Effects of COX-2 inhibitors on ROS produced by Chlamydia pneumoniae-primed human promonocytic cells (THP-1)

Chronic inflammation through foam cells and macrophages is important in atherosclerosis development, and can be considered as therapeutic targets. Cyclooxygenase and NADPH-oxidase were expressed within atherosclerotic lesions. Reactive oxygen species produced by NADPH oxidase were found to trigger the cyclooxygenase-2 expression. The effects of preferential COX-2 inhibitors on ROS produced by Chlamydia-primed human monocytes (THP-1 cells) were evaluated by fluorescence, chemiluminescence, oxymetry, and EPR spin trapping. Fluorescence assays showed an increased production of ROS with Chlamydia versus cells primed by 10(-8)M PMA. COX-2 inhibitors inhibited in a dose-dependent manner the luminol-enhanced CL while ibuprofen and diclofenac increased the chemiluminescence response. By EPR spin trapping, COX-2 inhibitors, ibuprofen, and diclofenac, exhibited a dose-dependent inhibiting effect (10 and 100muM) on the EPR signal appearance. Our cell model combining EPR, chemiluminescence, and oxymetry appeared relevant to study the modulating effects of preferential COX-2 inhibitors on the cell oxidant activity and chronic inflammatory diseases.     

Structural characterization of diC14-amidine, a pH-sensitive cationic lipid used for transfection

The structure of N-t-butyl-N'-tetradecyl-3-tetradecylaminopropionamidine (diC(14)-amidine) cationic vesicles, used for transfection, was investigated at different pH values and ionic strengths, through the analysis of the electron spin resonance (ESR) spectra of spin labels. Phospholipid derivatives, spin labeled at the 5th and 16th C-atoms along the hydrocarbon chain, incorporated in diC(14)-amidine bilayers, show that the bilayer structure is highly sensitive to the pH value of the medium, due to the two titratable groups present in the amphiphile. Compared with samples at higher pH values, the double charged diC(14)-amidine at pH 3 presents a rather non-organized bilayer gel phase, and a much lower gel-fluid temperature transition, in accord with a strong headgroup electrostatic repulsion. In addition, the structure was found to be highly dependent on the ionic strength of the medium. However, pH 3 diC(14)-amidine bilayer, in the fluid phase, was found to be slightly more closely packed than those at pH 7.4 or 9.0, which are less charged. Parallel to that, the larger isotropic hyperfine splitting measured for nitroxides in the center of the pH 3 diC(14)-amidine bilayer suggests a higher membrane polarity for the highly charged low pH sample.     

Comparative analysis of QTLs affecting domestication traits between two domesticated x wild pearl millet ( Pennisetum glaucum L., Poaceae) crosses

Comparative mapping of Quantitative trait loci (QTLs) involved in domestication of adaptative syndrome traits of pearl millet was realized at the intra-specific level using two F(2) populations derived from domesticated ( Pennisetum glaucum ssp. glaucum) x wild ( Pennisetum glaucum ssp. monodii) crosses. The two domesticated parents analyzed differ in their geographical origins, agronomic characteristics and life cycles. In both populations, two regions of the genome were identified on linkage groups 6 and 7, that controlled most of the key morphological differences. The importance of these two linkage groups reveals their central role both in the developmental control of spikelet structure and in the domestication process of this crop. In contrast, QTLs involved in traits that are components of yield and measure differences in resource allocation (such as the shape of the spike, the number of spikes per plant and plant height) show a low level of correspondence among our two crosses. The results of the comparative mapping between cereals, although preliminary, reveal that genes involved in seed-shattering could correspond in maize, rice and sorghum. The evolutionary significance of our results, and especially the relationships between genome organization and cereal domestication, are discussed. The potential use of these results in pearl millet genetic-resources enhancement are presented.     

Endotoxaemia, production of tumour necrosis factor alpha and polymorphonuclear neutrophil activation following strenuous exercise in humans

To examine whether endotoxaemia accompanying long-term, strenuous physical exercise is involved in exercise-induced increase in plasma tumour necrosis factor alpha (TNF-alpha) concentration and polymorphonuclear neutrophil (PMN) activation, 14 male recreational athletes [mean age 28 (SEM 1) years] were studied. Exercise consisted of a 1.5-km river swim, a 40-km bicycle race, and a 10-km road race. Mean time to complete the race was 149.8 (SEM 4.8) min. The plasma concentrations of granulocyte myeloperoxidase (MPO) and TNF-alpha were significantly higher than baseline values immediately and 1 h after exercise (P<0.001). Both variables returned to pre-race levels the day after exercise. Marked, transient decreases in plasma concentrations of anti-lipopolysaccharide (LPS) immunoglobulin G (IgG) and immunoglobulin M (IgM) antibodies directed against a panel of selected smooth gram-negative LPS were observed after the race, reaching in most cases minimal values in the blood sample drawn immediately following the completion of the triathlon. There was no significant correlation between the magnitude of PMN activation, as assessed by the increase in plasma concentrations of MPO, and the humoral markers of endotoxaemia and TNF-alpha. An inverse, highly significant relationship between the increase in plasma TNF-alpha concentrations and the changes in circulating anti-LPS IgM antibodies concentrations was observed (r = -0.7; P<0.01). These findings suggest that exercise-induced endotoxaemia was involved in the release of TNF-alpha, that the magnitude of the TNF-alpha response to exercise was down-regulated by anti-LPS antibodies of the IgM class, and that the production of TNF-alpha and endotoxaemia did not seem to play a role in the activation of circulating PMN in the exercising subjects.     

Daily cycles for emission of urticating hairs from the pine processionary caterpillar (Thaumetopoea pityocampa S.) and the brown tail moth (Euproctis chrysorrhoea L) (Lepidoptera) in laboratory conditions

Summary In laboratory conditions, urticating hairs from the pine processionary caterpillar (Thaumetopoea pityocampa S.) and from the brown tail moth (Euproctis chrysorrhoea L.) are detectable in the air using an apparatus designed for the capture of airborne microorganisms and pollen research studies. The hairs produced by the caterpillars of these two species are distributed either via air currents or moths (only forEuproctis). Daily cycles of hair emission were observed and were in relation with locomotion and feeding activities of the caterpillars and with flying and reproductive activities of moths.     

Effects of methylprednisolone on exercise-induced increases of plasma levels of polymorphonuclear elastase and myeloperoxidase in man. Preliminary results

The aim of the present study was to verify whether a single oral dose of methylprednisolone could modulate the exercise-induced release of polymorphonuclear neutrophil (PMN) elastase and myeloperoxidase. Four healthy, male subjects were submitted to a 20 min downhill run (-20%) at 60% VO(2) max, 3 h after oral absorption of a placebo or a single dose of 32 mg methylprednisolone. A marked neutrophilia (+103% of basal PMN count; p < 0.02) was observed 3 h after methylprednisolone ingestion. During both exercise trials, placebo and methylprednisolone, PMN counts were increased by 46% and 19% (p < 0.05), respectively. The running test caused marked and significant (p < 0.05) increases in plasma myeloperoxidase concentration (MPO). The magnitude of MPO changes was the same in the two trials (+110%). Exercise also resulted in significant changes in plasma elastase concentration (EL) in both experimental conditions (placebo: +104%, p < 0.05; methylprednisolone: +338%, p < 0.005). Plasma elastase levels reached at the end of exercise on methylprednisolone were significantly higher than after placebo (p < 0.05). A significant relationship was found between EL and PMN in methylprednisolone trial only (r = 0.72; l0 < 0.005). These results showed that the transient exercise-induced release of elastase and myeloperoxidase were not decreased by methylprednisolone.     

Three-Dimensional Immunoelectron Microscopy of Scorpion Hemocyanin Labeled with a Monoclonal Fab Fragment

An immunocomplex of the 4 × 6-meric hemocyanin of the scorpion Androctonus australis with the monoclonal Fab fragment L104 was reconstructed from electron micrographs of a negatively stained specimen, using the double-carbon-layer technique. The resulting structure enables a clear visualization of the Fab fragments bound to the four copies of the Aa6 subunit and directly confirms a previous localization of the L104 epitope deduced from two-dimensional image processing. Despite a strong flattening effect produced by the negative-staining technique the orientations of the Fab fragments are well characterized. Moreover, the observation of a central hole within the elbow bends of the Fab fragments provides information about the disposition of the Fabs around their main axis.     

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

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.     

Arthropod hemocyanin structure: isolation of eight subunits in the scorpion.

Eight immunologically pure subunits were isolated from Androctonus australis hemocyanin. Antisera specific against each of these were prepared. Two subunits associate to form a heteroöligomer which is probably one of the bridges visible in electron microscopy of the native molecule. There is no cross-reactivity between native subunits. When denatured by 7 m urea, the specific antigenic determinants disappear and a broader specificity appears. This is probably due to the unfolding of the molecule which unmasks deeply buried antigenic sites.