The inhibition of the iron responsive element RNA-protein interaction by heme does not mimic in vivo iron regulation

Hemin at greater than 1 microM concentrations inhibits the interaction of the iron responsive element (IRE) and the iron responsive element binding protein (IRE-BP) as measured by gel retardation and UV cross-linking. Heme has recently been proposed to inhibit the repression of translation of an IRE-containing mRNA (Lin, J. J., Daniels-McQueen, S., Patino, M. M., Gaffield, L., Walden, W. E., and Thach, R. E., (1990) Science 247, 74-76). Our binding inhibition provides structural support for these observations. The action of hemin, however, does not mimic the physiologically demonstrated inhibition of high affinity binding of the IRE to IRE-BP by the oxidation of a sulfhydryl of the IRE-BP. In addition to this effect, hemin also inhibits a wide variety of RNA and DNA binding proteins, restriction endonucleases, and nucleases. Therefore, in vitro, the inhibitory effects of hemin are not limited to the interaction of the IRE-BP and the IRE, but are nonspecific and affect a wide variety of nucleic acid-protein interactions. Any hypothesis on the effects on protein-nucleic acid interactions employing greater than 1 microM concentrations of hemin should be interpreted with caution.     

A Novel Zeolite Coating for Protection of Concrete Sewers from Biological Sulfuric Acid Attack

Microbial growth inhibition and resistance to biological deterioration of concrete specimens coated with silver-loaded zeolite was evaluated by measuring the time course of bacterial growth, biological sulfur oxidation, and sulfate production using Acidithiobacillus thiooxidans as a corrosive agent. Live bacterial cells declined from an initial inoculum concentration of 1.1 × 10₄ cell ml_-1 to zero in 10 days, during which only 0.5–1% of the initial sulfur concentration of 10 g l_-1 was biologically oxidized, corresponding to sulfate production rates of 35–42 mg SO ₄ ^{2 ?} g ^{? 1} S ^{? 1} . Leaching coefficients of calcium and silicon in the specimens coated with silver-loaded zeolite of 1.6 × 10 ^{? 4} to 4.6 × 10 ^{? 2} cm ² d ^{? 1} respectively, were only 0.8% and 1% of the uncoated specimens.     

Detection of Innate Immune Response Modulating Impurities in Therapeutic Proteins

Therapeutic proteins can contain multiple impurities, some of which are variants of the product, while others are derived from the cell substrate and the manufacturing process. Such impurities, even when present at trace levels, have the potential to activate innate immune cells in peripheral blood or embedded in tissues causing expression of cytokines and chemokines, increasing antigen uptake, facilitating processing and presentation by antigen presenting cells, and fostering product immunogenicity. Currently, while products are tested for host cell protein content, assays to control innate immune response modulating impurities (IIRMIs) in products are focused mainly on endotoxin and nucleic acids, however, depending on the cell substrate and the manufacturing process, numerous other IIRMI could be present. In these studies we assess two approaches that allow for the detection of a broader subset of IIRMIs. In the first, we use commercial cell lines transfected with Toll like receptors (TLR) to detect receptor-specific agonists. This method is sensitive to trace levels of IIRMI and provides information of the type of IIRMIs present but is limited by the availability of stably transfected cell lines and requires pre-existing knowledge of the IIRMIs likely to be present in the product. Alternatively, the use of a combination of macrophage cell lines of human and mouse origin allows for the detection of a broader spectrum of impurities, but does not identify the source of the activation. Importantly, for either system the lower limit of detection (LLOD) of impurities was similar to that of PBMC and it was not modified by the therapeutic protein tested, even in settings where the product had inherent immune modulatory properties. Together these data indicate that a cell-based assay approach could be used to screen products for the presence of IIRMIs and inform immunogenicity risk assessments, particularly in the context of comparability exercises.     

Relationship of Serum Vitamin D Concentrations and Allostatic Load as a Measure of Cumulative Biological Risk among the US Population: A Cross-Sectional Study

IntroductionThe allostatic load (AL) index is a multi-systemic measure of physiologic dysregulation known to be associated with chronic exposure to stress and adverse health outcomes. We examined the relationship between AL and serum 25-hydroxyvitamin D (25(OH)D) concentration in non-institutionalized US adults.MethodsData from the Third National Health and Nutrition Examination Survey (NHANES III, 1988–94) were used to calculate two versions of AL including 9 biomarkers and another two with 14 biomarkers (systolic and diastolic blood pressure, pulse rate, serum cholesterol, serum HDL-cholesterol, glycated hemoglobin, sex-specific waist-to-hip ratio, serum albumin, and serum C-reactive protein for AL1, and, additionally body mass index, serum triglyceride, serum creatinine, and serum herpes I & II antibodies for AL2), each set defined by predefined cut-offs or by quartiles. Serum vitamin D concentration was ranked into quartiles. Logistic regression, Poisson regression and linear regression were used to examine the association of serum 25(OH)D concentrations on AL, after adjusting for biological, physiological, socioeconomic, lifestyle, and health variables.ResultsOdds Ratios (OR) for high AL of the lowest 25(OH)D serum quartile were between 1.45 (95% CI: 1.28, 1.67) and 1.79 (95% CI: 1.39, 2.32) for the fully adjusted model, depending on AL version. Inverse relationships between vitamin D serum concentrations were observed for all AL versions and every adjustment. This relationship was consistent after stratification by sex, age or ethnic background. Sensitivity to low 25(OH)D concentrations was highest among the youngest group (20–39 years) with an OR of 2.11 (95% CI: 1.63, 2.73) for the lowest vitamin D quartile Q1.ConclusionsVitamin D had a consistent and statistically significant inverse association with all tested models of high AL, which remained consistent after adjusting for biological, socioeconomic, lifestyle and health variables. Our study adds evidence linking low 25(OH)D concentrations with poorer health, further-reaching than bone health.     

Genome-wide association mapping of genomic regions associated with phenotypic traits in Canadian western spring wheat

We recently reported genomic regions associated with resistance to four wheat diseases and insensitivity to three Pyrenophora tritici-repentis toxins in an association mapping panel consisting of 81 diverse Canadian western spring wheat (Triticum aestivum L.) cultivars. Here, we report genomic regions and SNPs associated with days to heading, plant maturity, plant height, test weight (grain volume weight), grain yield, and grain protein content in the same population using genome-wide association studies (GWAS). The 81 spring wheat cultivars were evaluated for the above six traits across six environments and genotyped with 19,919 polymorphic SNPs and 14 gene-specific markers. Using mixed liner model and a threshold of p ≤ 3.1 × 10^?4, we identified a total of 139 significant marker-trait associations that were mapped at 19 genomic regions on 11 chromosomes for heading (3 regions), maturity (2), plant height (3), test weight (3), grain yield (6), and grain protein (2). Each region consisted of clusters of markers ranging from 2 to 33 and individually explained from 4.5 to 26.1% of the phenotypic variation averaged over six environments. Some the genomic regions identified in the present study are novel, while others, such as the regions for grain protein on 1B, days to heading on 5A, plant height on 4B, and test weight on 7A, were located close to either known genes or QTLs reported in previous studies, but direct comparisons in some cases were challenging due to lack of common set of markers and reliable physical positions among the different studies. Results from this study provide additional information to wheat researchers developing improved spring wheat cultivars.     

Selection Practices of Bonga Sheep Reared in Southern Ethiopia

The study was conducted to assess the traits traditionally used to select the Bonga sheep reared in Southern Ethiopia. The breed was included for improvement under the community based breeding program (CBBP) launched in the year 2009. The results are based on the data collected from the project between 2009 till 2012. The findings are based on focus group discussions with the community elders who have been rearing the Bonga sheep for more than 20 years. The present findings are based on information obtained from 50% of the total respondents who were involved in CBBP. The Bonga sheep is a mutton type breed and the ewes are moderately prolific. In the past this breed of sheep were bartered to settle legal disputes and were traded against household commodities.. However, over the years (and also after the intervention of CBBP) the sheep have been selected for their body weight as well as for their distribution in many parts of Ethiopia The respondents selected the lambs at both pre weaning and post weaning stages. Traditionally the traits of rams are selected based on their body length, canon circumference, broad face, enlarged thyroid, while for the ewes, traits prolificacy, skin thickness and pelvic width are determining characters. Fat tail and brown coat color were preferred irrespective of the sexes in trait selection. The ram lambs selected for breeding purpose are locally known as “Dookoo”. These rams are selected based on some predetermined phenotypic traits and are initially selected at preweaning stage and further, again at the post weaning stage. These rams are preferentially cared and are provided with supplementary feed and comfortable housing.     

CCR2 receptor antagonists: optimization of biaryl sulfonamides to increase activity in whole blood

A series of biarylsulfonamides was identified as hCCR2 receptor antagonist but suffered from high plasma protein binding resulting in a >100 fold shift in activity in a functional GTPγS assay run in tandem in the presence and absence of human serum albumin. Introduction of an aryl amide with ethylenediamine linker led to compounds with reduced shifts and improved activity in whole blood.     

DNA-based faecal dietary analysis: a comparison of qPCR and high throughput sequencing approaches

The genetic analysis of faecal material represents a relatively non-invasive way to study animal diet and has been widely adopted in ecological research. Due to the heterogeneous nature of faecal material the primary obstacle, common to all genetic approaches, is a means to dissect the constituent DNA sequences. Traditionally, bacterial cloning of PCR amplified products was employed; less common has been the use of species-specific quantitative PCR (qPCR) assays. Currently, with the advent of High-Throughput Sequencing (HTS) technologies and indexed primers it has become possible to conduct genetic audits of faecal material to a much greater depth than previously possible. To date, no studies have systematically compared the estimates obtained by HTS with that of qPCR. What are the relative strengths and weaknesses of each technique and how quantitative are deep-sequencing approaches that employ universal primers? Using the locally threatened Little Penguin (Eudyptula minor) as a model organism, it is shown here that both qPCR and HTS techniques are highly correlated and produce strikingly similar quantitative estimates of fish DNA in faecal material, with no statistical difference. By designing four species-specific fish qPCR assays and comparing the data to the same four fish in the HTS data it was possible to directly compare the strengths and weaknesses of both techniques. To obtain reproducible quantitative data one of the key, and often overlooked, steps common to both approaches is ensuring that efficient DNA isolation methods are employed and that extracts are free of inhibitors. Taken together, the methodology chosen for long-term faecal monitoring programs is largely dependent on the complexity of the prey species present and the level of accuracy that is desired. Importantly, these methods should not be thought of as mutually exclusive, as the use of both HTS and qPCR in tandem will generate datasets with the highest fidelity.     

Development of SPECT imaging agents for the norepinephrine transporters: [123I]INER

A series of reboxetine analogs was synthesized and evaluated for in vitro binding as racemic mixtures. The best candidate (INER) was synthesized as the optically pure (S,S) enantiomer, labeled with iodine-123 and its in vivo binding determined by SPECT imaging in baboons. The in vivo specificity, selectivity, and kinetics of [123I]INER make it a promising agent for imaging NET in vivo by noninvasive SPECT imaging.