Oxygen consumption and electron spin resonance studies of free radical production by alveolar cells exposed to anoxia: inhibiting effects of the antibiotic ceftazidime.

in Redox Report : Communications in Free Radical Research (2002), 7(2), 85-94

By EPR spectroscopy, we investigated free radical production by cultured human alveolar cells subjected to anoxia/re-oxygenation (A/R), and tested the effects of ceftazidime, an antibiotic previously ... [more ▼]

By EPR spectroscopy, we investigated free radical production by cultured human alveolar cells subjected to anoxia/re-oxygenation (A/R), and tested the effects of ceftazidime, an antibiotic previously demonstrated to possess antioxidant properties. Two A/R models were performed on type II pneumocytes (A549 cell line), either on cells attached to culture dishes (monolayer A/R model; 3.5 h of anoxia, 30 min of re-oxygenation) or after cell detachment (suspension A/R model; 1 h of anoxia, 10 min of re-oxygenation). Ceftazidime and selective inhibitors (SOD, Tiron, L-NMMA) were added before anoxia. Free radical production was assessed by the EPR spin trapping technique. Oxygen consumption was monitored, in parallel with EPR studies, in the suspension A/R model. The production of free radical species was demonstrated by the generation of PBN-radical adducts: (a(N) = 15.2 G) in the monolayer A/R model and a six-line EPR spectrum (a(N) = 15.7 G and a(H) = 2.7 G) in the suspension A/R model. A kinetic study performed by oximetry, in parallel with EPR spectroscopy, demonstrated marked alterations of the cell respiratory function and that the free radical production started during anoxia and increased during re-oxygenation. In the suspension A/R model, the amplitude of EPR spectra were decreased upon the addition of 200 U/ml SOD (37% inhibition), 0.1 mM Tiron (67% inhibition) and 1 mM L-NMMA (43% inhibition). Addition of 1 mM ceftazidime decreased the amplitude of EPR spectra (37% inhibition) in both A/R models. Complementary in vitro EPR studies demonstrated that CAZ scavenged the hydroxyl radical (produced by the Fenton reaction). The protective effect of ceftazidime in the cell model could thus be linked to its ability to scavenge superoxide anions, nitrogen-derived species and hydroxyl radicals. [less ▲]

Interleukine-1 β regulation of transforming growth factor-β1 expression in articular chondrocytes : influence of aceclofenac

in Osteoarthritis and Cartilage (2001), 9SB

The expression of IL-1bêta, iNOS and COX-2 genes by human chondrocytes in differentially regulated by reactive oxygen species

in Clinical Rheumatology (2001), 20

The antioxidant properties of non stroidal anti-inflammatory drugs

in Osteoarthritis and Cartilage (2000), 8

Synthesis and evaluation of non-carboxylic pyridinic derivatives as cyclooxygenase inhibitors

in Mediators of Inflammation (2000), 9

Plasma Myeloperoxidase Level and Polymorphonuclear Leukocyte Activation in Horses Suffering from Large Intestinal Obstruction Requiring Surgery: Preliminary Results

in Canadian Journal of Veterinary Research = Revue Canadienne de Recherche Vétérinaire (1999), 63(2), 142-7

Myeloperoxidase (MPO) is a specific enzyme of neutrophil azurophilic granules with a strong oxidative activity. Thanks to a radioimmunoassay of equine myeloperoxidase, the authors have observed a ... [more ▼]

Myeloperoxidase (MPO) is a specific enzyme of neutrophil azurophilic granules with a strong oxidative activity. Thanks to a radioimmunoassay of equine myeloperoxidase, the authors have observed a significantly higher plasma level of MPO in horses operated for strangulation obstruction of the large intestine (n = 6) than in horses suffering from a non-strangulating displacement of the large intestine (n = 9). For the 2 groups, 3 phases were distinguished: reception (P1), intensive care (P2) and terminal phase (P3). The mean peak values of MPO for these phases were 121.6 ng/mL (P1), 168.6 ng/mL (P2), and 107.0 ng/mL (P3) for the non-strangulating group, and 242.6 ng/mL (P1); 426.0 ng/mL (P2), and 379.5 ng/mL (P3) for the strangulation group. The variations of the mean peak values of plasma MPO were significantly different between the 2 groups and between the different phases. A significant increase of the least square means of MPO was observed between P1 and P2. A significant decrease of the least square means of the number of circulating leukocytes was observed between P1 and P3. Polymorphonuclear neutrophil activation could play a major role in the pathogenesis of acute abdominal disease and endotoxic shock. [less ▲]

The Antibiotic Ceftazidime Is a Singlet Oxygen Quencher as Demonstrated by Ultra-Weak Chemiluminescence and by Inhibition of Aap Consumption

in Biochimica et Biophysica Acta (1998), 1379(1), 61-8

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 ... [more ▼]

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. [less ▲]

Effects of Perfluorocarbon Emulsions on Cultured Human Endothelial Cells

in Artificial Cells, Blood Substitutes, & Immobilization Biotechnology (1997), 25(6), 563-75

Perfluorocarbons (PFCs) and their emulsions (PFCEs) were used in organ preservation before transplantation, but not in organ perfusion. Our purpose was to achieve organ perfusion with a PFCE at room ... [more ▼]

Perfluorocarbons (PFCs) and their emulsions (PFCEs) were used in organ preservation before transplantation, but not in organ perfusion. Our purpose was to achieve organ perfusion with a PFCE at room temperature or at 37 degrees C, i. e. with oxygenation, to prevent damages related to reoxygenation after hypoxia. Therefore, we first investigated the effect of such emulsions on endothelial cells, the first cells to be in contact with the emulsion. A stem emulsion was prepared from perfluorooctyl bromide (90% w/v), emulsified with egg yolk phospholipids (2% w/v) and stabilized with a mixed fluorocarbon-hydrocarbon "molecular dowel" (1.4% w/v) (droplets of ca 0.2 micron in diameter). This emulsion was found to be stable when diluted with cell culture media or organ preservation fluids. Endothelial cells from human umbilical vein (HUVECs) were cultured in multiwell plates in M199 medium (with growth factors, 10% foetal calf serum and 5% human serum). Confluent cells were incubated overnight with 51Cr, washed and overlayed with M199 (control) or the above PFCE diluted 2x or 4x with M199 (test). After incubation, the cytotoxicity of the PFCEs was estimated by measuring 51Cr release and observing cell morphology by electron and light microscopy. The percentages of released 51Cr were identical to those of the control cells for the 2x, 3x or 4x diluted PFCEs at 4, 25 or 37 degrees C. After return to the M199 medium, the cells grew and multiplied normally. We conclude that the diluted PFCEs were devoid of cytotoxicity. The 2x diluted PFCE was however partially taken up by the cells: by microscopy, we observed intracellular PFC droplets and by density gradient analysis we found a slight increase in cellular density. The diluted PFCEs were compared to classical organ preservation solutions : HUVECs were incubated with UW (University of Wisconsin) or EC (EuroCollins) solutions at +4 and 37 degrees C (3, 17 or 24 h of incubation). The solutions were observed to be toxic to the cells under these conditions, with cell mortality after return to the M199 medium. This cytotoxicity may be attributed to the high K+ concentration of UW and EC, since similar assays performed on HUVECs with Hank's solution adjusted to 100 mM K+ showed a similar % of 51Cr release. UW and EC are therefore not acceptable as dilution media for PFCEs. [less ▲]

Inactivation of alpha(2)-Macroglobulin by Activated Human Polymorphonuclear Leukocytes.

in Mediators of Inflammation (1994), 3(2), 117-23

The proteolytic activity of trypsin releases the dye Remazol Brilliant Blue from its high molecular weight substrate, the skin powder (Hide Powder Azure, Sigma), with an increase in absorbance at 595 nm ... [more ▼]

The proteolytic activity of trypsin releases the dye Remazol Brilliant Blue from its high molecular weight substrate, the skin powder (Hide Powder Azure, Sigma), with an increase in absorbance at 595 nm. Active alpha(2)- macroglobulin (80 mug/ml) totally inhibits the proteolytic activity of trypsin (14 mug/ml) by trapping this protease. But after a 20 min incubation of alpha(2)-macroglobulin at 37 degrees C with 2 x 10(6) human polymorphonuclear leukocytes activated by N-formyl-L-methionyl-L-leucyl-L-phenylalanine (10(-7) M) and cytochalasin B (10(-8) M), 100% of trypsin activity was recovered, indicating a total inactivation of alpha(2)-macroglobuHn. Incubation with granulocyte myeloperoxidase also inactivates alpha(2)-macroglobulin. Hypochlorous acid, a by-product of myeloperoxidase activity, at a concentration of 10(-7) M also inactivates alpha(2)-macroglobulin, which indicates that an important cause of alpha(2)-macroglobulin inactivation by activated polymorphonuclear leukocytes could be the activity of myeloperoxidase. [less ▲]