Carbon disulphide absorption during xanthate reagent mixing in a gold mine concentrator.
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A xanthate reagent mixer at a gold mine concentrator was exposed to carbon disulphide by extensive skin contamination with xanthate powder and solution during the reagent mixing process. Absorption of carbon disulphide was confirmed by the detection of urinary 2-thiothiazolidine-4-carboxylic acid (TTCA). Drager colorimetric tube testing during subsequent mixing recorded a maximum concentration of at least 60 ppm carbon disulphide. An illness consisting of predominantly gastrointestinal symptoms began 20 h after the exposure. Although this may have been due to carbon disulphide toxicity this is by no means certain. The need for engineering controls, impervious protective clothing and full-face respirators with particulate and organic vapour cartridges is discussed. This episode occurred at another mine site, unrelated to Mount Isa Mines Limited. (+info)
A phosphatidylcholine-specific phospholipase C regulates activation of p42/44 mitogen-activated protein kinases in lipopolysaccharide-stimulated human alveolar macrophages.
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This study uses human alveolar macrophages to determine whether activation of a phosphatidylcholine (PC)-specific phospholipase C (PC-PLC) is linked to activation of the p42/44 (ERK) kinases by LPS. LPS-induced ERK kinase activation was inhibited by tricyclodecan-9-yl xanthogenate (D609), a relatively specific inhibitor of PC-PLC. LPS also increased amounts of diacylglycerol (DAG), and this increase in DAG was inhibited by D609. LPS induction of DAG was, at least in part, derived from PC hydrolysis. Ceramide was also increased in LPS-treated alveolar macrophages, and this increase in ceramide was inhibited by D609. Addition of exogenous C2 ceramide or bacterial-derived sphingomyelinase to alveolar macrophages increased ERK kinase activity. LPS also activated PKC zeta, and this activation was inhibited by D609. LPS-activated PKC zeta phosphorylated MAP kinase kinase, the kinase directly upstream of the ERK kinases. LPS-induced cytokine production (RNA and protein) was also inhibited by D609. As an aggregate, these studies support the hypothesis that one way by which LPS activates the ERK kinases is via activation of PC-PLC and that activation of a PC-PLC is an important component of macrophage activation by LPS. (+info)
In vitro endothelial differentiation of long-term cultured murine embryonic yolk sac cells induced by matrigel.
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The yolk sac of an early mammalian embryo contains progenitors of hematopoietic cells and vascular endothelial cells. We established a cell line, YS4, from murine embryonic yolk sac 10 years ago. The line has been successfully cultured since then. To determine whether these long-term cultured yolk sac cells still have the potential to differentiate into endothelial cells, an in vitro model of yolk sac cell differentiation into tubeforming endothelial cells was established in the present study by culturing the yolk sac cells on basement membrane proteins (Matrigel). The results indicate that upon plating onto Matrigel, YS4 cells attach quickly, align in tandem, and form a complete network of capillary structures within 12 h. By using antibodies against the known components of Matrigel in a tube formation inhibition assay, we found that extracellular matrix proteins such as laminin, collagen IV, vitronectin, and fibronectin are the most important components in the Matrigel which induce the yolk sac cells to undergo endothelial differentiation. New basement membrane proteins are also required for the endothelial differentiation process, as indicated by the fact that base membrane protein synthesis inhibitor, D609, can block the differentiation process. Furthermore, our experiments revealed the involvement of several signal transduction pathways, such as protein kinase A, C and protein tyrosine kinase in this differentiation process. (+info)
Selective mechanism-based inactivation of cytochromes P-450 2B1 and P-450 2B6 by a series of xanthates.
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Fifteen xanthates with carbon chains of different lengths or substitutions, including the antiviral compound D609 (O-tricyclo[5.2. 1.0(2,6)]dec-9-yl-dithiocarbonate), were tested for their ability to inactivate cytochromes P-450 (P-450s) 2B1 and 2B6. All of the xanthates tested were found to inactivate P-450 2B1 in a time- and concentration-dependent manner. The rates of inactivation at 30 degrees C ranged from 0.22 min-1 to 0.02 min-1. The concentrations required for half-maximal inactivation were between 2.4 and 69 microM. A general trend in the inactivation kinetics could be observed with an increasing chain length of the xanthates. Longer carbon chains resulted in slower rates of inactivation with longer half-times of inactivation and higher partition ratios. For P-450 2B1, the most effective inactivators were xanthates with substitutions of intermediate length. The best inactivator for P-450 2B1 was the C8 xanthate, with an inactivation potency (KI) of 2.4 microM, a rate of inactivation of 0.07 min-1, and a partition ratio of 4. Four xanthates were further examined for their effect on the 7-ethoxy-4-(trifluoromethyl)coumarin activity of P-450 2B6. The C8 xanthate was again the most effective inactivator, with a KI of 1 microM. Although the KI values were generally lower than those found with P-450 2B1, the rates of inactivation for P-450 2B6 with the various xanthates were 3- to 5-fold slower. In addition, the isozyme selectivity of xanthates was tested with P-450s 2E1, 1A1, 3A2, 3A4, 2C9, and 2D6. P-450 2E1 was inactivated by xanthates at concentrations 15- to 100-fold higher than those required to inactivate either P-450 2B1 or 2B6. P-450 1A1 was not inactivated by xanthates. However, all of the xanthates tested were able to inhibit the enzymatic activity of P-450 1A1 to a different extent, depending on the length of the xanthate carbon chain. Virtually no inactivation of P-450s 2D6 or 2C9 was seen, except that C8 and D609 were inhibitory at high concentrations (0.2-0.6 mM). None of the xanthates studied had any effect on the activities of P-450s 3A2 or 3A4. (+info)
Nuclei contain two differentially regulated pools of diacylglycerol.
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A number of recent studies have highlighted the presence of a nuclear pool of inositol lipids [1] [2] that is regulated during progression through the cell cycle [1] [3], differentiation [1] [2] and after DNA damage [2], suggesting that a number of different regulatory pathways impinge upon this pool of lipids. It has been suggested that the downstream consequence of the activation of one of these nuclear phosphoinositide (PI) regulatory pathways is the generation of nuclear diacylglycerol (DAG) [1] [3] [4], which is important in the activation of nuclear protein kinase C (PKC) [5] [6] [7]. Activation of PKC in turn appears to regulate the progression of cells through G1 and into S phase [4] and through G2 to mitosis [3] [8] [9] [10] [11]. Although the evidence is enticing, there is as yet no direct demonstration that nuclear PIs can be hydrolysed to generate nuclear DAG. Previous data in murine erythroleukemia (MEL) cells have suggested that nuclear phosphoinositidase Cbeta1 (PIC-beta1) activity is important in the generation of nuclear DAG. Here, we demonstrate that the molecular species of nuclear DAG bears little resemblance to the PI pool and is unlikely to be generated directly by hydrolysis of these inositol lipids. Further, we show that there are in fact two distinct subnuclear pools of DAG; one that is highly disaturated and mono-unsaturated (representing more than 90% of the total nuclear DAG) and one that is highly polyunsaturated and is likely to be derived from the hydrolysis of PI. Analysis of these pools, either after differentiation or during cell-cycle progression, suggests that the pools are independently regulated, possibly by the regulation of two different nuclear phospholipase Cs (PLCs). (+info)
Mutagenicity tests of 4-phenyl-1,3-dithia-2-thioxo-cyclopent-4-ene.
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The mutagenicity of 4-phenyl-1,3-dithia-2-thioxo-cyclopent-4-ene (DT827B) was examined in reverse mutation tests using Salmonella typhimurium and Escherichia coli, in the chromosomal aberration test with Chinese hamster ovary (CHO) cells, and in the micronucleus test using mice bone-marrow. In reverse mutation assay on DT827B according to Ames' method, DT827B was not mutagenic to S. typhimurium or E. coli when tested in dimethylsulfoxide to the limit of its solubility where precipitation occurred. In chromosomal aberration assay using CHO cells, DT827B was not clastogenic to induce structural chromosomal aberration but capable of inducing polyploidy. In micronucleus test, DT827B did not show micronucleus-inducing potential at the maximum dose. In conclusion of the three mutagenicity studies, DT827B was considered to cause no mutagenicity under the conditions used in the present experiments except the increase in polyploidy, which probably is due to a toxic effect of the compound. (+info)
Toluene-3,4-dithiol analysis of blood for assessing carbon disulfide exposure.
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Carbon disulfide is a neurotoxic compound used in the production of viscose rayon, and is a major decomposition product of dithiocarbamates used in industry, agriculture, and medicine. Methods used currently for assessing exposure to CS2 are limited in their ability to evaluate cumulative exposures and provide useful information for relatively short periods of time after exposure has ended. The present investigation evaluates a method for monitoring CS2 exposure that consists of cleaving the thiocarbonyl function of free CS2 or certain CS2-generated modifications on proteins using toluene-3,4-dithiol. The resulting toluene trithiocarbonate product is then quantified using reverse-phase high-performance liquid chromatography. The sensitivity, dose response, kinetics and specificity of this biomarker in blood were examined in rats administered CS2 by inhalation, intraperitoneal injection, or gavage for acute through subchronic periods. Dithiol reactive functions in plasma and hemolysate demonstrated a linear dose response over a wide range of exposure levels, were dependent upon the duration of exposure, and appeared to have an appropriate sensitivity for evaluating occupational levels of exposure. Elimination rates of dithiol reactive functions may also be dependent upon exposure duration and exhibit different kinetics for plasma and hemolysate suggesting that elimination rates may be useful for estimating cumulative exposure and intervals between exposure and sample procurement. Dithiol analysis, used in conjunction with previously established erythrocyte protein cross-linking biomarkers, may provide a means to characterize the internal dose of CS2 resulting from acute through chronic periods, and may provide insight into the level of CS2-mediated covalent protein modifications occurring within the nervous system. (+info)
Role of sphingolipids in the transport of prosaposin to the lysosomes.
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Prosaposin is the precursor of four lysosomal saposins that promote the degradation of glycosphingolipids (GSLs) by acidic hydrolases. GSLs contain a hydrophobic ceramide moiety, which acts as a membrane anchor, and a hydrophilic oligosaccharide chain that faces the lumen of the Golgi apparatus and extracellular spaces. By using fumonisin B1, PDMP and D609, we tested the hypothesis that sphingolipids mediate the transport of prosaposin to the lysosomes. Fumonisin B1 interferes with the synthesis of ceramide, PDMP blocks the formation of glucosylceramide and D609 blocks the formation of sphingomyelin. Fumonisin B1 produced a 59;-85% decrease in the density of gold particles in the lysosomes of CHO and NRK cells immunolabeled with anti-prosaposin antibody, and a 55% reduction in the lysosomes of CHO cells stably transfected with an expression vector containing a human prosaposin cDNA. To examine whether the mannose 6-phosphate receptor pathway was affected by this treatment, NRK and CHO cells treated or not with fumonisin B1 were labeled with anti-cathepsin A antibody. The results showed no significant differences in labeling of the lysosomes, suggesting that the effect of fumonisin B1 was specific. When fumonisin B1 and D609 were added to the media of transfected CHO cells, a decrease in immunofluorescence with anti-prosaposin antibody was observed by confocal microscopy. PDMP did not cause any reduction in immunoreactivity, indicating that sphingolmyelin appears to be involved in this process. In conclusion, our data support the hypothesis that sphingolipids, possibly sphingomyelin, are involved in the transport of prosaposin to the lysosomes. (+info)