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Find video protocols related to scientific articles indexed in Pubmed.
Biological Monitoring Method for Urinary Neonicotinoid Insecticides Using LC-MS/MS and Its Application to Japanese Adults.
J Occup Health
PUBLISHED: 11-07-2014
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Objectives: Agricultural use of neonicotinoid (NEO) insecticides has been increasing in recent years, but their biological monitoring methods have been scarcely reported. In this study, we developed and validated a rapid and sensitive method for quantifying urinary NEO concentrations using high-performance liquid chromatography-tandem mass spectrometry (LC-MS/MS). Methods: After phosphate-induced acidification of a urine sample, urinary NEOs were trapped by a solid-phase extraction column and eluted with methanol for acetamiprid, imidacloprid, thiacloprid, thiamethoxam, clothianidin and dinotefuran and with an acetonitrile and methanol solution (1:1, v/v) containing 5% NH3 for nitenpyram. A separation analysis was performed by LC-MS/MS within 10 min for the sample. This method was applied to first morning urine obtained from 52 Japanese (40.9 ± 10.5 years old, mean ± standard deviation) without occupational NEO exposure. Results: The linear dynamic ranges and their limit of quantification (LOQ, signal to noise ratio = 10) levels were 0.3-20 or 50 µg/l (r = 0.998-0.999) and 0.05-0.36 µg/l, respectively. The absolute recovery was above 64%, and the intra- and inter-day precisions were less than 16.4% (relative standard deviation, %RSD). This method was successfully applied for analysis of NEOs in human urine samples obtained from 52 adults. The frequencies of individuals who showed more than LOD levels was above 90% for imidacloprid, thiamethoxam, clothianidin and dinotefuran, more than 50% for acetamiprid and thiacloprid and 29% for nitenpyram. Conclusions: These results indicated that our new method could be applied to biological monitoring of NEO exposure even at environmental exposure levels in Japanese adults without occupational spraying histories.
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Ionic mechanisms underlying the negative chronotropic action of propofol on sinoatrial node automaticity in guinea-pig heart.
Br. J. Pharmacol.
PUBLISHED: 09-09-2014
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Propofol is a widely used intravenous anesthetic agent but has undesirable cardiac side effects, including bradyarrhythmia and its severe form asystole. This study examined the ionic and cellular mechanisms underlying propofol-induced bradycardia.
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Fenitrothion action at the endocannabinoid system leading to spermatotoxicity in Wistar rats.
Toxicol. Appl. Pharmacol.
PUBLISHED: 06-19-2014
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Organophosphate (OP) compounds as anticholinesterase agents may secondarily act on diverse serine hydrolase targets, revealing unfavorable physiological effects including male reproductive toxicity. The present investigation proposes that fenitrothion (FNT, a major OP compound) acts on the endocannabinoid signaling system in male reproductive organs, thereby leading to spermatotoxicity (sperm deformity, underdevelopment, and reduced motility) in rats. FNT oxon (bioactive metabolite of FNT) preferentially inhibited the fatty acid amide hydrolase (FAAH), an endocannabinoid anandamide (AEA) hydrolase, in the rat cellular membrane preparation from the testis in vitro. Subsequently, male Wistar rats were treated orally with 5 or 10mg/kg FNT for 9 weeks and the subchronic exposure unambiguously deteriorated sperm motility and morphology. The activity-based protein profiling analysis with a phosphonofluoridate fluorescent probe revealed that FAAH was selectively inhibited among the FNT-treated cellular membrane proteome in testis. Intriguingly, testicular AEA (endogenous substrate of FAAH) levels were elevated along with the FAAH inhibition caused by the subchronic exposure. More importantly, linear regression analyses for the FNT-elicited spermatotoxicity reveal a good correlation between the testicular FAAH activity and morphological indices or sperm motility. Accordingly, the present study proposes that the FNT-elicited spermatotoxicity appears to be related to inhibition of FAAH leading to overstimulation of the endocannabinoid signaling system, which plays crucial roles in spermatogenesis and sperm motility acquirement.
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Effects of exposure to Di(2-ethylhexyl)phthalate ?during fetal period on next generation.
Nihon Eiseigaku Zasshi
PUBLISHED: 05-27-2014
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The concept of the Developmental Origins of Health and Disease (DOHaD) is bringing new insights into the origin of lifestyle diseases: unbalanced nutrition in utero and during infancy is associated with an increased risk of lifestyle diseases. In order to clarify this association, experimental and epidemiological studies have been conducted. Maternal exposure to di(2-ethylhexyl)phthalate (DEHP), an agonist of peroxisome proliferator-activated receptor ? (PPAR?), decreases the number of live fetuses and newborn pups, and their body weights, and it enhances fetal desorption in wild-type mice. Similarly, these DEHP were also observed in mice expressing human PPAR?, but not in PPAR?-null mice. These results suggest that the DEHP toxicity in offspring is caused dependently on PPAR?. DEHP suppresses the increase in the levels of plasma triglyceride (TG)/fatty acids (FAs) only in wild-type pregnant mice, suggesting that the decreased lipid levels in utero may affect the fetus development, because TG/FAs are essential in the development of fetuses. Additionally, maternal DEHP exposure decreases estrogen and progesterone balances, which may also explain the effects on fetuses and pups mentioned above. Indeed, DEHP itself or metabolite(s) may induce the toxicity, because a difference in the metabolic route is observed between the wild-type and PPAR?-null mice. Thus, we were unable to conclude the causal factor(s) for the DEHP-induced offspring toxicity, that is, whether it is a direct or an indirect effect of the chemical or metabolite(s) via the toxic effects on maternal mice; however, PPAR? is indeed associated with in offspring toxicity.
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Pravastatin and olmesartan synergistically ameliorate renal failure-induced vascular calcification.
J. Atheroscler. Thromb.
PUBLISHED: 05-19-2014
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Vascular calcification is a critical problem in patients with chronic kidney disease (CKD). In this study, we examined the effects of a HMG Co-A reductase inhibitor (statin) and an angiotensin ? type 1 receptor blocker (ARB) on renal failure-induced vascular calcification.
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Engineered platform for bioethylene production by a cyanobacterium expressing a chimeric complex of plant enzymes.
ACS Synth Biol
PUBLISHED: 02-24-2014
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Ethylene is an industrially important compound, but more sustainable production methods are desirable. Since cellulosomes increase the ability of cellulolytic enzymes by physically linking the relevant enzymes via dockerin-cohesin interactions, in this study, we genetically engineered a chimeric cellulosome-like complex of two ethylene-generating enzymes from tomato using cohesin-dockerins from the bacteria Clostridium thermocellum and Acetivibrio cellulolyticus. This complex was transformed into Escherichia coli to analyze kinetic parameters and enzyme complex formation and into the cyanobacterium Synechococcus elongatus PCC 7942, which was then grown with and without 0.1 mM isopropyl ?-D-1-thiogalactopyranoside (IPTG) induction. Only at minimal protein expression levels (without IPTG), the chimeric complex produced 3.7 times more ethylene in vivo than did uncomplexed enzymes. Thus, cyanobacteria can be used to sustainably generate ethylene, and the synthetic enzyme complex greatly enhanced production efficiency. Artificial synthetic enzyme complexes hold great promise for improving the production efficiency of other industrial compounds.
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Direct negative chronotropic action of desflurane on sinoatrial node pacemaker activity in the guinea pig heart.
Anesthesiology
PUBLISHED: 02-13-2014
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Desflurane inhalation is associated with sympathetic activation and concomitant increase in heart rate in humans and experimental animals. There is, however, little information concerning the direct effects of desflurane on electrical activity of sinoatrial node pacemaker cells that determines the intrinsic heart rate.
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Preferential targeting of p39-activated Cdk5 to Rac1-induced lamellipodia.
Mol. Cell. Neurosci.
PUBLISHED: 01-28-2014
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Cdk5 is a member of the cyclin-dependent kinase (Cdk) family that plays a role in various neuronal activities including brain development, synaptic regulation, and neurodegeneration. Cdk5 requires the neuronal specific activators, p35 and p39 for subcellular compartmentalization. However, it is not known how active Cdk5 is recruited to F-actin cytoskeleton, which is a Cdk5 target. Here we found p35 and p39 localized to F-actin rich regions of the plasma membrane and investigated the underlying targeting mechanism in vitro by expressing them with Rho family GTPases in Neuro2A cells. Both p35 and p39 accumulated at the cell peripheral lamellipodia and perinuclear regions, where active Rac1 is localized. Interestingly, p35 and p39 displayed different localization patterns as p35 was found more at the perinuclear region and p39 was found more in peripheral lamellipodia. We then confirmed this distinct localization in primary hippocampal neurons. We also determined that the localization of p39 to lamellipodia requires myristoylation and Lys clusters within the N-terminal p10 region. Additionally, we found that p39-Cdk5, but not p35-Cdk5 suppressed lamellipodia formation by reducing Rac1 activity. These results suggest that p39-Cdk5 has a dominant role in Rac1-dependent lamellipodial activity.
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Defect of suppression of inflammasome-independent interleukin-8 secretion from SW982 synovial sarcoma cells by familial Mediterranean fever-derived pyrin mutations.
Mol. Biol. Rep.
PUBLISHED: 11-30-2013
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Familial Mediterranean fever (FMF) is a recessive inherited autoinflammatory syndrome. Patients with FMF have symptoms such as recurrent fever and abdominal pain, sometimes accompanied by arthralgia. Biopsy specimens have revealed substantial neutrophil infiltration into synovia. FMF patients have a mutation in the Mediterranean fever gene, encoding pyrin, which is known to regulate the inflammasome, a platform for processing interleukin (IL)-1?. FMF patients heterozygous for E148Q mutation, heterozygous for M694I mutation, or combined heterozygous for E148Q and M694I mutations, which were found to be major mutations in an FMF study group in Japan, suffer from arthritis, the severity of which is likely to be lower than in FMF patients with M694V mutations. Expression plasmids of wild-type (WT) pyrin and mutated pyrin, such as E148Q, M694I, M694V, and E148Q+M694I, were constructed, and SW982 synovial sarcoma cells were transfected with these expression plasmids. IL-8 and IL-6 were spontaneously secreted from the culture supernatant of SW982 cells without any stimulation, whereas IL-1? and TNF-? could not be detected even when stimulated with lipopolysaccharide. Notably, two inflammasome components, ASC and caspase-1, could not be detected in SW982 cells by Western blotting. IL-8 but not IL-6 secretion from SW982 cells was largely suppressed by WT pyrin, but less suppressed by mutated pyrin, which appeared to become weaker in the order of E148Q, M694I, E148Q+M694I, and M694V mutations. As for IL-8 and IL-6, similar results were obtained using stable THP-1 cells expressing the WT pyrin or mutated pyrins, such as M694V or E148Q, when stimulated by LPS. In addition, IL-8 secretion from mononuclear cells of FMF patients was significantly higher than that of healthy volunteers when incubated on a culture plate. Thus, our results suggest that IL-8 secretion from SW982 synovial sarcoma cells suppressed by pyrin independently of inflammasome is affected by pyrin mutations, which may reflect the activity in FMF arthritis.
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Organophosphate agents induce plasma hypertriglyceridemia in mouse via single or dual inhibition of the endocannabinoid hydrolyzing enzyme(s).
Toxicol. Lett.
PUBLISHED: 10-29-2013
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Diverse serine hydrolases including endocannabinoid metabolizing enzymes fatty acid amide hydrolase (FAAH) and monoacylglycerol lipase (MAGL) have been suggested as secondary targets for organophosphate (OP) agents to exert adverse toxic effects such as lipid homeostasis disruption. The goal of this investigation is to verify that a major OP insecticide fenitrothion (FNT) induces plasma hypertriglyceridemia through the inhibition of FAAH and/or MAGL in comparison with that elicited by isopropyl dodecylfluorophosphonate (IDFP), a potent FAAH/MAGL inhibitor. Fasted mice were treated intraperitoneally with FNT or IDFP and were subsequently sacrificed for evaluations of plasma triglyceride (TG) levels and liver FAAH/MAGL activities. Plasma TG levels were significantly enhanced by the FNT or IDFP treatment (1.7- or 4.8-fold, respectively) compared with that of vehicle control. The IDFP exposure reduced the liver FAAH and MAGL activities, whereas the FNT exposure led to the preferential FAAH inhibition. The brain acetylcholinesterase was almost unaffected by the FNT or IDFP treatment, thus leading to no neurotoxic sign. Intriguingly, the TG elevations were averted by concomitant administration with the cannabinoid receptor antagonist AM251. The present findings suggest that OP agents induce plasma hypertriglyceridemia in mouse through single or dual inhibition of FAAH or/and MAGL, apparently leading to overstimulation of cannabinoid signal regulating energy metabolism.
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Remifentanil has a minimal direct effect on sinoatrial node pacemaker activity in the Guinea pig heart.
Anesth. Analg.
PUBLISHED: 10-11-2013
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Whereas remifentanil administration is associated with severe bradycardia, it has yet to be fully investigated whether the negative chronotropic action of remifentanil is mediated by its direct action on sinoatrial (SA) node pacemaker activity in the heart versus indirect results of enhanced vagal activity.
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A potential target for organophosphate insecticides leading to spermatotoxicity.
J. Agric. Food Chem.
PUBLISHED: 10-01-2013
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Organophosphate (OP) insecticides as an anticholinesterase also act on the diverse serine hydrolase targets, thereby revealing secondary or unexpected toxic effects including male reproductive toxicity. The present investigation detects a possible target molecule(s) for OP-induced spermatotoxicity (sperm deformity, underdevelopment, and reduced motility) from a chemical standpoint. The activity-based protein profiling (ABPP) approach with a phosphonofluoridate fluorescent probe pinpointed the molecular target for fenitrothion (FNT, a major OP insecticide) oxon (bioactive metabolite of FNT) in the mouse testicular membrane proteome, i.e., FNT oxon phosphorylates the fatty acid amide hydrolase (FAAH), which plays pivotal roles in spermatogenesis and sperm motility acquirement. Subsequently, mice were treated orally with vehicle or FNT for 10 days, and FAAH activity in testis or epididymis cauda was markedly reduced by the subacute exposure. ABPP analysis revealed that FAAH was selectively inhibited among the FNT-treated testicular membrane proteome. Accordingly, FAAH is a potential target for OP-elicited spermatotoxicity.
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Sex Differences in Metabolism of Trichloroethylene and Trichloroethanol in Guinea Pigs.
J Occup Health
PUBLISHED: 09-10-2013
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Objectives: Trichloroethylene (TRI) has the potential to cause generalized dermatitis complicated with hepatitis. The guinea pig maximization test (GPMT) also suggests that both TRI and its metabolite trichloroethanol (TCE) exhibit immunogenicity and possible sex differences in guinea pigs. However, TRI and TCE metabolisms in guinea pigs have not been elucidated in detail. The first issue to clarify may be the sex differences in relation to the immunogenicity. Methods: We collected urine from Hartley male and female guinea pigs 24 hours after intracutaneous injection of TRI, TCE or trichloroacetic acid (TCA) during a GPMT and measured the urinary metabolites by gas chromatography-mass spectrometry. Results: After TRI treatment, the amount of TCA was significantly greater in females than males, while there was no sex difference in the total amount (TCA + TCE). TCA was only detected in urine after TCA treatment. Interestingly, not only TCE but also TCA was detected in urine of both sexes after TCE treatment, and the amount of TCA was also greater in females than males. An additional experiment showed that TCE treatment did not result in the detection of urinary TCA in cytochrome P450 (CYP)2E1-null mice but did in wild-type mice, suggesting the involvement of CYP2E1 in the metabolism from TCE to TCA. The constitutive expression of CYP2E1 in the liver of guinea pigs was greater in females than males. Conclusion: The sex difference in urinary TCA excretion after TRI and TCE treatments may be due to variation of the constitutive expression of CYP2E1.
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[The inflammasome].
Nippon Rinsho
PUBLISHED: 08-24-2013
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Inflammation is a protective response intended to eliminate primary causes of tissue injury, leading to tissue repair and adaptation in a focal lesion. It is thought that various factors are involved in the histopathological responses to tissue injury. One of the inflammatory cytokines, interleukin(IL)-1beta, is thought to play an important role in inflammatory responses. The inflammasome is a multi-protein complex, required for IL-1beta processing and activation to induce inflammation. Over the past decade, evidence has accumulated that the inflammasome contributes significantly to the pathogenesis of various diseases. In this review, we discuss the function of inflammasome and inflammasome-related diseases, that are now expanding beyond inflammation.
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Occupational trichloroethylene hypersensitivity syndrome: Human herpesvirus 6 reactivation and rash phenotypes.
J. Dermatol. Sci.
PUBLISHED: 07-08-2013
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Trichloroethylene (TCE) is an industrial solvent which can cause severe generalized dermatitis, i.e., occupational TCE hypersensitivity syndrome. Reactivation of latent human herpesvirus 6 (HHV6) can occur in such patients, which has made TCE known as a causative chemical of drug-induced hypersensitivity syndrome (DIHS).
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Species and inter-individual differences in metabolic capacity of di(2-ethylhexyl)phthalate (DEHP) between human and mouse livers.
Environ Health Prev Med
PUBLISHED: 06-28-2013
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This study was conducted to assess inter-species and inter-individual differences in the metabolism of di(2-ethylhexyl)phthalate (DEHP) in humans and mice.
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Pyramidanes.
J. Am. Chem. Soc.
PUBLISHED: 06-05-2013
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Pyramidane is an elusive but highly desirable target for synthetic chemists that has attracted a great deal of attention because of its nonclassical structure and unusual bonding features. Although well studied on theoretical grounds, neither the parent all-carbon pyramidane nor its derivatives containing heavier group 14 elements have ever been isolated and characterized. In this Communication, we report on the synthesis and structural elucidation of the first stable representatives of this class of highly strained polyhedral compounds: germa- and stannapyramidanes Ge[C4(SiMe3)4] and Sn[C4(SiMe3)4]. The peculiar structural and bonding features of these compounds are verified by combined experimental and computational analyses, showing these derivatives to be nonclassical neutral compounds with a very large contribution of ionic character.
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Anticholinesterase insecticide action at the murine male reproductive system.
Bioorg. Med. Chem. Lett.
PUBLISHED: 06-01-2013
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The present report describes for the first time that anticholinesterase type insecticides specifically inhibit the fatty acid amide hydrolase and/or monoacylglycerol lipase, as secondary target(s), in the murine male reproductive system (testis and epididymis cauda), thereby presumably being involved with spermatotoxicity such as deformity, underdevelopment, and reduced motility.
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Nectin-2 is a potential target for antibody therapy of breast and ovarian cancers.
Mol. Cancer
PUBLISHED: 05-31-2013
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Nectin-2 is a Ca(2+)-independent cell-cell adhesion molecule that is one of the plasma membrane components of adherens junctions. However, little has been reported about the involvement of Nectin-2 in cancer.
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Spirobis(pentagerma[1.1.1]propellane): a stable tetraradicaloid.
J. Am. Chem. Soc.
PUBLISHED: 04-23-2013
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In this contribution, we report a spirobis(pentagerma[1.1.1]propellane) derivative as a novel type of molecular architecture in cluster chemistry that features two spiro-fused [1.1.1]propellane units and represents a stable tetraradicaloid species. The crucial issue of the nature of the interaction between the germanium bridgeheads was probed computationally, revealing weak bonding interactions between the formally unpaired electrons.
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Importance of rabies virus nucleoprotein in viral evasion of interferon response in the brain.
Microbiol. Immunol.
PUBLISHED: 03-07-2013
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By using a cultured neuroblastoma cell line, the present authors recently showed that the N protein of virulent rabies virus fixed strain Nishigahara (Ni), but not that of the attenuated derivative Ni-CE, mediates evasion of induction of type I interferon (IFN). In this study, to determine whether Ni N protein indeed fulfills this function in vivo, the abilities to suppress IFN responses in the mouse brain of Ni-CE and the virulent chimeric virus CE(NiN), which has the N gene from Ni in the genetic background of Ni-CE, were compared. It was demonstrated that CE(NiN) propagates and spreads more efficiently than does Ni-CE in the brain and that IFN response in brains infected with CE(NiN) is weaker than in those infected with Ni-CE. It was also shown that amino acids at positions 273 and 394 in the N protein, which are known as pathogenic determinants, affect the ability of the viruses to suppress IFN response in the brain. These findings strongly suggest that, in the brain, rabies virus N protein plays important roles in evasion of innate immune responses and thereby in efficient propagation and spread of virus leading to lethal outcomes of infection.
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Exposure to DEHP decreased four fatty acid levels in plasma of prepartum mice.
Toxicology
PUBLISHED: 02-11-2013
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Maternal exposure to di(2-ethylhexyl) phthalate (DEHP) decreased the plasma triglyceride in prepartum mice. To identify the fatty acid (FA) species involved and to understand the underlying mechanisms, pregnant Sv/129 wild-type (mPPAR?), peroxisome proliferator-activated receptor ?-null (Ppar?-null) and humanized PPAR? (hPPAR?) mice were treated with diets containing 0%, 0.01%, 0.05% or 0.1% DEHP. Dams were dissected on gestational day 18 together with fetuses, and on postnatal day 2 together with newborns. n-3/n-6 polyunsaturated, saturated, and monounsaturated FAs in maternal plasma and in liver of wild-type offspring, and representative enzymes for FA desaturation and elongation in maternal liver, were measured. The plasma levels of linoleic acid, ?-linolenic acid, palmitic acid and oleic acid were higher in the pregnant control mPPARa mice than in Ppara-null and hPPARa mice. DEHP exposure significantly decreased the levels of these four FAs only in pregnant mPPAR? mice. Plasma levels of many FAs were higher in pregnant mice than in postpartum ones in a genotype-independent manner, while it was lower in the livers of fetuses than pups. DEHP exposure slightly increased hepatic arachidonic acid, ?-linolenic acid, palmitoleic acid and oleic acid in fetuses, but not in pups. However, DEHP exposure did not clearly influence FA desaturase 1 and 2 nor elongase 2 and 5 expressions in the liver of all maternal mice. Taken together, the levels of plasma four FAs with shorter carbon chains were higher in pregnant mPPAR? mice than in other genotypes, and DEHP exposure decreased these specific FA concentrations only in mPPAR? mice, similarly to triglyceride levels.
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Room-temperature synthesis of two-dimensional ultrathin gold nanowire parallel array with tunable spacing.
Langmuir
PUBLISHED: 01-24-2013
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A series of long-chain amidoamine derivatives with different alkyl chain lengths (CnAA where n is 12, 14, 16, or 18) were synthesized and studied with regard to their ability to form organogels and to act as soft templates for the production of Au nanomaterials. These compounds were found to self-assemble into lamellar structures and exhibited gelation ability in some apolar solvents. The gelation concentration, gel-sol phase transition temperature, and lattice spacing of the lamellar structures in organic solvent all varied on the basis of the alkyl chain length of the particular CnAA compound employed. The potential for these molecules to function as templates was evaluated through the synthesis of Au nanowires (NWs) in their organogels. Ultrathin Au NWs were obtained from all CnAA/toluene gel systems, each within an optimal temperature range. Interestingly, in the case of C12AA and C14AA, it was possible to fabricate ultrathin Au NWs at room temperature. In addition, two-dimensional parallel arrays of ultrathin Au NWs were self-assembled onto TEM copper grids as a result of the drying of dispersion solutions of these NWs. The use of CnAA compounds with differing alkyl chain lengths enabled precise tuning of the distance between the Au NWs in these arrays.
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Role of GalNAc4S-6ST in astrocytic tumor progression.
PLoS ONE
PUBLISHED: 01-17-2013
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N-Acetylgalactosamine 4-sulfate 6-O-sulfotransferase (GalNAc4S-6ST) is the sulfotransferase responsible for biosynthesis of highly sulfated chondroitin sulfate CS-E. Although involvements of CS-E in neuronal cell functions have been extensively analyzed, the role of GalNAc4S-6ST in astrocytic tumor progression remains unknown. Here, we reveal that GalNAc4S-6ST transcripts were detected in astrocytic tumors derived from all 30 patients examined using quantitative reverse transcription-PCR analysis. Patients with high GalNAc4S-6ST mRNA expression had significantly worse outcome compared with patients with low expression, and multivariate survival analysis disclosed that GalNAc4S-6ST is an independent poor prognostic factor for astrocytic tumors. We then tested whether CS-E enhanced haptotaxic migration of glioblastoma U251-MG cells that endogenously express both the CS-Es scaffold tyrosine phosphatase ? (PTP?) and GalNAc4S-6ST, in the presence of CS-Es preferred ligands, pleiotrophin (PTN) or midkine (MK), using a modified Boyden chamber method. Haptotaxic stimulation of cell migration by PTN was most robust on control siRNA-transfected U251-MG cells, while that enhancing effect was cancelled following transduction of GalNAc4S-6ST siRNA. Similar results were obtained using MK, suggesting that both PTN and MK enhance migration of U251-MG cells by binding to CS-E. We also found that PTP? as well as PTN and MK were frequently expressed in astrocytic tumor cells. Thus, our findings indicate that GalNAc4S-6ST mRNA expressed by astrocytic tumor cells is associated with poor patient prognosis likely by enhancing CS-E-mediated tumor cell motility in the presence of PTN and/or MK.
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Helicobacter pylori Cholesteryl ?-Glucosides Contribute to Its Pathogenicity and Immune Response by Natural Killer T Cells.
PLoS ONE
PUBLISHED: 01-01-2013
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Approximately 10-15% of individuals infected with Helicobacter pylori will develop ulcer disease (gastric or duodenal ulcer), while most people infected with H. pylori will be asymptomatic. The majority of infected individuals remain asymptomatic partly due to the inhibition of synthesis of cholesteryl ?-glucosides in H. pylori cell wall by ?1,4-GlcNAc-capped mucin O-glycans, which are expressed in the deeper portion of gastric mucosa. However, it has not been determined how cholesteryl ?-glucosyltransferase (?CgT), which forms cholesteryl ?-glucosides, functions in the pathogenesis of H. pylori infection. Here, we show that the activity of ?CgT from H. pylori clinical isolates is highly correlated with the degree of gastric atrophy. We investigated the role of cholesteryl ?-glucosides in various aspects of the immune response. Phagocytosis and activation of dendritic cells were observed at similar degrees in the presence of wild-type H. pylori or variants harboring mutant forms of ?CgT showing a range of enzymatic activity. However, cholesteryl ?-glucosides were recognized by invariant natural killer T (iNKT) cells, eliciting an immune response in vitro and in vivo. Following inoculation of H. pylori harboring highly active ?CgT into iNKT cell-deficient (J?18(-/-)) or wild-type mice, bacterial recovery significantly increased in J?18(-/-) compared to wild-type mice. Moreover, cytokine production characteristic of Th1 and Th2 cells dramatically decreased in J?18(-/-) compared to wild-type mice. These findings demonstrate that cholesteryl ?-glucosides play critical roles in H. pylori-mediated gastric inflammation and precancerous atrophic gastritis.
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[Exposure to nanoparticle-rich diesel exhaust may cause liver damage].
Nihon Eiseigaku Zasshi
PUBLISHED: 10-15-2011
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Diesel exhaust (DE) is one of the air pollutants in the world, and exposure to DE is an environmental health concern. Most studies amongst the limited number of studies on hepatotoxicity have focused on genotoxicity or mutagenicity. However, DE exposure may cause liver damage because one prospective study suggests that DE exposure is associated with increased mortality due to arteriosclerosis and cirrhosis of the liver. Peroxisome proliferator-activated receptor (PPAR) ? plays a role in the regulation of lipid homeostasis and inflammation and thereby may be involved in the progression of atherosclerosis. We investigated whether nanoparticle-rich diesel exhaust (NR-DE) affects the liver and how PPAR? is involved in the NR-DE induced effects. We report these results briefly in this minireview. Our results suggest NR-DE-induced hepatic inflammation and dyslipidemia. PPAR? may be involved in the development of these disorders.
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[Effects of nanoparticle-rich-diesel exhaust on steroidogenesis in rats and the mechanism underlying such effects].
Nihon Eiseigaku Zasshi
PUBLISHED: 10-15-2011
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Diesel exhaust (DE) is one of the major air pollutants in the world. DE disrupts steroid hormone levels, which may result from the disruption of spermatogenesis. Steroidogenesis occurs not only in the testis but also in the brain. Therefore, we investigated the effects of nanoparticle-rich DE (NR-DE) on steroidogenesis in both the testis and hippocampus. Exposure to NR-DE at concentrations comparable to the environmental standard for particulate matters 2.5 (PM(2.5)) in Japan increased plasma testosterone level. This exposure increased the expression levels of genes involved in steroidogenesis in the testis, but not in the hippocampus, suggesting that NR-DE disrupts steroid hormone balance. This finding suggests the need to reconsider the environmental limit of PM(2.5) in Japan.
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Differences in metabolite burden of di(2-ethylhexyl)phthalate in pregnant and postpartum dams and their offspring in relation to drug-metabolizing enzymes in mice.
Arch. Toxicol.
PUBLISHED: 09-13-2011
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Di(2-ethylhexyl)phthalate (DEHP) induced adverse effects on mice offspring, and the metabolite mono(2-ethylhexyl)phthalate (MEHP) may be essential to determine the toxicity. In this experiment, we measured liver MEHP levels and the factors determining the metabolism, two enzyme activities [lipase and uridine 5-diphosphate-glucuronosyltransferase (UGT)] or expression of cytochrome P450 4A14 (CYP4A14) in dams (on gestational day 18 and postnatal day 2) and their offspring. MEHP concentrations in the liver from pregnant dams were 1.5 times higher than those of postpartum dams at exposure to 0.05% DEHP. Accordingly, MEHP concentrations were 1.7 times higher in fetuses than in pups at the dose. Interestingly, lipase activity was 1.8-fold higher in pregnant dams than postpartum ones, but no such difference was noted in the activity between fetuses and pups. UGT activity was also 1.5-fold higher in pregnant dams than postpartum ones, whereas the activity in the fetuses was 1/2 that of pups. No difference was noted in CYP4A14 levels between pregnant and postpartum mice, whereas the levels in the fetuses were <1/10 those of pups. DEHP exposure did not influence lipase activity, whereas it slightly enhanced UGT activity and exclusively increased CYP4A14 levels in pregnant and/or postpartum dams. Taken together, the higher MEHP levels in pregnant dams than postpartum ones may be primarily due to higher lipase activities in pregnant dams, which may closely reflect those in fetuses and pups.
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Sirtuin 1 retards hyperphosphatemia-induced calcification of vascular smooth muscle cells.
Arterioscler. Thromb. Vasc. Biol.
PUBLISHED: 06-30-2011
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Arterial calcification is associated with cardiovascular disease as a complication of advanced atherosclerosis. Aged vascular cells manifest some morphological features of a senescent phenotype. Recent studies have demonstrated that mammalian sirtuin 1 (SIRT1), a histone deacetylase, is an exciting target for cardiovascular disease management. Here, we investigated the role of SIRT1 in a calcification model of vascular smooth muscle cells (SMCs).
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Amino acid substitution at position 95 in rabies virus matrix protein affects viral pathogenicity.
J. Vet. Med. Sci.
PUBLISHED: 06-07-2011
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We previously reported that rabies virus strain CE(NiM), but not the parental Ni-CE strain, killed mice after intracerebral inoculation. CE(NiM) and Ni-CE are genetically identical except for two amino acids at positions 29 and 95 in the M protein. In this study, to identify which residue determines the pathogenicity, we examined pathogenicities of two Ni-CE mutants, CE(NiM29) and CE(NiM95), which were established by replacement of an amino acid residue at position 29 or 95 in the Ni-CE M protein with the corresponding residue of CE(NiM), respectively. We found that CE(NiM95), but not CE(NiM29), killed mice, indicating that the amino acid at position 95 in the M protein is the pathogenic determinant.
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New analytical method for sensitive quantification of urinary 3-methyl-4-nitrophenol to assess fenitrothion exposure in general population and occupational sprayers.
Toxicol. Lett.
PUBLISHED: 04-12-2011
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The measurement of blood cholinesterase (ChE) activities is adopted worldwide for biological monitoring of exposure to organophosphorus insecticides (OPs). Recent development of analytical chemistry has made sensitive quantification possible of non-specific OP metabolites, dialkylphosphates, in urine as a biomarker of low-level OP exposure. In this study, we established a method for quantification of urinary 3-methyl-4-nitrophenol (MNP), a specific metabolite of fenitrothion (FNT), and a parathion metabolite p-nitrophenol (PNP), using gas chromatography-mass spectrometry. The limits of detection of MNP and PNP were 0.3 and 0.5?g/L, respectively. The method enabled the quantification of both free and conjugated metabolites. This method was actually applied to monitor human urine in summer and winter in FNT sprayers (N=29 and 9, respectively) and control workers (N=17 and 29, respectively). Geometric mean total MNP concentrations (?g/gcreatinine) in the FNT sprayers (28.8 in summer and 8.6 in winter) were significantly higher than those of the controls (3.1 in summer and 2.3 in winter) in both seasons. Among the sprayers, total MNP concentrations in summer were significantly higher than in winter. In contrast, no significant difference in total PNP concentrations was observed between FNT sprayers (geometric mean 3.4 in summer and 3.0 in winter) and controls (3.6 in summer and 2.1 in winter). No seasonal difference was observed in each group. In conclusion, the present new method is sensitive enough for biological monitoring of FNT and parathion metabolites even in a non-spraying population.
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Membrane-associated activation of cholesterol ?-glucosyltransferase, an enzyme responsible for biosynthesis of cholesteryl-?-D-glucopyranoside in Helicobacter pylori critical for its survival.
J. Histochem. Cytochem.
PUBLISHED: 04-09-2011
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Helicobacter pylori (H. pylori) is the causative pathogen underlying gastric diseases such as chronic gastritis and gastric cancer. Previously, the authors revealed that ?1,4-linked N-acetylglucosamine-capped O-glycan (?GlcNAc) found in gland mucin suppresses H. pylori growth and motility by inhibiting catalytic activity of cholesterol ?-glucosyltransferase (CHL?GcT), the enzyme responsible for biosynthesis of the major cell wall component cholesteryl-?-D-glucopyranoside (CGL). Here, the authors developed a polyclonal antibody specific for CHL?GcT and then undertook quantitative ultrastructural analysis of the enzymes localization in H. pylori. They show that 66.3% of CHL?GcT is detected in the cytoplasm beneath the H. pylori inner membrane, whereas 24.7% is present on the inner membrane. In addition, 2.6%, 5.0%, and 1.4% of the protein were detected in the periplasm, on the outer membrane, and outside microbes, respectively. By using an in vitro CHL?GcT assay with fractionated H. pylori proteins, which were used as an enzyme source for CHL?GcT, the authors demonstrated that the membrane fraction formed CGL, whereas other fractions did not. These data combined together indicate that CHL?GcT is originally synthesized in the cytoplasm of H. pylori as an inactive form and then activated when it is associated with the cell membrane. This article contains online supplemental material at http://www.jhc.org. Please visit this article online to view these materials.
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Ammonium perfluorooctanoate may cause testosterone reduction by adversely affecting testis in relation to PPAR?.
Toxicol. Lett.
PUBLISHED: 04-02-2011
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Perfluorooctanoate, a peroxisome proliferator-activated receptor alpha (PPAR?) agonist, has the potential to lower testosterone levels as a result of testicular toxicity. To elucidate the mechanism and impact of PPAR? on this reproductive toxicity, ammonium perfluorooctanoate (APFO) at doses of 0, 1.0 (low) mg/kg/day, or 5.0 (high) mg/kg/day was orally given daily to 129/sv wild-type (mPPAR?), Ppar?-null and PPAR?-humanized (hPPAR?) mice for 6 weeks. Both low- and high-dose APFO significantly reduced plasma testosterone concentrations in mPPAR? and hPPAR? mice, respectively. These decreases may, in part, be associated with decreased expression of mitochondrial cytochrome P450 side-chain cleavage enzyme, steroidogenic acute regulatory protein or peripheral benzodiazepine receptor as well as microsomal cytochrome P450(17?) involved in the steroidogenesis. Additionally, both doses increased abnormalities in sperm morphology and vacuolated cells in the seminiferous tubules of both mouse lines. In contrast, APFO caused only a marginal effect either on the testosterone synthesis system or sperm and testis morphology in Ppar?-null mice. These results suggest that APFO may disrupt testosterone biosynthesis by lowering the delivery of cholesterol into the mitochondria and decreasing the conversion of cholesterol to pregnenolone and androstandione in the testis of mPPAR? and hPPAR? mice, which may, in part, be related to APFO-induced mitochondrial damage.
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From tetragermacyclobutene to tetragermacyclobutadiene dianion to tetragermacyclobutadiene transition metal complexes.
J. Am. Chem. Soc.
PUBLISHED: 03-11-2011
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The reaction of 3,4-dichlorotetragermetene derivative 2 with Na(2)[Fe(CO)(4)] in THF produced a (tetragermacyclobutadiene)tricarbonyliron complex, [{?(4)-((t)Bu(2)MeSi)(4)Ge(4)}]Fe(CO)(3)4, which has a slightly folded Ge(4) ring perhaptocoordinated to the Fe center. Structural and spectral characteristics of 4 show a remarkable ?-donating ability of the tetragermacyclobutadiene ligand toward the transition metal, surpassing that of tetrasilacyclobutadiene and cyclobutadiene ligands. Reduction of 2 with KC(8) resulted in exclusive formation of the dipotassium salt of the tetragermacyclobutadiene dianion derivative 3(2-)·[K(+)(thf)(2)](2), representing a rare example of a 6?-electron compound that, on the basis of its structural and magnetic properties, was recognized as a nonaromatic species. Reaction of 3(2-)·[K(+)(thf)(2)](2) with CpCoI(2)(PPh(3)) produced a (cyclopentadienyl)(tetragermacyclobutadiene)cobalt complex, [{?(4)-((t)Bu(2)MeSi)(4)Ge(4)}]CoCp 7, as the first example of a sandwich compound featuring an all-germanium-containing cyclic polyene ligand.
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Lactacystin, a proteasome inhibitor, enhances BMP-induced osteoblastic differentiation by increasing active Smads.
Biochem. Biophys. Res. Commun.
PUBLISHED: 02-18-2011
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Proteasome inhibitors enhance bone formation and osteoblastic differentiation in vivo and in vitro. In the present study, we examined whether the molecular mechanisms of lactacystin, one of many proteasome inhibitors, stimulated the osteoblastic differentiation of C2C12 cells that is induced by bone morphogenetic proteins (BMPs). Pretreatment with lactacystin enhanced the alkaline phosphatase (ALP) activity induced by BMP2, BMP4 or BMP7, but lactacystin did not induce ALP in the absence of BMPs. In addition, lactacystin-stimulated BMP2 induced mRNA expression of ALP, type I collagen, osteonectin, osteocalcin, Id1, Osterix, and Runx2. Lactacystin maintained BMP2-induced phosphorylation of Smad1/5/8 and increased the length of time that these Smads were bound to target DNA. Moreover, lactacystin prevented BMP receptor-induced Smad degradation. This enhancement of BMP2-induced ALP activity and Smad phosphorylation by lactacystin was also observed in primary osteoblasts. These findings suggest that pretreatment with lactacystin accelerates BMP-induced osteoblastic differentiation by increasing the levels of phosphorylated Smads, which are maintained because BMP receptor-induced degradation is inhibited. We propose that optimized stimulation by proteasome inhibitors in a clinical setting may facilitate autogenous or BMP-induced bone formation in areas of defective bone.
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Modulation of ammonium perfluorooctanoate-induced hepatic damage by genetically different PPAR? in mice.
Arch. Toxicol.
PUBLISHED: 02-17-2011
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Perfluorooctanoic acid is a ligand for peroxisome proliferator-activated receptor (PPAR?). Ammonium perfluorooctanoate (APFO) at 0.1 and 0.3 mg/kg doses activated mouse PPAR?, but not human PPAR?. This study aimed to clarify whether milligram-order APFO can activate human PPAR?, and the receptor is involved in APFO-induced chronic hepatic damage. Male Sv/129 wild-type (mPPAR?), Ppar?-null, and humanized PPAR? (hPPAR?) mice (8 weeks old) were divided into three groups. The first was treated with water and the other two with 1.0 and 5.0 mg/kg APFO for 6 weeks, orally, respectively. Both doses activated mouse and human PPAR? to a similar or lower degree in the latter. APFO dose dependently increased hepatic triglyceride levels in Ppar?-null and hPPAR? mice, but conversely decreased those in mPPAR? ones. APFO-induced hepatic damage differed markedly among the three genotyped groups: single-cell necrosis was observed in all genotyped mice; inflammatory cells and macrovesicular steatosis only in Ppar?-null mice; and microvesicular steatosis and hydropic degenerations in hPPAR? and Ppar?-null mice. The molecular mechanism underlying these differences may be attributable to those of gene expressions involved in lipid homeostasis (PPAR?, ?- and ?-oxidation enzymes, and diacylglycerol acyltransferases) and uncoupling protein 2. Thus, milligram-order APFO activated both mouse and human PPAR? in a different manner, which may reflect histopathologically different types of hepatic damage.
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A blue digermene (t-Bu2MeSi)2Ge=Ge(SiMet-Bu2)2.
Chem. Commun. (Camb.)
PUBLISHED: 02-01-2011
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A novel tetrakis(trialkylsilyl)digermene, featuring rather unusual structural and chemical properties, was synthesized by a straightforward synthetic protocol: reduction of the corresponding dichlorobis(trialkylsilyl)germane precursor with potassium graphite.
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Hepatic peroxisome proliferator-activated receptor ? may have an important role in the toxic effects of di(2-ethylhexyl)phthalate on offspring of mice.
Toxicology
PUBLISHED: 01-08-2011
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Maternal exposure to di(2-ethylhexyl)phthalate (DEHP) is associated with adverse effects on offspring, and the metabolites are agonists of peroxisome proliferator-activated receptor (PPAR) ?, which exhibits species differences in expression and function. This study aimed to clarify the mechanism of DEHP-induced adverse effects on offspring in relation to maternal mouse and human PPAR?. Male and female Sv/129 wild-type (mPPAR?), Ppar?-null and humanized PPAR? (hPPAR?) mice were treated with diets containing 0%, 0.01%, 0.05% (medium) or 0.1% (high) DEHP. After 4 weeks, males and females were mated. Dams were killed on gestational day 18 and postnatal day (PND) 2. High-dose DEHP decreased the number of total and live fetuses, and increased resorptions in mPPAR? mice. In hPPAR? mice, resorptions were increased above the medium dose, and the number of births was decreased at the high dose. The number of live pups on PND2 was decreased over the medium dose in mPPAR? and at the high dose in hPPAR? mice. No such findings were observed in Ppar?-null mice. High-dose DEHP decreased plasma triglyceride in pregnant mPPAR? mice, but not in Ppar?-null and hPPAR? ones. Above the medium dose in mPPAR? mice significantly reduced hepatic microsomal triglyceride transfer protein (MTP) expression. Medium- and/or high-dose DEHP increased the levels of maternal PPAR? target genes in mPPAR? and hPPAR? mice. Taken together, PPAR? expression is required for the toxicity of DEHP in fetuses and pups and altered plasma triglyceride levels, through regulation of MTP may be important in mPPAR? mice and not in hPPAR? mice.
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Novel anti-carbohydrate antibodies reveal the cooperative function of sulfated N- and O-glycans in lymphocyte homing.
J. Biol. Chem.
PUBLISHED: 10-07-2010
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Cell surface glycans play pivotal roles in immune cell trafficking and immunity. Here we present an efficient method for generating anti-carbohydrate monoclonal antibodies (mAbs) using gene-targeted mice and describe critical glycans in lymphocyte homing. We immunized sulfotransferase GlcNAc6ST-1 and GlcNAc6ST-2 doubly deficient mice with sulfotransferase-overexpressing Chinese hamster ovary cells and generated two mAbs, termed S1 and S2. Both S1 and S2 bound high endothelial venules (HEVs) in the lymphoid organs of humans and wild-type mice, but not in those of doubly deficient mice. Glycan array analysis indicated that both S1 and S2 specifically bound 6-sulfo sialyl Lewis X and its defucosylated structure. Interestingly, S2 inhibited lymphocyte homing to peripheral lymph nodes by 95%, whereas S1 inhibited it by only 25%. S2 also significantly inhibited contact hypersensitivity responses and L-selectin-dependent leukocyte adhesion to HEVs. Immunohistochemical and Western blot analyses indicated that S1 preferentially bound sulfated O-glycans, whereas S2 bound both sulfated N- and O-glycans in HEVs. Furthermore, S2 strongly inhibited the N-glycan-dependent residual lymphocyte homing in mutant mice lacking sulfated O-glycans, indicating the importance of both sulfated N- and O-glycans in lymphocyte homing. Thus, the two mAbs generated by a novel method revealed the cooperative function of sulfated N- and O-glycans in lymphocyte homing and immune surveillance.
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Functional contributions of N- and O-glycans to L-selectin ligands in murine and human lymphoid organs.
Am. J. Pathol.
PUBLISHED: 09-08-2010
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L-selectin initiates lymphocyte interactions with high endothelial venules (HEVs) of lymphoid organs through binding to ligands with specific glycosylation modifications. 6-Sulfo sLe(x), a sulfated carbohydrate determinant for L-selectin, is carried on core 2 and extended core 1 O-glycans of HEV-expressed glycoproteins. The MECA-79 monoclonal antibody recognizes sulfated extended core 1 O-glycans and partially blocks lymphocyte-HEV interactions in lymphoid organs. Recent evidence has identified the contribution of 6-sulfo sLe(x) carried on N-glycans to lymphocyte homing in mice. Here, we characterize CL40, a novel IgG monoclonal antibody. CL40 equaled or surpassed MECA-79 as a histochemical staining reagent for HEVs and HEV-like vessels in mouse and human. Using synthetic carbohydrates, we found that CL40 bound to 6-sulfo sLe(x) structures, on both core 2 and extended core 1 structures, with an absolute dependency on 6-O-sulfation. Using transfected CHO cells and gene-targeted mice, we observed that CL40 bound its epitope on both N-glycans and O-glycans. Consistent with its broader glycan-binding, CL40 was superior to MECA-79 in blocking lymphocyte-HEV interactions in both wild-type mice and mice deficient in forming O-glycans. This superiority was more marked in human, as CL40 completely blocked lymphocyte binding to tonsillar HEVs, whereas MECA-79 inhibited only 60%. These findings extend the evidence for the importance of N-glycans in lymphocyte homing in mouse and indicate that this dependency also applies to human lymphoid organs.
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FGF receptor gene expression and its regulation by FGF signaling during early zebrafish development.
Genesis
PUBLISHED: 07-30-2010
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The expression of all four fgfr genes was extensively examined throughout early embryogenesis of the zebrafish (Danio rerio). fgfr1 alone was expressed maternally throughout the blastoderm, and then zygotically in the anterior neural plate and presomitic mesoderm. fgfr4 expression was first detected in late blastulae and was gradually restricted to the brain. fgfr2 and fgfr3 expression were initiated in early and late gastrulae, respectively; fgfr2 was expressed in the anterior neural plate and somitic mesoderm, whereas fgfr3 was activated in the axial mesoderm and then in the midbrain and somitic mesoderm. During somitogenesis, each of these fgfr genes was expressed in a characteristic manner in the brain. Using an FGF signal inhibitor, dominant-negative FGF receptors and fgf8.1/fgf8a mutants, we found that fgfr expression is directly or indirectly regulated by FGF signaling during epiboly and at the end of somitogenesis, revealing the presence of an autoregulatory mechanism.
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Amino acids at positions 273 and 394 in rabies virus nucleoprotein are important for both evasion of host RIG-I-mediated antiviral response and pathogenicity.
Virus Res.
PUBLISHED: 07-05-2010
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We previously reported that nucleoprotein (N) is related to the different pathogenicities of the virulent rabies virus strain Nishigahara (Ni) and avirulent strain Ni-CE and also that Ni N, but not Ni-CE N, functions to evade retinoic acid-inducible gene I (RIG-I)-mediated innate immunity. There are three amino acid differences between Ni and Ni-CE N (at positions 273, 394 and 395), indicating that one of these mutations or a combination of mutations is important for the pathogenicity and evasion of innate immunity. We generated Ni-CE mutants in which the amino acids in Ni-CE N were replaced with those of Ni in all combinations. Among the mutants, CE(NiN273/394) with mutations at positions 273 and 394 evaded activation of RIG-I-mediated signaling most efficiently and also showed the highest pathogenicity. This correlation reinforces the relation between evasion of host RIG-I-mediated innate immunity and pathogenicity of rabies virus.
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Endothelial heparan sulfate controls chemokine presentation in recruitment of lymphocytes and dendritic cells to lymph nodes.
Immunity
PUBLISHED: 06-24-2010
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Heparan sulfate can bind several adhesion molecules involved in lymphocyte trafficking. However, the in vivo function of endothelial heparan sulfate in lymphocyte homing and stimulation of the immune response has not been elucidated. Here, we generated mutant mice deficient in the enzyme Ext1, which is required for heparan sulfate synthesis, in a Tek-dependent and inducible manner. Chemokine presentation was diminished in the mutant mice, causing the lack of appropriate integrin-mediated adhesion, and resulted in a marked decrease in lymphocyte sticking to high endothelial venules and in recruitment of resident dendritic cells through lymphatic vessels to the lymph nodes. As a consequence, mutant mice displayed a severe impairment in lymphocyte homing and a compromised contact hypersensitivity response. By contrast, lymphocyte rolling was increased because of loss of electrostatic repulsion by heparan sulfate. These results demonstrate critical roles of endothelial heparan sulfate in immune surveillance and immune response generation.
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Role of interferon antagonist activity of rabies virus phosphoprotein in viral pathogenicity.
J. Virol.
PUBLISHED: 04-28-2010
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The fixed rabies virus (RV) strain Nishigahara kills adult mice after intracerebral inoculation, whereas the chicken embryo fibroblast cell-adapted strain Ni-CE causes nonlethal infection in adult mice. We previously reported that the chimeric CE(NiP) strain, which has the phosphoprotein (P protein) gene from the Nishigahara strain in the genetic background of the Ni-CE strain, causes lethal infection in adult mice, indicating that the P gene is responsible for the different pathogenicities of the Nishigahara and Ni-CE strains. Previous studies demonstrated that RV P protein binds to the interferon (IFN)-activated transcription factor STAT1 and blocks IFN signaling by preventing its translocation to the nucleus. In this study, we examine the molecular mechanism by which RV P protein determines viral pathogenicity by comparing the IFN antagonist activities of the Nishigahara and Ni-CE P proteins. The results, obtained from both RV-infected cells and cells transfected to express P protein only, show that Ni-CE P protein is significantly impaired for its capacity to block IFN-activated STAT1 nuclear translocation and, consequently, inhibits IFN signaling less efficiently than Nishigahara P protein. Further, it was demonstrated that a defect in the nuclear export of Ni-CE P protein correlates with a defect in its ability to cause the mislocalization of STAT1. These data provide the first evidence that the capacity of the RV P protein to inhibit STAT1 nuclear translocation and IFN signaling correlates with the viral pathogenicity.
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Amino acid substitutions at positions 242, 255 and 268 in rabies virus glycoprotein affect spread of viral infection.
Microbiol. Immunol.
PUBLISHED: 04-10-2010
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Rabies virus Nishigahara strain kills adult mice after intracerebral inoculation, whereas the derivative RC-HL strain does not. It has previously been reported by us that the R(G 242/255/268) strain, in which amino acids at positions 242, 255 and 268 on the G protein have been replaced by those from the Nishigahara strain in the genetic background of the RC-HL strain, kills adult mice. This indicates that these three amino acids of G protein are important for pathogenicity of the Nishigahara strain. In order to obtain insights into the mechanism by which these amino acids affect pathogenicity, in this study spread of viral infection and apoptosis-inducing ability of the attenuated RC-HL strain and the virulent R(G 242/255/268) strain were compared. RC-HL infection spread less efficiently in the mouse brain than did R(G 242/255/268) infection. However, the apoptosis-inducing abilities of both viruses were almost identical, as shown by both in vitro and in vivo experiments. It was demonstrated that cell-to-cell spread of RC-HL strain was less efficient than that of R(G 242/255/268) strain in mouse neuroblastoma cells. These results indicate that the three amino acid substitutions affect efficiency of cell-to-cell spread but not apoptosis-inducing ability, probably resulting in the distinct distributions of RC-HL and R(G 242/255/268) strain-infected cells in the mouse brain and, consequently, the different pathogenicities of these strains.
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Role of N-end rule ubiquitin ligases UBR1 and UBR2 in regulating the leucine-mTOR signaling pathway.
Genes Cells
PUBLISHED: 03-16-2010
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Of 20 natural amino acids, leucine is particularly important for promoting cellular protein synthesis. The effect of leucine involves mammalian target of rapamycin (mTOR), a key protein kinase controlling cell growth. Leucine enhances mTOR-mediated phosphorylation of S6K1 and 4E-BP, thereby promoting protein synthesis. However, how the presence of leucine is sensed and transmitted to mTOR is poorly understood. Here, we show evidence that UBR1 and UBR2 might be cellular targets of leucine. UBR1 and UBR2 are E3 ubiquitin ligases that recognize the identity of N-terminal residues and contribute to selective destabilization of target proteins according to the N-end rule. Using leucine-immobilized affinity beads, we identified UBR1 and UBR2 as leucine-binding proteins from leucine-responsive rat hepatoma H4IIE cells. Over-expression of UBR1 or UBR2 resulted in a reduction in mTOR-dependent S6K1 phosphorylation, whereas knockdown of UBR1 or UBR2 increased S6K1 phosphorylation in amino acid-starved human 293T cells. We also found that leucine binds to the substrate-recognition domain of UBR2 and inhibits degradation of N-end rule substrates in vitro. These findings suggest that UBR1 and UBR2 are negative regulators of the leucine-mTOR signaling pathway. Leucine might activate this pathway in part through inhibition of their ubiquitin ligase activity.
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Biogeography and biodiversity in sulfide structures of active and inactive vents at deep-sea hydrothermal fields of the Southern Mariana Trough.
Appl. Environ. Microbiol.
PUBLISHED: 03-12-2010
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The abundance, diversity, activity, and composition of microbial communities in sulfide structures both of active and inactive vents were investigated by culture-independent methods. These sulfide structures were collected at four hydrothermal fields, both on- and off-axis of the back-arc spreading center of the Southern Mariana Trough. The microbial abundance and activity in the samples were determined by analyzing total organic content, enzymatic activity, and copy number of the 16S rRNA gene. To assess the diversity and composition of the microbial communities, 16S rRNA gene clone libraries including bacterial and archaeal phylotypes were constructed from the sulfide structures. Despite the differences in the geological settings among the sampling points, phylotypes related to the Epsilonproteobacteria and cultured hyperthermophilic archaea were abundant in the libraries from the samples of active vents. In contrast, the relative abundance of these phylotypes was extremely low in the libraries from the samples of inactive vents. These results suggest that the composition of microbial communities within sulfide structures dramatically changes depending on the degree of hydrothermal activity, which was supported by statistical analyses. Comparative analyses suggest that the abundance, activity and diversity of microbial communities within sulfide structures of inactive vents are likely to be comparable to or higher than those in active vent structures, even though the microbial community composition is different between these two types of vents. The microbial community compositions in the sulfide structures of inactive vents were similar to those in seafloor basaltic rocks rather than those in marine sediments or the sulfide structures of active vents, suggesting that the microbial community compositions on the seafloor may be constrained by the available energy sources. Our findings provide helpful information for understanding the biogeography, biodiversity and microbial ecosystems in marine environments.
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Photoluminescent properties of chalcobromide-capped octahedral hexarhenium(III) complexes [{Re(6)Q(8-n)Br(n)}Br(6)](n-4) (Q = Se, n = 1-3; Q = S, n = 1, 2).
Inorg Chem
PUBLISHED: 03-06-2010
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Photoluminescent properties of chalcobromide-capped octahedral hexarhenium(III) complexes with terminal bromide ligands [{Re(6)Q(8-n)Br(n)}Br(6)](n-4) (Q = Se, n = 1 ([1-Se](3-)), n = 2 ([2a-Se](2-) and [2b-Se](2-)), and n = 3 ([3-Se](-)); Q = S, n = 1 ([1-S](3-)), n = 2 ([2a-S](2-), [2b-S](2-), and [2c-S](2-)) were studied. The Q(7)Br capped complex [{Re(6)Q(7)Br}Br(6)](3-) and Q(6)Br(2) [{Re(6)Q(6)Br(2)}Br(6)](2-) (both D(3d) and C(2v) symmetric geometrical isomers) were successfully separated by column chromatography. All of the chalcobromide-capped complexes studied showed photoluminescence in both crystalline and solution phases. The emission maximum wavelength of the complexes at 296 K spans 853-915 or 868-968 nm in the crystalline phase or in acetonitrile, respectively. The selenobromide-capped complexes showed more intense emission as compared with the thiobromide analogues. The emission quantum yield (Phi(em)) and emission lifetime (tau(em)) became smaller and shorter, respectively, with an increase in the number of a capping bromide ligand in [{Re(6)Q(8-n)Br(n)}Br(6)](n-4). In the crystalline phase at 80 K, the emission maximum of the chalcobromide-capped complex shifted to the longer wavelength relative to that at 296 K. The emissive excited-state of the chalcobromide-capped hexarhenium(III) complexes was concluded to originate from the {Re(6)Q(8-n)Br(n)}(n+2) core with a spin-triplet type. The Phi(em) and tau(em) values of the {Re(6)Q(8-n)Br(n)}(n+2) complex were dependent significantly on the symmetry of the hexarhenium core, showing more intense emission for the complex with the higher symmetric core. A linear correlation between natural logarithm of the nonradiative decay rate constant and the emission maximum energy was observed for [{Re(6)Q(6)Br(2)}Br(6)](2-).
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Structure-activity relationship of indolicidin, a Trp-rich antibacterial peptide.
J. Pept. Sci.
PUBLISHED: 03-03-2010
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A series of Trp and Arg analogs of antibacterial indolicidin (Ind) was synthesized and the antimicrobial and hemolytic activities were investigated. [L(9)]Ind, [L(11)]Ind, [K(8),L(9)]Ind and [K(6, 8),L(9)]Ind showed desirable characteristics, exhibiting negligible hemolytic activity while keeping strong antibacterial activity. The results indicated that the Trp residue at position 11 essentially contributes to both activities and one can not be exchanged for the other, whereas the Trp residues at positions 4 and 9 play important roles in antimicrobial and hemolytic activities, respectively. The Trp residues at positions 6 and 8 play no important roles in biological activities. We then found that the retro analog of Ind showed higher antibacterial activity than Ind against both Gram-positive and Gram-negative bacteria but remarkably lower hemolytic activity than that of Ind.
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Rabies virus nucleoprotein functions to evade activation of the RIG-I-mediated antiviral response.
J. Virol.
PUBLISHED: 02-03-2010
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The rabies virus Ni-CE strain causes nonlethal infection in adult mice after intracerebral inoculation, whereas the parental Nishigahara (Ni) strain kills mice. We previously reported that the chimeric CE(NiN) strain with the N gene from the Ni strain in the genetic background of the Ni-CE strain kills adult mice, indicating that the N gene is related to the different pathogenicities of Ni and Ni-CE strains. In the present study, to obtain an insight into the mechanism by which the N gene determines viral pathogenicity, we compared the effects of Ni, Ni-CE, and CE(NiN) infections on host gene expressions using a human neuroblastoma cell line. Microarray analysis of these infected cells revealed that the expression levels of particular genes in Ni- and CE(NiN)-infected cells, including beta interferon (IFN-beta) and chemokine genes (i.e., CXCL10 and CCL5) were lower than those in Ni-CE-infected cells. We also demonstrated that Ni-CE infection activated the interferon regulatory factor 3 (IRF-3)-dependent IFN-beta promoter and induced IRF-3 nuclear translocation more efficiently than did Ni or CE(NiN) infection. Furthermore, we showed that Ni-CE infection, but not Ni or CE(NiN) infection, strongly activates the IRF-3 pathway through activation of RIG-I, which is known as a cellular sensor of virus infection. These findings indicate that the N protein of rabies virus (Ni strain) has a function to evade the activation of RIG-I. To our knowledge, this is the first report that the Mononegavirales N protein functions to evade induction of host IFN and chemokines.
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Bisphenol A may cause testosterone reduction by adversely affecting both testis and pituitary systems similar to estradiol.
Toxicol. Lett.
PUBLISHED: 02-01-2010
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Bisphenol A (BPA) causes reproductive toxicities, but the mechanisms are still unclear. In the present study, we sought to clarify these mechanisms in comparison with those of 17beta-estradiol (E2). Prepubertal Wistar/ST male rats (4 weeks old) were subcutaneously administered BPA (0, 20, 100 and 200 mg/kg/day) or E2 (10 and 100 microg/kg/day) for 6 weeks. Both BPA and E2 treatments decreased plasma and testicular testosterone levels, and plasma luteinizing hormone (LH), but not E2 and follicle-stimulating hormone levels, though E2 treatment increased its plasma level. In relation to the decreased testosterone levels, BPA and E2 decreased expressions of steroidogenic enzymes and cholesterol carrier protein in Leydig cells. Thus, decreased testosterone levels in plasma might have resulted from decreased expressions of these enzymes and protein as well as from decreased plasma LH levels. Interestingly, the changes in steroidogenic enzymes and carrier protein were observed at lower levels of exposure to BPA or E2 than those inhibiting plasma LH levels. Microscopically, 200 mg/kg BPA and 100 microg/kg E2 significantly decreased Leydig cell numbers in the testis. In addition, BPA and E2 also decreased expression of estrogen receptor alpha-mRNA, which might be related to the decreased numbers of Leydig cells. Thus, BPA directly affects not only the Leydig cells but also the pituitary gland, but the former may be impaired at lower exposure concentrations than the latter.
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Differential response to trichloroethylene-induced hepatosteatosis in wild-type and PPARalpha-humanized mice.
Environ. Health Perspect.
PUBLISHED: 01-12-2010
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Trichloroacetic acid, an oxidative metabolite of trichloroethylene (TRI), is a ligand of the peroxisome proliferator-activated receptor alpha (PPAR) alpha, which is involved in lipid homeostasis and anti-inflammation.
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Magnified endoscopic observation using narrow-band imaging of periampullary adenoma in a patient with familial adenomatous polyposis.
Med. Sci. Monit.
PUBLISHED: 12-01-2009
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Adenoma of the major papilla carries a relatively high risk of malignant transformation to carcinoma, the leading cause of death in patients with familiar adenomatous polyposis (FAP) after colectomy.
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Indole derivatives from a marine sponge-derived yeast as DPPH radical scavengers.
J. Nat. Prod.
PUBLISHED: 12-01-2009
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Two new indole derivatives (3, 4) and three known compounds (1, 2, 5) were isolated as radical scavengers from the culture filtrate of a marine sponge-derived yeast. Their structures were determined to be tyrosol (1), tryptophol (2), 2-(1H-indol-3-yl)ethyl 2-hydroxypropanoate (3), 2-(1H-indol-3-yl)ethyl 5-hydroxypentanoate (4), and cyclo(L-Pro-L-Tyr) (5) on the basis of their spectroscopic data. The absolute configurations of compounds 3 and 5 were determined by chiral HPLC analysis combined with synthesis and Marfeys method, respectively. Each obtained compound was evaluated for DPPH (2,2-diphenyl-1-picrylhydrazyl) radical scavenging activity, and all compounds exhibited weak activities.
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Broken sperm, cytoplasmic droplets and reduced sperm motility are principal markers of decreased sperm quality due to organophosphorus pesticides in rats.
J Occup Health
PUBLISHED: 09-25-2009
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Recent reports have shown significant associations between organopshophorus pesticide (OP) exposure and decreased sperm motility in workers and laboratory animals. However, the notion that OPs possess spermatotoxicity has yet to be established. The aim of this study was to clarify the effects of OP exposure on detailed sperm toxicity markers, i.e., motility, morphology and sperm adenine nucleotide contents, and the histopathology of the testis and epididymis.
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"Hypothesis of seven balances": molecular mechanisms behind alcoholic liver diseases and association with PPARalpha.
J Occup Health
PUBLISHED: 08-26-2009
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The purpose of this review to collate current leading scientific advances of molecular mechanisms in alcoholic liver diseases and to propose a working "hypothesis of seven balances" in relation to peroxisome proliferator activated receptor alpha (PPARalpha), which has important roles in fatty acid oxidation, oxidative stress, inflammatory responses, and possibly liver fibrosis.
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Effects of inhaled nanoparticle-rich diesel exhaust on regulation of testicular function in adult male rats.
Inhal Toxicol
PUBLISHED: 08-06-2009
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We investigated the effects of nanoparticle-rich diesel exhaust (NR-DE) on reproductive function. Eight-week-old male F344 rats were divided into 12 experimental groups and exposed to either whole NR-DE at low (15.37 microg/m(3), 2.27 x 10(5) particles/cm(3)), middle (36.35 microg/m(3), 5.11 x 10(5) particles/cm(3)), or high (168.84 microg/m(3), 1.36 x 10(6) particles/cm(3)) concentrations or clean air for 4, 8, or 12 weeks (5 hours/day, 5 days/week). NR-DE exposure for 4 or 8 weeks did not affect body weight; however, body weight was significantly decreased in rats exposed to low- or high- concentration NR-DE for 12 weeks compared to the control group. Relative weights of testes, epididymides, seminal vesicles, and prostate had increased non-significantly in all NR-DE-exposed rats at 4, 8, and 12 weeks. Adrenal gland relative weights were significantly increased at 4 weeks in rats exposed to low-concentration NR-DE. Plasma luteinizing hormone and follicle stimulating hormone concentrations did not change significantly. Plasma testosterone concentrations were significantly increased after exposure to low- or middle-concentration NR-DE for 4 or 8 weeks compared to controls. Plasma immunoreactive (ir-) inhibin concentrations were significantly increased after exposure to high-concentration NR-DE for 4 weeks or middle- or high-concentration NR-DE for 12 weeks compared to controls. Testicular testosterone concentrations were significantly increased at 4, 8, and 12 weeks after exposure to low-concentration NR-DE compared to controls. In contrast, with exposure to low- or high-concentration NR-DE, testicular ir-inhibin concentrations were significantly greater than in controls, but only at 4 weeks. These results suggest that NR-DE inhalation disrupts the endocrine activity of the male reproductive system.
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Octachlorostyrene induces cytochrome P450, UDP-glucuronosyltransferase, and sulfotransferase via the aryl hydrocarbon receptor and constitutive androstane receptor.
Toxicol. Sci.
PUBLISHED: 06-12-2009
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Octachlorostyrene (OCS) is a byproduct produced in the process of synthesis of chlorinated compounds. There are some reports concerning environmental contamination by OCS, but few on the toxicological effects on human. Drug-metabolizing enzymes may play an important role in toxicity through metabolic activation or deactivation of OCS. In this study, we investigated whether OCS influences these enzymes using wild-type and aryl hydrocarbon receptor (Ahr)-null mice; AhR regulates cytochrome P450 (CYP) 1A, UDP-glucuronosyltransferase (UGT), or sulfotransferase (SULT). Both mouse lines were treated with OCS (0, 32, and 64 mumol/kg) for 4 days by gavage. As a reference, the mice were treated with 20 mg/kg 3-methylcholanthrene (3MC) for 4 days. OCS treatment increased the expression of CYP 1A1 and CYP1A2 mRNA and ethoxyresorfin O-deethylase activity only in the wild-type mice, similar to that of the AhR activator 3MC. OCS treatment increased expression of UGT1A6 and SULT 1A1 mRNA and their associated enzyme activities only in Ahr-null mice, whereas 3MC still influenced these enzymes only in wild-type mice. OCS induced constitutive androstane receptor (CAR) only in Ahr-null mice, and the target gene CYP2B10 mRNA was induced more strongly in Ahr-null mice than in wild-type mice. 3MC slightly induced CYP2B10 mRNA only in the wild-type mice. These results suggest that CAR is involved in regulation of the UGT and SULT genes by OCS. Thus, OCS may regulate CYP1A via AhR, whereas it controls UGT1A6 and SULT1A via CAR.
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Microgram-order ammonium perfluorooctanoate may activate mouse peroxisome proliferator-activated receptor alpha, but not human PPARalpha.
Toxicology
PUBLISHED: 06-09-2009
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Perfluorooctanoic acid (PFOA) is a ligand for peroxisome proliferator-activated receptor (PPAR) alpha, which exhibits marked species differences in expression and function, especially between rodents and humans. We investigated the functional difference in PFOA response between mice and humans, using a humanized PPARalpha transgenic mouse line. Three genotyped mice, 129/Sv wild-type (mPPARalpha), Pparalpha-null mice and humanized PPARalpha (hPPARalpha) mice (8-week-old males) were divided into three groups: the first was treated with water daily for 2 weeks by gavage (control group), and the remaining two groups were treated with 0.1 and 0.3mg/kg ammonium perflurooctanate (APFO), respectively, for 2 weeks by gavage. The APFO dosages used did not influence the plasma triglyceride or total cholesterol levels in any mouse line, but the high dose increased both hepatic lipid levels only in mPPARalpha mice. APFO increased mRNA and/or protein levels of PPARalpha target genes cytochrome P450 Cyp4a10, peroxisomal thiolase and bifunctional protein only in the liver of mPPARalpha mice, but not in Pparalpha-null or hPPARalpha mice. This chemical also increased expression of mitochondrial very long chain acyl-CoA dehydrogenase only in the liver of mPPARalpha mice. Taken together, human PPARalpha may be less responsive to PFOA than that of mice when a relatively low dose is applied. This information may be very valuable in considering whether PFOA influences the lipid metabolism in humans.
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Nanoparticle-rich diesel exhaust may disrupt testosterone biosynthesis and metabolism via growth hormone.
Toxicol. Lett.
PUBLISHED: 05-17-2009
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We previously reported that exposure to low (22.5+/-0.2 nm in diameter, 15.4+/-1.0 microg/m(3) in mass weight, 2.27x10(5)/cm(3) in mean number concentration), and medium (26.1+/-0.5 nm, 36.4+/-1.2 microg/m(3), 5.11x10(5)/cm(3)) concentrations of nanoparticle-rich diesel exhaust (NR-DE) for 1 and 2 months (5 h/day, 5 days/week) significantly increased plasma testosterone in male Fischer 344 rats, whereas exposure to a high concentration (27.1+/-0.5 nm, 168.8+/-2.7 microg/m(3), 1.36x10(6)/cm(3)) did not. The present study attempts to clarify the mechanism of this elevation. Low and medium exposures to NR-DE for 1 and 2 months significantly increased steroidogenic acute regulatory protein (StAR)- and cytochrome P450 side-chain cleavage (P450scc)-mRNA and their protein expressions in the testis of rats, in which the elevation pattern was very similar to that of plasma testosterone levels. Interestingly, both exposure levels for 1 month significantly increased growth hormone (GH) receptor expression in the testis, and low exposure also increased testicular insulin-like growth factor I-mRNA levels and hepatic microsomal cytochrome P450 2C11-mRNA and their protein levels in rats. These two factors are thought to be related to growth hormone secretion. Disruption of testosterone biosynthesis by NR-DE exposure may be a mode of action for reproductive toxicity, which may, in part, be regulated by increasing StAR and P450scc expressions via GH signalling.
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Identification of DNA-dependent protein kinase catalytic subunit (DNA-PKcs) as a novel target of bisphenol A.
PLoS ONE
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Bisphenol A (BPA) forms the backbone of plastics and epoxy resins used to produce packaging for various foods and beverages. BPA is also an estrogenic disruptor, interacting with human estrogen receptors (ER) and other related nuclear receptors. Nevertheless, the effects of BPA on human health remain unclear. The present study identified DNA-dependent protein kinase catalytic subunit (DNA-PKcs) as a novel BPA-binding protein. DNA-PKcs, in association with the Ku heterodimer (Ku70/80), is a critical enzyme involved in the repair of DNA double-strand breaks. Low levels of DNA-PK activity are previously reported to be associated with an increased risk of certain types of cancer. Although the Kd for the interaction between BPA and a drug-binding mutant of DNA-PKcs was comparatively low (137 nM), high doses of BPA were required before cellular effects were observed (100-300 ?M). The results of an in vitro kinase assay showed that BPA inhibited DNA-PK kinase activity in a concentration-dependent manner. In M059K cells, BPA inhibited the phosphorylation of DNA-PKcs at Ser2056 and H2AX at Ser139 in response to ionizing radiation (IR)-irradiation. BPA also disrupted DNA-PKcs binding to Ku70/80 and increased the radiosensitivity of M059K cells, but not M059J cells (which are DNA-PKcs-deficient). Taken together, these results provide new evidence of the effects of BPA on DNA repair in mammalian cells, which are mediated via inhibition of DNA-PK activity. This study may warrant the consideration of the possible carcinogenic effects of high doses of BPA, which are mediated through its action on DNA-PK.
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Evidence for diazinon-mediated inhibition of cis-permethrin metabolism and its effects on reproductive toxicity in adult male mice.
Reprod. Toxicol.
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The potential toxicity resulting from combinatorial effects of organophosphorus and pyrethroid insecticides are not completely known. We evaluated male reproductive toxicity in mice co-exposed to diazinon and cis-permethrin. Nine-week-old male Sv/129 mice were exposed to diazinon (10 ?mol/kg/day) or cis-permethrin (90 ?mol/kg/day) alone or in combination (100 ?mol/kg/day), or vehicle (corn oil), for 6 weeks. Diazinon and the diazinon-permethrin mixture inhibited plasma and liver carboxylesterase activities. In the mixture group, urinary excretion of cis-permethrin metabolite 3-phenoxybenzoic acid decreased along with increased plasma and testicular concentrations of cis-permethrin, while excretion of diazinon metabolites, diethylphosphate and diethylthiophosphate, did not change, versus mice exposed to each chemical alone, which suggested that inhibition of carboxylesterase decreased the metabolic capacity to cis-permethrin. Though the co-exposure decreased testosterone biosynthesis, increased degenerate germ cells in seminiferous tubule and sperm morphological abnormalities versus controls more clearly than exposure to cis-permethrin alone, the expected potentiation of toxicity was not evident.
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Plasticizers May Activate Human Hepatic Peroxisome Proliferator-Activated Receptor ? Less Than That of a Mouse but May Activate Constitutive Androstane Receptor in Liver.
PPAR Res
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Dibutylphthalate (DBP), di(2-ethylhexyl)phthalate (DEHP), and di(2-ethylhexyl)adipate (DEHA) are used as plasticizers. Their metabolites activate peroxisome proliferator-activated receptor (PPAR) ?, which may be related to their toxicities. However, species differences in the receptor functions between rodents and human make it difficult to precisely extrapolate their toxicity from animal studies to human. In this paper, we compared the species differences in the activation of mouse and human hepatic PPAR? by these plasticizers using wild-type (mPPAR?) and humanized PPAR? (hPPAR?) mice. At 12 weeks old, each genotyped male mouse was classified into three groups, and fed daily for 2 weeks per os with corn oil (vehicle control), 2.5 or 5.0?mmol/kg DBP (696, 1392?mg/kg), DEHP (977, 1953?mg/kg), and DEHA (926, 1853?mg/kg), respectively. Generally, hepatic PPAR? of mPPAR? mice was more strongly activated than that of hPPAR? mice when several target genes involving ?-oxidation of fatty acids were evaluated. Interestingly, all plasticizers also activated hepatic constitutive androstane receptor (CAR) more in hPPAR? mice than in mPPAR? mice. Taken together, these plasticizers activated mouse and human hepatic PPAR? as well as CAR. The activation of PPAR? was stronger in mPPAR? mice than in hPPAR? mice, while the opposite was true of CAR.
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Effect of nanoparticle-rich diesel exhaust on testicular and hippocampus steroidogenesis in male rats.
Inhal Toxicol
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Nanoparticle-rich diesel exhaust (NR-DE) has potentially adverse effects on testicular steroidogenesis. However, it is unclear whether NR-DE influences steroidogenic systems in the brain.
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The modulation of hepatic adenosine triphosphate and inflammation by eicosapentaenoic acid during severe fibrotic progression in the SHRSP5/Dmcr rat model.
Life Sci.
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Eicosapentaenoic acid (EPA) can ameliorate certain liver lesions involved in non-alcoholic steatohepatitis (NASH). A previous study has found that stroke-prone spontaneously hypertensive 5/Dmcr (SHRSP5/Dmcr) rats fed a high fat-cholesterol (HFC) diet developed fibrotic steatohepatitis with histological similarities to NASH. This study evaluated the potential effects and mechanisms of action of EPA supplementation using this rodent model.
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Generation of a live rabies vaccine strain attenuated by multiple mutations and evaluation of its safety and efficacy.
Vaccine
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An amino acid substitution at position 333 in rabies virus G protein is known to determine the pathogenicity: strains with Arg or Lys at that position kill adult mice after intracerebral inoculation, whereas strains with other amino acids cause non-lethal infection. Based on those findings, attenuated rabies virus strains have been established and used for oral vaccines mainly for wild animals. However, considering the possibility of back-mutation to the virulent phenotype, a strain that is attenuated by multiple mutations not only in the G protein but also in other viral proteins would be more appropriate as a safe live vaccine. We previously demonstrated that the fixed rabies virus Ni-CE strain, which causes only transient body weight loss in adult mice after intracerebral inoculation, is mainly attenuated by mutations in the N, P and M proteins, while this strain has virulent-type Arg at position 333 in the G protein. In this study, to obtain a live vaccine strain that is attenuated by multiple mutations, we generated Ni-CE mutant, Ni-CE(G333Glu) strain, which has an Arg-to-Glu mutation at position 333 in the G protein, and examined its pathogenicity and immunogenicity. We found that, in contrast to Ni-CE strain, Ni-CE(G333Glu) strain did not cause transient body weight loss in adult mice after intracerebral inoculation. The attenuated phenotype of Ni-CE(G333Glu) strain did not change even after 10 serial intracerebral passages in suckling mice. We also demonstrated that inoculation of Ni-CE(G333Glu) strain induced virus-neutralizing antibody in immunized mice and protected the mice from lethal challenge. These results indicate that Ni-CE(G333Glu) strain is a promising candidate for development of a live rabies vaccine with a high safety level.
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Urinary concentrations of organophosphorus insecticide metabolites in Japanese workers.
Chemosphere
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A recent development in analytical chemistry has enabled us to monitor systemic organophosphorus insecticide (OP) exposure at individual levels. At present, however, limited data are currently available on urinary OP metabolite levels worldwide. The purpose of this study was to assess urinary dialkylphosphate (DAP) concentrations in Japanese workers. Urine samples were collected in both summer and winter from 339 Japanese adults who worked as food distributors (FDs, n=164), apple farmers (AFs, n=147) and pest control operators (PCOs, n=28). DAPs were measured by gas chromatography-mass spectrometry after derivatization with pentafluorobenzylbromide. Dimethylphosphate (DMP), diethylphosphate (DEP), dimethylthiophosphate (DMTP) and diethylthiophosphate (DETP) were detected in the urine of over 87% of the studied populations in both seasons. The geometric mean values of total DAPs (nmol g(-1) creatinine), DMP, DMTP, DEP and DETP (?g g(-1) creatinine) in summer and winter were 106.7 and 98.3, 7.0 and 3.8, 3.4 and 4.5, 0.8 and 1.5, and 0.3 and 0.2 for the FDs, 440.8 and 197.7, 33.1 and 10.8, 10.1 and 5.8, 4.2 and 4.7 and 1.6 and 0.8 for the AFs, and 473.4 and 284.6, 28.9 and 22.2, 17.6 and 4.6, 3.5 and 4.4, and 0.5 and 0.6 for the PCOs, respectively, thereby revealing significantly higher concentrations in AFs and PCOs groups than in the FDs in both seasons except for winter DMTP. These DAP concentrations were approximately the same or at lower levels compared with those reported in the previous literature. This is one of the first studies to demonstrate urinary DAP concentrations in Japanese adults.
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[Relationship of maternal malnutrition caused by Di(2-ethylhexyl) phthalate exposure with lifestyle disease in offspring].
Nihon Eiseigaku Zasshi
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The hypothesis that offspring growing up malnourished during their fetal period have a high risk of lifestyle diseases in later life has been attracting great attention. Although animal experiments and epidemiological studies have been reported, most of them focused on the deficiency of maternal malnutritional elements or starvation. We found that di(2-ethylhexyl) phthalate (DEHP) decreased maternal plasma triglyceride levels, which is a significant source of nutrients for fetuses, in mice. Therefore, we analyzed how offspring exposed to malnutritional status during their fetal period develop potential adverse effects in later life. Male and female wild-type (mPPAR?), Ppar?-null, and hPPAR? mice were treated with diets containing 0 or 0.05% DEHP. After 4 weeks, males and females in the same genotype and dose group were mated. After continued exposure until weaning, each group was divided into two groups, and one of them was dissected. The remaining was further divided into two subgroups; one was fed normal feed (control-diet group), while the other was fed a high-fat diet (HFD group). After 8-week feeding, all the mice were dissected. In the control-diet group, DEHP exposure at the fetal and pup stages increased food consumption in mPPAR? and hPPAR? mice, but not in Ppara-null mice. In contrast, DEHP exposure decreased plasma leptin levels in mPPAR? and hPPAR? mice at the weaning stage. In the HFD group, DEHP exposure at the fetal and pup stages influenced neither food consumption nor leptin levels. These findings suggest that maternal malnutrition may be caused by not only nutritional deficiency but also exposure to some chemicals such as DEHP, and the latter case may also influence feeding behavior in offspring. These effects may be related to hepatic PPAR? and diminished by HFD feeding. Further study is warranted as to whether such feeding behavior influences the risk of lifestyle diseases such as obesity.
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Recovery of leptospires from miniature pigs experimentally infected with Leptospira interrogans serovar Manilae strain UP-MMC under immunosuppressive conditions by dexamethasone.
J. Vet. Med. Sci.
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Leptospira interrogans serovar Manilae strain UP-MMC was inoculated into miniature pigs to assess its pathogenicity. Leptospires were recovered from the whole blood, kidneys, and livers in the acute phase without showing any clinical signs. Under immunosuppressive conditions by dexamethasone, leptospires were recovered from the kidneys and their genes were detected from the urine in the chronic phase. These results indicate that leptospires persisted in the kidneys until the chronic phase, and excretion of leptospires in the urine was enhanced under immunosuppressive conditions, resulting in horizontal transmission among pigs on farms.
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