Evaluation of analgesic interaction between morphine, maropitant and dexmedetomidine in dogs undergoing ovariohysterectomy.

AIMS: To compare the analgesic effect of morphine combined with maropitant and/or dexmedetomidine to morphine alone but at a higher dose, and to evaluate the pharmacokinetics of the drug combinations, in dogs undergoing ovariohysterectomy (OHE). METHODS: Forty client-owned dogs were randomised into four treatment groups (n = 10 per group) each to receive a different analgesic protocol. After premedication with I/M acepromazine, anaesthesia was induced with propofol to effect and maintained with isoflurane in 100% oxygen delivered via a circle system. The heart rate, respiratory rate, blood pressure, haemoglobin oxygen saturation, end-tidal partial pressure of carbon dioxide, electrocardiogram and rectal temperature were monitored during anaesthesia. The test drugs (Mor: 0.6 mg/kg morphine; Maro + Mor: 0.3 mg/kg morphine and 1 mg/kg maropitant; Dex + Mor: 0.3 mg/kg morphine and 10 µg/kg dexmedetomidine; Dex + Maro + Mor: 0.2 mg/kg morphine, 7 µg/kg dexmedetomidine and 0.7 mg/kg maropitant) were administered I/M after induction of anaesthesia and 30 minutes before the expected start time of ovariohysterectomy, which was carried out by veterinary students under veterinary supervision. The short form of the Glasgow composite measure pain scale (CMPS-SF) and visual analogue scale (VAS) were used for pain assessment at 15 and 30 minutes and 1, 2, 3, 6, 9 and 24 hours after extubation. Dogs with CMPS-SF pain score ≥ 6 received rescue analgesia with S/C buprenorphine (0.02 mg/kg). Blood samples were collected before, 15, 30, 60 and 120 minutes after injection of the test drugs and concentration of the test drugs in plasma was determined by liquid chromatography-mass spectrometry. RESULTS: Dogs that received Dex + Mor had significantly lower CMPS-SF (estimate of difference = -1.53 (SE 0.58); p = 0.010) and VAS (estimate of difference = -0.67 (SE 0.25); p = 0.007) scores compared to the dogs that received morphine alone. There was no evidence of a difference in the number of dogs requiring rescue between groups. All dogs that received dexmedetomidine showed cardiac arrhythmia and second-degree heart block. Mean (SD) maximum concentrations (Cmax,) of morphine in plasma were 6.8 (4.56), 9.56 (8.29), 9.30 (3.35) and 18.99 (9.41) ng/mL for the groups Dex + Mor, Dex + Maro + Mor, Maro + Mor and Mor respectively. The Cmax of morphine was significantly lower in the Dex + Mor (p = 0.004), Dex + Maro + Mor (p = 0.034) and Maro + Mor (p = 0.018) groups compared to the Mor group. CONCLUSIONS: For dogs undergoing ovariohysterectomy, lower doses of morphine (0.2 and 0.3 mg/kg) combined with dexmedetomidine or maropitant may provide analgesia equivalent to or better than morphine when given alone at a higher dose (0.6 mg/kg).Abbreviations: AUC: Area under curve; Cmax: Maximum concentration in plasma; CMPS-SF: Glasgow composite measure pain scale - short form; NK1: Neurokinin-1; OHE: Ovariohysterectomy; Tmax: Time to Cmax; T1/2: Half-life of terminal elimination phase; VAS: Visual analogue scale.

Operating list composition and surgical performance.

BACKGROUND: Recent reviews suggest that the way in which surgeons prepare for a procedure (warm up) can affect performance. Operating lists present a natural experiment to explore this phenomenon. The aim was to use a routinely collected large data set on surgical procedures to understand the relationship between case list order and operative performance. METHOD: Theatre lists involving the 35 procedures performed most frequently by senior surgeons across 38 private hospitals in the UK over 26 months were examined. A linear mixed-effects model and matched analysis were used to estimate the impact of list order and the cost of switching between procedures on a list while controlling for key prognosticators. The influence of procedure method (open versus minimally invasive) and complexity was also explored. RESULTS: The linear mixed-effects model included 255 757 procedures, and the matched analysis 48 632 pairs of procedures. Repeating the same procedure in a list resulted in an overall time saving of 0·98 per cent for each increase in list position. Switching between procedures increased the duration by an average of 6·48 per cent. The overall reduction in operating time from completing the second procedure straight after the first was 6·18 per cent. This pattern of results was consistent across procedure method and complexity. CONCLUSION: There is a robust relationship between operating list composition and surgical performance (indexed by duration of operation). An evidence-based approach to structuring a theatre list could reduce the total operating time.

Evaluation of non-Saccharomyces yeasts for the reduction of alcohol content in wine.

Over recent decades, the average ethanol concentration of wine has increased, largely due to consumer preference for wine styles associated with increased grape maturity; sugar content increases with grape maturity, and this translates into increased alcohol content in wine. However, high ethanol content impacts wine sensory properties, reducing the perceived complexity of flavors and aromas. In addition, for health and economic reasons, the wine sector is actively seeking technologies to facilitate the production of wines with lower ethanol content. Nonconventional yeast species, in particular, non-Saccharomyces yeasts, have shown potential for producing wines with lower alcohol content. These yeast species, which are largely associated with grapes preharvest, are present in the early stages of fermentation but, in general, are not capable of completing alcoholic fermentation. We have evaluated 50 different non-Saccharomyces isolates belonging to 24 different genera for their capacity to produce wine with a lower ethanol concentration when used in sequential inoculation regimes with a Saccharomyces cerevisiae wine strain. A sequential inoculation of Metschnikowia pulcherrima AWRI1149 followed by an S. cerevisiae wine strain was best able to produce wine with an ethanol concentration lower than that achieved with the single-inoculum, wine yeast control. Sequential fermentations utilizing AWRI1149 produced wines with 0.9% (vol/vol) and 1.6% (vol/vol) (corresponding to 7.1 g/liter and 12.6 g/liter, respectively) lower ethanol concentrations in Chardonnay and Shiraz wines, respectively. In Chardonnay wine, the total concentration of esters and higher alcohols was higher for wines generated from sequential inoculations, whereas the total concentration of volatile acids was significantly lower. In sequentially inoculated Shiraz wines, the total concentration of higher alcohols was higher and the total concentration of volatile acids was reduced compared with those in control S. cerevisiae wines, whereas the total concentrations of esters were not significantly different.

KRAS and BRAF mutations are rare and related to DNA mismatch repair deficiency in gastric cancer from the East and the West: results from a large international multicentre study.

BACKGROUND: Inhibitors of the epidermal growth factor (EGFR) signaling pathway have a major role in the treatment of KRAS wild-type colorectal cancer patients. The EGFR pathway has been shown to be activated in gastric cancer (GC). However, published data on KRAS and BRAF mutation status is limited in GC and has not been compared between GC from different geographic regions. METHODS: The prevalence of KRAS and BRAF mutations was established in 712 GC: 278 GC from the United Kingdom, 230 GC from Japan and 204 GC from Singapore. The relationship between KRAS/BRAF mutation status, DNA mismatch repair (MMR) status, clinicopathological variables and overall survival was analysed. RESULTS: Overall, 30 (4.2%) GC carried a KRAS mutation. In total, 5.8% of the UK GC, 4% of Japan GC and 1.5% of Singapore GC were KRAS mutant. KRAS mutant GC had fewer lymph node metastases in the UK cohort (P=0.005) and were more frequent in elderly patients in the Japan cohort (P=0.034). KRAS mutations were more frequent in MMR-deficient GC in the UK and the Japanese cohort (P<0.05). A BRAF mutation was only detected in a single Japanese GC. CONCLUSIONS: This large multicentre study demonstrated that KRAS mutations and DNA MMR deficiency have a role in a small subgroup of GC irrespective of country of origin, suggesting that this subgroup of GC may have developed along a common pathway. Further studies need to establish whether concomitant mutations or amplifications of other EGFR signalling pathway genes may contribute to the activation of this pathway in GC.

Development and validation of a risk score (Delay-7) to predict the occurrence of a treatment delay following cycle 1 chemotherapy.

BACKGROUND: The risk of toxicity-related dose delays, with cancer treatment, should be included as part of pretreatment education and be considered by clinicians upon prescribing chemotherapy. An objective measure of individual risk could influence clinical decisions, such as escalation of standard supportive care and stratification of some patients, to receive proactive toxicity monitoring. PATIENTS AND METHODS: We developed a logistic regression prediction model (Delay-7) to assess the overall risk of a chemotherapy dose delay of 7 days for patients receiving first-line treatments for breast, colorectal and diffuse large B-cell lymphoma. Delay-7 included hospital treated, age at the start of chemotherapy, gender, ethnicity, body mass index, cancer diagnosis, chemotherapy regimen, colony stimulating factor use, first cycle dose modifications and baseline blood values. Baseline blood values included neutrophils, platelets, haemoglobin, creatinine and bilirubin. Shrinkage was used to adjust for overoptimism of predictor effects. For internal validation (of the full models in the development data) we computed the ability of the models to discriminate between those with and without poor outcomes (c-statistic), and the agreement between predicted and observed risk (calibration slope). Net benefit was used to understand the risk thresholds where the model would perform better than the 'treat all' or 'treat none' strategies. RESULTS: A total of 4604 patients were included in our study of whom 628 (13.6%) incurred a 7-day delay to the second cycle of chemotherapy. Delay-7 showed good discrimination and calibration, with c-statistic of 0.68 (95% confidence interval 0.66-0.7), following internal validation and calibration-in-the-large of -0.006. CONCLUSIONS: Delay-7 predicts a patient's individualised risk of a treatment-related delay at cycle two of treatment. The score can be used to stratify interventions to reduce the occurrence of treatment-related toxicity.

Chemotherapy dose reductions in obese patients with colorectal cancer.

BACKGROUND: In cancer patients with a large Body Surface Area, chemotherapy drug doses are often reduced, as studies have suggested that their pharmacokinetics may be altered. However, this strategy may result in underdosing obese patients. PATIENTS AND METHODS: In three Medical Research Council trials of chemotherapy for advanced colorectal cancer, dose reductions were not mandated. This provided the opportunity to compare the toxicity levels in those obese patients fully dosed and to investigate if those under dosed experienced a worse survival. Body Mass Index (BMI) was used to classify patients as normal weight (BMI < 25), overweight (BMI 25-29), or obese (BMI 30+). RESULTS: Of the 4781 patients, 2158 (45%) were classified as normal weight, 1753 (37%) as overweight, and 870 (18%) as obese. There was no evidence that, in those patients fully dosed, obese patients experienced more toxicity or that dose-reducing obese patients resulted in less toxicity. However, there was a suggestion that those obese patients who were given reduced doses had a worse progression-free survival [hazard ratio (HR) 1.21, 95% confidence interval (CI) 1.06-1.39, P = 0.006] and a slightly worse overall survival (HR 1.12, 95% CI 0.96-1.30, P = 0.152). CONCLUSION: These results, although not a randomised comparison, do not support the policy of reducing chemotherapy doses for obese patients with colorectal cancer.

Evaluation of gene modification strategies for the development of low-alcohol-wine yeasts.

Saccharomyces cerevisiae has evolved a highly efficient strategy for energy generation which maximizes ATP energy production from sugar. This adaptation enables efficient energy generation under anaerobic conditions and limits competition from other microorganisms by producing toxic metabolites, such as ethanol and CO(2). Yeast fermentative and flavor capacity forms the biotechnological basis of a wide range of alcohol-containing beverages. Largely as a result of consumer demand for improved flavor, the alcohol content of some beverages like wine has increased. However, a global trend has recently emerged toward lowering the ethanol content of alcoholic beverages. One option for decreasing ethanol concentration is to use yeast strains able to divert some carbon away from ethanol production. In the case of wine, we have generated and evaluated a large number of gene modifications that were predicted, or known, to impact ethanol formation. Using the same yeast genetic background, 41 modifications were assessed. Enhancing glycerol production by increasing expression of the glyceraldehyde-3-phosphate dehydrogenase gene, GPD1, was the most efficient strategy to lower ethanol concentration. However, additional modifications were needed to avoid negatively affecting wine quality. Two strains carrying several stable, chromosomally integrated modifications showed significantly lower ethanol production in fermenting grape juice. Strain AWRI2531 was able to decrease ethanol concentrations from 15.6% (vol/vol) to 13.2% (vol/vol), whereas AWRI2532 lowered ethanol content from 15.6% (vol/vol) to 12% (vol/vol) in both Chardonnay and Cabernet Sauvignon juices. Both strains, however, produced high concentrations of acetaldehyde and acetoin, which negatively affect wine flavor. Further modifications of these strains allowed reduction of these metabolites.

Adaptive evolution of Saccharomyces cerevisiae to generate strains with enhanced glycerol production.

The development of new wine yeast strains with improved characteristics is critical in the highly competitive wine market, which faces the demand of ever-changing consumer preferences. Although new strains can be constructed using recombinant DNA technologies, consumer concerns about genetically modified (GM) organisms strongly limit their use in food and beverage production. We have applied a non-GM approach, adaptive evolution with sulfite at alkaline pH as a selective agent, to create a stable yeast strain with enhanced glycerol production; a desirable characteristic for wine palate. A mutant isolated using this approach produced 41% more glycerol than the parental strain it was derived from, and had enhanced sulfite tolerance. Backcrossing to produce heterozygous diploids revealed that the high-glycerol phenotype is recessive, while tolerance to sulfite was partially dominant, and these traits, at least in part, segregated from each other. This work demonstrates the potential of adaptive evolution for development of novel non-GM yeast strains, and highlights the complexity of adaptive responses to sulfite selection.

Development of environmental thresholds for streams in agricultural watersheds.

Global increases in consumption of chemical nutrients, application of pesticides, and water withdrawal to enhance agricultural yield have resulted in degraded water quality and reduced water availability. Efforts to safeguard or improve environmental conditions of agroecosystems have usually focused on managing on-farm activities to reduce materials loss and conserve habitat. Another management measure for improving environmental quality is adoption of environmental performance standards (also called outcome-based standards). This special collection of six papers presents the results of four years of research to devise scientifically credible approaches for setting environmental performance standards to protect water quantity and quality in Canadian agriculturally dominated watersheds. The research, conducted as part of Canada's National Agri-Environmental Standards Initiative, aimed to identify Ideal Performance Standards (the desired environmental state needed to maintain ecosystem health) and Achievable Performance Standards (the environmental conditions achievable using currently available and recommended best available processes and technologies). Overviews of the papers, gaps in knowledge, and future research directions are presented. As humans, livestock, and wildlife (both terrestrial and aquatic) experience greater pressures to share the same limited water resources, innovative research is needed that incorporates a landscape perspective, economics, farm practices, and ecology to advance the development and application of tools for protecting water resources in agricultural watersheds.

Effects of the veterinary pharmaceutical salinomycin and its formulation on the plant Brassica rapa.

Veterinary pharmaceuticals are emerging contaminants found throughout the environment, and their presence and effects are a matter of concern. The purpose of this study was to compare the phytotoxicity of salinomycin (pure compound = 96 %) and Sacox 120 (formulated product = 120 g salinomycin/kg) to the plant species Brassica rapa as well as to investigate salinomycin persistence in soil. Calculated EC/IC(50) values for salinomycin and Sacox 120 were 1.10 and 2.88 and 2.19 and 18.03 mg/kg, respectively, based on salinomycin concentration. For exposure of B. rapa to salinomycin, significant adverse effects were observed for growth end points at the greater concentrations. For the reproduction end point (i.e., number of buds), as well as root length and wet mass, significant differences were observed at the lower concentrations (stimulating growth) and adverse effects at the greater concentrations. This study confirmed that the toxic effects of Sacox 120 are attributable to the active ingredient salinomycin. Liquid chromatography-electrospray ionization-mass spectrometry analyses confirmed that exposure concentrations of salinomycin were 90 and 83 % of the nominal concentrations, respectively, in the soils amended with either pure or formulated product. At the end of the experiment, after 14 days, salinomycin concentrations for both tests (salinomycin and Sacox 120) decreased to 6.2 and 5.8 % of the nominal exposure concentrations, respectively. Detected salinomycin concentrations in plant shoots ranged from 3.47 to 41.0 ng/g dry shoot. This study shows the importance of using plants as tools to evaluate environmental risk and as a bridge to relate environment and human health risks.

KRAS and BRAF mutations in Nigerian colorectal cancers.

PURPOSE: Activation of the KRAS oncogene is implicated in colorectal carcinogenesis and mutations have been reported in 30-50% of cases. BRAF mutation, though less common, is also reported and importantly associated with shorter progression-free interval. This study aims to determine the KRAS and BRAF mutation statuses of Nigerian colorectal cancers (CRC). METHODS: Mutation analysis was carried out on archival paraffin-embedded blocks of CRC tissues. KRAS codons 12, 13 and 61 and BRAF V600E were assessed by pyrosequencing after DNA extraction from 200 cases at the Leeds Institute of Molecular Medicine, St. James's University Hospital, UK. Mutation rates and the spectra were determined. RESULTS: Pyrosequencing was successful in 112 of 200 cases. KRAS mutation in codons 12 and 13 was demonstrated in 23 of 112 cases (21%); none in codon 61. BRAF mutation in codon 600 was demonstrated in 4.5%. CONCLUSION: This study shows that 21% of Nigerian CRC patients carry a KRAS mutation; half the rate in Caucasians; and that BRAF mutation also occurs in Nigerian CRC cancers.

A novel methodology independent of fermentation rate for assessment of the fructophilic character of wine yeast strains.

The yeast Saccharomyces cerevisiae has a fundamental role in fermenting grape juice to wine. During alcoholic fermentation its catabolic activity converts sugars (which in grape juice are a near equal ratio of glucose and fructose) and other grape compounds into ethanol, carbon dioxide and sensorily important metabolites. However, S. cerevisiae typically utilises glucose and fructose with different efficiency: glucose is preferred and is consumed at a higher rate than fructose. This results in an increasing difference between the concentrations of glucose and fructose during fermentation. In this study 20 commercially available strains were investigated to determine their relative abilities to utilise glucose and fructose. Parameters measured included fermentation duration and the kinetics of utilisation of fructose when supplied as sole carbon source or in an equimolar mix with glucose. The data were then analysed using mathematical calculations in an effort to identify fermentation attributes which were indicative of overall fructose utilisation and fermentation performance. Fermentation durations ranged from 74.6 to over 150 h, with clear differences in the degree to which glucose utilisation was preferential. Given this variability we sought to gain a more holistic indication of strain performance that was independent of fermentation rate and therefore utilized the area under the curve (AUC) of fermentation of individual or combined sugars. In this way it was possible to rank the 20 strains for their ability to consume fructose relative to glucose. Moreover, it was shown that fermentations performed in media containing fructose as sole carbon source did not predict the fructophilicity of strains in wine-like conditions (equimolar mixture of glucose and fructose). This work provides important information for programs which seek to generate strains that are faster or more reliable fermenters.

Application of nitrogen and phosphorus criteria for streams in agricultural landscapes.

Efforts to control eutrophication of water resources in agriculturally dominated ecosystems have focused on managing on-farm activities to reduce nutrient loss; however, another management measure for improving water quality is adoption of environmental performance criteria (or 'outcome-based standards'). Here, we review approaches for setting environmental quality criteria for nutrients, summarize approaches developed in Canada for setting 'ideal' and 'achievable' nutrient criteria for streams in agricultural watersheds, and consider how such criteria could be applied. As part of a 'National Agri-Environmental Standards Initiative', the Government of Canada committed to the development of non-regulatory environmental performance standards that establish total P (TP) and total N (TN) concentrations to protect ecological condition of agricultural streams. Application of four approaches for defining ideal standards using only chemistry data resulted in values for TP and TN spanning a relatively narrow range of concentrations within a given ecoregion. Cross-calibration of these chemically derived standards with information on biological condition resulted in recommendations for TP and TN that would likely protect aquatic life from adverse effects of eutrophication. Non-point source water quality modelling was then conducted in a specific watershed to estimate achievable standards, i.e. chemical conditions that could be attained using currently available and recommended management practices. Our research showed that, taken together, short-term achievable standards and ultimate ideal standards could be used to set policy targets that should, if realized, lower N and P concentrations in Canadian agricultural streams and improve biotic condition.

The ethanol stress response and ethanol tolerance of Saccharomyces cerevisiae.

Saccharomyces cerevisiae is traditionally used for alcoholic beverage and bioethanol production; however, its performance during fermentation is compromised by the impact of ethanol accumulation on cell vitality. This article reviews studies into the molecular basis of the ethanol stress response and ethanol tolerance of S. cerevisiae; such knowledge can facilitate the development of genetic engineering strategies for improving cell performance during ethanol stress. Previous studies have used a variety of strains and conditions, which is problematic, because the impact of ethanol stress on gene expression is influenced by the environment. There is however some commonality in Gene Ontology categories affected by ethanol assault that suggests that the ethanol stress response of S. cerevisiae is compromised by constraints on energy production, leading to increased expression of genes associated with glycolysis and mitochondrial function, and decreased gene expression in energy-demanding growth-related processes. Studies using genome-wide screens suggest that the maintenance of vacuole function is important for ethanol tolerance, possibly because of the roles of this organelle in protein turnover and maintaining ion homoeostasis. Accumulation of Asr1 and Rat8 in the nucleus specifically during ethanol stress suggests S. cerevisiae has a specific response to ethanol stress although this supposition remains controversial.

How does climate variability affect water quality dynamics in Canada's oil sands region?

In Canada's oil sands region, classic boreal hydrology (i.e., winter low flow followed by peaks during spring freshet and then summer flow recession) combined with erosion of both natural and anthropogenically-exposed bitumen results in seasonal and inter-annual variability in stream water chemistry. Using data collected from all seasons over three years (2012-2015), we investigated the mechanisms driving spatial and temporal change in the concentration of 26 water quality parameters for six rivers draining Canada's oil sands region. Mantel tests showed a strong spatial aggregation of climatic drivers (average daily precipitation, accumulated precipitation, snow water equivalent) associated with west versus east discharge patterns. Wavelet analysis highlighted unique watershed attributes, in particular the importance of developed area in lowering responsiveness to seasonal precipitation. Concentrations of most chemical parameters (20 of 23) showed distinct temporal patterns that were correlated with seasonal changes in hydrology which, in turn, were related to changes in weather. Comparison of concentrations observed in this study with those reported in the scientific literature for the same watersheds showed 81% of comparisons differed significantly. This was likely due to the short duration of previous field campaigns and thus the sampling of a very narrow window of the annual streamflow regime.

Differential CpG DNA methylation in peripheral naïve CD4+ T-cells in early rheumatoid arthritis patients.

BACKGROUND: The genetic risk associated with rheumatoid arthritis (RA) includes genes regulating DNA methylation, one of the hallmarks of epigenetic re-programing, as well as many T-cell genes, with a strong MHC association, pointing to immunogenetic mechanisms as disease triggers leading to chronicity. The aim of our study was to explore DNA methylation in early, drug-naïve RA patients, towards a better understanding of early events in pathogenesis. RESULT: Monocytes, naïve and memory CD4+ T-cells were sorted from 6 healthy controls and 10 RA patients. DNA methylation was assessed using a genome-wide Illumina 450K CpG promoter array. Differential methylation was confirmed using bisulfite sequencing for a specific gene promoter, ELISA for several cytokines and flow cytometry for cell surface markers. Differentially methylated (DM) CpGs were observed in 1047 genes in naïve CD4+ T-cells, 913 in memory cells and was minimal in monocytes with only 177 genes. Naive CD4+ T-cells were further investigated as presenting differential methylation in the promoter of > 500 genes associated with several disease-relevant pathways, including many cytokines and their receptors. We confirmed hypomethylation of a region of the TNF-alpha gene in early RA and differential expression of 3 cytokines (IL21, IL34 and RANKL). Using a bioinformatics package (DMRcate) and an in-house analysis based on differences in ß values, we established lists of DM genes between health and RA. Publicly available gene expression data were interrogated to confirm differential expression of over 70 DM genes. The lists of DM genes were further investigated based on a functional relationship database analysis, which pointed to an IL6/JAK1/STAT3 node, related to TNF-signalling and engagement in Th17 cell differentiation amongst many pathways. Five DM genes for cell surface markers (CD4, IL6R, IL2RA/CD25, CD62L, CXCR4) were investigated towards identifying subpopulations of CD4+ T-cells undergoing these modifications and pointed to a subset of naïve T-cells, with high levels of CD4, IL2R, and CXCR4, but reduction and loss of IL6R and CD62L, respectively. CONCLUSION: Our data provided novel conceptual advances in the understanding of early RA pathogenesis, with implications for early treatment and prevention.

Influence of alginate backbone on efficacy of thermo-responsive alginate-g-P(NIPAAm) hydrogel as a vehicle for sustained and controlled gene delivery.

Alginate grafted poly(N-isopropylacrylamide) hydrogels (Alg-g-P(NIPAAm)) form three-dimensional networks in mild conditions, making them suitable for incorporation of labile macromolecules, such as DNA. The impact of P(NIPAAm) on copolymer characteristics has been well studied, however the impact of alginate backbone characteristics on copolymer properties has to-date not been investigated. Six different Alg-g-P(NIPAAm) hydrogels were synthesised with 10% alginate, which varied in terms of molecular weight (MW) and mannuronate/guluronate (M/G) monomer ratio, and with 90% NIPAAm in order to develop an injectable and thermo-responsive hydrogel formulation for localised gene delivery. Hydrogel stiffness was directly proportional to MW and the M/G ratio of the alginate backbone. Hydrogels with a high MW or low M/G ratio alginate backbone demonstrated a greater stiffness than those hydrogels comprising low MW alginates and high M/G ratio. Hydrogels with a high M/G ratio also produced a complexed and meshed hydrogel network while hydrogels with a low M/G ratio produced a simplified structure with the superposition of Alg-g-P(NIPAAm) sheets. This study was designed to produce the optimal Alg-g-P(NIPAAm) hydrogel with respect to localised delivery of DNA nanoparticles as a potential medical device for those with castrate resistant prostate cancer (CRPC). Given that CRPC typically disseminates to bone causing pain, morbidity and a plethora of skeletal related events, a copolymer based hydrogel was designed to for long term release of therapeutic DNA nanoparticles. The nanoparticles were comprised of plasmid DNA (pDNA), complexed with an amphipathic cell penetrating peptide termed RALA that is designed to enter cells with high efficiency. Alginate MW and M/G ratio affected stiffness, structure, injectability and degradation of the Alg-g-P(NIPAAm) hydrogel. Algogel 3001, had the optimal characteristics for long-term application and was loaded with RALA/pDNA NPs. From the release profiles, it was evident that RALA protected the pDNA from degradation over a 30-day period and conferred a sustained and controlled release profile from the hydrogels compared to pDNA only. Taken together, we have designed a slowly degrading hydrogel suitable for sustained delivery of nucleic acids when incorporated with the RALA delivery peptide. This now opens up several opportunities for the delivery of therapeutic pDNA from this thermo-responsive hydrogel with numerous medical applications.

Catchment properties and the photosynthetic trait composition of freshwater plant communities.

Unlike in land plants, photosynthesis in many aquatic plants relies on bicarbonate in addition to carbon dioxide (CO2) to compensate for the low diffusivity and potential depletion of CO2 in water. Concentrations of bicarbonate and CO2 vary greatly with catchment geology. In this study, we investigate whether there is a link between these concentrations and the frequency of freshwater plants possessing the bicarbonate use trait. We show, globally, that the frequency of plant species with this trait increases with bicarbonate concentration. Regionally, however, the frequency of bicarbonate use is reduced at sites where the CO2 concentration is substantially above the air equilibrium, consistent with this trait being an adaptation to carbon limitation. Future anthropogenic changes of bicarbonate and CO2 concentrations may alter the species compositions of freshwater plant communities.

PTCH mutations in basal cell carcinomas from azathioprine-treated organ transplant recipients.

The immunosuppressant azathioprine is used to prevent graft rejection after organ transplantation. To investigate whether azathioprine-associated mutagenesis contributes to the high incidence of skin tumours in organ transplant recipients (OTRs), we analysed PTCH gene mutations in 60 basal cell carcinomas (BCC); 39 from OTRs receiving azathioprine and 21 from individuals never exposed to azathioprine. PTCH was mutated in 55% of all tumours, independent of azathioprine treatment. In both the azathioprine and non-azathioprine groups, transitions at dipyrimidine sequences, considered to indicate mutation by ultraviolet-B radiation, occurred frequently in tumours from chronically sun-exposed skin. In BCC from non-sun-exposed skin of azathioprine-treated patients, there was an over-representation of unusual G:C to A:T transitions at non-dipyrimidine sites. These were exclusive to the azathioprine-exposed group and all in the same TGTC sequence context at different positions within PTCH. Meta-analysis of 247 BCCs from published studies indicated that these mutations are rare in sporadic BCC and had never previously been reported in this specific sequence context. This study of post-transplant BCC provides the first indication that azathioprine exposure may be associated with PTCH mutations, particularly in tumours from non-sun-exposed skin.

Skeletal and Appendage Diversity as Design Elements in the Synthesis of a Discovery Library of Nonaromatic Polycyclic 5-Iminooxazolidin-2-ones, Hydantoins, and Acylureas.

Amino acid-derived cross-conjugated trienes were used as a starting point for the synthesis of a discovery library of over 200 polycyclic 5-iminooxazolidin-2-ones, hydantoins, and acylureas. The main feature of this library synthesis is a triple branching strategy which provides efficient access to five skeletally diverse scaffolds. In addition, four sets of building blocks were applied in both a front end and a back end diversification strategy. Multiple fused rings were obtained by cyclization of diamides with phosgene and stereoselective Diels-Alder reactions with maleimides. The 5-iminooxazolidin-2-one scaffold was rearranged into the isomeric hydantoin scaffold through a sequence of ring opening and ring closing reactions.