Cardiopulmonary toxicity of peat wildfire particulate matter and the predictive utility of precision cut lung slices

BackgroundEmissions from a large peat fire in North Carolina in 2008 were associated with increased hospital admissions for asthma and the rate of heart failure in the exposed population. Peat fires often produce larger amounts of smoke and last longer than forest fires, however few studies have reported on their toxicity. Moreover, reliable alternatives to traditional animal toxicity testing are needed to reduce the number of animals required for hazard identification and risk assessments.MethodsSize-fractionated particulate matter (PM; ultrafine, fine, and coarse) were obtained from the peat fire while smoldering (ENCF-1) or when nearly extinguished (ENCF-4). Extracted samples were analyzed for chemical constituents and endotoxin content. Female CD-1 mice were exposed via oropharyngeal aspiration to 100μg/mouse, and assessed for relative changes in lung and systemic markers of injury and inflammation. At 24h post-exposure, hearts were removed for ex vivo functional assessments and ischemic challenge. Lastly, 8mm diameter lung slices from CD-1 mice were exposed (11μg) ± co-treatment of PM with polymyxin B (PMB), an endotoxin-binding compound.ResultsOn an equi-mass basis, coarse ENCF-1PM had the highest endotoxin content and elicited the greatest pro-inflammatory responses in the mice including: increases in bronchoalveolar lavage fluid protein, cytokines (IL-6, TNF-α, and MIP-2), neutrophils and intracellular reactive oxygen species (ROS) production. Exposure to fine or ultrafine particles from either period failed to elicit significant lung or systemic effects. In contrast, mice exposed to ENCF-1 ultrafine PM developed significantly decreased cardiac function and greater post-ischemia-associated myocardial infarction. Finally, similar exposures to mouse lung slices induced comparable patterns of cytokine production; and these responses were significantly attenuated by PMB.ConclusionsThe findings suggest that exposure to coarse PM collected during a peat fire causes greater lung inflammation in association with endotoxin and ROS, whereas the ultrafine PM preferentially affected cardiac responses. In addition, lung tissue slices were shown to be a predictive, alternative assay to assess pro-inflammatory effects of PM of differing size and composition. Importantly, these toxicological findings were consistent with the cardiopulmonary health effects noted in epidemiologic reports from exposed populations

Trafficking-Deficient G572R-hERG and E637K-hERG Activate Stress and Clearance Pathways in Endoplasmic Reticulum

Background: Long QT syndrome type 2 (LQT2) is the second most common type of all long QT syndromes. It is well-known that trafficking deficient mutant human ether-a-go-go-related gene (hERG) proteins are often involved in LQT2. Cells respond to misfolded and trafficking-deficient proteins by eliciting the unfolded protein response (UPR) and Activating Transcription Factor (ATF6) has been identified as a key regulator of the mammalian UPR. In this study, we investigated the role of ER chaperone proteins (Calnexin and Calreticulin) in the processing of G572R-hERG and E637K-hERG mutant proteins. Methods: pcDNA3-WT-hERG, pcDNA3-G572R-hERG and pcDNA3-E637K-hERG plasmids were transfected into U2OS and HEK293 cells. Confocal microscopy and western blotting were used to analyze subcellular localization and protein expression. Interaction between WT or mutant hERGs and Calnexin/Calreticulin was tested by coimmunoprecipitation. To assess the role of the ubiquitin proteasome pathway in the degradation of mutant hERG proteins, transfected HEK293 cells were treated with proteasome inhibitors and their effects on the steady state protein levels of WT and mutant hERGs were examined. Conclusion: Our results showed that levels of core-glycosylated immature forms of G572R-hERG and E637K-hERG in association with Calnexin and Calreticulin were higher than that in WT-hERG. Both mutant hERG proteins could activate the UPR by upregulating levels of active ATF6. Furthermore, proteasome inhibition increased the levels of core-glycosylated immature forms of WT and mutant hERGs. In addition, interaction between mutant hERGs and Calnexin/Calreticulin wa

De Novo Transcriptome of Safflower and the Identification of Putative Genes for Oleosin and the Biosynthesis of Flavonoids

Safflower (Carthamus tinctorius L.) is one of the most extensively used oil crops in the world. However, little is known about how its compounds are synthesized at the genetic level. In this study, Solexa-based deep sequencing on seed, leaf and petal of safflower produced a de novo transcriptome consisting of 153,769 unigenes. We annotated 82,916 of the unigenes with gene annotation and assigned functional terms and specific pathways to a subset of them. Metabolic pathway analysis revealed that 23 unigenes were predicted to be responsible for the biosynthesis of flavonoids and 8 were characterized as seed-specific oleosins. In addition, a large number of differentially expressed unigenes, for example, those annotated as participating in anthocyanin and chalcone synthesis, were predicted to be involved in flavonoid biosynthesis pathways. In conclusion, the de novo transcriptome investigation of the unique transcripts provided candidate gene resources for studying oleosin-coding genes and for investigating genes related to flavonoid biosynthesis and metabolism in safflower

Coupling dynamics of a geared multibody system supported by Elastohydrodynamic lubricated cylindrical joints

A comprehensive computational methodology to study the coupling dynamics of a geared multibody system supported by ElastoHydroDynamic (EHD) lubricated cylindrical joints is proposed throughout this work. The geared multibody system is described by using the Absolute-Coordinate-Based (ACB) method that combines the Natural Coordinate Formulation (NCF) describing rigid bodies and the Absolute Nodal Coordinate Formulation (ANCF) characterizing the flexible bodies. Based on the finite-short bearing approach, the EHD lubrication condition for the cylindrical joints supporting the geared system is considered here. The lubrication forces developed at the cylindrical joints are obtained by solving the Reynolds’ equation via the finite difference method. For the evaluation of the normal contact forces of gear pair along the Line Of Action (LOA), the time-varying mesh stiffness, mesh damping and Static Transmission Error (STE) are utilized. The time-varying mesh stiffness is calculated by using the Chaari’s methodology. The forces of sliding friction along the Off-Line-Of-Action (OLOA) are computed by using the Coulomb friction models with a time-varying coefficient of friction under the EHD lubrication condition of gear teeth. Finally, two numerical examples of application are presented to demonstrate and validate the proposed methodology.National Natural Science Foundations of China under Grant 11290151, 11221202 and 11002022, Beijing Higher Education Young Elite Teacher Project under Grant YETP1201

Brown Carbon Aerosol in Urban Xi’an, Northwest China: TheComposition and Light Absorption Properties

Light-absorbing organic carbon (i.e., brown carbon or BrC) in the atmospheric aerosol has significant contribution to light absorption and radiative forcing. However, the link between BrC optical properties and chemical composition remains poorly constrained. In this study, we combine spectrophotometric measurements and chemical analyses of BrC samples collected from July 2008 to June 2009 in urban Xi'an, Northwest China. Elevated BrC was observed in winter (5 times higher than in summer), largely due to increased emissions from wintertime domestic biomass burning. The light absorption coefficient of methanol-soluble BrC at 365 nm (on average approximately twice that of water-soluble BrC) was found to correlate strongly with both parent polycyclic aromatic hydrocarbons (parent-PAHs, 27 species) and their carbonyl oxygenated derivatives (carbonyl-OPAHs, 15 species) in all seasons (r(2) > 0.61). These measured parent-PAHs and carbonyl-OPAHs account for on average similar to 1.7% of the overall absorption of methanol-soluble BrC, about 5 times higher than their mass fraction in total organic carbon (OC, similar to 0.35%). The fractional solar absorption by BrC relative to element carbon (EC) in the ultraviolet range (300-400 nm) is significant during winter (42 +/- 18% for water-soluble BrC and 76 +/- 29% for methanol-soluble BrC), which may greatly affect the radiative balance and tropospheric photochemistry and therefore the climate and air quality

Regulation of MntH by a Dual Mn(II)- and Fe(II)-Dependent Transcriptional Repressor (DR2539) in Deinococcus radiodurans

The high intracellular Mn/Fe ratio observed within the bacteria Deinococcus radiodurans may contribute to its remarkable resistance to environmental stresses. We isolated DR2539, a novel regulator of intracellular Mn/Fe homeostasis in D. radiodurans. Electrophoretic gel mobility shift assays (EMSAs) revealed that DR2539 binds specifically to the promoter of the manganese acquisition transporter (MntH) gene, and that DR0865, the only Fur homologue in D. radiodurans, cannot bind to the promoter of mntH, but it can bind to the promoter of another manganese acquisition transporter, MntABC. β-galactosidase expression analysis indicated that DR2539 acts as a manganese- and iron-dependent transcriptional repressor. Further sequence alignment analysis revealed that DR2539 has evolved some special characteristics. Site-directed mutagenesis suggested that His98 plays an important role in the activities of DR2539, and further protein-DNA binding activity assays showed that the activity of H98Y mutants decreased dramatically relative to wild type DR2539. Our study suggests that D. radiodurans has evolved a very efficient manganese regulation mechanism that involves its high intracellular Mn/Fe ratio and permits resistance to extreme conditions

Genetic Variations and Haplotype Diversity of the UGT1 Gene Cluster in the Chinese Population

Vertebrates require tremendous molecular diversity to defend against numerous small hydrophobic chemicals. UDP-glucuronosyltransferases (UGTs) are a large family of detoxification enzymes that glucuronidate xenobiotics and endobiotics, facilitating their excretion from the body. The UGT1 gene cluster contains a tandem array of variable first exons, each preceded by a specific promoter, and a common set of downstream constant exons, similar to the genomic organization of the protocadherin (Pcdh), immunoglobulin, and T-cell receptor gene clusters. To assist pharmacogenomics studies in Chinese, we sequenced nine first exons, promoter and intronic regions, and five common exons of the UGT1 gene cluster in a population sample of 253 unrelated Chinese individuals. We identified 101 polymorphisms and found 15 novel SNPs. We then computed allele frequencies for each polymorphism and reconstructed their linkage disequilibrium (LD) map. The UGT1 cluster can be divided into five linkage blocks: Block 9 (UGT1A9), Block 9/7/6 (UGT1A9, UGT1A7, and UGT1A6), Block 5 (UGT1A5), Block 4/3 (UGT1A4 and UGT1A3), and Block 3′ UTR. Furthermore, we inferred haplotypes and selected their tagSNPs. Finally, comparing our data with those of three other populations of the HapMap project revealed ethnic specificity of the UGT1 genetic diversity in Chinese. These findings have important implications for future molecular genetic studies of the UGT1 gene cluster as well as for personalized medical therapies in Chinese