38 research outputs found
Effects of sphingosine and sphingosine analogues on the free radical production by stimulated neutrophils: ESR and chemiluminescence studies
Sphingolipids inhibit the activation of the neutrophil (PMN) NADPH oxidase by protein kinase C pathway. By electron spin resonance spectroscopy (ESR) and chemiluminescence (CL), we studied the effects of sphingosine (SPN) and ceramide analogues on phorbol 12-myristate 13-acetate (PMA, 5 × 10-7M) stimulated PMN (6 × 106 cells). By ESR with spin trapping (100 mM DMPO: 5,5-dimethyl-1-pyrroline-Noxide), we showed that SPN (5 to 8 × 10-6M), C2-ceramide (N-acetyl SPN) and C6-ceramide (N-hexanoyl SPN) at the final concentration of 2 × 10-5 and 2 × 10-4M inhibit the production of free radicals by stimulated PMN. The ESR spectrum of stimulated PMN was that of DMPO-superoxide anion spin adduct. Inhibition by 5 × 10-6M SPN was equivalent to that of 30 U/ml SOD. SPN (5 to 8 × 10-6M) has no effect
on in vitro systems generating superoxide anion (xanthine 50 mM/xanthine oxidase 110 mU/ml) or hydroxyl radical (Fenton reaction: 88 mM H2O2, 0.01 mM Fe2+ and 0.01 mM EDTA). SPN and N-acetyl SPN also inhibited the CL of PMA stimulated PMN in a dose dependent manner (from 2 × 10-6 to 10-5M), but N-hexanoyl SPN was less active (from 2 × 10-5 to 2 × 10-4M). These effects were compared with those of known PMN inhibitors, superoxide dismutase, catalase and azide. SPN was a better inhibitor compared with these agents. The complete inhibition by SPN of ESR signal and CL of stimulated PMN confirms that this compound or one of its metabolites act at the level of NADPH-oxidase, the key enzyme responsible for production of oxygen-derived free radicals
Synthesis and pharmacological evaluation of 6-piperidino- and 6-piperazinoalkyl-2(3H)-benzothiazolones as mixed sigma/5-HT1A ligands
In an effort to produce new pharmacological probes with mixed sigma/5-HT1A affinity, we have synthesized a series of 12 original 6-piperidino- or piperazino-alkyl-2(3H)-benzothiazolones and their receptor binding profile (sigma, 5-HT1A, 5-HT2A, 5-HT3, D-2, H-1, and M-1) was determined. The best mixed sigma/5-HT1A affinity profile was found within the piperidine series with 4-benzyl substitution associated to linker methylene chain n = 2 (K-i 5 and 4 nM, respectively). Moreover, a highly selective sigma2 ligand was obtained with a 3,4-dichlorobenzyl substitution associated to n = 4 (K-i 2 nM, selectivity ratio sigma1/sigma2 = 70). (C) 2002 Elsevier Science Ltd. All rights reserved
Polyphenols from Silybum marianum inhibit in vitro the oxidant response of equine neutrophils and myeloperoxidase activity.
A recent study showed that silymarin, a standardized extract of S. marianum might be used in the prevention of equine laminitis. We investigated the effects of quercetin and some compounds found in silymarin (silybin, taxifolin and dehydrosilybin) on reactive oxygen species (ROS) production and myeloperoxidase (MPO) release by stimulated equine neutrophils (PMNs) and on MPO activity. All compounds (tested between 100 nm and 100 mum) inhibited superoxide anion production by stimulated PMNs in a dose-dependent manner. Dehydrosilybin and quercetin inhibited superoxide production and MPO release from 10 mum. Classical MPO assay showed quercetin as the most potent inhibitor, followed by taxifolin, dehydrosilybin and silybin. SIEFED MPO assay highlighting the binding of tested compounds to MPO showed that only quercetin and taxifolin maintained an efficient inhibition above 90% at 10 mum. Altogether, our results showed a strong inhibition of PMN activation by planar compounds such as quercetin and dehydrosilybin and a strong inhibition of MPO activity by the smallest molecules, quercetin and taxifolin. In conclusion, the compounds from silymarin may be useful for modulating the oxidative response of PMNs, involved in the pathogenesis of laminitis, but further in vivo studies are needed
Antioxidant and anti-inflammatory like properties of benzoic acid analogs on the oxidant response of neutrophils: structure/redox potential relationship study
High Concentrations of Histamine Stimulate Equine Polymorphonuclear Neutrophils to Produce Reactive Oxygen Species
OBJECTIVE AND DESIGN: Because high concentrations of histamine are locally released in inflammation, we investigated the effects of supraphysiological doses of histamine on the production of reactive oxygen species (ROS) by neutrophils. MATERIALS AND METHODS: Isolated equine neutrophils were activated by 10(-4) to 5 x 10(-3) M histamine. The production of ROS and free radicals was estimated by luminol-enhanced chemiluminescence (CL) and electron spin resonance (ESR) with spin trapping technique. In this model of histamine-stimulated neutrophils, we tested the antagonists of H1 and H2 histamine receptors, the role of Ca2+ and Mg2+, the role of staurosporine and pertussis toxin (inhibitors of protein kinase C and proteins G) and the effects of superoxide dismutase, catalase, hydroxyl radical scavengers (phenylalanine and mannitol) and N(G)-monomethyl-L-arginine (L-NMMA), inhibitor of NO-synthase. RESULTS: Histamine (from 10(-5) to 10(-3) M) stimulated neutrophils to produce CL and ESR signals characterized by spin adducts of superoxide anion and/or hydroxyl radicals. The CL response was inhibited by 10(-4) and 10(-3) M H1 receptor antagonists (promethazine, pyrilamine, and diphenhydramine), by Ca2+ and Mg2+ depletion and by 10 nmoles staurosporine. CL was partially inhibited by pertussis toxin (4 microg/ mL). The ESR signals were practically suppressed by pyrilamine (an H1 receptor antagonist) and superoxide dismutase, and partially inhibited by catalase, hydroxyl radical scavengers and L-NMMA (respectively 59, +/- 30% and 68% inhibition). CONCLUSIONS: High concentrations of histamine stimulated the neutrophils to product ROS and free radicals via H1 receptors and the NADPH-oxidase pathway
Reactivity towards singlet oxygen of propofol inside liposomes and neuronal cells
Singlet oxygen (1O2), a reactive oxygen species, has been found to be implicated in many cellular events and pathological disorders.Herein, we investigated the reactivity of 1O2 towards the anaesthetic agent propofol (PPF) encapsulated within DMPC liposomes. By time
resolved luminescence, the rate constant of 1O2 quenching by PPF was evaluated, depending on the location of the sensitizer. The nature of the oxidation product, resulting from the reaction of
1O2 with PPF, was determined using absorption and HPLC techniques. Finally, the in vitro protective effect of PPF towards the1O2-induced neuronal cell toxicity was evaluated in terms of cell viability
The antibiotic ceftazidime is a singlet oxygen quencher as demonstrated by ultra-weak chemiluminescence and by inhibition of AAP consumption.
We demonstrated that the cephalosporin antibiotic ceftazidime (CAZ) deactivated singlet oxygen (1O2). We then studied the mechanisms of the CAZ effects on the ultra weak chemiluminescence (uwCL) associated with the energy decay of 1O2 generated by the Mallet reaction (H2O2 + HOCl --> HCl + H2O + 1O2), and on the anthracene-9,10-dipropionic acid (AAP) consumption by 1O2 generated by irradiation of Rose Bengal (RB). The uwCL generated by the Mallet reaction was amplified (6.2 times) by CAZ. The use of red and blue filters, which absorb radiation below 610 nm and between 470 and 700 nm respectively, demonstrated that CAZ increased the uwCL by a radiation emission at wavelengths shorter than the 633 and 704 nm wavelength emissions of 1O2. CAZ was excited by scavenging the energy excess of 1O2, which so returned to its fundamental state, while CAZ deactivated with light emission between 430-480 nm. CAZ also inhibited in a dose-dependent manner the consumption of AAP by 1O2 generated by the irradiation of RB. The protection of AAP by 5 x 10(-3) M CAZ was equivalent to that of 10(-3) M histidine and 3 X 10(-6) M sodium azide. This process of 1O2 deactivation will be useful in diseases characterized by an excessive PMN activation with a release of activated oxygen species
ESR technique for non invasive way to quantify cyclodextrins effect on cell membranes
A new way to study the action of cyclodextrin was developed to quantify the damage caused on cell membrane and lipid bilayer. The Electron Spin Resonance (ESR) spectroscopy was used to study the action of Randomly methylated-beta-cyclodextrin (Rameb) on living cells (HCT-116). The relative anisotropy observed in ESR spectrum of nitroxide spin probe (5-DSA and cholestane) is directly related to the rotational mobility of the probe,which can be further correlated with themicroviscosity. The use of ESR probes clearly shows a close correlation between cholesterol contained in cells and cellular membrane microviscosity. This study also demonstrates the Rameb ability to extract cholesterol and phospholipids in time- and dose-dependent ways. In addition, ESR spectra enabled to establish that cholesterol is extracted from lipid rafts to form stable aggregates. The present work supports that ESR is an easy, reproducible and noninvasive technique to study the effect of cyclodextrins on cell membranes