Daytime pattern of post-exercise protein intake affects whole-body protein turnover in resistance-trained males

Background The pattern of protein intake following exercise may impact whole-body protein turnover and net protein retention. We determined the effects of different protein feeding strategies on protein metabolism in resistance-trained young men. Methods: Participants were randomly assigned to ingest either 80g of whey protein as 8x10g every 1.5h (PULSE; n=8), 4x20g every 3h (intermediate, INT; n=7), or 2x40g every 6h (BOLUS; n=8) after an acute bout of bilateral knee extension exercise (4x10 repetitions at 80% maximal strength). Whole-body protein turnover (Q), synthesis (S), breakdown (B), and net balance (NB) were measured throughout 12h of recovery by a bolus ingestion of [ 15N]glycine with urinary [15N]ammonia enrichment as the collected end-product. Results PULSE Q rates were greater than BOLUS (?19%, P<0.05) with a trend towards being greater than INT (?9%, P=0.08). Rates of S were 32% and 19% greater and rates of B were 51% and 57% greater for PULSE as compared to INT and BOLUS, respectively (P<0.05), with no difference between INT and BOLUS. There were no statistical differences in NB between groups (P=0.23); however, magnitude-based inferential statistics revealed likely small (mean effect90%CI; 0.590.87) and moderate (0.800.91) increases in NB for PULSE and INT compared to BOLUS and possible small increase (0.421.00) for INT vs. PULSE. Conclusion We conclude that the pattern of ingested protein, and not only the total daily amount, can impact whole-body protein metabolism. Individuals aiming to maximize NB would likely benefit from repeated ingestion of moderate amounts of protein (?20g) at regular intervals (?3h) throughout the day.

Facile mutant identification via a single parental backcross method and application of whole genome sequencing based mapping pipelines

Forward genetic screens have identified numerous genes involved in development and metabolism, and remain a cornerstone of biological research. However, to locate a causal mutation, the practice of crossing to a polymorphic background to generate a mapping population can be problematic if the mutant phenotype is difficult to recognize in the hybrid F2 progeny, or dependent on parental specific traits. Here in a screen for leaf hyponasty mutants, we have performed a single backcross of an Ethane Methyl Sulphonate (EMS) generated hyponastic mutant to its parent. Whole genome deep sequencing of a bulked homozygous F2 population and analysis via the Next Generation EMS mutation mapping pipeline (NGM) unambiguously determined the causal mutation to be a single nucleotide polymorphisim (SNP) residing in HASTY, a previously characterized gene involved in microRNA biogenesis. We have evaluated the feasibility of this backcross approach using three additional SNP mapping pipelines; SHOREmap, the GATK pipeline, and the samtools pipeline. Although there was variance in the identification of EMS SNPs, all returned the same outcome in clearly identifying the causal mutation in HASTY. The simplicity of performing a single parental backcross and genome sequencing a small pool of segregating mutants has great promise for identifying mutations that may be difficult to map using conventional approaches.

Tissue viability imaging of skin microcirculation following exposure to whole body cryotherapy (-110°C) and cold water immersion (8°C)

Cryotherapy is currently used in various clinical, rehabilitative, and sporting settings. However, very little is known regarding the impact of cooling on the microcirculatory response. Objectives: The present study sought to examine the influence of two commonly employed modalities of cryotherapy, whole body cryotherapy (WBC; -110°C) and cold water immersion(CWI; 8±1°C), on skin microcirculation in the mid- thigh region. Methods: The skin area examined was a 3 × 3 cm located between the most anterior aspect of the inguinal fold and the patella. Following 10 minutes of rest, 5 healthy, active males were exposed to either WBC for 3 minutes or CWI for 5 minutes in a randomised order. Volunteers lay supine for five minutes after treatment, in order to monitor the variation of red blood cell (RBC) concentration in the region of interest for a duration of 40 minutes. Microcirculation response was assessed using a non-invasive, portable instrument known as a Tissue Viability imaging system. After a minimum of seven days, the protocol was repeated. Subjective assessment of the volunteer’s thermal comfort and thermal sensation was also recorded. Results: RBC was altered following exposure to both WBC and CWI but appeared to stabilise approximately 35 minutes after treatments. Both WBC and CWI affected thermal sensation (p < 0.05); however no betweengroup differences in thermal comfort or sensation were recorded (p > 0.05). Conclusions: As both WBC and CWI altered RBC, further study is necessary to examine the mechanism for this alteration during whole body cooling.

The effect of three different (-135°C) whole body cryotherapy exposure durations on elite rugby league players

Background: Whole body cryotherapy (WBC) is the therapeutic application of extreme cold air for a short duration. Minimal evidence is available for determining optimal exposure time. Purpose: To explore whether the length of WBC exposure induces differential changes in inflammatory markers, tissue oxygenation, skin and core temperature, thermal sensation and comfort. Method: This study was a randomised cross over design with participants acting as their own control. Fourteen male professional first team super league rugby players were exposed to 1, 2, and 3 minutes of WBC at -135°C. Testing took place the day after a competitive league fixture, each exposure separated by seven days. Results: No significant changes were found in the inflammatory cytokine interleukin six. Significant reductions (p<0.05) in deoxyhaemoglobin for gastrocnemius and vastus lateralis were found. In vastus lateralis significant reductions (p<0.05) in oxyhaemoglobin and tissue oxygenation index (p<0.05) were demonstrated. Significant reductions (p<0.05) in skin temperature were recorded. No significant changes were recorded in core temperature. Significant reductions (p<0.05) in thermal sensation and comfort were recorded. Conclusion: Three brief exposures to WBC separated by 1 week are not sufficient to induce physiological changes in IL-6 or core temperature. There are however significant changes in tissue oxyhaemoglobin, deoxyhaemoglobin, tissue oxygenation index, skin temperature and thermal sensation. We conclude that a 2 minute WBC exposure was the optimum exposure length at temperatures of -135°C and could be applied as the basis for future studies.

Treating the whole not the hole : necessary coupling of technologies for diabetic foot ulcer treatment

Highlights • Diabetic foot ulcers (DFUs) are a major complication of diabetes. • We describe the development of next-generation technologies for DFU repair. • We highlight the modest success of growth factor-, scaffold-, and cell-based DFU therapies. • We rationalize that combination therapies will be necessary to enable effective and reliable DFU repair.

Framework of whole life cycle costing for Malaysian high rise residential property development

This project was an initial stage in formulating and management of the optimum budget allocation during the operational, maintenance and rehabilitation phases in high rise residential property development in Malaysia. The principal objective of this project is to develop a framework of Whole Life Cycle Costing for high rise residential property development that will enhance the quality and cost effectiveness of this building type in Malaysia. The researcher investigated 13 building components from 6 high rise residential property developments in Johor, Malaysia to determine the affect and economic impact of component initial cost and quality by applying them to a Whole Life Cycle Cost model approach. The results provide valuable data in respect to the overall cost of specific components over the whole life of a large high rise building. In addition, Dr. Mat Noor also determined the impact and satisfaction of quality of building components through WLCC.

Stepwise engineering of a Pichia pastoris D-amino acid oxidase whole cell catalyst

Trigonopsis variabilis D-amino acid oxidase (TvDAO) is a well characterized enzyme used for cephalosporin C conversion on industrial scale. However, the demands on the enzyme with respect to activity, operational stability and costs also vary with the field of application. Processes that use the soluble enzyme suffer from fast inactivation of TvDAO while immobilized oxidase preparations raise issues related to expensive carriers and catalyst efficiency. Therefore, oxidase preparations that are more robust and active than those currently available would enable a much broader range of economically viable applications of this enzyme in fine chemical syntheses. A multi-step engineering approach was chosen here to develop a robust and highly active Pichia pastoris TvDAO whole-cell biocatalyst. As compared to the native T. variabilis host, a more than seven-fold enhancement of the intracellular level of oxidase activity was achieved in P. pastoris through expression optimization by codon redesign as well as efficient subcellular targeting of the enzyme to peroxisomes. Multi copy integration further doubled expression and the specific activity of the whole cell catalyst. From a multicopy production strain, about 1.3 x 103 U/g wet cell weight (wcw) were derived by standard induction conditions feeding pure methanol. A fed-batch cultivation protocol using a mixture of methanol and glycerol in the induction phase attenuated the apparent toxicity of the recombinant oxidase to yield final biomass concentrations in the bioreactor of >or= 200 g/L compared to only 117 g/L using the standard methanol feed. Permeabilization of P. pastoris using 10% isopropanol yielded a whole-cell enzyme preparation that showed 49% of the total available intracellular oxidase activity and was notably stabilized (by three times compared to a widely used TvDAO expressing Escherichia coli strain) under conditions of D-methionine conversion using vigorous aeration. Stepwise optimization using a multi-level engineering approach has delivered a new P. pastoris whole cell TvDAO biocatalyst showing substantially enhanced specific activity and stability under operational conditions as compared to previously reported preparations of the enzyme. The production of the oxidase through fed-batch bioreactor culture and subsequent cell permeabilization is high-yielding and efficient. Therefore this P. pastoris catalyst has been evaluated for industrial purposes.

10,000 steps Rockhampton : (1) building a whole community approach to increasing levels of physical activity

10,000 Steps Rockhampton is a multi-strategy health promotion program which aims to develop sustainable community-based strategies to increase physical activity.The central coordinating focus of the project is the use of pedometers to raise awareness of and provide motivation for physical activity, around the theme of '10,000 steps/day - Every step counts.' To date, five key strategies have been implemented: (1) a media-based awareness raising campaign; (2) promotion of physical activity by health professionals; (3) improving social support for physical activity through group-based programs; (4) working with local council to improve environmental support for physical activity, and; (5) establishment of a ‘micro-grants’ fund to which community groups could apply for assistance with small, innovative physical activity enhancing projects. Strategies were introduced on a rolling basis beginning in February 2002 with 'layering' of interventions designed to address the multi-level individual social and environmental determinants of physical activity. The project was quasi-experimental in design, involving collection of baseline and two year follow-up data from community based surveys in Rockhampton and in a matched regional Queensland town. In August 2001,the baseline CATI survey (N=1281)found that 47.9% of men and 33.0% of women were meeting the national guidelines for physical activity. In August 2002, a smaller survey (N=400) found an increase in activity levels among women (39.7% active) but not in men (48.5%). Data from the two year follow up survey, to be conducted in August 2003, will be presented, with discussion of the major successes and challenges of this landmark physical activity intervention. Acknowledgement: This project is supported by a grant from Health Promotion Queensland.

10,000 steps Rockhampton: a whole community approach to increasing levels of physical activity

10,000 Steps Rockhampton is a multi-strategy health promotion program which aims to develop sustainable community-based strategies to increase physical activity.The central coordinating focus of the project is the use of pedometers to raise awareness of and provide motivation for physical activity, around the theme of '10,000 steps/day - Every step counts.' To date, five key strategies have been implemented: (1) a media-based awareness raising campaign; (2) promotion of physical activity by health professionals; (3) improving social support for physical activity through group-based programs; (4) working with local council to improve environmental support for physical activity, and; (5) establishment of a ‘micro-grants’ fund to which community groups could apply for assistance with small, innovative physical activity enhancing projects. Strategies were introduced on a rolling basis beginning in February 2002 with 'layering' of interventions designed to address the multi-level individual social and environmental determinants of physical activity. The project was quasi-experimental in design, involving collection of baseline and two year follow-up data from community based surveys in Rockhampton and in a matched regional Queensland town. In August 2001,the baseline CATI survey (N=1281)found that 47.9% of men and 33.0% of women were meeting the national guidelines for physical activity. In August 2002, a smaller survey (N=400) found an increase in activity levels among women (39.7% active) but not in men (48.5%). Data from the two year follow up survey, to be conducted in August 2003, will be presented, with discussion of the major successes and challenges of this landmark physical activity intervention. Acknowledgement: This project is supported by a grant from Health Promotion Queensland

Integration of science and monitoring of river ecosystem health to guide investments in catchment protection and rehabilitation

1. Stream ecosystem health monitoring and reporting need to be developed in the context of an adaptive process that is clearly linked to identified values and objectives, is informed by rigorous science, guides management actions and is responsive to changing perceptions and values of stakeholders. To be effective, monitoring programmes also need to be underpinned by an understanding of the probable causal factors that influence the condition or health of important environmental assets and values. This is often difficult in stream and river ecosystems where multiple stressors, acting at different spatial and temporal scales, interact to affect water quality, biodiversity and ecosystem processes. 2. In this article, we describe the development of a freshwater monitoring programme in South East Queensland, Australia, and how this has been used to report on ecosystem health at a regional scale and to guide investments in catchment protection and rehabilitation. We also discuss some of the emerging science needs to identify the appropriate scale and spatial arrangement of rehabilitation to maximise river ecosystem health outcomes and, at the same time, derive other benefits downstream. 3. An objective process was used to identify potential indicators of stream ecosystem health and then test these across a known catchment land-use disturbance gradient. From the 75 indicators initially tested, 22 from five indicator groups (water quality, ecosystem metabolism, nutrient cycling, invertebrates and fish) responded strongly to the disturbance gradient, and 16 were subsequently recommended for inclusion in the monitoring programme. The freshwater monitoring programme was implemented in 2002, funded by local and State government authorities, and currently involves the assessment of over 120 sites, twice per year. This information, together with data from a similar programme on the region's estuarine and coastal marine waters, forms the basis of an annual report card that is presented in a public ceremony to local politicians and the broader community. 4. Several key lessons from the SEQ Healthy Waterways Programme are likely to be transferable to other regional programmes aimed at improving aquatic ecosystem health, including the importance of a shared common vision, the involvement of committed individuals, a cooperative approach, the need for defensible science and effective communication. 5. Thematic implications: this study highlights the use of conceptual models and objective testing of potential indicators against a known disturbance gradient to develop a freshwater ecosystem health monitoring programme that can diagnose the probable causes of degradation from multiple stressors and identify the appropriate spatial scale for rehabilitation or protection. This approach can lead to more targeted management investments in catchment protection and rehabilitation, greater public confidence that limited funds are being well spent and better outcomes for stream and river ecosystem health.

Whole-genome multiparametric screening to identify modulators of epithelial-to-mesenchymal transition

Metastasis accounts for the poor prognosis of the majority of solid tumors. The phenotypic transition of nonmotile epithelial tumor cells to migratory and invasive “mesenchymal” cells (epithelial-to-mesenchymal transition [EMT]) enables the transit of cancer cells from the primary tumor to distant sites. There is no single marker of EMT; rather, multiple measures are required to define cell state. Thus, the multiparametric capability of high-content screening is ideally suited for the comprehensive analysis of EMT regulators. The aim of this study was to generate a platform to systematically identify functional modulators of tumor cell plasticity using the bladder cancer cell line TSU-Pr1-B1 as a model system. A platform enabling the quantification of key EMT characteristics, cell morphology and mesenchymal intermediate filament vimentin, was developed using the fluorescent whole-cell-tracking reagent CMFDA and a fluorescent promoter reporter construct, respectively. The functional effect of genome-wide modulation of protein-coding genes and miRNAs coupled with those of a collection of small-molecule kinase inhibitors on EMT was assessed using the Target Activation Bioapplication integrated in the Cellomics ArrayScan platform. Data from each of the three screens were integrated to identify a cohort of targets that were subsequently examined in a validation assay using siRNA duplexes. Identification of established regulators of EMT supports the utility of this screening approach and indicated capacity to identify novel regulators of this plasticity program. Pathway analysis coupled with interrogation of cancer-related expression profile databases and other EMT-related screens provided key evidence to prioritize further experimental investigation into the molecular regulators of EMT in cancer cells.

Some comparison on whole-proteome phylogeny of large dsDNA viruses based on dynamical language approach and feature frequency profiles method

There has been a growing interest in alignment-free methods for phylogenetic analysis using complete genome data. Among them, CVTree method, feature frequency profiles method and dynamical language approach were used to investigate the whole-proteome phylogeny of large dsDNA viruses. Using the data set of large dsDNA viruses from Gao and Qi (BMC Evol. Biol. 2007), the phylogenetic results based on the CVTree method and the dynamical language approach were compared in Yu et al. (BMC Evol. Biol. 2010). In this paper, we first apply dynamical language approach to the data set of large dsDNA viruses from Wu et al. (Proc. Natl. Acad. Sci. USA 2009) and compare our phylogenetic results with those based on the feature frequency profiles method. Then we construct the whole-proteome phylogeny of the larger dataset combining the above two data sets. According to the report of The International Committee on the Taxonomy of Viruses (ICTV), the trees from our analyses are in good agreement to the latest classification of large dsDNA viruses.