Estrogen and progesterone induce persistent increases in p53-dependent apoptosis and suppress mammary tumors in BALB/c-Trp53+/- mice

INTRODUCTION Treatment with estrogen and progesterone (E+P) mimics the protective effect of parity on mammary tumors in rodents and depends upon the activity of p53. The following experiments tested whether exogenous E+P primes p53 to be more responsive to DNA damage and whether these pathways confer resistance to mammary tumors in a mouse model of Li-Fraumeni syndrome. METHODS Mice that differ in p53 status (Trp53+/+, Trp53+/-, Trp53-/-) were treated with E+P for 14 days and then were tested for p53-dependent responses to ionizing radiation. Responses were also examined in parous and age-matched virgins. The effects of hormonal exposures on tumor incidence were examined in BALB/c-Trp53+/- mammary tissues. RESULTS Nuclear accumulation of p53 and apoptotic responses were increased similarly in the mammary epithelium from E+P-treated and parous mice compared with placebo and age-matched virgins. This effect was sustained for at least 7 weeks after E+P treatment and did not depend on the continued presence of ovarian hormones. Hormone stimulation also enhanced apoptotic responses to ionizing radiation in BALB/c-Trp53+/- mice but these responses were intermediate compared with Trp53+/+ and Trp-/- tissues, indicating haploinsufficiency. The appearance of spontaneous mammary tumors was delayed by parity in BALB/c-Trp53+/- mice. The majority of tumors lacked estrogen receptor (ER), but ER+ tumors were observed in both nulliparous and parous mice. However, apoptotic responses to ionizing radiation and tumor incidence did not differ among outgrowths of epithelial transplants from E+P-treated donors and nulliparous donors. CONCLUSION Therefore, E+P and parity confer a sustained increase in p53-mediated apoptosis within the mammary epithelium and suppress mammary tumorigenesis, but this effect was not retained in epithelial outgrowths.This work was supported by grants from the US Army Medical Research and Materiel Command (W81XWH0410385 to KAD and DAMD17-01-1-0315 to ACB) and the National Institutes of Health (RO1-CA095164 to DJJ)

Development of a Method for the Isolation and Quantitation of Albumin for Use in the Analysis of Protein Adducts

In order to improve exposure assessment in epidemiological studies, alternatives to traditional methods such as air sampling are under investigation. The ability to detect individual exposures through the use of biomarkers is thought to be a way to improve the accuracy and specificity of exposure assessment. Protein adducts are biomarkers which show potential for giving more specific information about exposure. Since 1985, investigators have been validating methods that measure adducts of potentially genotoxic species with serum albumin. One of the major difficulties encountered by these researchers is the isolation and quantitation of albumin. This research involved testing two procedures for the isolation of albumin from plasma (affinity chromatography and ammonium sulfate precipitation), two methods for desalting the isolated albumin (gel filtration and dialysis), and two methods for albumin quantitation (Bradford assay and absorbance at 280nm). Based on purity of albumin obtained (as judged by SDS-PAGE), comments in the literature, and applicability of the methods to screening large numbers of plasma samples, a rapid method of albumin isolation and quantitation was developed for use in analyzing protein adducts.Master of Science in Public Healt

Viral suppression of multiple escape mutants by de novo CD8+ T cell responses in a human immunodeficiency virus-1 Infected elite suppressor

Elite suppressors or controllers (ES) are HIV-1 infected patients who maintain undetectable viral loads without treatment. While HLA-B*57-positive ES are usually infected with virus that is unmutated at CTL epitopes, a single, dominant variant containing CTL escape mutations is typically seen in plasma during chronic infection. We describe an ES who developed seven distinct and rare escape variants at an HLA-B*57-restricted Gag epitope over a five year period. Interestingly, he developed proliferative, de novo CTL responses that suppressed replication of each of these variants. These responses, in combination with low viral fitness of each variant, may contribute to sustained elite control in this ES

A study of protein and amino nutrition of growing pigs.

End of Project ReportProtein nutrition of the pig is concerned primarily with supplying the amino acid requirements for fast, efficient growth and development of a lean carcass. In addition, surplus protein contributes to a high level of nitrogen excretion in manure which is a problem in complying with the Nitrates Directive. Metabolism of the excess protein / nitrogen in the pig involves creation of urea and this process depresses the efficiency of energy utilisation. As the pig grows, its requirement for individual amino acids falls but the optimum ratio changes. Providing a diet with the correct levels and balance of the principal amino acids is expected to improve performance. Improvements in genetics and changes in management such as slaughter weight require that the amino acid requirements be reassessed periodically. The objective of this study was to examine response of pigs to variation in dietary lysine in several weight ranges with the concentrations of the other principal amino acids held constant. Entire males had superior FCR to females in all trials except 15 to 30kg, but differences in dietary lysine requirements did not occur until the finishing stage. At heavier weights, response of male and female pigs began to diverge at lysine concentrations greater than 10.7 g/kg (ADG) and 9.7 g/kg (FCR). There appeared to be a need to increase the threonine to lysine ratio in the diet from 0.60 to 0.70 when lysine concentration was reduced from 12.0 to 9.5 g/kg as weight of the pig increased. Providing the same mean lysine content (11.1 g/kg) to pigs from 38 to 97 kg in a series of five diets declining in lysine concentration compared with a single diet did not affect performance, or reduce N excretion. However, lowering the overall mean lysine concentration from 11g/kg to 10.0 g/kg reduced overall N excretion by 13 %, without a negative effect on pig performance. Pigs which were offered a low lysine diet in the early stage of growth exhibited a compensatory response during realimentation on a high lysine diet but it was not sufficient to equal the overall performance of pigs previously offered a lysine-adequate diet. Nitrogen excretion was reduced by 23 % while the low lysine diet was fed in the initial period but there was no residual effect on N excretion during realimentation.Teagasc Walsh Fellowship ProgrammeNational Development Programme Funds (NDP

Spatial and temporal variability in costs and effectiveness in phosphorus loss mitigation at farm scale: A scenario analysis

Current policy instruments under the EU Water Framework Directive (WFD) to mitigate phosphorus (P) loss require that P use on farms is managed through regulation of farm gate P balances. Regulation at farm scale does not account for spatial variability in nutrient use and soil fertility at field scale, affecting the costs and effectiveness of farm gate measures. This study simulated the implementation of a P loss mitigation measure coupled with improving soil fertility so that farm productivity would not be compromised. The measure was simulated at field scale and the costs and effectiveness assessed at farm scale. Effectiveness was expressed as the time taken for excessive soil P levels to decline to levels that matched off-takes and this varied temporally and spatially within and between farms ranging from 1 to 8 years. Sub-optimum soil fertility was corrected on all fields across both farms, with applications of other soil nutrients and lime to protect productivity. An increase in costs ranging from 1.5 to 116% was predicted in the first two years of the measure on both farms after-which savings of 15-31% were predicted for each subsequent year until the measure was effective in year 9. Despite initial cost increase, there was no statistically significant difference in costs over the time taken for the measure to be effective, when compared to baseline costs. Successful implementation of measures should consider the impact on farm costs and time taken for measures to environmentally effective. Adoption of measures could improve if demonstrating to farmers that costs will not vary significantly from current practice and in time may results in savings if measures are paired with correcting soil fertility and increasing yields. This 'win-win' approach could be used into the future to ensure successful implementation and uptake of measures within the farming community

Measurements of the acid-binding capacity of ingredients used in pig diets

peer-reviewedSome feed ingredients bind more acid in the stomach than others and for this reason may be best omitted from pig starter foods if gastric acidity is to be promoted. The objective of this study was to measure the acid-binding capacity (ABC) of ingredients commonly used in pig starter foods. Ingredients were categorised as follows: (i) milk products (n = 6), (ii) cereals (n = 10), (iii) root and pulp products (n = 5), (iv) vegetable proteins (n = 11), (v) meat and fish meal (n = 2), (vi) medication (n = 3), (vii) amino acids (n = 4), (viii) minerals (n = 16), (ix) acid salts (n = 4), (x) acids (n = 10). A 0.5 g sample of food was suspended in 50 ml distilled de-ionised water with continuous stirring. This suspension was titrated with 0.1 mol/L HCl or 0.1 mol/L NaOH so that approximately 10 additions of titrant was required to reach pH 3.0. The pH readings after each addition were recorded following equilibration for three minutes. ABC was calculated as the amount of acid in milliequivalents (meq) required to lower the pH of 1 kg food to (a) pH 4.0 (ABC-4) and (b) pH 3.0 (ABC-3). Categories of food had significantly different (P < 0.01) ABC values. Mean ABC-4 and ABC-3 values of the ten categories were: (i) 623 (s.d. 367.0) and 936 (s.d. 460.2), (ii) 142 (s.d. 79.2) and 324 (s.d. 146.4), (iii) 368 (s.d. 65.3) and 804 (s.d. 126.7), (iv) 381 (s.d. 186.1) and 746 (s.d. 227.0), (v) 749 (s.d. 211.6) and 1508 (s.d. 360.8), (vi) 120 (s.d. 95.6) and 261 (s.d. 163.2), (vii) 177 (s.d. 60.7) and 1078 (s.d. 359.0), (viii) 5064 (s.d. 5525.1) and 7051 (s.d. 5911.6), (ix) 5057 (s.d. 1336.6) and 8945 (s.d. 2654.1) and (x) -5883 (s.d. 4220.5) and -2591 (s.d. 2245.4) meq HCl per kg, respectively. Within category, ABC-3 and ABC- 4 values were highly correlated: R2 values of 0.80 and greater for food categories i, iv, v, vi, vii and viii. The correlation between predicted and observed ABC values of 34 mixed diets was 0.83 for ABC-4 and 0.71 for ABC-3. It was concluded that complete diets with low ABC values may be formulated through careful selection of ingredients. The final pH to which ABC is measured should matter little as ABC-3 and ABC-4 are highly correlated

Evolution of the HIV-1 nef gene in HLA-B*57 Positive Elite Suppressors

Elite controllers or suppressors (ES) are HIV-1 infected patients who maintain viral loads of < 50 copies/ml without antiretroviral therapy. CD8+ T cells are thought to play a key role in the control of viral replication and exert selective pressure on gag and nef in HLA-B*57 positive ES. We previously showed evolution in the gag gene of ES which surprisingly was mostly due to synonymous mutations rather than non-synonymous mutation in targeted CTL epitopes. This finding could be the result of structural constraints on Gag, and we therefore examined the less conserved nef gene. We found slow evolution of nef in plasma virus in some ES. This evolution is mostly due to synonymous mutations and occurs at a rate similar to that seen in the gag gene in the same patients. The results provide further evidence of ongoing viral replication in ES and suggest that the nef and gag genes in these patients respond similarly to selective pressure from the host

Inadvertent paralog inclusion drives artifactual topologies and timetree estimates in phylogenomics

Increasingly, large phylogenomic datasets include transcriptomic data from non-model organisms. This has allowed controversial and unexplored evolutionary relationships in the tree of life to be addressed but also increases the risk of inadvertent inclusion of paralogs in the analysis. While this may be expected to result in decreased phylogenetic support it is not clear if it could also drive highly supported artefactual relationships. Many groups, including the hyper-diverse Lissamphibia, are especially susceptible to these issues due to ancient gene duplication events, small numbers of sequenced genomes and because transcriptomes are increasingly applied to resolve historically conflicting taxonomic hypotheses. We tested the potential impact of paralog inclusion on the topologies and timetree estimates of the Lissamphibia using published and de novo sequencing data including 18 amphibian species, from which 2,656 single-copy gene families were identified. A novel paralog filtering approach resulted in four differently curated datasets, which were used for phylogenetic reconstructions using Bayesian inference, maximum likelihood and quartet-based supertrees. We found that paralogs drive strongly supported conflicting hypotheses within the Lissamphibia (Batrachia and Procera) and older divergence time estimates even within groups where no variation in topology was observed. All investigated methods, except Bayesian inference with the CAT-GTR model, were found to be sensitive to paralogs, but with filtering convergence to the same answer (Batrachia) was observed. This is the first large-scale study to address the impact of orthology selection using transcriptomic data and emphasises the importance of quality over quantity particularly for understanding relationships of poorly sampled taxa

Identifying incident colorectal and lung cancer cases in health service utilisation databases in Australia: a validation study

Data from centralised, population-based statutory cancer registries are generally considered the 'gold standard' for confirming incident cases of cancer. When these are not available, or more current information is needed, hospital or other routinely collected population-level data may be feasible alternative sources. We aimed to determine the validity of various methods using routinely collected administrative health data for ascertaining incident cases of colorectal or lung cancer in participants from the 45 and Up Study in New South Wales (NSW), Australia. METHODS: For 266,844 participants in the 45 and Up Study (recruited 2006-2009) ascertainment of incident colorectal or lung cancers was assessed using diagnosis and treatment records in linked administrative health datasets (hospital, emergency department, Medicare and pharmaceutical claims, death records). This was compared with ascertainment via the NSW Cancer Registry (NSWCR, the 'gold standard') for a period for which both data sources were available for participants. RESULTS: A total of 2253 colorectal and 1019 lung cancers were recorded for study participants in the NSWCR over the period 2006-2010. A diagnosis of primary cancer recorded in the statewide Admitted Patient Data Collection identified the majority of NSWCR colorectal and lung cancers, with sensitivities and positive predictive values (PPV) of 95% and 91% for colorectal cancer and 81% and 85% for lung cancer, respectively. Using additional information on lung cancer deaths from death records increased sensitivity to 84% (PPV 83%) for lung cancer, but did not improve ascertainment of colorectal cancers. Hospital procedure codes for colorectal cancer surgery identified cases with sensitivity 81% and PPV 54%. No other individual indicator had sensitivity >50% or PPV >65% for either cancer type and no combination of indicators increased both the sensitivity and PPV above that achieved using the hospital cancer diagnosis data. All specificities were close to 100%; 95% confidence intervals for sensitivity and PPV were generally +/-2%. CONCLUSIONS: In NSW, identifying new cases of colorectal and lung cancer from administrative health datasets, such as hospital records, is a feasible alternative when cancer registry data are not available. However, the strengths and limitations of the different data sources should be borne in mind

Spatial and temporal variability in costs and effectiveness in phosphorus loss mitigation at farm scale: A scenario analysis

Current policy instruments under the EU Water Framework Directive (WFD) to mitigate phosphorus (P) loss require that P use on farms is managed through regulation of farm gate P balances. Regulation at farm scale does not account for spatial variability in nutrient use and soil fertility at field scale, affecting the costs and effectiveness of farm gate measures. This study simulated the implementation of a P loss mitigation measure coupled with improving soil fertility so that farm productivity would not be compromised. The measure was simulated at field scale and the costs and effectiveness assessed at farm scale. Effectiveness was expressed as the time taken for excessive soil P levels to decline to levels that matched off-takes and this varied temporally and spatially within and between farms ranging from 1 to 8 years. Sub-optimum soil fertility was corrected on all fields across both farms, with applications of other soil nutrients and lime to protect productivity. An increase in costs ranging from 1.5 to 116% was predicted in the first two years of the measure on both farms after-which savings of 15–31% were predicted for each subsequent year until the measure was effective in year 9. Despite initial cost increase, there was no statistically significant difference in costs over the time taken for the measure to be effective, when compared to baseline costs. Successful implementation of measures should consider the impact on farm costs and time taken for measures to environmentally effective. Adoption of measures could improve if demonstrating to farmers that costs will not vary significantly from current practice and in time may results in savings if measures are paired with correcting soil fertility and increasing yields. This ‘win-win’ approach could be used into the future to ensure successful implementation and uptake of measures within the farming community