Chronic apocynin treatment attenuates beta amyloid plaque size and microglial number in hAPP(751)(SL) mice

Background

NADPH oxidase is implicated in neurotoxic microglial activation and the progressive nature of Alzheimer''s Disease (AD). Here, we test the ability of two NADPH oxidase inhibitors, apocynin and dextromethorphan (DM), to reduce learning deficits and neuropathology in transgenic mice overexpressing human amyloid precursor protein with the Swedish and London mutations (hAPP(751)_SL).

Methods

Four month old hAPP(751)_SL mice were treated daily with saline, 15 mg/kg DM, 7.5 mg/kg DM, or 10 mg/kg apocynin by gavage for four months.

Results

Only hAPP(751)_SL mice treated with apocynin showed reduced plaque size and a reduction in the number of cortical microglia, when compared to the saline treated group. Analysis of whole brain homogenates from all treatments tested (saline, DM, and apocynin) demonstrated low levels of TNFα, protein nitration, lipid peroxidation, and NADPH oxidase activation, indicating a low level of neuroinflammation and oxidative stress in hAPP(751)_SL mice at 8 months of age that was not significantly affected by any drug treatment. Despite in vitro analyses demonstrating that apocynin and DM ameliorate Aβ-induced extracellular superoxide production and neurotoxicity, both DM and apocynin failed to significantly affect learning and memory tasks or synaptic density in hAPP(751)_SL mice. To discern how apocynin was affecting plaque levels (plaque load) and microglial number in vivo, in vitro analysis of microglia was performed, revealing no apocynin effects on beta-amyloid (Aβ) phagocytosis, microglial proliferation, or microglial survival.

Conclusions

Together, this study suggests that while hAPP(751)_SL mice show increases in microglial number and plaque load, they fail to exhibit elevated markers of neuroinflammation consistent with AD at 8 months of age, which may be a limitation of this animal model. Despite absence of clear neuroinflammation, apocynin was still able to reduce both plaque size and microglial number, suggesting that apocynin may have additional therapeutic effects independent of anti-inflammatory characteristics.     

Characterizing hospital workers' willingness to respond to a radiological event

Introduction

Terrorist use of a radiological dispersal device (RDD, or “dirty bomb”), which combines a conventional explosive device with radiological materials, is among the National Planning Scenarios of the United States government. Understanding employee willingness to respond is critical for planning experts. Previous research has demonstrated that perception of threat and efficacy is key in the assessing willingness to respond to a RDD event.

Methods

An anonymous online survey was used to evaluate the willingness of hospital employees to respond to a RDD event. Agreement with a series of belief statements was assessed, following a methodology validated in previous work. The survey was available online to all 18,612 employees of the Johns Hopkins Hospital from January to March 2009.

Results

Surveys were completed by 3426 employees (18.4%), whose demographic distribution was similar to overall hospital staff. 39% of hospital workers were not willing to respond to a RDD scenario if asked but not required to do so. Only 11% more were willing if required. Workers who were hesitant to agree to work additional hours when required were 20 times less likely to report during a RDD emergency. Respondents who perceived their peers as likely to report to work in a RDD emergency were 17 times more likely to respond during a RDD event if asked. Only 27.9% of the hospital employees with a perception of low efficacy declared willingness to respond to a severe RDD event. Perception of threat had little impact on willingness to respond among hospital workers.

Conclusions

Radiological scenarios such as RDDs are among the most dreaded emergency events yet studied. Several attitudinal indicators can help to identify hospital employees unlikely to respond. These risk-perception modifiers must then be addressed through training to enable effective hospital response to a RDD event.     

Arabidopsis glt1-T mutant defines a role for NADH-GOGAT in the non-photorespiratory ammonium assimilatory pathway

The physiological role of the NADH-dependent glutamine-2-oxoglutarate aminotransferase (NADH-GOGAT) enzyme was addressed in Arabidopsis using gene expression analysis and by the characterization of a knock-out T-DNA insertion mutant (glt1-T) in the single NADH-GOGAT GLT1 gene. The NADH-GOGAT GLT1 mRNA is expressed at higher levels in roots than in leaves. This expression pattern contrasts with GLU1, the major gene encoding Fd-GOGAT, which is most highly expressed in leaves and is involved in photorespiration. These distinct organ-specific expression patterns suggested a non-redundant physiological role for the NADH-GOGAT and Fd-GOGAT gene products. To test the in vivo function of NADH-GOGAT, we conducted molecular and physiological analysis of the glt1-T mutant, which is null for NADH-GOGAT, as judged by mRNA level and enzyme activity. Metabolic analysis showed that the glt1-T mutant has a specific defect in growth and glutamate biosynthesis when photorespiration was repressed by 1% CO2. Under these conditions, the glt1-T mutant displayed a 20% decrease in growth and a dramatic 70% reduction in glutamate levels. Herein, we discuss the significance of NADH-GOGAT in non-photorespiratory ammonium assimilation and in glutamate synthesis required for plant development.     

Maximum tag to body size ratios for an endangered coho salmon (<Emphasis Type="Italic">O. kisutch</Emphasis>) stock based on physiology and performance

Many coho salmon stocks (Oncorhynchus kisutch) have been in decline during the past three decades. Canada’s most endangered salmon stock, the Thompson River coho salmon, is being studied extensively as managers attempt to reverse these population declines. Investigators are using acoustic telemetry to track the migratory behaviour and survival of the Thompson River (and other) coho salmon stocks. Coho salmon pre-smolts are relatively small compared with salmonid species that are typically studied using acoustic telemetry; therefore the identification of the appropriate sizes of fish and tags to use is critical. This study tested the effects of surgically implanting the three smallest sizes of acoustic tags currently available on the growth, survival, tag retention, swimming performance and physical condition of coho salmon pre-smolts for 300 days post-surgery. Maximum tag size to body size ratios ranged from 15–17% by fork length and 7–8% by mass for the three tag sizes (11 cm fork length for a 6 × 19 mm tag, 12.5 cm for a 7 × 19 mm tag, and 14 cm for a 9 × 21 mm tag). Based on our results, it is unlikely that coho salmon pre-smolts implanted with acoustic transmitters following these size guidelines would have poor survival in studies of freshwater migratory behaviour as a result of the surgery or the tag.     

Fluorescent Amplified Fragment Length Polymorphism Analysis of Norwegian Bacillus cereus and Bacillus thuringiensis Soil Isolates

We examined 154 Norwegian B. cereus and B. thuringiensis soil isolates (collected from five different locations), 8 B. cereus and 2 B. thuringiensis reference strains, and 2 Bacillus anthracis strains by using fluorescent amplified fragment length polymorphism (AFLP). We employed a novel fragment identification approach based on a hierarchical agglomerative clustering routine that identifies fragments in an automated fashion. No method is free of error, and we identified the major sources so that experiments can be designed to minimize its effect. Phylogenetic analysis of the fluorescent AFLP results reveals five genetic groups in these group 1 bacilli. The ATCC reference strains were restricted to two of the genetic groups, clearly not representative of the diversity in these bacteria. Both B. anthracis strains analyzed were closely related and affiliated with a B. cereus milk isolate (ATCC 4342) and a B. cereus human pathogenic strain (periodontitis). Across the entire study, pathogenic strains, including B. anthracis, were more closely related to one another than to the environmental isolates. Eight strains representing the five distinct phylogenetic clusters were further analyzed by comparison of their 16S rRNA gene sequences to confirm the phylogenetic status of these groups. This analysis was consistent with the AFLP analysis, although of much lower resolution. The innovation of automated genotype analysis by using a replicated and statistical approach to fragment identification will allow very large sample analyses in the future.     

Reactivity of new adhesion molecules on lymphocytes from patients with chronic graft versus host disease

Reaction patterns of the 7th Human Leukocyte Differentiation Antigen Workshop blind panel adhesion molecules were studied on CD3/CD4, CD3/CD8, CD3/TCR gamma delta double positive T cells from peripheral blood of patients with chronic graft versus host disease (n = 8) and healthy controls (n = 4). Reactivity of 14 adhesion antibodies was tested by three-colour immunophenotyping. The mean proportion of CD3+ T cells (69 +/- 19%). CD3/CD8++ (31 +/- 13%) and CD3/TCR gamma delta++ (4 +/- 2%) T sub-populations of patients were comparable with the healthy controls. However, the mean percentage of CD3/CD4++ T cell subset in patients (14 +/- 12%) proved to be significantly decreased in comparison with the normal control value (34 +/- 16%) presumably due to secondary immunodeficiency. The workshop antibodies proved to be reactive with three T cell subsets expressing the examined antigens. Based on the results of the adhesion molecule workshop new CD categories have been introduced: CD156b as a transmembrane protein, CD167a as an epithelial tyrosin kinase receptor, CD168 as a receptor for hyaluronan mediated motility (RHAMM) and CD171 as a co-stimulatory adhesion molecule. There were significant differences in the expression of the CD167a and CD156b antigens on the CD3/CD4++ subset between the samples of patients compared with the controls characterizing the CD4+ T lymphocyte subpopulation in chronic graft versus host disease.     

Cross-beta order and diversity in nanocrystals of an amyloid-forming peptide

The seven-residue peptide GNNQQNY from the N-terminal region of the yeast prion protein Sup35, which forms amyloid fibers, colloidal aggregates and highly ordered nanocrystals, provides a model system for characterizing the elusively protean cross-beta conformation. Depending on preparative conditions, orthorhombic and monoclinic crystals with similar lath-shaped morphology have been obtained. Ultra high-resolution (<0.5A spacing) electron diffraction patterns from single nanocrystals show that the peptide chains pack in parallel cross-beta columns with approximately 4.86A axial spacing. Mosaic striations 20-50 nm wide observed by electron microscopy indicate lateral size-limiting crystal growth related to amyloid fiber formation. Frequently obtained orthorhombic forms, with apparent space group symmetry P2(1)2(1)2(1), have cell dimensions ranging from /a/=22.7-21.2A, /b/=39.9-39.3A, /c/=4.89-4.86A for wet to dried states. Electron diffraction data from single nanocrystals, recorded in tilt series of still frames, have been mapped in reciprocal space. However, reliable integrated intensities cannot be obtained from these series, and dynamical electron diffraction effects present problems in data analysis. The diversity of ordered structures formed under similar conditions has made it difficult to obtain reproducible X-ray diffraction data from powder specimens; and overlapping Bragg reflections in the powder patterns preclude separated structure factor measurements for these data. Model protofilaments, consisting of tightly paired, half-staggered beta strands related by a screw axis, can be fit in the crystal lattices, but model refinement will require accurate structure factor measurements. Nearly anhydrous packing of this hydrophilic peptide can account for the insolubility of the crystals, since the activation energy for rehydration may be extremely high. Water-excluding packing of paired cross-beta peptide segments in thin protofilaments may be characteristic of the wide variety of anomalously stable amyloid aggregates.