Differential expression of interferon responsive genes in rodent models of transmissible spongiform encephalopathy disease

Background

Alzheimer's disease (AD) is characterized by a decline in cognitive function and accumulation of amyloid-β peptide (Aβ) in extracellular plaques. Mutations in amyloid precursor protein (APP) and presenilins alter APP metabolism resulting in accumulation of Aβ42, a peptide essential for the formation of amyloid deposits and proposed to initiate the cascade leading to AD. However, the role of Aβ40, the more prevalent Aβ peptide secreted by cells and a major component of cerebral Aβ deposits, is less clear. In this study, virally-mediated gene transfer was used to selectively increase hippocampal levels of human Aβ42 and Aβ40 in adult Wistar rats, allowing examination of the contribution of each to the cognitive deficits and pathology seen in AD.

Results

Adeno-associated viral (AAV) vectors encoding BRI-Aβ cDNAs were generated resulting in high-level hippocampal expression and secretion of the specific encoded Aβ peptide. As a comparison the effect of AAV-mediated overexpression of APPsw was also examined. Animals were tested for development of learning and memory deficits (open field, Morris water maze, passive avoidance, novel object recognition) three months after infusion of AAV. A range of impairments was found, with the most pronounced deficits observed in animals co-injected with both AAV-BRI-Aβ40 and AAV-BRI-Aβ42. Brain tissue was analyzed by ELISA and immunohistochemistry to quantify levels of detergent soluble and insoluble Aβ peptides. BRI-Aβ42 and the combination of BRI-Aβ40+42 overexpression resulted in elevated levels of detergent-insoluble Aβ. No significant increase in detergent-insoluble Aβ was seen in the rats expressing APPsw or BRI-Aβ40. No pathological features were noted in any rats, except the AAV-BRI-Aβ42 rats which showed focal, amorphous, Thioflavin-negative Aβ42 deposits.

Conclusion

The results show that AAV-mediated gene transfer is a valuable tool to model aspects of AD pathology in vivo, and demonstrate that whilst expression of Aβ42 alone is sufficient to initiate Aβ deposition, both Aβ40 and Aβ42 may contribute to cognitive deficits.     

Efficacy of hyaluronic acid binding assay in selecting motile spermatozoa with normal morphology at high magnification

Background  

The present study aimed to evaluate the efficacy of the hyaluronic acid (HA) binding assay in the selection of motile spermatozoa with normal morphology at high magnification (8400x).     

Regulation of JH epoxide hydrolase versus JH esterase activity in the cabbage looper, Trichoplusia ni, by juvenile hormone and xenobiotics

JH III esterase and JH III epoxide hydrolase (EH) in vitro activity was compared in whole body Trichoplusia ni homogenates at each stage of development (egg, larva, pupa and adult). While activity of both enzymes was detected at all ages tested, JH esterase was significantly higher than EH activity except for day three of the fifth (last) stadium (L5D3). For both enzymes, activity was highest in eggs. Adult virgin females had 4.6- and 4.0-fold higher JH esterase and EH activities, respectively, than adult virgin males. JH III metabolic activity also was measured in whole body homogenates of fifth stadium T. ni that were fed a nutritive diet (control) or starved on a non-nutritive diet of alphacel, agar and water. With larvae that were starved for 6, 28 and 52 h, EH activity per insect equivalent was 48%, 5% and 1%, respectively, of the control insects. At the same time points, JH esterase activity levels in starved T. ni were 29%, 4% and 3% of that of insects fed the nutritive diet. Selected insect hormones and xenobiotics were administered topically or orally to fifth stadium larvae for up to 52 h, and the effects on whole body EH and JH esterase activity analyzed. JH III increased the JH III esterase activity as high as 2.2-fold, but not the JH III EH activity. The JH analog, methoprene, increased both JH esterase and EH activity as high as 2.5-fold. The JH esterase inhibitor, 3-octylthio-1,1,1-trifluoropropan-2-one (OTFP), had no impact on EH activity. The epoxides trans- and cis-stilbene oxide (TSO and CSO) in separate experiments increased the EH activity approximately 2.0-fold. TSO did not alter JH esterase levels when topically applied, but oral administration reduced activity to 70% of the control at 28 h, and then increased the activity 1.8-fold at 52 h after the beginning of treatment. CSO had no effect on JH esterase activity. Phenobarbital increased EH activity by 1.9-fold, but did not change JH esterase levels. Clofibrate and cholesterol 5alpha,6alpha-epoxide had no effect on EH. JH esterase activity also was not affected by clofibrate, but cholesterol 5alpha,6alpha-epoxide reduced the JH esterase activity to 60-80% of the control. The biological significance of these results is discussed.     

2004 update of dosimetry for the Utah Thyroid Cohort Study

In the 1980s, individual thyroid doses and uncertainties were estimated for members of a cohort of children identified in 1965 in Utah and Nevada who had potentially been exposed to fallout from the Nevada Test Site. That reconstruction represented the first comprehensive assessment of doses received by the cohort and was the first large effort to assess the uncertainty of dose on an individual person basis. The data on dose and thyroid disease prevalence during different periods were subsequently used in an analysis to determine risks of radiogenic thyroid disease. This cohort has received periodic medical follow-up to observe changes in disease frequency and to reassess the previously reported radiation-related risks, most recently after a Congressional mandate in 1998. In a recent effort to restore the databases and computer codes used to estimate doses in the 1980s, various deficiencies were found in the estimated doses due to improperly operating computer codes, corruption of secondary data files, and lack of quality control procedures. From 2001 through 2004, the dosimetry system was restored and corrected and all doses were recalculated. In addition, two parameter values were updated. While the mean of all doses has not changed significantly, many individual doses have changed by more than an order of magnitude.     

Reconstruction of the contamination of the Techa River in 1949–1951 as a result of releases from the “MAYAK” Production Association

More accurate reconstruction of the radioactive contamination of the Techa River system in 1949–1951 has been made on the basis of refined data on the amounts and the rate of discharge of radionuclides into the Techa River from the Mayak Production Association; this has led to the development of a modified Techa River model that describes the transport of radionuclides through the up-river ponds and along the Techa River and deposition of radionuclides in the river-bottom sediments and flooded areas. The refined Techa River source-term data define more precisely the time-dependent rates of release and radionuclide composition of the releases that occurred during 1949–1951. The Techa River model takes into account the time-dependent characteristics of the releases and considers (a) the transport of radionuclides adsorbed on solid particles originally contained in the discharges or originating in the up-river ponds as a result of stirring up of contaminated bottom sediments and (b) the transport of radionuclides in soluble form. The output of the Techa River model provides concentrations of all source-term radionuclides in the river water, bottom sediments, and floodplain soils at different distances from the site of radioactive releases for the period of major contamination in 1950–1951. The outputs of the model show good agreement with historical measurements of water and sediment contamination. In addition, the river-model output for ⁹⁰Sr concentration in the river water is harmonized with retrospective estimates derived from the measurements of ⁹⁰Sr in the residents of the Techa Riverside villages. Modeled contamination of the floodplain soils by ¹³⁷Cs is shown to be in agreement with the values reconstructed from late measurements of this radionuclide. Reconstructed estimates of the Techa River contamination are being used for the quantification of internal and external doses received by residents of the Techa Riverside communities.     

Routes to improving the reliability of low level DNA analysis using real-time PCR

Background  

Accurate quantification of DNA using quantitative real-time PCR at low levels is increasingly important for clinical, environmental and forensic applications. At low concentration levels (here referring to under 100 target copies) DNA quantification is sensitive to losses during preparation, and suffers from appreciable valid non-detection rates for sampling reasons. This paper reports studies on a real-time quantitative PCR assay targeting a region of the human SRY gene over a concentration range of 0.5 to 1000 target copies. The effects of different sample preparation and calibration methods on quantitative accuracy were investigated.