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2005-01-01
Photodynamic therapy (PDT) is an established anticancer modality and hypericin is a promising photosensitizer for the treatment of bladder tumors. We show that exposure of bladder cancer cells to hypericin PDT leads to a rapid rise in the cytosolic calcium concentration which is followed by the generation of arachidonic acid by phospholipase A2 (PLA2). PLA2 inhibition significantly protects cells from the PDT-induced intrinsic apoptosis and attenuates the activation of p38 MAPK, a survival signal mediating the up-regulation of cyclooxygenase-2 that converts arachidonic acid into prostanoids. Importantly, inhibition of p38α MAPK blocks the release of vascular endothelial growth factor and suppresses tumor-promoted endothelial cell migration, a key step in angiogenesis. Hence, targeted inhi...
http://www4.gu.edu.au:8080/adt-root/public/adt-QGU20060810.144732
The Langendorff perfused murine heart has become an increasingly important research model in cardiovascular physiology and pharmacology. However, the model remains relatively poorly characterised when compared with the widely employed rat preparation. The purpose of the research within this thesis was initially two-fold: 1) to characterise the functional and substrate-dependent properties of the murine model; and 2) to characterise the relationships between glycolysis, ischaemic tolerance and adenosine-mediated cardioprotection in the mouse. Initial studies, confirmed by simultaneous/subsequent work in other laboratories, revealed the frequent occurrence of regular cyclic oscillations in contractile function and coronary flow in glucose-perfused isovolumically contracting hearts. This phenomenon (labelled 'cycling') was unaltered by inhibition of ?-adrenergic receptors, prostaglandins, and nitric oxide synthase. However, A1/A2 adenosine receptor agonism did abolish the oscillations in flow and reduced contractile oscillations by 50%. Importantly, cycling was eliminated by addition of 50 IU/l insulin to perfusion fluid, or provision of 5 mM pyruvate as a co-substrate with glucose. These data suggest that functional 'cycling' in glucose-perfused murine hearts likely occurs as a result of a mismatch between substrate metabolism (energy supply) and myocardial energy demand. It may be that glycolysis with exogenous glucose is insufficient to ensure appropriate matching of myocardial energy supply and demand. For this reason, it is advisable to employ a co-substrate such as pyruvate in studies of murine hearts. Further studies performed within this thesis generally employ this co-substrate addition. Addition of pyruvate as co-substrate removes 'cycling' but is also known to inhibit/modify glycolysis, which may affect ischaemic tolerance and/or cardioprotection mediated by adenosine. Experiments throughout this thesis demonstrated that pyruvate-perfusion improved tolerance to both ischaemia (delayed time to onset of ischaemic contracture; TOC) and reperfusion (reduced diastolic dysfunction and cell death). The delay in TOC as a result of pyruvate-perfusion also suggests that contracture is not solely influenced by anaerobic glycolysis (as outlined in current paradigms). To test the relevance of glycolysis to ischaemic injury hearts were subjected to various forms of glycolytic inhibition. Glycolysis was inhibited by use of 10 mM pyruvate, (iodoacetic acid) IAA treatment, and glycogen depletion by pre-ischaemic substrate-free perfusion (all groups employing pyruvate as sole-substrate). Each form of glycolytic modification resulted in significant delays in TOC, in complete contrast to findings from other models and species. Glycogen depletion also reduced the peak level of contracture. These findings indicate that the mouse is either unique in terms of substrate metabolism and mechanisms of contracture (an unlikely possibility), or raise serious questions regarding current models of contracture development during ischaemia (theorised to be delayed by prolonging anaerobic glycolysis). Modification of glycolysis also altered post-ischaemic outcome, with pyruvate perfusion and glycogen depletion both enhancing functional recoveries. However, IAA treated hearts, despite near-identical ischaemic tolerance (ie contracture development) to pyruvate-perfused hearts, displayed very poor functional recovery, which was below that for all other groups. These data clearly reveal that blocking glycolysis improves tolerance to ischaemia (as evidenced by reduced contracture), provide evidence of dissociation of ischaemic injury or contracture from post-ischaemic recovery, and confirm the key importance of glycolysis in enhancing recovery from ischaemia. Since tolerance to ischaemia/reperfusion was shown to be glycolysis dependent, and since it has been theorised that adenosine protects hearts through modulating glycolysis, the relationships between glycolytic inhibition and adenosine-mediated cardioprotection was tested. In a number of studies, exogenously applied adenosine was shown to protect both glucose- and pyruvate-perfused hearts (supporting no dependence of adenosinergic protection on glycolysis). However, to more equivocally test the role of glycolysis effects of IAA were studied and were shown to markedly limit protection with adenosine. The effects of adenosine during ischaemia were abolished by IAA treatment, and effects on post-ischaemic recovery were reduced (but not eliminated). Similar results were acquired for protection with endogenous adenosine (using iodotubercidin to block adenosine phosphorylation). Collectively, these data reveal that adenosinergic protection during ischaemia depends entirely upon glycolysis while protection during reperfusion likely involves glycolysis dependent and independent processes. However, glycolysis is required for full recovery of function during reperfusion. Further studies assessed the involvement of glycolysis in cardioprotection afforded by transgenic A1 adenosine receptor (A1AR) overexpression. It was found that pyruvate-perfusion provided the same protection as A1AR overexpression, and the two responses (to pyruvate and A1AR overexpression) were not additive. Thus, it is probable that common mechanisms are targeted in both responses (likely glycolysis). Finally, the effects of adenosine and pyruvate on oxidant injury were studied, testing whether interactions between adenosine and pyruvate observed in prior work within this thesis could be explained by alterations in anti-oxidant responses. It was found that adenosine has quite profound anti-oxidant responses in glucose-perfused hearts, with very selective effects on markers of damage. Pyruvate also had some anti-oxidant effects but interestingly it reduced the anti-oxidant effects of adenosine. In conclusion, the work entailed within this thesis demonstrates that the isolated mouse heart model may possess unique properties and should be further characterised by potential users in order to improve its utility, and the reliability of experimental findings (chiefly when studying ischaemia-reperfusion). Other work within thesis demonstrates that modification of glycolysis is important in dictating recovery from ischaemia-reperfusion, and also impacts on adenosine-mediated protection (principally but not exclusively during ischaemia itself). The manner in which glycolysis is modified and contributes to protection remains unclear. Publisher: Griffith University. School of Health Science Language: en Rights: http://www.gu.edu.au/disclaimer.html); Copyright Benjamin Daniel Hack
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http://ro.uow.edu.au/theses/602
The Gram positive bacterium Streptococcus pyogenes (group A streptococcus; GAS) is the major etiological agent of a variety of skin and mucosal infections in humans. Whilst the majority of GAS infection results in only mild, uncomplicated disease, the migration of GAS from superficial to deep tissue sites can result in invasive infection.In recent years, there has been a resurgence in severe GAS disease, however, the details of GAS pathogenesis have yet to be fully elucidated. Increasingly, subversion of the host plasminogen activation system is being implicated in the virulence of S. pyogenes. GAS display receptors for the human zymogen plasminogen on the cell surface, one of which is the plasminogen-binding group A streptococcal M-like protein (PAM). PAM has been implicated in the pathogenesis of certain GAS isolates, but the mechanism of plasminogen binding by PAM, and the role of this interaction in the pathogenesis of GAS, requires further investigation. Thus, the focus of this thesis has been to characterise plasminogen binding by PAM and a number of naturally occurring PAM variants.Characterisation of PAM genes from 13 GAS isolates revealed that whilst thesemolecules are highly conserved in the c and d repeat domains, they display significant variation within the plasminogen binding repeat motifs (a1/a2). Percent identity to the prototype PAM a1/a2 repeat sequence ranged from 52% to 100% amongst the variants studied here. No correlation was seen between the presence of a PAM gene, or variationwithin the sequence of PAM, and site of GAS isolation. In order to determine theimpact of sequence variation on protein function, recombinant proteins representing six naturally occurring variants of PAM, together with a recombinant M1 protein were expressed and purified. Equilibrium dissociation constants for the interaction of PAMvariants with biotinylated glu-plasminogen ranged from 1.58 nM to 7.58 nM. Effective concentrations of prototype PAM required for 50% inhibition of plasminogen binding to immobilised PAM variants ranged from 0.34 nM to 22.06 nM. These results suggest that while variation in the a1/a2 region of the PAM protein does affect the comparative affinity of PAM variants, the functional capacity to bind plasminogen at physiologicallyrelevant concentrations is conserved. Additionally, a potential role for the a region of PAM in eliciting a protective immune response was investigated using a mouse model for GAS infection. The a1 region of PAM was found to protect immunised mice challenged with a homologous PAM-positive GAS strain. These data suggest a link between selective immune pressure against the plasminogen-binding repeats and thefunctional conservation of the binding domain in PAM variants. Site-directed mutagenesis of full length PAMNS13 protein from an invasive GAS isolatewas undertaken to assess the contribution of residues in the a1 and a2 repeat domains to plasminogen binding function. Mutagenesis to alanine of key plasminogen binding site lysine residues in the a1 and a2 repeats (Lys98 and Lys111) did not abrogate plasminogenbinding by PAM, nor did additional mutagenesis of Arg101, His102 and Glu104, which have previously been implicated in plasminogen binding by PAM. Plasminogen binding was only abolished with the additional mutagenesis of Arg114 and His115 to alanine. Furthermore, mutagenesis of both arginine (Arg101 and Arg114) and histidine (His102 and His115) residues abolished interaction with plasminogen despite the presence of Lys98 and Lys111 in the binding repeats. This study shows for the first time that residues Arg101, Arg114, His102 and His115 in both the a1 and a2 repeat domains of PAM canmediate high affinity plasminogen binding. These data suggest that highly conserved arginine and histidine residues may compensate for variation elsewhere in the a1 and a2 plasminogen binding repeats, and may explain the maintenance of high affinity plasminogen binding by naturally occurring variants of PAM. Initial sequence characterisation of PAM variants in this study revealed aphylogenetically distinct PAM variant, PAMNS88.2. This variant binds plasminogen with high affinity (Kd = 7.58 nM), despite displaying only 52% identity to the classical a1/a2 repeat domain of PAM. It was therefore of interest to characterise the putative plasminogen binding domain of PAMNS88.2. Additionally, the association of GAS strainNS88.2, from which PAMNS88.2 was isolated, with the invasive disease bacteraemia, makes it a candidate for virulence studies employing the recently developed human plasminogen transgenic mouse. Site-directed mutagenesis of the putative plasminogen binding site indicated that as with PAMNS13, PAMNS88.2 does not interact with plasminogen exclusively via lysine residues. Mutagenesis to alanine of lysine residues Lys96 and Lys101 reduced but did not abrogate plasminogen binding by PAMNS88.2. Plasminogen binding was only abolished with the additional mutagenesis of Arg107 and His108 to alanine. Furthermore, mutagenesis of Arg107 and His108 abolished plasminogen binding by PAMNS88.2 despite the presence of Lys96 and Lys101 in the binding site. Given that GAS strain NS88.2 is associated with the invasive disease bacteraemia, and is virulent in the humanised plasminogen transgenic mouse, the successful abrogation of plasminogen binding by PAMNS88.2 may facilitate the development of a PAMNS88.2 allelic replacement isogenic mutant for use in future studies involving this model.This study examines in detail the interaction of PAM and PAM variants with the human zymogen plasminogen. The maintenance of plasminogen-binding function in spite of binding site sequence variation suggests that the ability to interact with plasminogen is evolutionarily advantageous to a subset of GAS isolates. Additionally, this studyprovides previously unreported details of the ability of PAM to interact withplasminogen independently of binding site lysine residues. These findings haveimplications for both the future identification of novel plasminogen binding proteins, and may facilitate both the understanding of the role of PAM in GAS disease, and the development of therapeutics to assist in the treatment and prevention of streptococcal infection. Publisher: School of Biological Sciences - Faculty of Science Source: University of Wollongong Thesis Collection
http://espace.library.uq.edu.au/view/UQ:158144
Cardiovascular diseases are responsible for over 50% of all deaths in Western society, with the underlying cause of most a single disease - atherosclerosis. Several classes of drugs have been developed to treat the clinical sequelae of atherosclerosis, such as myocardial infarction, including the HMG-CoA-reductase inhibitors (statins) which lower serum cholesterol and anti-platelet drugs such as aspirin and clopidogrel. These drugs are often used as combination therapy. As the activation of thromboxane prostanoid (TP) receptors by thromboxane A2 (TXA2) is implicated in secondary thrombotic events following myocardial infarction, TP receptor antagonists are a potential therapy. S18886 is a recently developed potent TP receptor antagonist under investigation. It inhibits platelet aggregation and vasoconstriction induced by the agonists of TP receptors, with no effect on the production of prostacyclin. S18886, like statins, has pleiotropic effects in addition to its demonstrated anti-thrombotic properties. Previous studies in our laboratory and others have found that treatment with S18886 prevents the development and progression of atherosclerosis in animal models by significantly decreasing the quantity of lipid-laden macrophages in fatty streaks and advanced lesions. In vitro, S18886 inhibits monocyte adherence to stimulated endothelial cells consistent with its inhibitory effects on ICAM-1 expression in vivo. Since macrophage-related proteolysis within atherosclerotic plaques contributes to the weakening of the protective fibrous cap and therefore promotes the susceptibility of those plaques to rupture, we hypothesised that S18886 may stabilise the vulnerable plaque. Thus, in this thesis the effect of S18886 on the stability of pre-formed atherosclerotic plaques was examined in the cholesterol-fed rabbit model, and its effects compared with aspirin, clopidogrel, and pravastatin alone and in combination. S18886 was shown to inhibit the atherogenic process in both uninjured and injured arteries, and provided additional, and indeed, superior benefits to those achieved with the currently available therapies. S18886 completely inhibited pre-existing fatty streaks from becoming thicker in spite of the continued presence of hyperlipidaemia. Animals treated with S18886 alone had stainable lipid in the fatty streaks no different from the control, indicating that it completely inhibited further lipid accumulation within the wall. S18886 prevented progression of lesion formation by controlling macrophage accumulation and subsequent MMP-9 expression. This was further indicated by the inhibition of ICAM-1 and VCAM-1 expression in S18886-treated Groups. Combining S18886 with aspirin, clopidogrel or pravastatin yielded some beneficial effects when compared to the comparator drugs alone, although even combined with S18886, aspirin barely improved in efficacy. S18886 in combination with clopidogrel or pravastatin prevented progression of advanced lesions. The addition of S18886 to pravastatin also resulted in a significantly smaller accumulation of macrophages in the injured femoral artery compared to pravastatin alone. S18886 combined with clopidogrel significantly reduced the macrophage accumulation and MMP-9 content when compared to clopidogrel alone. Notably, S18886 and clopidogrel proved to be more beneficial than clopidogrel and aspirin in several parameters. The mechanism by which S18886 exerts its plaque stabilisation effects is through antagonism of TP receptors. TP receptors are stimulated not only by thromboxane, but by other products of oxidative stress such as hydroxyeicosatetraenoic acids (HETES) and F2-isoprostane. HETES and F2- isoprostane are nonenzymatic peroxidation products of arachidonic acid that are not inhibited by aspirin. Antagonism of TP receptors prevents expression of ICAM-1 and VCAM-1 on endothelial cells, which in turn does not allow monocyte-derived macrophage entry into the vessel wall and thus decreases the potential for MMP production and plaque destabilisation. In summary, we suggest that S18886 may prove to be effective clinically in preventing patients ‘at risk’ from further atherosclerotic plaque progression and the development of clinical sequelae such as plaque rupture, when administered either alone or when added to the regimen of lipid-lowering statins or anti-platelet therapies.
http://espace.library.uq.edu.au/view/UQ:178442
Phosphine, hydrogen phosphide (PH3), gas is a fumigant that is used worldwide to protect stored grain from infestation by insect pests. Despite a long history of phosphine use, little is known about either the mode of action of this compound or the mechanisms whereby insect pests have become resistant. To better understand phosphine toxicity and resistance mechanisms, a genetically well-characterised model organism, Caenorhabditis elegans, was used in my PhD project. Three previously created phosphine resistant C. elegans mutants (pre-1, pre-7 and pre-33) developed from the wild type N2 strain were used in this study, though analysis of pre-33 was the primary focus. The three mutants were determined to be 2, 5 and 9 times more resistant toward phosphine than was the parental N2 strain by comparison of LC50 values. Molecular oxygen was shown to be an extremely effective synergist with phosphine as, under hyperoxic conditions, 100% mortality was observed in wild-type nematodes exposed to 0.1 mg/l phosphine, a non-lethal concentration in air. All three mutants were resistant to the synergistic effects of oxygen in proportion to their resistance to phosphine with one mutant, pre-33, showing complete resistance to this synergism. I take the proportionality of cross-resistance between phosphine and the synergistic effect of oxygen to imply that all three mutants circumvent a mechanism of phosphine toxicity that is directly coupled to oxygen metabolism. Compared with the wild-type strain, each of the three mutants has an extended average life expectancy of 12.5 to 25.3%. This is consistent with the proposed involvement of oxidative stress in both phosphine toxicity and ageing. Indeed, a correlation between phosphine resistance and resistance to other stressors (e.g. heavy metal, heat and UV) was also detected. On the other hand, no significant difference in methyl viologen sensitivity was found between pre-33 and N2 strains, suggesting that pre-33 mutant does not seem to provide resistance to phosphine via protection against oxidative damage. Additionally, to test for possible involvement of the DAF-2/DAF-16 signalling pathway in the phosphine response, the levels of phosphine sensitivity of mutants in this pathway were tested. Phosphine resistance levels were increased in daf-2 and age-1 mutants but decreased in daf-16 nematodes, which mirrors the longevity phenotypes of these mutants, suggesting some congruence in glucose signalling between the phosphine resistance and longevity traits. In contrast, no congruence is observed between phosphine resistance and oxidative metabolism as the clk-mutation, which disrupts oxidative metabolism does not cause phosphine resistance and neither do the phosphine resistant mutants cause the severe developmental delay of the clk-1 mutation. The phosphine induced time-dependent mortality was assessed in both N2 and pre-33 nematodes at two fixed phosphine concentrations (0.3 and 3.0 mg/l), allowing the determination of minimum exposure periods required for any mortality as well as the exposure time required to achieve 50% mortality. As a result, it was determined that 15 hours of exposure was needed for significant mortality in N2 and pre-33 strain when exposed to 0.3 and 3.0 mg/l of phosphine, respectively; whereas this period is 5 hours for N2 when treated with 3.0 mg/l phosphine. The fact that the LT50 value for N2 at 0.3 mg/l phosphine is indistinguishable from that of pre-33 at 3.0 mg/l (24.6 and 24.5 respectively) suggests that 0.3 and 3.0 mg/l of phosphine have the same toxic effects on N2 and pre-33 nematodes respectively. This result is consistent with the finding that pre-33 is ~9 fold more resistant to phosphine than is the N2 strain. Moreover, the LT50 was determined to be 8.4 hours for N2 when treated with 3.0 mg/l of phosphine, which is only three times faster than pre-33 when exposed to the same level of phosphine. In contrast to the differential toxicity of phosphine between the N2 and pre-33 lines, the delay in reaching reproductive maturity caused by phosphine exposure is indistinguishable between WT and pre-33 nematodes. This indicates that the phosphine induced delay in maturation is independent of the toxic effects of phosphine. Since the inhibition of complex IV (cytochrome c oxidase) in the mitochondrial electron transport chain has been proposed as a mechanism of phosphine toxicity, the phosphine effects on cellular ATP metabolism, presented as ATP+ADP content and ATP/ADP ratio, were also assessed. Phosphine exposure (0.3 mg/l, 25 hours) led to a significant decrease in ATP+ADP levels as well as the ATP/ADP ratio in N2 nematodes. Similar results were also detected in pre-33 nematodes when exposed to 3.0 mg/l phosphine for 25 hours. These observations indicate that phosphine can interrupt cellular ATP metabolism, which is associated with phosphine induced mortality. Additionally, the fact that mutant pre-33 can maintain its ATP levels under phosphine exposure at 0.3 mg/l suggests it has a greater ability to maintain mitochondrial function than does the N2 strain. To better understand the mechanism of phosphine toxicity in the wild type N2 strain, gene expression profiling by DNA microarray analysis was employed. A significant overlap between phosphine and DAF-16 regulated genes was detected, supporting the previous finding that the DAF-2/DAF-16 pathway can contribute to phosphine resistance. Phosphine exposure also strongly induced xenobiotic detoxification and stress responses, indicating nematodes are able to sense phosphine induced toxic effects and protect themselves by switching on native detoxification mechanisms. Furthermore, glycolysis and gluconeogenesis were also up-regulated by phosphine, possibly due to an increase in energy demand caused by increased xenobiotic detoxification activities. Consistent with the previous findings that phosphine delays median reproductive age and reduces fertility, expressions of a large number of genes involved in growth, embryonic development and reproduction were suppressed by phosphine. Moreover, the microarray results of seven genes whose expression levels were significantly altered by phosphine were validated using RT-PCR, confirming the robustness of the microarray results. The most direct way to determine the phosphine resistance mechanism in mutant pre-33 is to identify and characterise the mutation itself. Using a classic F1 test, the resistance mutation in pre-33 was determined to be incompletely recessive. Additionally, using three mapping strategies, the resistance mutation was mapped to Chromosome IV between 12,591,683 and 12,879,637 bp with 45 genes located in this small region. In an attempt to identify the resistance gene, the effect of suppressing each of 28 of the 45 genes in the interval was determined using a commercially available gene suppression library. It was observed that only knockdown of gene vha-7 resulted in a slight decrease in phosphine sensitivity (84.6%) compared to N2 (97.6%). However, this result does not clearly implicate vha-7 as the resistance gene in pre-33. The microarray results indicated that linoleate and arachidonate signalling pathways might be activated by phosphine. This was observed as induction of a phospholipase A2 gene that regulates the release of arachidonic acid from the C-2 position of membrane phospholipids, as well as several CYP genes predicted to catalyse the oxidation of linoleate and arachidonate. Therefore, phosphine effects on the linoleate and arachidonate dependent signalling pathways were assessed. It was found that, in the presence of phosphine, the pre-33 mutant has a greater ability to transform linoleate and arachidonate epoxides to diols than does N2. This activity may help pre-33 to better maintain mitochondrial function and, therefore, ATP metabolism than N2 during phosphine exposure. The microarray results also showed that phosphine exposure caused up-regulation of glycolysis and gluconeogenesis, indicating phosphine regulation of carbohydrate metabolism. As expected, a preliminary metabonomic analysis by 1H nuclear magnetic resonance (NMR) into the effect of phosphine exposure on metabolism in N2 nematodes revealed significant alteration of the metabonomic profile.
http://ro.uow.edu.au/theses/79
Individual proteins have a specific three-dimensional structure that gives them their unique function. However, a protein must be folded from a linear string of amino acids in order to gain this native conformation and thus function. There are many hurdles to a protein attaining and maintaining its native conformation. Stresses that are encountered in the life of a protein, such as changes in pH, temperature and oxidative stress, can promote protein misfolding or unfolding. In addition, some genetic mutations can modify a protein such that a non-native conformation is more energetically favourable than the native state. The unfolding or misfolding of a protein makes it more likely that it will aggregate with itself. There are more than 40 human diseases associated with the inappropriate deposition of aggregated protein. It is well known that there is a welldefined and efficient quality control system to deal with intracellular proteins that have either unfolded or are misfolded. Cells have a range of molecular chaperones to inhibit inappropriate aggregation and if this fails the cell labels the proteins with ubiquitin for degradation via the proteasome. However, many proteins are secreted from cells into the extracellular environment and there are many protein deposition disorders associated with extracellular protein deposits, outside the reach of the well-characterised intracellular quality control system. This thesis reports that the secreted proteins haptoglobin and a2-macroglobulin have small heat shock protein-like chaperone activity. Both haptoglobin and a2- macroglobulin specifically inhibited the precipitation of a variety of proteins induced by either heat or oxidation, including proteins in unfractionated human serum. In addition, it was demonstrated that haptoglobin and a2-macroglobulin inhibit the precipitation of stressed proteins by forming solubilized complexes with them, cannot protect enzymes from heat-induced loss of function, and lack ATPase activity and the ability to independently refold proteins following stresses. In addition, data presented here shows that clusterin, haptoglobin and a2-macroglobulin exert potent effects on amyloid formation, and provide evidence to suggest that these effects are exerted via interactions with pre-fibrillar species. These findings suggest that clusterin, haptoglobin and a2- macroglobulin are an important element in the control of extracellular protein misfolding. Publisher: School of Biological Sciences - Faculty of Science Source: University of Wollongong Thesis Collection
2008-06-02
{beta}1 integrin signaling has been shown to mediate cellular resistance to apoptosis after exposure to ionizing radiation (IR). Other signaling molecules that increase resistance include Akt, which promotes cell survival downstream of {beta}1 integrin signaling. We showed previously that {beta}1 integrin inhibitory antibodies, AIIB2, enhance apoptosis and decrease growth in human breast cancer cells in 3 dimensional laminin-rich extracellular matrix (3D lrECM) cultures and in vivo. Here we asked whether AIIB2 could synergize with IR to modify Akt-mediated IR resistance. We used 3D lrECM cultures to test the optimal combination of AIIB2 with IR treatment of two breast cancer cell lines, MCF-7 and HMT3522-T4-2, as well as T4-2 myr-Akt breast cancer colonies or HMT3522-S-1, which form normal organotypic structures in 3D lrECM. Colonies were assayed for apoptosis and {beta}1 integrin/Akt signaling pathways were evaluated using western blot. In addition, mice bearing MCF-7 xenografts were used to validate the findings in 3D lrECM. We report that AIIB2 increased apoptosis optimally post-IR by down regulating Akt in breast cancer colonies in 3D lrECM. In vivo, addition of AIIB2 after IR significantly enhanced tumor growth inhibition and apoptosis compared to either treatment alone. Remarkably, the degree of tumor growth inhibition using AIIB2 plus 2 Gy radiation was similar to that of 8 Gy alone. We showed previously that AIIB2 had no discernible toxicity in mice; here, its addition allowed for a significant reduction in the IR dose that was necessary to achieve comparable growth inhibition and apoptosis in breast cancer xenografts in vivo.
The problem of living in a world contaminated with chemicals
1990-12-31
The proliferation of xenobiotic chemicals in the global environment poses living problems for each of us aboard {open_quotes}spaceship earth.{close_quotes} Seven case studies are presented that illustrate the magnitude of the problem that can result from waiting to identify toxic hazards until there have been decades of {open_quotes}human guinea pig{close_quotes} exposure. 25 refs., 5 tabs.
2005-01-01
We demonstrate that cells derived from primary cultures of rabbit proximal tubules (RPTC), human embryonic kidney (HEK293) and human kidney carcinomas (Caki-1) express microsomal Ca2+-independent phospholipase A2 (iPLA2g) and cytosolic Ca2+-independent phospholipase A2 (iPLA2b). Inhibition of iPLA2 activity in these cells using the iPLA2 inhibitor bromoenol lactone (BEL) (0-5.0mM) for 24h did not induce cell death as determined by annexin V and propidium iodide (PI) staining. However, BEL treatment prior to cisplatin (50mM) or vincristine (2mM) exposure reduced apoptosis 30-50% in all cells tested (RPTC, HEK293 and Caki-1 cells). To identify the phospholipids altered during cell death electrospray ionization-mass spectrometry and lipidomic analysis of HEK293 and Caki-1 cells was performed....
2007-01-01
Background and ObjectivesInhibition of cyclooxygenase (COX) and prostaglandin E2 (PGE2) protects cells against cell injury in specific pathophysiological situations: inflammation and oxidative stress. Although the anti-inflammatory effects have been reported in clinical fields for specific wavelength irradiation during wound healing, the physiological mechanism has not been clarified yet. The aim of the present study is to investigate the anti-inflammatory mechanism of 635 nm light-emitting-diode (LED) irradiation compared with existing COX inhibitors.Study Design/Materials and MethodsThe present study investigated anti-inflammatory effects of 635 nm irradiation on PGE2 release, COX and phospholipase A2 (PLA2) expression, and reactive oxygen species (ROS) dissociation in arachidonic acid (...
The Role of Rho Kinase in Sex-Dependent Vascular Dysfunction in Type 1 Diabetes
2010-01-01
We hypothesized that rho/rho kinase plays a role in sex differences in vascular dysfunction of diabetics. Contractions to serotonin were greater in isolated aortic rings from nondiabetic males versus...Full Text Available
The Role of Rho Kinase in Sex-Dependent Vascular Dysfunction in Type 1 Diabetes
2010-01-01
Full Text Available.We hypothesized that rho/rho kinase plays a role in sex differences in vascular dysfunction of diabetics. Contractions to serotonin were greater in isolated aortic rings from nondiabetic males versus females and increased further in streptozotocin-induced diabetic males but not females. The increased contractions to serotonin in males were reduced by inhibitors of rho kinase (fasudil, Y27632 and H1152) despite no change in expression of rhoA or rho kinase. Contractions to U46619 were not altered by fasudil or Y27632 or the presence of diabetes. In contrast to acute effects of fasudil, chronic treatment with fasudil increased contractions to serotonin in aorta from both non-diabetic and diabetic males. In summary, serotonin-induced contractions were increased in aorta from diabetic males but not females. Although administration of rho kinase inhibitors acutely decreased contractions to serotonin, long-term treatment with fasudil increased contractions. Long-term fasudil treatment may increase compensatory mechanisms to enhance vasoconstrictions.
The Role of Cyclooxygenase-2 in Cell Proliferation and Cell Death in Human Malignancies
2010-01-01
It is well admitted that the link between chronic inflammation and cancer involves cytokines and mediators of inflammatory pathways, which act during the different steps of tumorigenesis. The cyclooxygenases...Full Text Available
The Role of Cyclooxygenase-2 in Cell Proliferation and Cell Death in Human Malignancies
2010-01-01
Full Text Available.It is well admitted that the link between chronic inflammation and cancer involves cytokines and mediators of inflammatory pathways, which act during the different steps of tumorigenesis. The cyclooxygenases (COXs) are a family of enzymes, which catalyze the rate-limiting step of prostaglandin biosynthesis. This family contains three members: ubiquitously expressed COX-1, which is involved in homeostasis; the inducible COX-2 isoform, which is upregulated during both inflammation and cancer; and COX-3, expressed in brain and spinal cord, whose functions remain to be elucidated. COX-2 was described to modulate cell proliferation and apoptosis mainly in solid tumors, that is, colorectal, breast, and prostate cancers, and, more recently, in hematological malignancies. These findings prompt us to analyze here the effects of a combination of COX-2 inhibitors together with different clinically used therapeutic strategies in order to further improve the efficiency of future anticancer treatments. COX-2 modulation is a promising field investigated by many research groups.
The Dual Role of Calcium as Messenger and Stressor in Cell Damage, Death, and Survival
2010-01-01
Full Text Available.Ca2+ is an important second messenger participating in many cellular activities; when physicochemical insults deregulate its delicate homeostasis, it acts as an intrinsic stressor, producing/increasing cell damage. Damage elicits both repair and death responses; intriguingly, in those responses Ca2+ also participates as second messenger. This delineates a dual role for Ca2+ in cell stress, making difficult to separate the different and multiple mechanisms required for Ca2+-mediated control of cell survival and apoptosis. Here we attempt to disentangle the two scenarios, examining on the one side, the events implicated in deregulated Ca2+ toxicity and the mechanisms through which this elicits reparative or death pathways; on the other, reviewing the role of Ca2+ as a messenger in the transduction of these same signaling events.
The Dual Role of Calcium as Messenger and Stressor in Cell Damage, Death, and Survival
2010-01-01
Ca2+ is an important second messenger participating in many cellular activities; when physicochemical insults deregulate its delicate homeostasis, it acts as an intrinsic stressor, producing/increasing...Full Text Available
T Cell-Dependence of Lassa Fever Pathogenesis
2010-03-01
Lassa virus (LASV), the causative agent of Lassa fever (LF), is endemic in West Africa, accounting for substantial morbidity and mortality. In spite of ongoing research efforts, LF pathogenesis and...Full Text Available
T Cell-Dependence of Lassa Fever Pathogenesis
2010-03-01
Full Text Available.Lassa virus (LASV), the causative agent of Lassa fever (LF), is endemic in West Africa, accounting for substantial morbidity and mortality. In spite of ongoing research efforts, LF pathogenesis and mechanisms of LASV immune control remain poorly understood. While normal laboratory mice are resistant to LASV, we report that mice expressing humanized instead of murine MHC class I (MHC-I) failed to control LASV infection and develop severe LF. Infection of MHC-I knockout mice confirmed a key role for MHC-I-restricted T cell responses in controlling LASV. Intriguingly we found that T cell depletion in LASV-infected HHD mice prevented disease, irrespective of high-level viremia. Widespread activation of monocyte/macrophage lineage cells, manifest through inducible NO synthase expression, and elevated IL-12p40 serum levels indicated a systemic inflammatory condition. The absence of extensive monocyte/macrophage activation in T cell-depleted mice suggested that T cell responses contribute to deleterious innate inflammatory reactions and LF pathogenesis. Our observations in mice indicate a dual role for T cells, not only protecting from LASV, but also enhancing LF pathogenesis. The possibility of T cell-driven enhancement and immunopathogenesis should be given consideration in future LF vaccine development.
Radiation-induced p53 protein response in the A549 cell line is culture growth-phase dependent
1995-12-01
One role of the p53 tumor suppressor protein has been recently revealed. Kastan, M.B. reported that p53 protein accumulates in cells exposed to ionizing radiation. The accumulation of p53 protein is in response to DNA damage, most importantly double-strand breaks, that results from exposure to ionizing radiation. The rise in cellular p53 levels is necessary for an arrest in the G{sub 1} phase of the cell cycle to provide additional time for DNA repair. The p53 response has also been demonstrated to enhance PCNA-dependent repair. p53 is thus an important regulator of the cellular response to DNA-damaging radiation. From this data, it can be concluded that the magnitude of the p53 response is not dependent on the phase of culture growth.
Quinacrine protects neuronal cells against heat-induced injury
2009-01-01
The effects of quinacrine (QA) on heat-induced neuronal injury have been explored, with the intention of understanding the mechanisms of QA protection. Primary cultivated striatum neurons from newborn rats were treated with QA 1h before heat treatment at 43degreeC which lasted for another 1h, and necrosis and apoptosis were detected by Annexin-V-FITC and propidium iodide (PI) double staining. Neuronal apoptosis was determined using terminal deoxynucleotidyl transferase-mediated biotinylated UTP nick end labeling (TUNEL) techniques. Cell membrane fluidity, activity of cytosolic phospholipase A2 (cPLA2) and the level of arachidonic acid (AA) were also examined. Membrane surface ultrastructure of striatum neurons was investigated by atomic force microscopy (AFM). Results showed that heat trea...
Proteolytic resistance conferred to fibrinogen by von Willebrand factor1
2010-02-01
SummaryThe formation of platelet-rich thrombi under high shear rates requires both fibrinogen and von Willebrand factor (VWF) as molecular adhesives between platelets. We attempted...Full Text Available
Proteolytic resistance conferred to fibrinogen by von Willebrand factor1
2010-02-01
Full Text Available.SummaryThe formation of platelet-rich thrombi under high shear rates requires both fibrinogen and von Willebrand factor (VWF) as molecular adhesives between platelets. We attempted to describe the role of VWF as a potential substrate and modulator of the fibrinolytic system using binding assays, as well as kinetic measurements on the cleavage of fibrin(ogen) and a synthetic plasmin substrate (Spectrozyme-PL). The similar dissociation constants for the binding of plasminogen, plasmin, and active site-blocked plasmin onto immobilized VWF suggest that the primary binding site in plasmin(ogen) is not the active site. The progressive loss of clottability and generation of degradation products during fibrinogen digestion with plasmin were delayed in the presence of VWF at physiological concentrations, while VWF cleavage was not detectable. Determination of kinetic parameters for fibrinogen degradation by plasmin, miniplasmin and microplasmin showed that VWF did not modify the Km, whereas kcat values decreased with increasing VWF concentrations following the kinetic model of non-competitive inhibition. Inhibitory constants calculated for VWF were in the range of its physiological plasma concentration (5.4 μg/ml, 5.7 μg/ml and 10.0 μg/ml for plasmin, miniplasmin and microplasmin, respectively) and their values suggested a modulating role of the kringle 5 domain in the interaction between VWF and (mini)plasmin. VWF had no effect on the amidolytic activity of plasmin on Spectrozyme-PL, or on fibrin dissolution by (mini)plasmin. Our data suggest that VWF, while a poor plasmin substrate relative to fibrinogen, protects fibrinogen against degradation by plasmin preserving its clottability in plasma and its adhesive role in platelet-rich thrombi.
Ozone as an ecotoxicological problem
1996-11-01
Ozone is quantitatively the dominating oxidant in photochemical air pollution. Other compounds like hydrogen peroxide, aldehydes, formate, peroxyacetyl nitrate (PAN) and nitrogen dioxide are present too, and several of these are known to be phytotoxic, but under Danish conditions the concentration of these gases are without significance for direct effects on vegetation. Therefore, it is the effects of ozone on plant growth that will be described below. (EG) 65 refs.
Full Text Available.BackgroundRecent studies suggest an important role for neurotransmitters as modulators of inflammation. Neuroinflammatory mediators such as cytokines and molecules of the arachidonic acid pathway are generated and released by microglia. The monoamine norepinephrine reduces the production of cytokines by activated microglia in vitro. However, little is known about the effects of norepinephrine on prostanoid synthesis. In the present study, we investigate the role of norepinephrine on cyclooxygenase- (COX-)2 expression/synthesis and prostaglandin (PG)E2 production in rat primary microglia.ResultsInterestingly, norepinephrine increased COX-2 mRNA, but not protein expression. Norepinephrine strongly enhanced COX-2 expression and PGE2 production induced by lipopolysaccharide (LPS). This effect is likely to be mediated by β-adrenoreceptors, since β-, but not α-adrenoreceptor agonists produced similar results. Furthermore, β-adrenoreceptor antagonists blocked the enhancement of COX-2 levels induced by norepinephrine and β-adrenoreceptor agonists.ConclusionsConsidering that PGE2 displays different roles in neuroinflammatory and neurodegenerative disorders, norepinephrine may play an important function in the modulation of these processes in pathophysiological conditions.
BackgroundRecent studies suggest an important role for neurotransmitters as modulators of inflammation. Neuroinflammatory mediators such as cytokines and molecules of the arachidonic...Full Text Available
The availability of nutrients influences cellular growth and survival by affecting gene transcription. Glucocorticoids also influence gene transcription and have diverse activities on cell growth,...Full Text Available
Full Text Available.The availability of nutrients influences cellular growth and survival by affecting gene transcription. Glucocorticoids also influence gene transcription and have diverse activities on cell growth, energy expenditure, and survival. We found that the growth arrest-specific 5 (Gas5) noncoding RNA, which is abundant in cells whose growth has been arrested due to lack of nutrients or growth factors, sensitized cells to apoptosis by suppressing glucocorticoid-mediated induction of several responsive genes, including the one encoding cellular inhibitor of apoptosis 2. Gas5 bound to the DNA-binding domain of the glucocorticoid receptor (GR) by acting as a decoy “glucocorticoid response element (GRE)”, thus, competing with DNA GREs for binding to the GR. We conclude that Gas5 is a ribo-repressor of the GR, influencing cell survival and metabolic activities during starvation by modulating the transcriptional activity of the GR.
Neuroprotectin D1 Attenuates Laser-induced Choroidal Neovascularization in Mouse
PurposeTo examine the effects of neuroprotectin D1 (NPD1), a stereospecific derivative of docosahexaenoic acid, on choroidal neovascularization (CNV) in a laser-induced mouse model....Full Text Available
Neuroprotectin D1 Attenuates Laser-induced Choroidal Neovascularization in Mouse
Full Text Available.PurposeTo examine the effects of neuroprotectin D1 (NPD1), a stereospecific derivative of docosahexaenoic acid, on choroidal neovascularization (CNV) in a laser-induced mouse model. Specifically, this was assessed by clinically grading laser-induced lesions, measuring leakage area, and volumetrically quantifying vascular endothelial cell proliferation.MethodsC57Bl/6 mice were treated with vehicle control or NPD1, and choroidal neovascularization was induced by laser rupture of Bruch's membrane; treatment was administered throughout the first week of recovery. One and two weeks after CNV induction, fundus fluorescein angiography was performed. Angiograms were clinically graded to assess leakage severity, while leakage area was measured by image analysis of angiograms. Proliferation of vascular endothelial cells was evaluated volumetrically by three-dimensional laser confocal immunofluorescent microscopy of cytoskeletal, nuclear, and endothelial cell markers.ResultsAt seven days after CNV induction, NPD1-treated mice had 60% fewer clinically relevant lesions than controls, dropping to 80% fewer by 14 days. NPD1 mice exhibited 25% smaller leakage area than controls at 7 days and 44% smaller area at 14 days. Volumetric immunofluorescence revealed 46% less vascular endothelial cell volume in 7-day NPD1-treated mice than in 7-day controls, and by 14 days NPD1 treatment was 68% lower than controls. Furthermore, comparison of 7- and 14-day volumes of NPD1-treated mice revealed a 50% reduction at 14 days.ConclusionsNPD1 significantly inhibits choroidal neovascularization. There are at least two possible mechanisms that could explain the neuroprotective action of NPD1. Ultimately, nuclear factor-κB could be inhibited with a reduction in cyclooxygenase-2 (COX-2) to reduce vascular endothelial growth factor (VEGF) expression, and/or activation of the resolution phase of the inflammatory response/survival pathways could be upregulated. Moreover, NPD1 continues to be effective after treatment is concluded, suggesting sustained protection and highlighting the potential applicability of this lipid mediator in preventing or ameliorating endothelial cell growth in pathoangiogenesis.
2008-01-01
Hyperglycemia encountered during diabetes triggers abnormalities of vascular function associated with cell acidosis and calcium overload. The purpose of this study was to determine, whether Na+/H+ exchanger (NHE-1) inhibition by cariporide protects coronary cells against the deleterious effect of hyperglycemia in the rat. In vivo hyperglycemia was triggered by streptozotocin injection. One week after, the glycemia was checked and the control and diabetic animals were treated or not with cariporide (2.5 mg/kg/day) for two weeks. Glycemia was again estimated and the hearts were perfused according to the Langendorff mode at forced flow. The left ventricle developed pressure (LVDP) and heart rate (HR) were determined with a latex balloon inserted into the left ventricle. Coronary pressure wa...
Full Text Available.BackgroundAlthough donor dopamine treatment reduces the requirement for post transplantation dialysis in renal transplant recipients, implementation of dopamine in donor management is hampered by its hemodynamic side-effects. Therefore novel dopamine derivatives lacking any hemodynamic actions and yet are more efficacious in protecting tissue from cold preservation injury are warranted. We hypothesized that variation of the molecular structure would yield more efficacious compounds avoid of any hemodynamic effects.Methodology/Principal FindingsTo this end, we assessed protection against cold preservation injury in HUVEC by the attenuation of lactate dehydrogenase (LDH) release. Modification of dopamine by an alkanoyl group increased cellular uptake and significantly improved efficacy of protection. Further variation revealed that only compounds bearing two hydroxy groups in ortho or para position at the benzene nucleus, i.e. strong reductants, were protective. However, other reducing agents like N-acetyl cysteine and ascorbate, or NADPH oxidase inhibition did not prevent cellular injury following cold storage. Unlike dopamine, a prototypic novel compound caused no hemodynamic side-effects.Conclusions/SignificanceIn conclusion, we demonstrate that protection against cold preservation injury by catecholamines is exclusively governed by strong reducing capacity and sufficient lipophilicity. The novel dopamine derivatives might be of clinical relevance in donor pre-conditioning as they are completely devoid of hemodynamic action, their increased cellular uptake would reduce time of treatment and therefore also may have a potential use for non-heart beating donors.
BackgroundAlthough donor dopamine treatment reduces the requirement for post transplantation dialysis in renal transplant recipients, implementation of dopamine in donor management...Full Text Available
2010-02-01
Tuberculosis (TB) is an escalating global health problem and improved vaccines against TB are urgently needed. HLA-E restricted responses may be of interest for vaccine development since HLA-E displays...Full Text Available
2010-02-01
Full Text Available.Tuberculosis (TB) is an escalating global health problem and improved vaccines against TB are urgently needed. HLA-E restricted responses may be of interest for vaccine development since HLA-E displays very limited polymorphism (only 2 coding variants exist), and is not down-regulated by HIV-infection. The peptides from Mycobacterium tuberculosis (Mtb) potentially presented by HLA-E molecules, however, are unknown. Here we describe human T-cell responses to Mtb-derived peptides containing predicted HLA-E binding motifs and binding-affinity for HLA-E. We observed CD8+ T-cell proliferation to the majority of the 69 peptides tested in Mtb responsive adults as well as in BCG-vaccinated infants. CD8+ T-cells were cytotoxic against target-cells transfected with HLA-E only in the presence of specific peptide. These T cells were also able to lyse M. bovis BCG infected, but not control monocytes, suggesting recognition of antigens during mycobacterial infection. In addition, peptide induced CD8+ T-cells also displayed regulatory activity, since they inhibited T-cell proliferation. This regulatory activity was cell contact-dependent, and at least partly dependent on membrane-bound TGF-β. Our results significantly increase our understanding of the human immune response to Mtb by identification of CD8+ T-cell responses to novel HLA-E binding peptides of Mtb, which have cytotoxic as well as immunoregulatory activity.
Mechanisms of energy transfer and conversion in plant Light-Harvesting Complex II
2009-09-24
The light-harvesting complex of photosystem II (LHC-II) is the major antenna complex in plant photosynthesis. It accounts for roughly 30% of the total protein in plant chloroplasts, which makes it arguably the most abundant membrane protein on Earth, and binds about half of plant chlorophyll (Chl). The complex assembles as a trimer in the thylakoid membrane and binds a total of 54 pigment molecules, including 24 Chl a, 18 Chl b, 6 lutein (Lut), 3 neoxanthin (Neo) and 3 violaxanthin (Vio). LHC-II has five key roles in plant photosynthesis. It: (1) harvests sunlight and transmits excitation energy to the reaction centres of photosystems II and I, (2) regulates the amount of excitation energy reaching each of the two photosystems, (3) has a structural role in the architecture of the photosynthetic supercomplexes, (4) contributes to the tight appression of thylakoid membranes in chloroplast grana, and (5) protects the photosynthetic apparatus from photo damage by non photochemical quenching (NPQ). A major fraction of NPQ is accounted for its energy-dependent component qE. Despite being critical for plant survival and having been studied for decades, the exact details of how excess absorbed light energy is dissipated under qE conditions remain enigmatic. Today it is accepted that qE is regulated by the magnitude of the pH gradient ({delta}pH) across the thylakoid membrane. It is also well documented that the drop in pH in the thylakoid lumen during high-light conditions activates the enzyme violaxanthin de-epoxidase (VDE), which converts the carotenoid Vio into zeaxanthin (Zea) as part of the xanthophyll cycle. Additionally, studies with Arabidopsis mutants revealed that the photosystem II subunit PsbS is necessary for qE. How these physiological responses switch LHC-II from the active, energy transmitting to the quenched, energy-dissipating state, in which the solar energy is not transmitted to the photosystems but instead dissipated as heat, remains unclear and is the subject of this thesis. From the results obtained during this doctoral work, five main conclusions can be drawn concerning the mechanism of qE: 1. Substitution of Vio by Zea in LHC-II is not sufficient for efficient dissipation of excess excitation energy. 2. Aggregation quenching of LHC-II does not require Vio, Neo nor a specific Chl pair. 3. With one exception, the pigment structure in LHC-II is rigid. 4. The two X-ray structures of LHC-II show the same energy transmitting state of the complex. 5. Crystalline LHC-II resembles the complex in the thylakoid membrane. Models of the aggregation quenching mechanism in vitro and the qE mechanism in vivo are presented as a corollary of this doctoral work. LHC-II aggregation quenching in vitro is attributed to the formation of energy sinks on the periphery of LHC-II through random interaction with other trimers, free pigments or impurities. A similar but unrelated process is proposed to occur in the thylakoid membrane, by which excess excitation energy is dissipated upon specific interaction between LHC-II and a PsbS monomer carrying Zea. At the end of this thesis, an innovative experimental model for the analysis of all key aspects of qE is proposed in order to finally solve the qE enigma, one of the last unresolved problems in photosynthesis research. (orig.)
Information needs for risk management/communication
1990-12-31
The hazardous waste cleanup program under the Comprehensive Environmental Response, Compensation, and Liability Act (Superfund) is delegated to the ten Regions of the US Environmental Protection Agency (EPA) and has, to date, identified more than 33,000 sites for consideration. The size and complexity of the program places great demands on those who would provide information to achieve national consistency in application of risk assessment while meeting site-specific needs for risk management and risk communication.
Information needs for risk assessment
1990-12-31
Risk assessment can be thought of as a conceptual approach to bridge the gap between the available data and the ultimate goal of characterizing the risk or hazard associated with a particular environmental problem. To lend consistency to and to promote quality in the process, the US Environmental Protection Agency (EPA) published Guidelines for Risk Assessment of Carcinogenicity, Developmental Toxicity, Germ Cell Mutagenicity and Exposure Assessment, and Risk Assessment of Chemical Mixtures. The guidelines provide a framework for organizing the information, evaluating data, and for carrying out the risk assessment in a scientifically plausible manner. In the absence of sufficient scientific information or when abundant data are available, the guidelines provide alternative methodologies that can be employed in the risk assessment. 4 refs., 3 figs., 2 tabs.
Information and technology: A coexistence without limits, a beginning with no apparent ending
1990-12-31
A variety of issues must be addressed in development of software for information resources. One is accessibility and use of information. Another is that to properly design, abstract, index, and do quality control on a database requires the effort of well-trained and knowledgeable personnel as well as substantial financial resources. Transferring data to other locations has inherent difficulties, including those related to incompatibility. The main issue in developing health risk assessment databases is the needs of the user.
2000-11-15
Influence of melanocytes in skin pigmentation is well documented, however its photo-protective role has given rise to controversy. The role of melanocytes have been investigated on reconstructed epidermis with 100 % of keratinocytes or 95 % of keratinocytes and 5 % of melanocytes. In a first time, the effect of an acute UVB dose has been studied on both reconstructed epidermis, next we have investigated UVA and UVA+B effects on these epidermis. Following irradiation, the presence of melanocytes in reconstructed epidermis protects against apoptosis without protecting significantly against DNA damage formation (CPD, 6-4PP) and protects against UV-induced unbalance of the SOD/catalase ratio (antioxidants enzymes). On the contrary, the presence of melanocytes in reconstructed epidermis amplifies lipids and proteins oxidations but seems to protect against DNA oxidations. Melanocytes differ from keratinocytes by their melanin content and their more important concentration in polyunsaturated fatty acids. To evaluate what is the part of melanin and the part of polyunsaturated fatty acids in epidermal UV responses, reconstructed epidermis with keratinocytes have been supplemented with polyunsaturated fatty acid. This study indicates that polyunsaturated fatty acids are responsible for lipids and proteins oxidations and that melanin protect against DNA oxidation induced by lipid peroxidation. All these studies demonstrate that, model of reconstructed epidermis and epidermis in-vivo have the same behaviour following UV irradiation. In the last part, sunscreens and antioxidants have been tested on reconstructed epidermis and have demonstrated that model of reconstructed epidermis is suitable for photo-protective molecules screening. (author)
Immuno-modulation and anti-inflammatory benefits of antibiotics: The example of tilmicosin
2010-01-01
Full Text Available.Exagerated immune responses, such as those implicated in severe inflammatory reactions, are costly to the metabolism. Inflammation and pro-inflammatory mediators negatively affect production in the food animal industry by reducing growth, feed intake, reproduction, milk production, and metabolic health. An ever-increasing number of findings have established that antibiotics, macrolides in particular, may generate anti-inflammatory effects, including the modulation of pro-inflammatory cytokines and the alteration of neutrophil function. The effects are time- and dose-dependent, and the mechanisms responsible for these phenomena remain incompletely understood. Recent studies, mostly using the veterinary macrolide tilmicosin, may have shed new light on the mode of action of some macrolides and their anti-inflammatory properties. Indeed, research findings demonstrate that this compound, amongst others, induces neutrophil apoptosis, which in turn provides anti-inflammatory benefits. Studies using tilmicosin model systems in vitro and in vivo demonstrate that this antibiotic has potent immunomodulatory effects that may explain why at least parts of its clinical benefits are independent of anti-microbial effects. More research is needed, using this antibiotic and others that may have similar properties, to clarify the biological mechanisms responsible for antibiotic-induced neutrophil apoptosis, and how this, in turn, may provide enhanced clinical benefits. Such studies may help establish a rational basis for the development of novel, efficacious, anti-microbial compounds that generate anti-inflammatory properties in addition to their antibacterial effects.
Immuno-modulation and anti-inflammatory benefits of antibiotics: The example of tilmicosin
2010-01-01
Exagerated immune responses, such as those implicated in severe inflammatory reactions, are costly to the metabolism. Inflammation and pro-inflammatory mediators negatively affect production in the...Full Text Available
Human SolCD39 Inhibits Injury-induced Development of Neointimal Hyperplasia
2010-02-01
SUMMARYBlood platelets provide the initial response to vascular endothelial injury, becoming activated as they adhere to the injured site. Activated platelets recruit leukocytes,...Full Text Available
Human SolCD39 Inhibits Injury-induced Development of Neointimal Hyperplasia
2010-02-01
Full Text Available.SUMMARYBlood platelets provide the initial response to vascular endothelial injury, becoming activated as they adhere to the injured site. Activated platelets recruit leukocytes, and initiate proliferation and migration of vascular smooth muscle cells (SMC) within the injured vessel wall, leading to development of neointimal hyperplasia. Endothelial CD39/NTPDase1 and recombinant solCD39 rapidly metabolize nucleotides, including stimulatory ADP released from activated platelets, thereby suppressing additional platelet reactivity. Using a murine model of vascular endothelial injury, we investigated whether circulating human solCD39 could reduce platelet activation and accumulation, thus abating leukocyte infiltration and neointimal formation following vascular damage. Intraperitoneally-administered solCD39 ADPase activity in plasma peaked 1 hr post-injection, with an elimination half-life of 43 hr. Accordingly, mice were administered solCD39 or saline 1 hr prior to vessel injury, then either sacrificed 24 hr post-injury or treated with solCD39 or saline (3X weekly) for an additional 18 days. 24 hr post-injury, solCD39-treated mice displayed a reduction in platelet activation and recruitment, P-selectin expression, and leukocyte accumulation in the arterial lumen. Furthermore, repeated administration of solCD39 modulated the late stage of vascular injury by suppressing leukocyte deposition, macrophage infiltration and SMC proliferation/migration, resulting in abrogation of neointimal thickening. In contrast, injured femoral arteries of saline-injected mice exhibited massive platelet thrombus formation, marked P-selectin expression, and leukocyte infiltration. Pronounced neointimal growth with macrophage and SMC accretion was also observed (intimal-to-medial area ratio 1.56±0.34 at 19 days). Thus, systemic administration of solCD39 profoundly affects injury-induced cellular responses, minimizing platelet deposition and leukocyte recruitment, and suppressing neointimal hyperplasia.
2007-01-01
Among the enzymes involved in the hydrolysis of membrane phospholipids, type-IIA secreted phospholipase A2 (sPLA2-IIA) plays a major role in the development of a number of inflammatory diseases. Indeed, this enzyme has been shown to release arachidonic acid, the precursor of pro-inflammatory eicosanoids, and to hydrolyze phospholipids of pulmonary surfactant. This hydrolysis, which alters the tensioactive properties of surfactant phospholipids, leading to alveolar collapse, plays a critical role in the pathogenesis of acute respiratory distress syndrome. sPLA2-IIA has also been shown to bind to specific receptors located on cell surface membranes, although the biological significance of this binding remains unclear. However, the most well-established biological role of sPLA2-IIA is related...
Full Text Available.BackgroundA hallmark of the prion diseases is the conversion of the host-encoded cellular prion protein (PrPC) into a disease related, alternatively folded isoform (PrPSc). The accumulation of PrPSc within the brain is associated with synapse loss and ultimately neuronal death. Novel therapeutics are desperately required to treat neurodegenerative diseases including the prion diseases.Principal FindingsTreatment with glimepiride, a sulphonylurea approved for the treatment of diabetes mellitus, induced the release of PrPC from the surface of prion-infected neuronal cells. The cell surface is a site where PrPC molecules may be converted to PrPSc and glimepiride treatment reduced PrPSc formation in three prion infected neuronal cell lines (ScN2a, SMB and ScGT1 cells). Glimepiride also protected cortical and hippocampal neurones against the toxic effects of the prion-derived peptide PrP82–146. Glimepiride treatment significantly reduce both the amount of PrP82–146 that bound to neurones and PrP82–146 induced activation of cytoplasmic phospholipase A2 (cPLA2) and the production of prostaglandin E2 that is associated with neuronal injury in prion diseases. Our results are consistent with reports that glimepiride activates an endogenous glycosylphosphatidylinositol (GPI)-phospholipase C which reduced PrPC expression at the surface of neuronal cells. The effects of glimepiride were reproduced by treatment of cells with phosphatidylinositol-phospholipase C (PI-PLC) and were reversed by co-incubation with p-chloromercuriphenylsulphonate, an inhibitor of endogenous GPI-PLC.ConclusionsCollectively, these results indicate that glimepiride may be a novel treatment to reduce PrPSc formation and neuronal damage in prion diseases.
BackgroundA hallmark of the prion diseases is the conversion of the host-encoded cellular prion protein (PrPC) into a disease related, alternatively folded isoform (PrPSc)....Full Text Available
Four-color laser irradiation system for laser-plasma interaction experiments
1996-06-01
Since 1986, optical smoothing of the laser irradiance on targets for Inertial Confinement Fusion (ICF) has gained increasing attention. Optical smoothing can significantly reduce wavefront aberrations that produce nonuniformities in the energy distribution of the focal spot. Hot spots in the laser irradiance can induce local self focusing of the light, producing filamentation of the plasma. Filamentation can have detrimental consequences on the hydrodynamics of an ICF plasma, and can affect the growth of parametric instabilities, as well as add to the complexity of the study of such instabilities as stimulated Brillouin scattering (SBS) and stimulated Raman scattering (SRS). As experiments approach and exceed breakeven (i.e., where driver energy = fusion yield), the likelihood of significant excitation of these processes increases. As a result, the authors are including a scheme for implementing optical-beam smoothing for target experiments in the baseline design for the proposed next-generation ICF facility--the National Ignition Facility (NIF). To verify the efficacy of this design for the suppression of parametric instabilites in NIF-like indirect-drive targets, the authors successfully modified a Nova beamline to simulate the proposed NIF conditions. In this article, they discuss the laser science associated with a four-color target campaign on Nova to test the effect of f-number (ratio of focal length to beam diameter) and temporal smoothing on the scaling of SBS with a four-segment interaction beam using NIF-like parameters. The results of the target series associated with the four-color configuration are discussed elsewhere.
First operation of a dielectric-loaded double-stripline free-electron maser experiment
1995-12-31
A tabletop free-electron maser (FEM) experiment based on a dielectric-loaded double-stripline waveguide is presented. It employs a low-energy (8 keV, 0.5 A) electron beam and a folded-foil wiggler ({lambda}w = 2 cm). Metal striplines protects the dielectric slabs from the electron beam and support quasi-TEM modes in the waveguide. Radiation output is observed at f = 3.5 GHz, in agreement with the dielectric-loaded FEM tuning relation.
Expression of the p16{sup INK4a} tumor suppressor gene in rodent lung tumors
1995-12-01
Aberrations on the short arm of chromosome 9 are among the earliest genetic changes in human cancer. p16{sup INK4a} is a candidate tumor suppressor gene that lies within human 9p21, a chromosome region associated with frequent loss of heterozygosity in human lung tumors. The p16{sup INK4a} protein functions as an inhibitor of cyclin D{sub 1}-dependent kinases that phosphorylate the retinoblastoma (Rb) tumor suppressor gene product enabling cell-cycle progression. Thus, overexpression of cyclin D{sub 1}, mutation of cyclin-dependent kinase genes, or loss of p16{sup INK4a} function, can all result in functional inactivation of Rb. Inactivation of Rb by mutation or deletion can result in an increase in p16{sup INK4a} transcription, suggesting that an increased p16{sup INK4a} expression in a tumor cell signals dysfunction of the pathway. The p16{sup (INK4a)} gene, unlike some tumor suppressor genes, is rarely inactivated by mutation. Instead, the expression of this gene is suppressed in some human cancers by hypermethylation of the CpG island within the first exon or by homozygous deletion: 686. Chromosome losses have been observed at 9p21 syntenic loci in tumors of the mouse and rat, two species often used as animal models for pulmonary carcinogenesis. Expression of p16{sup INK4a} is lost in some mouse tumor cell lines, often due to homozygous deletion. These observations indicate that p16{sup INK4a} dysfunction may play a role in the development of neoplasia in rodents as well as humans. The purpose of the current investigation was to define the extent to which p16{sup INK4a} dysfunction contributes to the development of rodent lung tumors and to determine the mechanism of inactivation of the gene. There is no evidence to suggest a loss of function of the p16{sup INK4a} tumor suppressor gene in these primary murine lung tumors by mutation, deletion, or methylation.
1995-12-01
Tumors arising from epithelial cells, including lung cancers are frequently resistant to factors that regulate growth and differentiation in normal in normal cells. Once such factor is transforming growth factor-{Beta} (TGF-{Beta}). Escape from the growth-inhibitory effects of TGF-{Beta} is thought to be a key step in the transformation of airway epithelial cells. most lung cancer cell lines require serum for growth. In contrast, normal human bronchial epithelial (NHBE) cells are exquisitely sensitive to growth-inhibitory and differentiating effects of TGF-{Beta}. The recent identification of a novel cyclin-dependent kinase inhibitor, p15{sup INK4B}, which is regulated by TGF-{Beta}, suggests a mechanism by which TGF-{Beta} mediates growth arrest in NHBE cells. The purpose of this study was two-fold: (1) to determine if p15{sup INK4B} is induced by TGF-{Beta} in NHBE cells or immortalized bronchial epithelial (R.1) cells and if that induction corresponds to a G1/S cell-cycle arrest; (2) to determine the temporal relationship between p15{sup INK4B} induction, cell-cycle arrest, and the phosphorylation state of the pRB because it is thought that p15{sup INK4B} acts indirectly by preventing phosphorylation of the RB gene product. In this study, expression of p15{sup INK4B} was examined in NHBE cells and R.1 cells at different time intervals following TGF-{Beta} treatment. The expression of this kinase inhibitor and its relationship to the cell and the pRb phosphorylation state were examined in cells that were both sensitive (NHBE) and resistant (R.1) to the effects of TGF-{Beta}. These results suggest that the cyclin-dependent kinase inhibitor, p15{sup INK4B}, is involved in airway epithelial cell differentiation and that loss or reduction of expression plays a role in the resistance of transformed or neoplastic cells to the growth-inhibitory effects of TGF-{Beta}.
Evolution of toxicology information systems
1990-12-31
Society today is faced with new health risk situations that have been brought about by recent scientific and technical advances. Federal and state governments are required to assess the many potential health risks to exposed populations from the products (chemicals) and by-products (pollutants) of these advances. Because a sound analysis of any potential health risk should be based on the use of relevant information, it behooves those individuals responsible for making the risk assessments to know where to obtain needed information. This paper reviews the origins of toxicology information systems and explores the specialized information center concept that was proposed in 1963 as a means of providing ready access to scientific and technical information. As a means of illustrating this concept, the operation of one specialized information center (the Environmental Mutagen Information Center at Oak Ridge National Laboratory) will be discussed. Insights into how toxicological information resources came into being, their design and makeup, will be of value to those seeking to acquire information for risk assessment purposes. 7 refs., 1 fig., 4 tabs.
1990-12-31
Three areas are addressed in this paper: generic issues that arise simply in the process of decision-making under environmental statutes; different decision-making standards under various environmental statutes; and efforts to legislate a {open_quotes}safe{close_quotes} or {open_quotes}acceptable{close_quotes} risk from exposure to carcinogenic chemicals.
2010-03-01
The human epidermis serves 2 crucial barrier functions: it protects against water loss and prevents penetration of infectious agents and allergens. The physiology of the epidermis is maintained by a...Full Text Available
2010-03-01
Full Text Available.The human epidermis serves 2 crucial barrier functions: it protects against water loss and prevents penetration of infectious agents and allergens. The physiology of the epidermis is maintained by a balance of protease and antiprotease activities, as illustrated by the rare genetic skin disease Netherton syndrome (NS), in which impaired inhibition of serine proteases causes severe skin erythema and scaling. Here, utilizing mass spectrometry, we have identified elastase 2 (ELA2), which we believe to be a new epidermal protease that is specifically expressed in the most differentiated layer of living human and mouse epidermis. ELA2 localized to keratohyalin granules, where it was found to directly participate in (pro-)filaggrin processing. Consistent with the observation that ELA2 was hyperactive in skin from NS patients, transgenic mice overexpressing ELA2 in the granular layer of the epidermis displayed abnormal (pro-)filaggrin processing and impaired lipid lamellae structure, which are both observed in NS patients. These anomalies led to dehydration, implicating ELA2 in the skin barrier defect seen in NS patients. Thus, our work identifies ELA2 as a major new epidermal protease involved in essential pathways for skin barrier function. These results highlight the importance of the control of epidermal protease activity in skin homeostasis and designate ELA2 as a major protease driving the pathogenesis of NS.
Full Text Available.BackgroundHuntington's disease (HD) is a polyglutamine-expanded related neurodegenerative disease. Despite the ubiquitous expression of expanded, polyQ-Huntingtin (ExpHtt) in the brain, striatal neurons present a higher susceptibility to the mutation. A commonly admitted hypothesis is that Dopaminergic inputs participate to this vulnerability. We previously showed that D2 receptor stimulation increased aggregate formation and neuronal death induced by ExpHtt in primary striatal neurons in culture, and chronic D2 antagonist treatment protects striatal dysfunctions induced by ExpHtt in a lentiviral-induced model system in vivo. The present work was designed to elucidate the signalling pathways involved, downstream D2 receptor (D2R) stimulation, in striatal vulnerability to ExpHtt.Methodology/Principal FindingsUsing primary striatal neurons in culture, transfected with a tagged-GFP version of human exon 1 ExpHtt, and siRNAs against D2R or D1R, we confirm that DA potentiates neuronal dysfunctions via D2R but not D1R stimulation. We demonstrate that D2 agonist treatment induces neuritic retraction and growth cone collapse in Htt- and ExpHtt expressing neurons. We then tested a possible involvement of the Rho/ROCK signalling pathway, which plays a key role in the dynamic of the cytoskeleton, in these processes. The pharmacological inhibitors of ROCK (Y27632 and Hydroxyfasudil), as well as siRNAs against ROCK-II, reversed D2-related effects on neuritic retraction and growth cone collapse. We show a coupling between D2 receptor stimulation and Rho activation, as well as hyperphosphorylation of Cofilin, a downstream effector of ROCK-II pathway. Importantly, D2 agonist-mediated potentiation of aggregate formation and neuronal death induced by ExpHtt, was totally reversed by Y27632 and Hydroxyfasudil and ROCK-II siRNAs.Conclusions/SignificanceOur data provide the first demonstration that D2R-induced vulnerability in HD is critically linked to the activation of the Rho/ROCK signalling pathway. The inclusion of Rho/ROCK inhibitors could be an interesting therapeutic option aimed at forestalling the onset of the disease.
BackgroundHuntington's disease (HD) is a polyglutamine-expanded related neurodegenerative disease. Despite the ubiquitous expression of expanded, polyQ-Huntingtin (ExpHtt) in the...Full Text Available
Corrosion inhibition by control of gas composition during mist drilling
1981-05-01
Chemical compositional specifications have been generated for inert gases which reduce drill string corrosion when used in conjunction with mist drilling processes. These specifications are based on the assumption that the corrosion rate is dependent on the dissolved gaseous species concentrations. Data taken both from the literature and from a mist drilling field test with nitrogen in Valle Grande, NM, relate corrosion rates to fluid compositions. These solution compositions are then associated with gas phase compositions using equilibrium data available from the literature and material balances. Two sources of gas were considered: cryogenically purified nitrogen from air and exhaust gas from a diesel engine, which contain (in addition to N/sub 2/ and O/sub 2/) CO/sub 2/, NO/sub x/, SO/sub 2/, H/sub 2/O, and CO. A maximum concentration of 50 ppM O/sub 2/ in the gas phase is recommended to alleviate pitting corrosion.
Coherent transient grating effects and auger inhibition in InAsSb systems
1995-12-31
Pump-probe measurements of interband recombination lifetimes have been performed with the Free Electron Laser (CLIO) at room temperature undoped bulk InSb. Significant bleaching near and below the fundamental absorption edge at 7{mu}m was seen near the excitation frequency, with recovery times in the range 0.2-5 ns which were found to be strongly dependent on the pump photon energy. The scattering is dominated by Auger processes, which have rates following quadratic or linear carrier density dependence in low excitation and highly degenerate regimes respectively. The coefficients for Auger recombination in InSb at room temperature were found to be 1.1{+-}0.5x10{sup -26} cm{sup 6}s{sup -1} and 4.0{+-}0.5x 10{sup -9} cm{sup 3}s{sup -1} in these two regimes. These experiments also reveal associated coherent transient grating effects for the first time in these systems. A parametric scattering of the pump pulse into the probe beam is observed at delay times smaller than the coherence length of the FEL which allows us to determine the third-order nonlinear susceptibility and the coherence length of the laser system. A preliminary bleaching experiment on an undoped strained layer superlattice (SLS) sample of InAs/InAs{sub 0.61} Sb{sub 0.39} is also reported. It is well known that the narrower the bandgap in HgCdTe alloys the easier energy and momentum conservation becomes. This SLS structure (band edge 11I{mu}m) shows strong inhibition of the Auger recombination process with lifetimes up to 30 times longer than even the bulk InSb sample (7{mu}m). This opens the possibility of a major leap into the IR for III-V semiconductor light-emitting and detection device applications.
Cancer Vaccine by Fusions of Dendritic and Cancer Cells
2009-01-01
Full Text Available.Dendritic cells (DCs) are potent antigen-presenting cells and play a central role in the initiation and regulation of primary immune responses. Therefore, their use for the active immunotherapy against cancers has been studied with considerable interest. The fusion of DCs with whole tumor cells represents in many ways an ideal approach to deliver, process, and subsequently present a broad array of tumor-associated antigens, including those yet to be unidentified, in the context of DCs-derived costimulatory molecules. DCs/tumor fusion vaccine stimulates potent antitumor immunity in the animal tumor models. In the human studies, T cells stimulated by DC/tumor fusion cells are effective in lysis of tumor cells that are used as the fusion partner. In the clinical trials, clinical and immunological responses were observed in patients with advanced stage of malignant tumors after being vaccinated with DC/tumor fusion cells, although the antitumor effect is not as vigorous as in the animal tumor models. This review summarizes recent advances in concepts and techniques that are providing new impulses to DCs/tumor fusions-based cancer vaccination.
Cancer Vaccine by Fusions of Dendritic and Cancer Cells
2009-01-01
Dendritic cells (DCs) are potent antigen-presenting cells and play a central role in the initiation and regulation of primary immune responses. Therefore, their use for the active immunotherapy against...Full Text Available
COX-2 in liver, from regeneration to hepatocarcinogenesis: What we have learned from animal models?
2010-03-28
The use of animals lacking genes or expressing genes under the control of cell-specific promoters has significantly increased our knowledge of the genetic and molecular basis of physiopathology, allowing...Full Text Available
COX-2 in liver, from regeneration to hepatocarcinogenesis: What we have learned from animal models?
2010-03-28
Full Text Available.The use of animals lacking genes or expressing genes under the control of cell-specific promoters has significantly increased our knowledge of the genetic and molecular basis of physiopathology, allowing testing of functional hypotheses and validation of biochemical and pharmacologic approaches in order to understand cell function. However, with unexpected frequency, gene knockout animals and, more commonly, animal models of transgenesis give experimental support to even opposite conclusions on gene function. Here we summarize what we learned on the role of cyclooxygenase 2 (COX-2) in liver and revise the results obtained in 3 independent models of mice expressing a COX-2 transgene specifically in the hepatocyte. Upon challenge with pro-inflammatory stimuli, the animals behave very differently, some transgenic models having a protective effect but others enhancing the injury. In addition, one transgene exerts differential effects on normal liver physiology depending on the transgenic animal model used.
Behçet’s Disease as a Model of Venous Thrombosis
Full Text Available.Behçet’s disease (BD) is a chronic inflammatory disease of unknown aetiology characterized by recurrent oral, genital aphthous ulcerations, uveitis, skin lesions and other multisystem affections associated with vasculitis. Different types of vessels, predominantly veins, can be affected in BD. The frequency of vascular lesions in BD, such as superficial and deep venous thromboses, arterial aneurysms and occlusions, ranges between 7-29%. In this review, various factors of thrombogenesis in BD, particularly pro- and antithrombotic endothelial and non-endothelial factors, factors of coagulation, platelet activation and rheological changes are presented and discussed from positions of Virchow’s triad of venous thrombosis. Despite advances in understanding of thrombogenesis in BD, still many issues of diagnosis and targeted preventive and therapeutic measures remain unresolved. Further studies are needed to clarify the pathobiology of BD-related thrombosis and to provide the clinicians with recommendations over the utility, safety and effectiveness of the antithrombotic therapy in BD.
Behçet’s Disease as a Model of Venous Thrombosis
Behçet’s disease (BD) is a chronic inflammatory disease of unknown aetiology characterized by recurrent oral, genital aphthous ulcerations, uveitis, skin lesions and other multisystem...Full Text Available
1995-12-01
The existence of a nose-brain barrier that functions to protect the central nervous system (CNS) from inhaled toxicants has been postulated. Just as a blood-brain barrier protects the CNS from systemic toxicants, the nose-brain barrier may have similar characteristic functions. One component of interest is nasal xenobiotic metabolism and its effect on the transport of pollutants into the CNS at environmentally plausible levels of exposure. Previous results have shown that inhaled xylene are dimethyl phenol (DMP) and methyl benzyl alcohol (MBA), and the nonvolatile metabolites are toluic acid (TA) and methyl hippuric acid (MHA). The nonvolatile metabolites of xylene, along with a small quantity of volatiles, representing either parent xylene or volatile metabolites, are transported via the olfactory epithelium to the glomeruli within the olfactory bulbs of the brain. Further work will be done to establish the linearity for each analyte at the actual highest detection limit of the GC/MS.
Altered expression of the IQGAP1 gene in human lung cancer cell lines
1995-12-01
IQGAP1 is a GTPase activation protein that accelerates GTP hydrolysis by normal p21 ras proteins. Therefore, IQGAP1 could act as an upstream affector of p21 ras activity by convert in excess amounts of active GTP-21 ras to inactive GDP-21 ras. IQGAP1 displays extensive sequence similarity to the catalytic domain of all previously reported ras GAPs, including the tumor suppressor gene protein neurofibromatosis type 1 (NF1). It has been shown that abnormal NF1 protein cannot negatively regulate the activity of ras proteins in neuroblast cells. This observation supports the hypothesis that NF1 is a tumor suppressor gene whose product acts upstream of ras. IQGAP1 is primarily expressed in lung, where it may play a role similar to NF1 in regulating the activity of H-ras or K-ras proteins. IQGAP1 functions as other GAPs by controlling the activity of ras.
Alleviation of High-Fat Diet-Induced Fatty Liver Damage in Group IVA Phospholipase A2-Knockout Mice
Hepatic fat deposition with hepatocellular damage, a feature of non-alcoholic fatty liver disease, is mediated by several putative factors including prostaglandins. In the present study, we examined...Full Text Available
Alleviation of High-Fat Diet-Induced Fatty Liver Damage in Group IVA Phospholipase A2-Knockout Mice
Full Text Available.Hepatic fat deposition with hepatocellular damage, a feature of non-alcoholic fatty liver disease, is mediated by several putative factors including prostaglandins. In the present study, we examined whether group IVA phospholipase A2 (IVA-PLA2), which catalyzes the first step in prostanoid biosynthesis, is involved in the development of fatty liver, using IVA-PLA2-knockout mice. Male wild-type mice on high-fat diets (20% fat and 1.25% cholesterol) developed hepatocellular vacuolation and liver hypertrophy with an increase in the serum levels of liver damage marker aminotransferases when compared with wild-type mice fed normal diets. These high-fat diet-induced alterations were markedly decreased in IVA-PLA2-knockout mice. Hepatic triacylglycerol content was lower in IVA-PLA2-knockout mice than in wild-type mice under normal dietary conditions. Although high-fat diets increased hepatic triacylglycerol content in both genotypes, the degree was lower in IVA-PLA2-knockout mice than in wild-type mice. Under the high-fat dietary conditions, IVA-PLA2-knockout mice had lower epididymal fat pad weight and smaller adipocytes than wild-type mice. The serum level of prostaglandin E2, which has a fat storage effect, was lower in IVA-PLA2-knockout mice than in wild-type mice, irrespective of the kind of diet. In both genotypes, high-fat diets increased serum leptin levels equally between the two groups, but did not affect the serum levels of adiponectin, resistin, free fatty acid, triacylglycerol, glucose, or insulin. Our findings suggest that a deficiency of IVA-PLA2 alleviates fatty liver damage caused by high-fat diets, probably because of the lower generation of IVA-PLA2 metabolites, such as prostaglandin E2. IVA-PLA2 could be a promising therapeutic target for obesity-related diseases including non-alcoholic fatty liver disease.
Air pollution and ecology. From local to global impacts
1996-11-01
Human impact on nature is increasing - not only in magnitude, but also in geographical scale. It has been known for centuries, that vegetation does not thrive near air pollution sources, but it was not before after the Second World War that the importance of long range transport of pollutants was realized - first for sulphur and nitrogen compounds, later for photochemical oxidants. The results have been acidification of rivers and lakes, forest dieback and eutrophication of inner waters. In recent decades the attention has been focused on global effects: Ozone depletion and increased greenhouse effect. Here air pollution threatens to alter the conditions of life on the entire Earth. In the political and public debate - and sometimes in science as well - the problems are treated separately. Since however, the basic phenomena all take place in the same atmosphere, they are more or less interrelated. Also the environmental effects must be considered a result of a complex impact. This complexity should be taken into account in the planning of an effective abatement strategy. (au) 11 refs.
2010-01-01
Full Text Available. Aim. Investigate the promoter methylation of the Thrombospondin-1 (TSP1) gene in gastric cardia adenocarcinoma (GCA). Methods. MSP approach, immunohistochemistry method, and RT-PCR were used respectively to examine the promoter methylation of TSP1, its protein and mRNA expression in tumors and corresponding normal tissues. The expression and concentration of TGF-β1 were examined respectively by immunohistochemistry and ELISA method. The status of T cell immunity was examined by Flow cytometry analysis. Results. TSP1 was methylated in 34/96 (35.4%) tumor specimens, which was significantly higher than that in corresponding normal tissues (P < .001). Protein and mRNA expression of TSP1 in GCA tumor tissues were reduced significantly and were associated with TSP1 methylation. The protein expression of TGF-β1 was significantly higher in tumor tissues (P < .001) and was associated with TNM stage and histological differentiation. The concentration of active and total TGF-β1 did not show significant difference between the GCA patients with hypermethylation of TSP1 and without methylation of TSP1 (P > .05). The function of T cell immunity was significantly different between the GCA patients with hypermethylation of TSP1 and without methylation of TSP1. Conclusions. Epigenetic silencing of TSP1 gene by promoter hypermethylation may play an important role in GCA.
2010-01-01
Aim. Investigate the promoter methylation of the Thrombospondin-1 (TSP1) gene in gastric cardia adenocarcinoma (GCA). Methods. MSP approach, immunohistochemistry...Full Text Available
2008-01-01
Abstract Background and Aim: We previously reported that hypothermic machine perfusion (HMP) for liver preservation is feasible, but hepatic microcirculatory dysfunction and significant liver damage remain major obstacles in its application when the preservation is extended to 24 h. The underlying injury mechanism is not well understood. The present study sought to investigate the role of thromboxane A2 (TXA2) in the pathogenesis of liver injury after prolonged HMP. Methods: Livers isolated from Sprague-Dawley rats were subjected to continuous machine perfusion with University of Wisconsin (UW) solution at a flow rate of 0.4 mL/min/g liver at 4degreeC for 24 h. A specific TXA2 synthase inhibitor, OKY-046 (OKY), was added to UW solution during the preservation period and to the Krebs-Hensel...
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