scholarly journals Phagocytosis-induced chemotaxis receptor cycling in neutrophils is mediated by thiol oxidation

Blood ◽  
1982 ◽  
Vol 59 (6) ◽  
pp. 1337-1343
Author(s):  
TA Lane ◽  
GE Lamkin
Keyword(s):  
Important Determinant ◽  
Receptor Occupancy ◽  
Thiol Groups ◽  
Thiol Oxidation ◽  
Chemotactic Peptide ◽  
Oxidation Reduction ◽  
Dependent Inhibition ◽  
Dose Dependent ◽  
Physiologic Role ◽  
Mediated Reduction

We previously demonstrated that neutrophil (PMN) phagocytosis of opsonized zymosan (OPZ) caused oxygen-dependent inhibition of chemotactic peptide receptor (CPR) binding using the ligand 3H-formyl- methionyl-leucyl-phenylalanine (3H-FMLP). In the current studies we sought to determine the mechanism of CPR inhibition by OPZ. We found that 3 mM cysteine and 5 mM dithiothreitol (DTT) did not decrease PMN phagocytosis, but abolished CPR inhibition by OPZ. Furthermore, incubation of PMN with DTT after OPZ partially restored the suppressed CPR. PMN CPR saturation studies with 3H-FMLP on PMN incubated with or without DTT after phagocytosis indicated that DTT restored receptor numbers to 92% +/- 6% of prephagocytosis values and also improved CPR affinity for 3H-FMLP. In additional studies we found that the cell penetrating thiol reagents mercuric chloride (HgCl2), N-ethylmaleimide (NEM), and diamide, but not the nonpenetrating agents p- chloromercuriphenylsulfonic acid and p-chloromercuribenzoate, caused rapid, dose-dependent, DTT-inhibitable suppression of up to 85% CPR binding. CPR inhibition by HgCl2 and NEM was irreversible. PMN CPR saturation studies showed that NEM decreased total receptor numbers, not CPR affinity. The effect of NEM was not inhibited by receptor occupancy at 4 degree C or at 37 degree C. These studies indicate that: (1) intact thiol groups are a requirement for CPR ligand-binding; (2) the thiol groups are presumptively located on the interior side of, or within hydrophobic portions of, the plasma membrane and are not part of the 3H-FMLP binding site; (3) thiol oxidation-reduction induces reversible alteration in CPR binding; (4) phagocytosis-induced modulation of the CPR is due, in part, to thiol oxidation. These studies suggest that postphagocytic, thiol-mediated reduction in CPR binding may play a physiologic role in the inhibition of PMN chemotaxis at inflammatory sites. The state of CPR-associated thiols may be an important determinant of CPR binding under physiologic conditions other than phagocytosis.

scholarly journals Phagocytosis-induced chemotaxis receptor cycling in neutrophils is mediated by thiol oxidation

Blood ◽  
1982 ◽  
Vol 59 (6) ◽  
pp. 1337-1343 ◽  
Author(s):  
TA Lane ◽  
GE Lamkin
Keyword(s):  
Important Determinant ◽  
Receptor Occupancy ◽  
Thiol Groups ◽  
Thiol Oxidation ◽  
Chemotactic Peptide ◽  
Oxidation Reduction ◽  
Dependent Inhibition ◽  
Dose Dependent ◽  
Physiologic Role ◽  
Mediated Reduction

Abstract We previously demonstrated that neutrophil (PMN) phagocytosis of opsonized zymosan (OPZ) caused oxygen-dependent inhibition of chemotactic peptide receptor (CPR) binding using the ligand 3H-formyl- methionyl-leucyl-phenylalanine (3H-FMLP). In the current studies we sought to determine the mechanism of CPR inhibition by OPZ. We found that 3 mM cysteine and 5 mM dithiothreitol (DTT) did not decrease PMN phagocytosis, but abolished CPR inhibition by OPZ. Furthermore, incubation of PMN with DTT after OPZ partially restored the suppressed CPR. PMN CPR saturation studies with 3H-FMLP on PMN incubated with or without DTT after phagocytosis indicated that DTT restored receptor numbers to 92% +/- 6% of prephagocytosis values and also improved CPR affinity for 3H-FMLP. In additional studies we found that the cell penetrating thiol reagents mercuric chloride (HgCl2), N-ethylmaleimide (NEM), and diamide, but not the nonpenetrating agents p- chloromercuriphenylsulfonic acid and p-chloromercuribenzoate, caused rapid, dose-dependent, DTT-inhibitable suppression of up to 85% CPR binding. CPR inhibition by HgCl2 and NEM was irreversible. PMN CPR saturation studies showed that NEM decreased total receptor numbers, not CPR affinity. The effect of NEM was not inhibited by receptor occupancy at 4 degree C or at 37 degree C. These studies indicate that: (1) intact thiol groups are a requirement for CPR ligand-binding; (2) the thiol groups are presumptively located on the interior side of, or within hydrophobic portions of, the plasma membrane and are not part of the 3H-FMLP binding site; (3) thiol oxidation-reduction induces reversible alteration in CPR binding; (4) phagocytosis-induced modulation of the CPR is due, in part, to thiol oxidation. These studies suggest that postphagocytic, thiol-mediated reduction in CPR binding may play a physiologic role in the inhibition of PMN chemotaxis at inflammatory sites. The state of CPR-associated thiols may be an important determinant of CPR binding under physiologic conditions other than phagocytosis.

scholarly journals μ-opioids activate phospholipase C in SH-SY5Y human neuroblastoma cells via calcium-channel opening

1995 ◽  
Vol 305 (2) ◽  
pp. 577-581 ◽  
Author(s):  
D Smart ◽  
G Smith ◽  
D G Lambert
Keyword(s):  
Phospholipase C ◽  
Opioid Receptor ◽  
Neuroblastoma Cells ◽  
Receptor Occupancy ◽  
Mu Opioid Receptor ◽  
Human Neuroblastoma ◽  
Mu Opioid ◽  
Inositol 1,4,5 Trisphosphate ◽  
Dependent Inhibition ◽  
Dose Dependent

We have recently reported that, in SH-SY5Y cells, mu-opioid receptor occupancy activates phospholipase C via a pertussis toxin-sensitive G-protein. In the present study we have further characterized the mechanisms involved in this process. Fentanyl (0.1 microM) caused a monophasic increase in inositol 1,4,5-trisphosphate mass formation, with a peak (20.5 +/- 3.6 pmol/mg of protein) at 15 s. Incubation in Ca(2+)-free buffer abolished this response, while Ca2+ replacement 1 min later restored the stimulation of inositol 1,4,5-trisphosphate formation (20.1 +/- 0.6 pmol/mg of protein). In addition, nifedipine (1 nM-0.1 mM), an L-type Ca(2+)-channel antagonist, caused a dose-dependent inhibition of inositol 1,4,5-trisphosphate formation, with an IC50 of 60.3 +/- 1.1 nM. Elevation of endogenous beta/gamma subunits by selective activation of delta-opioid and alpha 2 adrenoceptors failed to stimulate phospholipase C. Fentanyl also caused a dose-dependent (EC50 of 16.2 +/- 1.0 nM), additive enhancement of carbachol-induced inositol 1,4,5-trisphosphate formation. In summary, we have demonstrated that in SH-SY5Y cells activation of the mu-opioid receptor allows Ca2+ influx to activate phospholipase C. However, the possible role of this mechanism in the process of analgesia remains to be elucidated.

Differences between antigenic determinants of pig and cat zona pellucida proteins

Reproduction ◽  
2000 ◽  
pp. 15-23 ◽  
Author(s):  
K Jewgenow ◽  
M Rohleder ◽  
I Wegner
Keyword(s):  
In Vitro Fertilization ◽  
Zona Pellucida ◽  
Antigenic Determinants ◽  
Vitro Fertilization ◽  
Contraceptive Vaccine ◽  
Sperm Binding ◽  
Dependent Inhibition ◽  
Dose Dependent Inhibition ◽  
Dose Dependent

Despite many efforts, the control of reproduction in feral cat populations is still a problem in urban regions around the world. Immunocontraception is a promising approach; thus the present study examined the suitability of the widely used pig zona pellucida proteins (pZP) for contraception in feral domestic cats. Purified zona pellucida proteins obtained from pig and cat ovaries were used to produce highly specific antisera in rabbits. Antibodies against pZP raised in rabbits or lions were not effective inhibitors of either in vitro sperm binding (cat spermatozoa to cat oocytes) or in vitro fertilization in cats, whereas antibodies against feline zona pellucida proteins (fZP) raised in rabbits showed a dose-dependent inhibition of in vitro fertilization. Immunoelectrophoresis, ELISA and immunohistology of ovaries confirmed these results, showing crossreactivity of anti-fZP sera to fZP and to a lesser extent to pZP, but no interaction of anti-pZP sera with fZP. It is concluded that cat and pig zonae pellucidae express a very small number of shared antigenic determinants, making the use of pZP vaccine in cats questionable. A contraceptive vaccine based on feline zona pellucida determinants will be a better choice for the control of reproduction in feral cats if immunogenity can be achieved.

Antioxidant effects and in vitro cytotoxicity on human cancer cell lines of flavonoid-rich Flamboyant (Delonix regia (Bojer) Raf.) flower extract

2020 ◽  
Vol 21 ◽  
Author(s):  
Putthiporn Khongkaew ◽  
Phanphen Wattanaarsakit ◽  
Konstantinos I. Papadopoulos ◽  
Watcharaphong Chaemsawang
Keyword(s):  
Cell Line ◽  
Cell Lines ◽  
Cancer Cell ◽  
Leukemia Cell ◽  
Leukemia Cell Line ◽  
Flower Extract ◽  
Dependent Inhibition ◽  
Dose Dependent Inhibition ◽  
Murine Leukemia Cell ◽  
Dose Dependent

Background: Cancer is a noncommunicable disease with increasing incidence and mortality rates both worldwide and in Thailand. Its apparent lack of effective treatments is posing challenging public health issues. Introduction: Encouraging research results indicating probable anti-cancer properties of the Delonix regia flower extract (DRE) have prompted us to evaluate the feasibility of developing a type of product for future cancer prevention or treatment. Methods and Results: In the present report, using High Performance Liquid Chromatography (HPLC), we demonstrate in the DRE, the presence of high concentrations of three identifiable flavonoids, namely rutin 4.15±0.30 % w/w, isoquercitrin 3.04±0.02 %w/w, and myricetin 2.61±0.01 % w/w respectively while the IC50 of DPPH and ABTS assay antioxidation activity was 66.88±6.30 µg/ml and 53.65±7.24 µg/ml respectively. Discussion: Our cancer cell line studies using the MTT assay demonstrated DREs potent and dose dependent inhibition of murine leukemia cell line (P-388: 35.28±4.07% of cell viability remaining), as well as of human breast adenocarcinoma (MCF-7), human cervical carcinoma (HeLa), human oral cavity carcinoma (KB), and human colon carcinoma (HT-29) cell lines in that order of magnitude. Conclusion: Three identifiable flavonoids (rutin, isoquercitrin and myricetin) with high antioxidation activity and potent and dose dependent inhibition of murine leukemia cell line and five other cancer cell lines were documented in the DRE. The extract’s lack of cytotoxicity in 3 normal cell lines is a rare advantage not usually seen in current antineoplastic agents. Yet another challenge of the DRE was its low dissolution rate and long-term storage stability, issues to be resolved before a future product can be formulated.

Investigation on the antibacterial activity of electronic cigarette liquids (ECLs): a proof of concept study

2020 ◽  
Vol 21 ◽  
Author(s):  
Virginia Fuochi ◽  
Massimo Caruso ◽  
Rosalia Emma ◽  
Aldo Stivala ◽  
Riccardo Polosa ◽  
...  
Keyword(s):  
Antibacterial Activity ◽  
Bacterial Species ◽  
A549 Cells ◽  
Bactericidal Effect ◽  
Minimal Bactericidal Concentration ◽  
Viability Assay ◽  
Dependent Inhibition ◽  
Dose Dependent Inhibition ◽  
Dose Dependent

Background: The key ingredients of e-cigarettes liquid are commonly propane-1,2-diol (also called propylene glycol) and propane-1,2,3-triol (vegetal glycerol) and their antimicrobial effects are already established. The nicotine and flavors which are often present in e-liquids can interfere with the growth of some microorganisms. Objective: The effect of the combining these elements in e-liquids is unknown. The aim of the study was to investigate the possible effects of these liquids on bacterial growth in the presence or absence of nicotine and flavors. Methods: Susceptibilities of pathogenic strains (Klebsiella pneumoniae, Staphylococcus aureus, Pseudomonas aeruginosa, Acinetobacter baumannii, Escherichia coli, Enterococcus faecalis and Sarcina lutea) were studied by means of a multidisciplinary approach. Cell viability and antioxidant assays were also evaluated. Results: All e-liquids investigated showed antibacterial activity against at least one pathogenic strain. A higher activity was correlated to the presence of flavors and nicotine. Discussion: In most cases the value of minimal bactericidal concentration is equal to the value of minimal inhibitory concentration showing that these substances have a bactericidal effect. This effect was observed in concentrations up to 6.25% v/v. Antioxidant activity was also correlated to presence of flavors. Over time, the viability assay in human epithelial lung A549 cells showed a dose-dependent inhibition of cell growth. Conclusion: Our results have shown that flavors considerably enhance the antibacterial activity of propane-1,2-diol and propane-1,2,3-triol. This study provides important evidence that should be taken into consideration in further investigative approaches, to clarify the different sensitivity of the various bacterial species to e-liquids, including the respiratory microbiota, to highlight the possible role of flavors and nicotine.

scholarly journals In VitroActivities of Hexaazatrinaphthylenes against Leishmania spp.

2015 ◽  
Vol 59 (5) ◽  
pp. 2867-2874 ◽  
Author(s):  
Atteneri López-Arencibia ◽  
Daniel García-Velázquez ◽  
Carmen M. Martín-Navarro ◽  
Ines Sifaoui ◽  
María Reyes-Batlle ◽  
...  
Keyword(s):  
Therapeutic Agents ◽  
Content Type ◽  
Inhibition Effect ◽  
Intracellular Amastigotes ◽  
Dependent Inhibition ◽  
Leishmania Spp ◽  
Dose Dependent Inhibition ◽  
Dose Dependent ◽  
Potential Use

ABSTRACTThein vitroactivity of a novel group of compounds, hexaazatrinaphthylene derivatives, against two species ofLeishmaniais described in this study. These compounds showed a significant dose-dependent inhibition effect on the proliferation of the parasites, with 50% inhibitory concentrations (IC50s) ranging from 1.23 to 25.05 μM against the promastigote stage and 0.5 to 0.7 μM against intracellular amastigotes. Also, a cytotoxicity assay was carried out to in order to evaluate the possible toxic effects of these compounds. Moreover, different assays were performed to determine the type of cell death induced after incubation with these compounds. The obtained results highlight the potential use of hexaazatrinaphthylene derivatives againstLeishmaniaspecies, and further studies should be undertaken to establish them as novel leishmanicidal therapeutic agents.

μ-Opiate Receptors in Neuronal Primary Cultures from Cerebral Cortex

1990 ◽  
Vol 17 (3) ◽  
pp. 177-181
Author(s):  
Peter S. Eriksson ◽  
Elisabeth Hansson ◽  
Lars Rönnbäck
Keyword(s):  
Cerebral Cortex ◽  
Primary Cultures ◽  
Opiate Receptors ◽  
Neuronal Cultures ◽  
Camp Accumulation ◽  
Dependent Inhibition ◽  
Dose Dependent Inhibition ◽  
Neuronal Primary Cultures ◽  
Dose Dependent ◽  
Μ Receptor

The presence of μ-opioid receptors was demonstrated as effects of receptor stimulation on PGE1-induced cAMP accumulation in neuronal-enriched primary cultures from rat cerebral cortex. Morphine was used as a μ-receptor agonist. There was a dose-dependent inhibition of the PGE1-stimulated cAMP accumulation by morphine, blocked by the μ-receptor antagonist naloxone. These findings suggest that these neuronal cultures express μ-receptors, possibly connected to adenylate cyclase via an inhibitory Gi-protein. The probable use of functional μ-receptors in neurotoxicological tests is discussed.

Central glucagon-like peptide 1 receptor-induced anorexia requires glucose metabolism-mediated suppression of AMPK and is impaired by central fructose

2013 ◽  
Vol 304 (7) ◽  
pp. E677-E685 ◽  
Author(s):  
Melissa A. Burmeister ◽  
Jennifer Ayala ◽  
Daniel J. Drucker ◽  
Julio E. Ayala
Keyword(s):  
Food Intake ◽  
Glucose Metabolism ◽  
Dependent Manner ◽  
Anorectic Effect ◽  
Dependent Inhibition ◽  
Glycolytic Inhibitor ◽  
Glucagon Like Peptide ◽  
Glucagon Like Peptide 1 ◽  
Amp Activated Protein Kinase ◽  
Mediated Reduction

Glucagon-like peptide-1 (GLP-1) suppresses food intake via activation of a central (i.e., brain) GLP-1 receptor (GLP-1R). Central AMP-activated protein kinase (AMPK) is a nutrient-sensitive regulator of food intake that is inhibited by anorectic signals. The anorectic effect elicited by hindbrain GLP-1R activation is attenuated by the AMPK stimulator AICAR. This suggests that central GLP-1R activation suppresses food intake via inhibition of central AMPK. The present studies examined the mechanism(s) by which central GLP-1R activation inhibits AMPK. Supporting previous findings, AICAR attenuated the anorectic effect elicited by intracerebroventricular (icv) administration of the GLP-1R agonist exendin-4 (Ex-4). We demonstrate that Ex-4 stimulates glycolysis and suppresses AMPK phosphorylation in a glucose-dependent manner in hypothalamic GT1-7 cells. This suggests that inhibition of AMPK and food intake by Ex-4 requires central glucose metabolism. Supporting this, the glycolytic inhibitor 2-deoxyglucose (2-DG) attenuated the anorectic effect of Ex-4. However, icv glucose did not enhance the suppression of food intake by Ex-4. AICAR had no effect on Ex-4-mediated reduction in locomotor activity. We also tested whether other carbohydrates affect the anorectic response to Ex-4. Intracerebroventricular pretreatment with the sucrose metabolite fructose, an AMPK activator, attenuated the anorectic effect of Ex-4. This potentially explains the increased food intake observed in sucrose-fed mice. In summary, we propose a model whereby activation of the central GLP-1R reduces food intake via glucose metabolism-dependent inhibition of central AMPK. We also suggest that fructose stimulates food intake by impairing central GLP-1R action. This has significant implications given the correlation between sugar consumption and obesity.

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