ELISA analysis of β-secretase cleavage of the Swedish amyloid precursor protein in the secretory and endocytic pathways

Limiting beta amyloid (Aβ) production could become an important therapeutic target in Alzheimer's disease (AD). Aβ is derived by the sequential cleavage of amyloid precursor protein (APP) by β- and γ-secretases. A double missense mutation in APP found in a Swedish pedigree (APPsw) elevates Aβ40 and Aβ42 production. Aβ production and, therefore, β-secretase cleavage of APPsw reportedly occur in the endoplasmic reticulum (ER), Golgi and endocytic compartments. However, the relative contribution of β-secretase cleavage occurring in each compartment has not been determined. Experiments described here use a novel ELISA to measure the β-cleaved product, APPswβ. Using this ELISA, we provide new information regarding the relative amount of β-secretase cleavage of APPsw that occurs in secretory and endocytic pathways. Using a dilysine retrieval motif to retain APPsw in the ER, we discovered that less than 15% of the β-secretase cleavage was still detected. Experiments utilizing endocytosis-impaired mutants of APPsw revealed that little or no β-secretase cleavage of APPsw appears to take place through endocytosis. Surprisingly, deletion of the entire cytoplasmic tail increased both APPswβ and Aβ secretion, suggesting that protein interactions with this region normally impede β-secretase cleavage. These results suggest that APPsw is cleaved by β-secretase late in the secretory pathway.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/66393/1/j.0022-3042.2002.00764.x.pd

Survey of sediment quality in Sabine Lake, Texas and vicinity

The toxicity of sediments in Sabine Lake, Texas, and adjoining Intracoastal Waterway canals was determined as part of bioeffects assessment studies managed by NOAA’s National Status and Trends Program. The objectives of the survey were to determine: (1) the incidence and degree of toxicity of sediments throughout the study area; (2) the spatial patterns (or gradients) in chemical contamination and toxicity, if any, throughout the study area; (3) the spatial extent of chemical contamination and toxicity; and (4) the statistical relationships between measures of toxicity and concentrations of chemicals in the sediments. Surficial sediment samples were collected during August, 1995 from 66 randomly-chosen locations. Laboratory toxicity tests were performed as indicators of potential ecotoxicological effects in sediments. A battery of tests was performed to generate information from different phases (components) of the sediments. Tests were selected to represent a range in toxicological endpoints from acute to chronic sublethal responses. Toxicological tests were conducted to measure: reduced survival of adult amphipods exposed to solid-phase sediments; impaired fertilization success and abnormal morphological development in gametes and embryos, respectively, of sea urchins exposed to pore waters; reduced metabolic activity of a marine bioluminescent bacteria exposed to organic solvent extracts; and induction of a cytochrome P-450 reporter gene system in exposures to solvent extracts of the sediments. Chemical analyses were performed on portions of each sample to quantify the concentrations of trace metals, polynuclear aromatic hydrocarbons, and chlorinated organic compounds. Correlation analyses were conducted to determine the relationships between measures of toxicity and concentrations of potentially toxic substances in the samples. Based upon the compilation of results from chemical analyses and toxicity tests, the quality of sediments in Sabine Lake and vicinity did not appear to be severely degraded. Chemical concentrations rarely exceeded effects-based numerical guidelines, suggesting that toxicant-induced effects would not be expected in most areas. None of the samples was highly toxic in acute amphipod survival tests and a minority (23%) of samples were highly toxic in sublethal urchin fertilization tests. Although toxic responses occurred frequently (94% of samples) in urchin embryo development tests performed with 100% pore waters, toxicity diminished markedly in tests done with diluted pore waters. Microbial bioluminescent activity was not reduced to a great degree (no EC50 <0.06 mg/ml) and cytochrome P-450 activity was not highly induced (6 samples exceeded 37.1 ug/g benzo[a]pyrene equivalents) in tests done with organic solvent extracts. Urchin embryological development was highly correlated with concentrations of ammonia and many trace metals. Cytochrome P450 induction was highly correlated with concentrations of a number of classes of organic compounds (including the polynuclear aromatic hydrocarbons and chlorinated compounds). (PDF contains 51 pages

T cell epitope clustering in the highly immunogenic BZLF1 antigen of Epstein-Barr virus

Polymorphism in the human leukocyte antigen (HLA) loci ensures that the CD8 T cell response to viruses is directed against a diverse range of antigenic epitopes, thereby minimizing the impact of virus escape mutation across the population. The BZLF1 antigen of Epstein-Barr virus is an immunodominant target for CD8 T cells, but the response has been characterized only in the context of a limited number of HLA molecules due to incomplete epitope mapping. We have now greatly expanded the number of defined CD8 T cell epitopes from BZLF1, allowing the response to be evaluated in a much larger proportion of the population. Some regions of the antigen fail to be recognized by CD8 T cells, while others include clusters of overlapping epitopes presented by different HLA molecules. These highly immunogenic regions of BZLF1 include polymorphic sequences, such that up to four overlapping epitopes are impacted by a single amino acid variation common in different regions of the world. This focusing of the immune response to limited regions of the viral protein could be due to sequence similarity to human proteins creating "immune blind spots" through self-tolerance. This study significantly enhances the understanding of the immune response to BZLF1, and the precisely mapped T cell epitopes may be directly exploited in vaccine development and adoptive immunotherapy

Quantum oscillations in the parent pnictide BaFe2_2As2_2 : itinerant electrons in the reconstructed state

We report quantum oscillation measurements that enable the direct observation of the Fermi surface of the low temperature ground state of \ba122. From these measurements we characterize the low energy excitations, revealing that the Fermi surface is reconstructed in the antiferromagnetic state, but leaving itinerant electrons in its wake. The present measurements are consistent with a conventional band folding picture of the antiferromagnetic ground state, placing important limits on the topology and size of the Fermi surface.Comment: 5 pages, 3 figure

Efficacy of antimicrobial 405 nm blue-light for inactivation of airborne bacteria

Airborne transmission of infectious organisms is a considerable concern within the healthcare environment. A number of novel methods for ‘whole room’ decontamination, including antimicrobial 405 nm blue light, are being developed. To date, research has focused on its effects against surface-deposited contamination; however, it is important to also establish its efficacy against airborne bacteria. This study demonstrates evidence of the dose-response kinetics of airborne bacterial contamination when exposed to 405 nm light and compares bacterial susceptibility when exposed in three different media: air, liquid and surfaces. Bacterial aerosols of Staphylococcus epidermidis, generated using a 6-Jet Collison nebulizer, were introduced into an aerosol suspension chamber. Aerosolized bacteria were exposed to increasing doses of 405 nm light, and air samples were extracted from the chamber using a BioSampler liquid impinger, with viability analysed using pour-plate culture. Results have demonstrated successful aerosol inactivation, with a 99.1% reduction achieved with a 30 minute exposure to high irradiance (22 mWcm-2) 405 nm light (P=0.001). Comparison to liquid and surface exposures proved bacteria to be 3-4 times more susceptible to 405 nm light inactivation when in aerosol form. Overall, results have provided fundamental evidence of the susceptibility of bacterial aerosols to antimicrobial 405 nm light treatment, which offers benefits in terms of increased safety for human exposure, and eradication of microbes regardless of antibiotic resistance. Such benefits provide advantages for a number of applications including ‘whole room’ environmental decontamination, in which reducing levels of airborne bacteria should reduce the number of infections arising from airborne contamination

Characterising rhamnolipid production in Burkholderia thailandensis E264, a non-pathogenic producer

Burkholderia thailandensis E264 is a rhamnolipid (RL)-producing gram-negative bacterium first isolated from the soils and stagnant waters of central and north-eastern Thailand. Growth of B. thailandensis E264 under two different incubation temperatures (25 and 30 °C) resulted in a significantly higher dry cell biomass production at 30 °C (7.71 g/l) than at 25 °C (4.75 g/l) after 264 h; however, incubation at the lower temperature resulted in consistently higher concentration of RL production throughout the growth period. After 264 h, the concentration of crude RL extract for the 25 °C culture was 2.79 g/l compared to 1.99 g/l for the 30 °C culture. Overall RL production concentration after 264 h was 0.258 g/g dry cell biomass (DCB) for the 30 °C culture compared to 0.587 g/g DCB for the 25 °C culture. Real-time PCR (qPCR) was also used to analyse expression of the RL biosynthesis genes throughout the incubation period at 25 °C showing that the expression of the rhlA, rhlB and rhlC genes is continuous. During the log and early stationary phases of growth, expression levels remain low and are increased upon entry to the late stationary phase. B. thailandensis E264 produces mostly di-RLs and the Di-RL C14-C14 in most abundance (41.88 %). Fermentations were also carried out in small-scale bioreactors (4 l working volume) under controlled conditions, and results showed that RL production was maintained. Our findings show that B. thailandensis E264 has excellent potential for industrial scale RL production

Niche shifts and energetic condition of songbirds in response to phenology of food-resource availability in a high-elevation sagebrush ecosystem

Seasonal fluctuations in food availability can affect diets of consumers, which in turn may influence the physiological state of individuals and shape intra- and inter-specific patterns of resource use. High-elevation ecosystems often exhibit a pronounced seasonal “pulse” in productivity, although few studies document how resource use and energetic condition by avian consumers change in relation to food-resource availability in these ecosystems. We tested the hypothesis that seasonal increases (pulses) in food resources in high-elevation sagebrush ecosystems result in 2 changes after the pulse, relative to the before-pulse period: (1) reduced diet breadth of, and overlap between, 2 sympatric sparrow species; and (2) enhanced energetic condition in both species. We tracked breeding-season diets using stable isotopes and energetic condition using plasma metabolites of Brewer\u27s Sparrows (Spizella breweri), Vesper Sparrows (Pooecetes gramineus), and their food resources during 2011, and of only Brewer\u27s Sparrows and their food resources during 2013. We quantify diet breadth and overlap between both species, along with coincident physiological consequences of temporal changes in resource use. After invertebrate biomass increased following periods of rainfall in 2011, dietary breadth decreased by 35% in Brewer\u27s Sparrows and by 48% in Vesper Sparrows, while dietary overlap decreased by 88%. Energetic condition of both species increased when dietary overlap was lower and diet breadth decreased, after the rapid rise of food-resource availability. However, energetic condition of Brewer\u27s Sparrows remained constant in 2013, a year with low precipitation and lack of a strong pulse in food resources, even though the species\u27 dietary breadth again decreased that year. Our results indicate that diet breadth and overlap in these sparrow species inhabiting sagebrush ecosystems generally varied as predicted in relation to intra- and interannual changes in food resources, and this difference in diet was associated with improved energetic condition of sparrows at least in one year