Nutrient signalling in the regulation of human muscle protein synthesis

The mammalian target of rapamycin (mTOR) and AMP-activated protein kinase (AMPK) are important nutrient- and energy-sensing and signalling proteins in skeletal muscle. AMPK activation decreases muscle protein synthesis by inhibiting mTOR signalling to regulatory proteins associated with translation initiation and elongation. On the other hand, essential amino acids (leucine in particular) and insulin stimulate mTOR signalling and protein synthesis. We hypothesized that anabolic nutrients would be sensed by both AMPK and mTOR, resulting in an acute and potent stimulation of human skeletal muscle protein synthesis via enhanced translation initiation and elongation.
We measured muscle protein synthesis and mTOR-associated upstream and downstream signalling proteins in young male subjects ( n = 14) using stable isotopic and immunoblotting techniques. Following a first muscle biopsy, subjects in the 'Nutrition' group ingested a leucine-enriched essential amino acid–carbohydrate mixture (EAC). Subjects in the Control group did not consume nutrients. A second biopsy was obtained 1 h later. Ingestion of EAC significantly increased muscle protein synthesis, modestly reduced AMPK phosphorylation, and increased Akt/PKB (protein kinase B) and mTOR phosphorylation ( P < 0.05). mTOR signalling to its downstream effectors (S6 kinase 1 (S6K1) and 4E-binding protein 1 (4E-BP1) phosphorylation status) was also increased ( P < 0.05). In addition, eukaryotic elongation factor 2 (eEF2) phosphorylation was significantly reduced ( P < 0.05). Protein synthesis and cell signalling (phosphorylation status) was unchanged in the control group ( P > 0.05).
We conclude that anabolic nutrients alter the phosphorylation status of both AMPK- and mTOR-associated signalling proteins in human muscle, in association with an increase in protein synthesis not only via enhanced translation initiation but also through signalling promoting translation elongation.     

Co-detection and discrimination of six human herpesviruses by multiplex PCR-ELAHA.

BACKGROUND: Herpesviruses are a significant cause of human morbidity. Traditional approaches to the identification of these viruses require infectious or at least antigenic virus. Multiplex PCR (mPCR) is capable of simultaneously amplifying a range of targets from a single preparation of nucleic acids and when combined with a suitable detection assay, it is capable of discriminating each of the amplicons. OBJECTIVES: Several methods have been described in the literature, however, they lack one or more significant design features required to suitably control a routinely applied nucleic acid amplification assay. We aimed to design a multiplex herpesvirus PCR that could co-amplify eight human herpesvirus targets plus an internal control (IC) molecule in a single tube. STUDY DESIGN: Primers were designed to target the DNA polymerase genes of each of the human herpesviruses. Synthetic controls were developed to act as templates for the evaluation of assay sensitivity and specificity and for development of an in-house competitive quantitative PCR. Amplicon was discriminated using a simplified enzyme linked amplicon hybridisation assay (ELAHA). RESULTS AND CONCLUSIONS: For routine diagnostic use we reduced the number of herpesviral targets from 8 to 6 in order to maintain adequate clinical sensitivity. The ELAHA proved more sensitive than agarose gel electrophoresis. Additionally, 36 cytomegalovirus positive patients were examined with an in-house quantitative PCR-ELAHA which was developed to confirm that that the mPCR's co-detection limit of 10(2) copy of synthetic template per millilitre was relevant for use in detecting virus from clinical samples. The mPCR-ELAHA was then applied to the screening of 174 patient specimens resulting in a specificity of 98% and a sensitivity of 93%. This preliminary study demonstrated that the mPCR-ELAHA was a complete approach to the detection of herpesviruses from a range of clinical samples and disease states.     

Hypotonicity stimulates translocation of ICln in neonatal rat cardiac myocytes

 Cell volume expansion stimulates the efflux of solutes, including the amino acid taurine, to accomplish a regulatory volume decrease (RVD). One protein that may play a role in taurine efflux is the cytosolic protein ICln. In rat neonatal cardiac myocytes under isotonic conditions, ICln is found predominantly (greater than 90%) in the cytosol. However, after cell volume expansion by exposure to hypotonic medium, ICln rapidly translocates to the particulate fraction (the Triton X-114-insoluble fraction). After 2 min in hypotonic medium the percentage of ICln in the particulate fraction increases to 30%, 46% at 5 min, 40% at 10 min, and 25% at 30 min. The time course of this response is similar to that of hypotonicity-stimulated taurine efflux. Hypotonicity-stimulated taurine efflux as well as ICln translocation parallel the reduction in medium osmolarity. As osmolarity decreases, taurine efflux and ICln movement increase. The movement of ICln from the particulate back to the cytosolic fraction is accelerated when volume-expanded cells are returned to isotonic medium. When ICln is analyzed under non-denaturing conditions, a dimer is detected in the particulate fraction of volume-expanded cells, along with the monomer. This dimer is not detected in the cytosol. Treatment of the particulate fraction from volume-expanded cells with the lyotropic agent KSCN caused release of ICln but not Na-K-ATPase into the soluble fraction, indicating that translocated ICln associates with membranes in the particulate fraction rather than inserting into them. Received: 31 October 1997 / Received after revision and accepted: 23 March 1998     

Delay between pregnancy confirmation and sickle cell and [corrected] thalassaemia screening: a population-based cohort study.

BACKGROUND: Antenatal sickle cell and thalassaemia screening sometimes occurs too late to allow couples a choice regarding termination of affected fetuses. The target gestational age for offering the test in the UK is 10 weeks. AIM: To describe the proportion of women screened before 70 days' (10 weeks') gestation and the delay between pregnancy confirmation in primary care and antenatal sickle cell and thalassaemia screening. DESIGN OF STUDY: Cohort study of reported pregnancies. SETTING: Twenty-five general practices in two UK inner-city primary care trusts offering universal screening. METHOD: Anonymised data on all pregnancies reported to participating general practices was collected for a minimum of 6 months. RESULTS: There were 1441 eligible women intending to proceed with their pregnancies, whose carrier status was not known. The median (interquartile range [IQR]) gestational age at pregnancy confirmation was 7.6 weeks (6.0-10.7 weeks) and 74% presented before 10 weeks. The median gestational age at screening was 15.3 weeks (IQR = 12.6-18.0 weeks), with only 4.4% being screened before 10 weeks. The median delay between pregnancy confirmation and screening was 6.9 weeks (4.7-9.3 weeks) After allowing for practice level variation, there was no association between delay times and maternal age, parity, and ethnic group. CONCLUSION: About 74% of women consulted for pregnancy before 10 weeks' gestation but fewer than 5% of women were screened before the target time of 10 weeks. Reducing the considerable delay between pregnancy confirmation in primary care and antenatal sickle cell and thalassaemia screening requires methods of organising and delivering antenatal care that facilitate earlier screening to be developed and evaluated.     

Physical activity and motor skill outcomes of a 10-week intervention for children with intellectual and developmental disabilities ages 4–13: A pilot study

BackgroundChildren with intellectual and developmental disabilities (IDD) often experience increased barriers to engaging in physical activity (PA) which can stem from lack of gross motor function (GMF) development. Intervening on GMF at an early age can create better opportunities for children with IDD to engage in regular PA. In turn, increased PA can improve health outcomes and increase social skills.ObjectiveThe primary objective of this pilot study was to explore the effectiveness of a community-based GMF-focused PA intervention for improving overall motor skills and PA for children with IDD.MethodsAll study participants (n = 24) engaged in 10 weeks of programming for 1 h each week. A convenience sample was utilized.ResultsResults indicated no statistically significant changes pre to post for motor skill scores. However, a visual analysis of mean changes showed a consistent pattern of increased scores from pre to post on most skills. Additionally, we found that a change in participant locomotor skills significantly predicted change in Moderate to Vigorous Physical Activity (MVPA), F (1,11) = 5.16, Adj R² = .26, p = .04.ConclusionsThese results suggest individualized attention on GMF may help to increase motor skills for children with IDD. This study adds to the small but growing amount of research examining the efficacy of community based adapted PA interventions. Further, study results should support continued exploration of effective approaches to address the motor delays experienced by children with IDD.     

The expectations and realities of nutrigenomic testing in australia: A qualitative study

BackgroundConsumer genomic testing for nutrition and wellness, (nutritional genomics), is becoming increasingly popular. Concurrently, health‐care practitioners (HPs) working in private practice (including doctors interested in integrative medicine, private genetic counsellors, pharmacists, dieticians, naturopaths and nutritionists) are involved as test facilitators or interpreters.ObjectiveTo explore Australian consumers’ and HPs’ experiences with nutrigenomic testing.MethodSemi‐structured in‐depth interviews were conducted using predominantly purposive sampling. The two data sets were analysed individually, then combined, using a constant comparative, thematic approach.ResultsOverall, 45 interviews were conducted with consumers (n = 18) and HPs (n = 27). Many of the consumer interviewees experienced chronic ill‐health. Nutrigenomic testing was perceived as empowering and a source of hope for answers. While most made changes to their diet/supplements post‐test, self‐reported health improvements were small. A positive relationship with their HP appeared to minimize disappointment. HPs’ adoption and views of nutrigenomic testing varied. Those enthusiastic about testing saw the possibilities it could offer. However, many felt nutrigenomic testing was not the only ‘tool’ to utilize when offering health care.DiscussionThis research highlights the important role HPs play in consumers’ experiences of nutrigenomics. The varied practice suggests relevant HPs require upskilling in this area to at least support their patients/clients, even if nutrigenomic testing is not part of their practice.Patient or public contributionAdvisory group included patient/public group representatives who informed study design; focus group participants gave feedback on the survey from which consumer interviewees were sourced. This informed the HP data set design. Interviewees from HP data set assisted with snowball sampling.