共查询到20条相似文献,搜索用时 15 毫秒
1.
R J Smith F F Sun B J Bowman S S Iden H W Smith J C McGuire 《Biochemical and biophysical research communications》1982,109(3):943-949
A23187, a calcium ionophore, stimulated a time-dependent generation of 5(S), 12(R)-dihydroxy-6,8,10,14-eicosatetraenoic acid (leukotriene B4), production of superoxide anion (O2?) and release of granule-associated β-glucuronidase and lysozyme by human neutrophils. Leukotriene B4 also elicited the selective release of granule enzymes from cytochalasin B-treated neutrophils. U-60,257, a recently identified inhibitor of leukotriene (LT) C4 and D4 synthesis, caused a dose-related (1–10 μM) suppression of LTB4 production by A23187-activated neutrophils. Degranulation and O2? generation by neutrophils exposed to A23187 and the chemotactic oligopeptide, N-formyl-methionyl-leucyl-phenylalanine (FMLP), were also inhibited with U-60,257. 相似文献
2.
Catherine Matron Arpita Chakravarti Isabelle Allaeys Patrice E. Poubelle 《Free radical biology & medicine》2010,48(2):207-215
Antimicrobial defense by neutrophils implicates the production of reactive oxygen species. Neutrophil responses can be modulated by agonists such as bacterial peptides and proinflammatory factors that modulate neutrophil activity and survival. We evaluated the production of superoxide anions (O2?) in response to fMLF by normal human neutrophils after 3 days of preincubation with GM-CSF + IL-4 + TNF-α (survival medium). After 3 days of incubation in survival medium, long-lived neutrophils produced up to six times more O2? relative to control neutrophils in response to fMLF and WKYMVM. This augmented response to fMLF was preferentially linked to formyl peptide receptor (FPR), whereas the response to WKYMVM was dependent on formyl peptide receptor-like 1 (FPRL-1). Real-time RT-PCR revealed a diminution of FPR and FPRL-1 expression in neutrophils incubated in survival medium. fMLF-induced overproduction of O2? by long-lived neutrophils was independent of intracellular calcium mobilization. The protein tyrosine phosphorylation profile of long-lived neutrophils was modified with respect to control cells. Pharmacological inhibitors of intracellular signals indicated that mechanisms of the excessive fMLF-induced production of O2? by long-lived neutrophils implicated the protein kinase C (PKC) pathway, preferentially the PKC-δ isoform, whose protein was augmented in these cells. Thus, long-term cytokine exposure modifies molecular pathways and functional characteristics of the neutrophil. 相似文献
3.
Charles N. Serhan M.Johan Broekman Helen M. Korchak James E. Smolen Aaron J. Marcus Gerald Weissmann 《Biochimica et Biophysica Acta (BBA)/Molecular Cell Research》1983,762(3):420-428
Human neutrophils aggregate and release mediators of inflammation, such as active oxygen species and lysosomal enzymes, when exposed to the chemoattractant, fMet-Leu-Phe, or the tumor promotor, phorbol myristate acetate. In order to ‘stage’ events which may lead to such neutrophil responses, we determined the temporal relationship between stimulus-induced changes in the endogenous phospholipids phosphatidylinositol (PI) and phosphatidic acid, the mobilization of calcium, and the onset of aggregation and generation of superoxide anion during the initial 2 min of cell activation. Within 5 s after addition of fMet-Leu-Phe (10?7 M) neutrophils accumulated phosphatidic acid and the levels of PI decreased, as determined by two-dimensional thin-layer chromatography and phosphorus determinations. By 5 s, phosphatidic acid levels rose approximately 3.5-fold and at 15 s the loss of PI exceeded the quantity of phosphatidic acid generated. In response to phorbol myristate acetate (1 μg/ml), however, changes in PI or phosphatidic acid were not observed until after 60 s. Accumulation of phosphatidic acid in fMet-Leu-Phe-stimulated cells was not inhibited by chelation of extracellular calcium. Neutrophils exposed to either fMet-Leu-Phe or phorbol myristate acetate also showed rapid decrements in fluorescence of cell-associated chlorotetracycline (used as an indirect probe of mobilization of intracellular membrane-associated calcium) and took up 45Ca2+ from the extracellular medium (under 60 s). The results indicate that changes in calcium mobilization, together with the alterations in phospholipid metabolism (under 5 s) anteceded aggregation and the generation of O?2 (10–15 s) induced by fMet-Leu-Phe. In contrast, when neutrophils were exposed to phorbol myristate acetate, changes in PI and phosphatidic acid (over 60 s) were observed after the mobilization of calcium (under 5 s) and the onset of O?2 generation and aggregation (30–35 s). 相似文献
4.
In this work, the effects of H2O2 at concentrations of 10?8–10?2 mol/l on the neutrophil ability to generate reactive oxygen and chlorine species (ROCS) and to secrete myeloperoxidase (MPO) were studied, as well as the H2O2 damaging action on neutrophils. It was found that H2O2 at concentrations of 2 × 10?3–10?2 mol/l led to disturbances of neutrophil membrane barrier properties and to a lactate dehydrogenase release. Incubation of neutrophils with an addition of 10?4–10?7 mol/l H2O2 was accompanied by an increase of the cell ability to generate ROCS during phagocytosis and a decrease of neutrophil ability to secrete MPO and ROCS into the extracellular medium during adhesion. Mechanisms of the H2O2 action are coupled with arachidonic acid metabolism. Inhibition of the 5-lipoxygenase or cyclooxygenase metabolism pathways produced an enhancement of the H2O2 destructive effect. Block of 5-lipoxygenase pathway led to elimination of the H2O2 action on MPO and ROCS secretion and to an enhancement of the H2O2 effect on the neutrophil ability to generate ROCS during phagocytosis. The obtained data indicate a high blood neutrophil resistance to the H2O2 destructive action and confirm the H2O2 regulatory role with respect to the neutrophil functions. 相似文献
5.
J. Kelley Bentley Peter W. Reed 《Biochimica et Biophysica Acta (BBA)/General Subjects》1981,678(2):238-244
All of the common cytochalasins activate superoxide anion release and exocytosis of β-N-acetylglucosaminidase and lysozyme from guinea-pig polymorphonuclear leukocytes (neutrophils) incubated in a buffered sucrose medium. Half-maximal activation of both processes is produced by approx. 2 μM cytochalasin A, C >μM cytochalasin B ? 4–5 μM cytochalasin D, E. While maximal rates of O2? release and extents of exocytosis require extracellular calcium (1–2 mM), replacing sucrose with monovalent cation chlorides is inhibitory to neutrophil activation by cytochalasins. Na+, K+ or choline inhibited either cytochalasin B- or E-stimulated O2? production with IC50 values of 5–10 mM and inhibition occurs whether Cl?, NO3? or SCN? is the anion added with Na+ or K+. Release of β-N-acetylglucosaminidase in control or cytochalasin B-stimulated cells is inhibited by NaCl (IC50 ≈ 10 mM), while cytochalasin E-stimulated exocytosis is reduced less and K+ or choline chloride are ineffective in inhibiting either cytochalasin B- or E-stimulated exocytosis. Release of β-glucuronidase, myeloperoxidase or acid phosphatase from neutrophils incubated in buffered sucrose is not stimulated by cytochalasin B. Stimulation of either O2? or β-N-acetylglucosaminidase release by low concentrations of cytochalasin A is followed by inhibition of each at higher concentrations. It appears that all cytochalasins can activate both NAD(P)H oxidase and selective degranulation of neutrophils incubated in salt-restricted media and that differential inhibition of these two processes by monovalent cations and/or anions is produced at some step(s) subsequent to cytochalasin interaction with the cell. 相似文献
6.
The correlation between an increased production of reactive oxygen species (ROS) and an enhanced calcium entry in primed neutrophils
stimulated with fMLP suggests that endogenous ROS could serve as an agonist to reinforce calcium signaling by positive feedback.
This work shows that exogenous H2O2 produced a rapid influx of Mn2+ and an increase of intracellular calcium. The H2O2 was insufficient to produce significant changes in the absence of extracellular calcium but addition of Ca2+ to H2O2-treated cells suspended in a free Ca2+/EGTA buffer resulted in a great increase in [Ca2+]i reflecting influx of Ca2+ across the cell membrane. The increase of intracellular calcium was inhibited by Ni2+, La3+, and hyperosmotic solutions of mannitol and other osmolytes. This raises the possibility that the secretion of H2O2 by activated neutrophils could act as an autocrine regulator of neutrophil function through the activation of calcium entry. 相似文献
7.
To more closely examine the role of the cell surface in transmembrane signal transduction in human neutrophils, sealed right-side-membrane vesicles free of organellar membrane components were used as models of the plasma membrane. These vesicles, incubated with a fluorescent analogue of the chemotactic peptide fMLP, bound this ligand similarly in extent and kinetics to intact neutrophils. Vesicles responded to this stimulation with a slow increase in internal [Ca++] which was inhibited by EGTA but not by verapamil; the cytosolic Ca++ transient seen in intact cells within 10 sec of stimulation was absent in vesicles. The vesicles also maintained a transmembrane potential (ψ) and were depolarized by the K+ ionophore valinomycin. However, unlike intact cells which hyperpolarized and then depolarized in response to fMLP, the vesicles demonstrated only a sustained hyperpolarization. Vesicles also differed from intact cells by not producing superoxide (O2?) in response to fMLP. Finally, fMLP caused dramatic alterations in membrane vesicle lipid metabolism: at early time points (within 5–10 sec), there was a transient production of diacylglycerol (DAG) concomitant with inositol lipid breakdown, with no apparent hydrolysis of non-inositol phospholipids. For up to 5 min after stimulation, there was no increase in the levels of phosphatidic acid or of inositol lipids. Thus, a significant portion of the signalling pathway in neutrophils is located at the cell surface or in the plasma membrane and functions independently of intracellular components. Furthermore, the plasma membrane is intimately involved in events occurring during both the early (DAG generation) and late (slow, prolonged rise in [Ca++]) phases of cellular response. In contrast, several of the responses to fMLP (the Ca++ transient, depolarization, generation of O2?, recycling of lipid metabolites) involve signalling machinery not constitutively resident on the neutrophil surface. © 1993 Wiley-Liss, Inc. 相似文献
8.
Kuender D. Yang Nancy H. Augustine Men-Fang Shaio John F. Bohnsack Harry R. Hill 《Journal of cellular physiology》1994,158(2):347-353
Previous studies have shown that fibronectin (Fn) enhances phagocytosis and killing of antibody-coated bacteria by neutrophils and macrophages. In an attempt to understand the mechanism of this enhancement, we have investigated the effects of Fn on phagocytosis-related actin organization as well as respiratory burst activity in neutrophils, monocytes and culture-derived macrophages. Employing an NBD-phallacidin flow cytometric analysis of filamentous actin formation, we found that Fn promotes rapid actin polymerization within 30 seconds in neutrophils, monocytes, and macrophages, but not lymphocytes. Enhancement of actin polymerization by Fn was concentration-dependent and mediated by a pertussis toxin- but not cholera toxin- sensitive G protein. Inhibition of protein kinase C by sphingosine (20 μM), calcium influx by verapamil (0.1 mM), or intracellular calcium mobilization by 8-(N, N-diethyl-amino) octyl-3,4,5-trimethoxybenzoate HCI (TMB-8; 0.1 mM) did not block Fn-enhanced actin polymerization in phagocytes. Incubation of neutrophils and macrophages on microtiter plates precoated with Fn suppressed superoxide (O2?) production induced by IgG- and IgA- opsonized group B streptococci. In contrast, Fn significantly enhanced IgA- and IgG-mediated O2? production by freshly isolated monocytes. These data suggest that Fn enhances phagocytosis, presumably through G protein-coupled cytoskeleton reorganization and augments O2? production by circulating monocytes. In contrast, it appears to suppress O2? production by the active phagocytic cells, neutrophils and macrophages. This may result in enhanced phagocytosis and intracellular killing of microorganisms without damaging interstitial tissues. © 1994 Wiley-Liss, Inc. 相似文献
9.
Pholasin, the photoprotein of the common piddock Pholas dactylus, emits an intense luminescence upon oxidation. The contribution of superoxide anion radicals and myeloperoxidase (MPO) to Pholasin luminescence in stimulated neutrophils was investigated. Data on Pholasin luminescence were compared with results of superoxide anion radical generation detected by the cytochrome c test as well as with the release of elastase and MPO. In N-formyl-methionyl-leucyl-phenylalanine (fMLP) stimulated neutrophils, most of the luminescence is caused by superoxide anion radicals, whereas MPO shows only a small effect as shown by coincubation with superoxide dismutase (SOD) as well as potassium cyanide (KCN), an inhibitor of MPO. However, both, O2- and MPO contribute to light emission in fMLP/cytochalasin B and phorbol myristoyl acetate (PMA) stimulated cells. Thus, the kinetics of O2- generation and MPO release can be very well detected by Pholasin luminescence in stimulated neutrophils.Degranulation of azurophilic granules was assessed using an ELISA test kit for released MPO or detection of elastase activity with MeO-Suc-Ala-Ala-Pro-Val-p-nitroanilide in the supernatant of stimulated cells. Both approaches revealed concurrently similar results concerning the amount and kinetics of enzyme release with data of Pholasin luminescence. Both, cytochrome c measurements and Pholasin luminescence indicate that fMLP/cytochalasin B and PMA stimulated neutrophils produce more O2- than fMLP stimulated cells. Thus, Pholasin luminescence can be used to detect, sensitively and specifically, O2- production and MPO release from stimulated neutrophils. 相似文献
10.
《Free radical research》2013,47(5):309-318
The ability of neutrophils from a Holstein-Friesian calf with bovine leukocyte adhesion deficiency (the proband with a genetic deficiency of the Mac-1 (CD11b/CD18) glycoprotein corresponding to the receptor of complement iC3b) to generate oxygen radicals was examined using electron spin resonance spectrometry (ESR) combined with a spin-trapping technique and luminol-dependent chemiluminescence spectrometry. When the neutrophils were stimulated with phorbol 12-myristate 13-acetate (PMA), an ESR spectrum confirming the generation of superoxide anions (O2?) was clearly observed in both healthy and diseased calves. However, when the neutrophils were stimulated by opsonized zymosan, appearance of the ESR spectrum was recognized in the healthy calves but not in the diseased calf. Similar results were obtained from chemiluminescence experiments. 相似文献
11.
Free radicals are reportedly involved in mucosal injury, including NH4OH-induced gastric lesions, but the kind, location and origin of radical generation have yet to be clarified. We developed the non-invasive measurement of reactive oxygen species (ROS) in stomach, and applied to mucosal injury. NH4OH-induced gastric lesions were prepared in rats, which were then given a nitroxyl probe intragastrically or intravenously, and the spectra of the gastric region were obtained by in vivo 300?MHz electron spin resonance (ESR) spectroscopy. The spectral change of the nitroxyl probe administered intragastrically was significantly enhanced 30?min after NH4OH administration, but no change occurred when the probe was given by intravenous injection. The enhanced change was confirmed to be due to ?OH generation, because it was completely suppressed by mannitol, catalase and desferrioxamine (DFO), and was not observed in neutropenic rats. NH4OH-induced neutrophil infiltration of the gastric mucosa was suppressed by intravenous injection of superoxide dismutase (SOD) or catalase, or by administration of allopurinol. The present study provided the direct evidence in NH4OH-treated living rats that ?OH produced from O2?- derived from neutrophils caused gastric lesion formation, while O2?- or H2O2 derived from the xanthine oxidase system in endothelial cells was involved in neutrophil infiltration. 相似文献
12.
Chiara Dianzani Marinella Parrini Cinzia Ferrara Roberto Fantozzi 《Cell biochemistry and function》1996,14(3):193-200
HNE (4-hydroxy-2,3-trans-nonenal), an aldehydic product of lipid peroxidation, has been reported to modulate different functional parameters of human and rat neutrophils (PMNs), such as chemiluminescence, migration and some enzymatic activities, thus exerting effects that varied according to the concentration tested. Experiments were done to evaluate the effects of HNE on superoxide anion (O2?.) production from human PMNs, isolated from healthy volunteers. After having tested that HNE by itself was not able to activate the cells, comparisons were made between its effects on PMNs, stimulated by either a single stimulus, N-formyl-methionyl-leucyl-phenylalanine (FMLP), or a combination of stimuli, such as FMLP and the neuropeptide substance P (SP; primed PMNs). In the concentration range tested (10?12–10?4 M ), HNE inhibited FMLP-evoked O2?. production with an IC50 of 11·6 ± 1·5 × 10?6 M ; at concentrations ≤10?6 M , HNE enhanced O2?. production elicited by FMLP + SP, while higher concentrations were inhibitory. There was a bell-shaped dose–response curve to the enhancing effects of HNE, depending on the incubation time being recorded after only short periods (≤5 min) of the exposure of the cells to HNE; this was not shown by structurally-related aldehydes, such as 2-nonenal and nonanal. These results suggest that low concentrations of HNE may participate in the evolution of the inflammatory process, by contributing to the activation of PMNs. The effects of high concentrations of the aldehyde may represent a mechanism which contributes to the regulation of the extent of the inflammatory response. 相似文献
13.
Bejarano I Terrón MP Paredes SD Barriga C Rodríguez AB Pariente JA 《Molecular and cellular biochemistry》2007,296(1-2):77-84
We have studied the effect of exogenous administration of hydrogen peroxide (H2O2) on phagocytic activity of human neutrophils. The treatment of cells with increasing concentrations of H2O2 evoke a significant elevation of phagocytic function assayed as phagocytic index, percentage and efficiency; and was similar
to that induced by the calcium mobilising agonist formyl-methionyl-leucyl-phenylalanine (fMLP). This stimulatory effect was
reduced by pre-treatment of neutrophils with catalase and abolished in neutrophils loaded with the intracellular calcium quelator
dimethyl BAPTA. In the absence of extracellular calcium, treatment of cells with H2O2 resulted in a increase in [Ca2+]
i
, indicating the release of calcium from intracellular stores. H2O2 abolished the typical calcium release stimulated by the physiological agonist fMLP, while depletion of agonist-sensitive
calcium pools by fMLP was able to prevent H2O2-induced calcium release. We conclude that H2O2 induces calcium release from agonist-sensitive stores and consequently increase the phagocytosis process. 相似文献
14.
Inhibition of the neutrophil oxidative response to a chemotactic peptide by inhibitors of arachidonic acid oxygenation 总被引:5,自引:0,他引:5
N-formylmethionylphenylalanine stimulates a short burst of antimycin A-insensitive O2 uptake, O2? production and hexosemonophosphate shunt oxidation of glucose by guinea pig peritoneal neutrophils. The stimulated oxidative metabolism, as well as release of lysosomal enzymes ± cytochalasin B, are inhibited by 5,8,11,14-eicosatetraynoic acid (ID50 1.5 × 10?5 M). High concentrations of indomethacin inhibit the peptide-stimulated oxidations (ID50 1.6 × 10?4 M) while acetylsalicylic acid (2.5 × 10?3 M) does not. Digitonin-stimulated oxidative metabolism and enzyme release are not inhibited by 5,8,11,14-eicosatetraynoic acid or indomethacin at concentrations that depress effects of the N-formylated peptide. 相似文献
15.
Eve de Lamirande Geneviève Lamothe Michèle Villemure 《Free radical biology & medicine》2009,46(10):1420-1427
We studied the modulation of superoxide anion (O2·?) and nitric oxide (NO·) generation during human sperm capacitation (changes needed for the acquisition of fertility). The production of NO· (diaminofluorescein-2 fluorescence assay), but not that of O2·? (luminescence assay), related to sperm capacitation was blocked by inhibitors of protein kinase C, Akt, protein tyrosine kinase, etc., but not by those of protein kinase A. Extracellular calcium (Ca2+) controlled O2·? synthesis but extra- and intracellular Ca2+ regulated NO· formation. Zinc inhibited capacitation and formation of O2·? and NO·. Zinc chelators (TPEN and EDTA) and sulfhydryl-targeted compounds (diamide and N-ethylmaleimide) stimulated capacitation and formation of O2·? and NO·; superoxide dismutase (SOD) and nitric oxide synthase inhibitor (L-NMMA) prevented these events. Diphenyliodonium (flavoenzyme inhibitor) blocked capacitation and related O2·? synthesis but promoted NO· formation, an effect canceled by SOD and L-NMMA. NADPH induced capacitation and NO· (but not O2·?) synthesis and these events were blocked by L-NMMA and not by SOD. Integration of these data on O2·? and NO· production during capacitation reinforces the concept that a complex, but flexible, network of factors is involved and probably is associated with rescue mechanisms, so that spermatozoa can achieve successful fertilization. 相似文献
16.
Chunqing Cai Shihao Tang Xubu Wang Shaoxi Cai Xiaojing Meng Wenying Zou Fei Zou 《Biochemical and biophysical research communications》2013,430(2):816-821
Tissue penetration of neutrophils is a key process in many inflammatory diseases. In response to inflammatory stimuli such as N-formyl-methionine-leucine-phenylalanine (fMLP), neutrophils polarize and migrate towards the chemotactic gradient of the stimulus. Elevated intracellular Ca2+ concentration is known to play a critical role in neutrophil polarization and migration; however, the exact mechanism remains elusive. Here, we demonstrated that fMLP stimulation caused not only store-operated calcium entry (SOCE), but also receptor-operated calcium entry (ROCE) in neutrophils by using both pharmacological and neutralizing monoclonal antibody approaches. We also investigated neither Rac2 nor Cdc42 activation could take place if either SOCE or ROCE was inhibited. This study thus provides the first evidence for coordination of Ca2+ influx by SOCE and ROCE to regulate neutrophil polarization. 相似文献
17.
Paul G. Heyworth John M. Robinson J. Ding Beverly A. Ellis J. A. Badwey 《Histochemistry and cell biology》1997,108(3):221-233
Neutrophils contain a 21-kDa phosphoprotein that undergoes rapid dephosphorylation upon stimulation of these cells with the
chemoattractant N-fMet-Leu-Phe (fMLP), activators of protein kinase C [e.g., 4β-phorbol 12-myristate 13-acetate (PMA)] or
the calcium ionophore A23187. This phosphoprotein was identified as the non-muscle form of cofilin by peptide sequencing and
immunoblotting with specific antibodies. Evidence is presented that in neutrophils cofilin is regulated by a continual cycle
of phosphorylation and dephosphorylation, and that the phosphatase undergoes activation during cell stimulation. Experiments
with a wide variety of antagonists further suggested that the protein kinase that participates in these reactions may be a
novel enzyme. The kinetics of cofilin dephosphorylation in neutrophils stimulated with fMLP or PMA were very similar to those
observed for superoxide (O2
–) release. Immunofluorescent studies revealed that cofilin was present thouroughout the cytosol of resting neutrophils and
underwent rapid translocation to the F-actin-rich, ruffled membranes of stimulated cells. Cytochemical analysis further revealed
that the ruffled membranes also contained large amounts of hydrogen peroxide (H2O2), a product of the O2
–/H2O2-generating activity of stimulated neutrophils (NADPH oxidase). Cofilin is therefore well placed to participate in the continual
polymerization and depolymerization of F-actin that is thought to give rise to the oscillatory pattern of H2O2 production observed under certain conditions.
Accepted: 22 April 1997 相似文献
18.
Impaired cardiovascular function during acute myocardial infarction (MI) is partly associated with recruitment of activated polymorphonuclear neutrophils. The protective role of arjunolic acid (AA; 2,3,23-trihydroxy olean-12-en-28-oic acid) is studied in the modulation of neutrophil functions in vitro by measuring the reactive oxygen species (ROS) generation. Neutrophils were isolated from normal and acute MI mice to find out the efficacy of AA in reducing oxidative stress. Stimulation of neutrophils with phorbol-12-myristate-13-acetate (PMA) resulted in an oxidative burst of superoxide anion (O2−) and enhanced release of lysosomal enzymes. The treatment of neutrophils with PMA induced phosphorylation of Ser345 on p47phox, a cytosolic component of NADPH oxidase. Furthermore, we observed activated ERK induced phosphorylation of Ser345 in MI neutrophils. Treatment with AA significantly inhibited the phosphorylation of P47phox and ERK in the stimulated controls and MI neutrophils. Oxidative phosphorylation activities in MI cells were lower than in control, while the glycolysis rates were elevated in MI cells compared to the control. In addition, we observed AA decreased intracellular oxidative stress and reduced the levels of O2− in neutrophils. This study therefore identifies targets for AA in activated neutrophils mediated by the MAPK pathway on p47phox involved in ROS generation. 相似文献
19.
Maria Armida Rossi Clelia Di Mauro Hermann Esterbauer Fulvio Fidale Mario Umberto Dianzani 《Cell biochemistry and function》1994,12(4):275-280
A comparison has been made between the effects of 4-hydroxy-2,3-trans-nonenal (HNE) and 4-hydroxy-2,3-trans-octenal (HOE), two lipid peroxidation products, on the basal and GTPgammaS-stimulated activities of phosphoinositide-specific phospholipase C (PL-C) of rat polymorphonuclear leukocytes. PL-C activity was determined in vitro by measuring the hydrolysis of [3H] phosphatidylinositol-4,5-bis-phosphate (PtdIns-P2) added as exogenous substrate to neutrophil plasma membranes. PL-C was activated by concentrations of HNE ranging from 10?8 to 10?6 M both in the presence and in the absence of 2 × 10?5 M GTPgammaS; HOE stimulated the enzymatic activity between 10?11 and 10?8 M ; maximal stimulation was given by 10?11 M HOE plus GTPgammaS. The aldehyde concentrations able to accelerate PtdIns-P2 breakdown displayed a good correspondence with those which have been reported to stimulate the oriented migration of rat neutrophils. Pretreatment of neutrophils with pertussis toxin prevented the stimulation of PL-C by 10?11 M HOE and by HOE plus GTPgammaS. Our results suggest that the chemotactic action of HNE and HOE might depend on the activation of PL-C; furthermore a regulatory G protein appears to be involved in the acceleration of PtdIns-P2 turnover by HOE. 相似文献
20.
《Free radical research》2013,47(3-6):341-345
Human neutrophils can aggregate, degranulate, and release mediators of inflammation including oxygen radicals and lipoxygenase (LO)-derived products of arachidonic acid. The regulation of 5– and 15-lipoxy-genases appears to be important since their products (e.g. leukotrienes and lipoxins) display unique spectra of bioactions. Addition of 15-HETE. a product of the 15-LO, to neutrophils in suspension dramatically shifted the LO products generated and led to a dose-dependent increase in lipoxins, while the production of leukotriene B4 and its μ-oxidation products (i.e. 20-COOH-LTB4 and 20-OH-LTB4) was inhibited. Exogenous 15-HETE also dose-dependently inhibited the generation of superoxide anions induced by either the chemotactic peptide f-met-leu-phe or the divalent cation ionophore A23187. Neither lipoxin A, nor lipoxin B4 (10?8?10?6M) inhibited O2?? generation induced by either f-met-leu-phe or A23187. These results indicate that in addition to serving as a substrate for lipoxin generation, 15-HETE also inhibits superoxide anion generation by human neutrophils. Together they provide further evidence to suggest that products of the 15-lipoxygenase may serve a regulatory role at inflammatory loci. 相似文献