Acute metobromuron poisoning with severe associated methemoglobinemia. Identification of four metabolites in plasma and urine by LC-DAD, LC-ESI-MS, and LC-ESI-MS-MS.
(65/1809)
A case of self poisoning with metobromuron, a urea derivative used as a herbicide, is reported. Severe methemoglobinemia observed at the admission (80%) disappeared only at day 11, and hemolysis appeared at day 4 and decreased slowly to day 12. Metobromuron was analyzed by liquid chromatography with diode-array detection. Initial plasma concentration and elimination half-life were 4.9 mg/L and 5 h, respectively. Several metabolites were also detected, and four of those were identified by liquid chromatography-electrospray mass spectrometry. Normetobromuron, bromophenylurea, and bromoacetanilide were detected in plasma, but only N-methyl bromophenylurea was detected in urine. Bromoacetanilide probably results from acetylation of the intermediate bromoaniline. Methemoglobinemia could result from metabolization of metobromuron to bromoaniline and bromoacetanilide. (+info)
Secretory phospholipases A2 induce beta-glucuronidase release and IL-6 production from human lung macrophages.
(66/1809)
Secretory phospholipases A2 (sPLA2s) are a group of extracellular enzymes that release fatty acids at the sn-2 position of phospholipids. Group IIA sPLA2 has been detected in inflammatory fluids, and its plasma level is increased in inflammatory diseases. To investigate a potential mechanism of sPLA2-induced inflammation we studied the effect of group IA (from cobra venom) and group IIA (human synovial) sPLA2s on human macrophages. Both sPLA2s induced a concentration- and Ca2+-dependent, noncytotoxic release of beta-glucuronidase (16.2 +/- 2.4% and 13.1 +/- 1.5% of the total content with groups IA and IIA, respectively). Both sPLA2s also increased the rate of secretion of IL-6 and enhanced the expression of IL-6 mRNA. Preincubation of macrophages with inhibitors of the hydrolytic activity of sPLA2 or cytosolic PLA2 did not influence the release of beta-glucuronidase. Incubation of macrophages with p-aminophenyl-mannopyranoside-BSA (mp-BSA), a ligand of the mannose receptor, also resulted in beta-glucuronidase release. However, while preincubation of macrophages with mp-BSA had no effect on beta-glucuronidase release induced by group IIA sPLA2, it enhanced that induced by group IA sPLA2. A blocking Ab anti-mannose receptor inhibited both mp-BSA- and group IIA-induced beta-glucuronidase release. Taken together, these data indicate that group IA and IIA sPLA2s activate macrophages with a mechanism independent from their enzymatic activities and probably related to the activation of the mannose receptor or sPLA2-specific receptors. The secretion of enzymes and cytokines induced by sPLA2s from human macrophages may play an important role in inflammation and tissue damage associated with the release of sPLA2s. (+info)
Expression of ryanodine receptor RyR3 produces Ca2+ sparks in dyspedic myotubes.
(67/1809)
Discrete, localized elevations of myoplasmic [Ca2+], Ca2+ 'sparks', were readily detected using the fluorescent Ca2+ indicator fluo-3 and laser scanning confocal microscopy in 'dyspedic' 1B5 myotubes, i.e. myotubes which do not express ryanodine receptors (RyRs), transduced with virions containing cDNA for RyR type 3 that were saponin permeabilized to allow dye entry. Ca2+ sparks were never observed in non-transduced RyR null myotubes. The spatial locations of sparks observed in permeabilized myotubes roughly corresponded to regions of RyR protein expression in the same myotube as detected after subsequent fixation and antibody staining. Permeabilized RyR3-transduced myotubes exhibited similar punctate peripheral RyR3 protein immunohistochemical patterns as myotubes fixed before permeabilization indicating that permeabilization did not affect the structural organization of the triad. Ca2+ sparks, recorded in line scan mode, in permeabilized myotubes expressing RyR3 exhibited mean amplitudes (change in fluorescence/mean fluorescence, DeltaF/F: 1.20 +/- 0.04) and temporal rise times (10-90%; 6.31 +/- 0.12 ms) similar to those of sparks recorded in permeabilized frog skeletal muscle fibres (0.98 +/- 0.01; 6.11 +/- 0.07, respectively) using the same confocal system. Spatial extent and temporal duration of the Ca2+ sparks were approximately 40% larger in the RyR3-expressing myotube cultures than in frog fibres. Ca2+ sparks recorded in line scan mode often occurred repetitively at the same spatial location in RyR3-expressing myotubes. Such repetitive events were highly reproducible in amplitude and spatio-temporal properties, as previously observed for repetitive mode sparks in frog skeletal muscle. Ca2+ sparks recorded in xy mode were frequently compressed in the y (slower scan) direction compared to the x direction. This asymmetry was reproduced assuming spatially symmetric events having the time course of Ca2+ sparks recorded in line scan (xt) mode. These expression studies demonstrate that the presence of RyR3 is sufficient for the production of Ca2+ sparks in a skeletal muscle system lacking the expression of any other RyR isoform. (+info)
Two Ca2+ entry pathways mediate InsP3-sensitive store refilling in guinea-pig colonic smooth muscle.
(68/1809)
Sarcolemma Ca2+ influx, necessary for store refilling, was well maintained, over a wide range (-70 to + 40 mV) of membrane voltages, in guinea-pig single circular colonic smooth muscle cells, as indicated by the magnitude of InsP3-evoked Ca2+ transients. This apparent voltage independence of store refilling was achieved by the activity of sarcolemma Ca2+ channels some of which were voltage gated while others were not. At negative membrane potentials (e.g. -70 mV), Ca2+ influx through channels which lacked voltage gating provided for store refilling while at positive membrane potentials (e.g. +40 mV) voltage-gated Ca2+ channels were largely responsible. Sarcolemma voltage-gated Ca2+ currents were not activated following store depletion. Removal of external Ca2+ or the addition of the Ca2+ channel blocker nimodipine (1 microM) inhibited store refilling, as assessed by the magnitude of InsP3-evoked Ca2+ transients, with little or no change in bulk average cytoplasmic Ca2+ concentration. One hypothesis for these results is that the store may refill from a high subsarcolemma Ca2+ gradient. Influx via channels, some of which are voltage gated and others which lack voltage gating, may permit the establishment of a subsarcolemma Ca2+ gradient. Store access to the gradient allows InsP3-evoked Ca2+ signalling to be maintained over a wide voltage range in colonic smooth muscle. (+info)
Nutrient modulation of polarized and sustained submembrane Ca2+ microgradients in mouse pancreatic islet cells.
(69/1809)
The intracellular calcium concentration ([Ca2+]i) near the plasma membrane was measured in mouse pancreatic islet cells using confocal spot detection methods. Whereas small cytosolic Ca2+ gradients were observed with 3 mM glucose, a steeper sustained gradient restricted to domains beneath the plasma membrane (space constant, 0.67 micrometer) appeared with 16.7 mM glucose. When the membrane potential was clamped with increasing K+ concentrations (5, 20 and 40 mM), no [Ca2+]i gradients were observed in any case. Increasing glucose concentration (0, 5 and 16.7 mM) in the presence of 100 microM diazoxide, a K+ channel opener, plus 40 mM K+ induced steeper [Ca2+]i gradients, confirming the role of membrane potential-independent effects of glucose. Prevention of Ca2+ store refilling with 30 microM cyclopiazonic acid (CPA) or blockade of uniporter-mediated Ca2+ influx into the mitochondria with 1 microM carbonyl cyanide m-chlorophenyl hydrazone (CCCP) or 1 microM Ru-360 significantly reduced the steepness of the 16.7 mM glucose-induced [Ca2+]i gradients. Measured values of [Ca2+]i reached 6.74 +/- 0.67 microM at a distance of 0.5 micrometer from the plasma membrane and decayed to 0.27 +/- 0.03 microM at a distance of 2 micrometer. Mathematically processed values at 0.25 and 0 micrometer gave a higher [Ca2+]i, reaching 8.18 +/- 0.86 and 10.05 +/- 0.98 microM, respectively. The results presented indicate that glucose metabolism generates [Ca2+]i microgradients, which reach values of around 10 microM, and whose regulation requires the involvement of both mitochondrial Ca2+ uptake and endoplasmic reticulum Ca2+ stores. (+info)
Description of Sphingomonas xenophaga sp. nov. for strains BN6T and N,N which degrade xenobiotic aromatic compounds.
(70/1809)
The taxonomic position of two bacterial strains, BN6T and N,N, with the ability to degrade xenobiotic aromatic compounds (naphthalenesulfonates or N,N-dimethylaniline) was investigated. The 165 rRNA gene sequence, the G+C content of the DNA (62-63 mol%) and the detection of ubiquinone Q-10, 2-hydroxymyristic acid and the sphingoglycolipid present clearly placed the two strains into the genus Sphingomonas. Both strains are representatives of one species according to the level of DNA relatedness (70.7%). The strains could be separated from all validly described taxa of the genus Sphingomonas, according to the 16S rRNA gene sequence (the highest sequence similarity observed was 96 % to Sphingomonas yanoikuyae), the pattern of the polar lipids and physiological characteristics. Therefore, the new species Sphingomonas xenophaga is proposed to accommodate strains BN6T (= DSM 6383T) and N,N (= DSM 8566). (+info)
Three distinct types of Ca(2+) waves in Langendorff-perfused rat heart revealed by real-time confocal microscopy.
(71/1809)
Although Ca(2+) waves in cardiac myocytes are regarded as arrhythmogenic substrates, their properties in the heart in situ are poorly understood. On the hypothesis that Ca(2+) waves in the heart behave diversely and some of them influence the cardiac function, we analyzed their incidence, propagation velocity, and intercellular propagation at the subepicardial myocardium of fluo 3-loaded rat whole hearts using real-time laser scanning confocal microscopy. We classified Ca(2+) waves into 3 types. In intact regions showing homogeneous Ca(2+) transients under sinus rhythm (2 mmol/L [Ca(2+)](o)), Ca(2+) waves did not occur. Under quiescence, the waves occurred sporadically (3.8 waves. min(-1) x cell(-1)), with a velocity of 84 microm/s, a decline half-time (t(1/2)) of 0.16 seconds, and rare intercellular propagation (propagation ratio <0.06) (sporadic wave). In contrast, in presumably Ca(2+)-overloaded regions showing higher fluorescent intensity (113% versus the intact regions), Ca(2+) waves occurred at 28 waves x min(-1) x cell(-1) under quiescence with a higher velocity (116 microm/s), longer decline time (t(1/2) = 0.41 second), and occasional intercellular propagation (propagation ratio = 0.23) (Ca(2+)-overloaded wave). In regions with much higher fluorescent intensity (124% versus the intact region), Ca(2+) waves occurred with a high incidence (133 waves x min(-1) x cell(-1)) and little intercellular propagation (agonal wave). We conclude that the spatiotemporal properties of Ca(2+) waves in the heart are diverse and modulated by the Ca(2+)-loading state. The sporadic waves would not affect cardiac function, but prevalent Ca(2+)-overloaded and agonal waves may induce contractile failure and arrhythmias. (+info)
Evaluation of cytotoxicity, cell proliferation, and genotoxicity induced by p-cresidine in hetero- and nullizygous transgenic p53 mice.
(72/1809)
The heterozygous p53 knockout mouse is being used as a short-term alternative model for carcinogenicity screening of chemicals. In most cases, these mice develop tumors within 6 months of exposure to genotoxic carcinogens. The bladder and liver carcinogen, p-cresidine, is recommended as a positive control chemical for these assays. To evaluate early effects of p53 deficiency on bladder and liver histopathology and genotoxicity induced by p-cresidine, we treated 4-week-old heterozygous and nullizygous p53 male mice with p-cresidine by gavage (100, 200, 400, and 800 mg/kg/day) 5 days/week for 7 weeks. Tissue sections were prepared for hematoxylin-eosin staining and immunohistochemistry for PCNA protein or 3'-OH DNA fragments to assess cell proliferation and apoptosis, respectively. Blood and bone marrow were examined for methemoglobin and micronuclei in polychromatic erythrocytes (MN-PCE), respectively. Individual cell necrosis of the bladder transitional epithelium was evident in both p53 heterozygous and nullizygous mice at all doses. In addition, diffuse hyperplasia of the bladder epithelium was observed at 400 and 800 mg/kg in both genotypes. In the liver, both genotypes exhibited similar increases in hepatocyte apoptosis (10-fold increase) and cell proliferation (20-fold increase) at 800 mg/kg/day. Methemoglobin levels were increased 6-fold in both genotypes at 800 mg/kg. Background MN-PCE rates were similar in both genotypes and there were no treatment-related increases. Also, no point mutations were observed in codon 12 of the c-Ha-ras gene from urinary bladder DNA from p-cresidine treated p53 mice. These results suggest that loss of p53 allele(s) in mice does not influence the early markers of carcinogenic activity induced by subchronic treatment with p-cresidine. Increased tumor susceptibility associated with a reduction in p53 dosage may be dependent on neoplastic progression rather than initiation and promotional events elicited by p-cresidine. (+info)