isopropyl fluorophosphate). Syk kinase autophosphorylation assays were done on anti-FcRγ immunoprecipitates 'captured' with protein A/G PLUSAgarose beads (Santa Cruz Biotechnology) from Nonidet-P40 lysates (20 mM HEPES, pH 7.4, 150 mM NaCl and 1% (volume/volume) Nonidet-P40, plus the inhibitors listed above).…”

Section: Immunoblot and Immunoprecipitationmentioning

confidence: 99%

Integrin signaling in neutrophils and macrophages uses adaptors containing immunoreceptor tyrosine-based activation motifs

et al. 2006

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At sites of inflammation, ligation of leukocyte integrins is critical for the activation of cellular effector functions required for host defense. However, the signaling pathways linking integrin ligation to cellular responses are poorly understood. Here we show that integrin signaling in neutrophils and macrophages requires adaptors containing immunoreceptor tyrosine-based activation motifs (ITAMs). Neutrophils and macrophages lacking two ITAM-containing adaptor proteins, DAP12 and FcRγ, were defective in integrin-mediated responses. Activation of the tyrosine kinase Syk by integrins required that DAP12 and FcRγ were first phosphorylated by Src family kinases. Retroviral transduction of neutrophils and macrophages with wild-type and mutant Syk or DAP12 demonstrated that the Src homology 2 domains of Syk and the ITAM of DAP12 were required for integrin signaling. Our data show that integrin signaling for the activation of cellular responses in neutrophils and macrophages proceeds by an immunoreceptor-like mechanism.Integrins are transmembrane adhesion receptors that coordinate cellular responses with the extracellular environment. Integrin function is especially important in neutrophils and macrophages, key effector cells that kill or suppress invading microorganisms during the innate immune response. In neutrophils and macrophages, integrin signaling is critical for cellular functions such as firm adhesion, cell spreading, chemotaxis, the production of COMPETING INTERESTS STATEMENTThe authors declare that they have no competing financial interests. Although several molecules required for relaying signals 'downstream' of leukocyte integrins (often called 'outside-in' signaling) have been identified, the initial steps of β 2 integrin signaling remain poorly understood. Src family kinases are involved in an early step of integrin signaling in neutrophils 6 and macrophages 7,8 . Also, the Syk tyrosine kinase is essential for integrin signaling in neutrophils 9 , macrophages 10 and platelets 11 . As Syk is probably involved in a receptor-proximal event during integrin signal transduction, the mechanism of activation of Syk by integrins and its relationship to Src family kinases may be the key to understanding the initiation of integrin signaling. Unfortunately, despite attempts to clarify that issue, the mechanism of activation of Syk by integrins remains poorly understood. HHS Public AccessSyk and the related kinase Zap70 are also essential for signaling downstream of immunoreceptors, such as B cell and T cell receptors and Fc receptors. In contrast to integrin signal transduction, the mechanism of Syk activation initiated by ligation of these immunoreceptors is well characterized. Engagement of immunoreceptors leads to Src family kinase-mediated phosphorylation of immunoreceptor tyrosine-based activation motifs (ITAMs) on receptor-associated transmembrane adaptor proteins 12 . Those adaptors provide docking sites for the tandem Src homology 2 (SH2) domains of the Syk or Zap70 tyrosine kinases, which leads...

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Differential effects of tyrosine kinase inhibitors and an inhibitor of the mitogen-activated protein kinase cascade on degranulation and superoxide production of human neutrophil granulocytes

Cited by 45 publications

References 33 publications

L-Selectin Signaling of Neutrophil Adhesion and Degranulation Involves p38 Mitogen-activated Protein Kinase

L-Selectin Signaling of Neutrophil Adhesion and Degranulation Involves p38 Mitogen-activated Protein Kinase

A Ca2+-activated NADPH Oxidase in Testis, Spleen, and Lymph Nodes

Integrin signaling in neutrophils and macrophages uses adaptors containing immunoreceptor tyrosine-based activation motifs

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