Mimicking exercise in vitro: effects of myotube contractions and mechanical stretch on omics.

The number of studies using skeletal muscle (SkM) cell culture models to study exercise in vitro are rapidly expanding. Progressively, more comprehensive analysis methods, such as different omics approaches including transcriptomics, proteomics, and metabolomics have been used to examine the intra- and extracellular molecular responses to exercise mimicking stimuli in cultured myotubes. Among other techniques, exercise-like electrical pulse stimulation (EL-EPS) and mechanical stretch of SkM cells are the two most commonly used methods to mimic exercise in vitro. In this mini-review, we focus on these two approaches and their effects on the omics of myotubes and/or cell culture media. Furthermore, besides traditional two-dimensional (2-D) methods, the use of three-dimensional (3-D) SkM approaches are increasing in the field of in vitro exercise mimicry. Our aim with this mini-review is to provide the reader with an up-to-date overview of the 2-D and 3-D models and the use of omics approaches to study the molecular response to exercise in vitro.

PGC-1α alternative promoter (Exon 1b) controls augmentation of total PGC-1α gene expression in response to cold water immersion and low glycogen availability.

This investigation sought to determine whether post-exercise cold water immersion and low glycogen availability, separately and in combination, would preferentially activate either the Exon 1a or Exon 1b Peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α) promoter. Through a reanalysis of sample design, we identified that the systemic cold-induced augmentation of total PGC-1α gene expression observed previously (Allan et al. in J Appl Physiol 123(2):451-459, 2017) was largely a result of increased expression from the alternative promoter (Exon 1b), rather than canonical promoter (Exon 1a). Low glycogen availability in combination with local cooling of the muscle (Allan et al. in Physiol Rep 7(11):e14082, 2019) demonstrated that PGC-1α alternative promoter (Exon 1b) expression continued to rise at 3 h post-exercise in all conditions; whilst, expression from the canonical promoter (Exon 1a) decreased between the same time points (post-exercise-3 h post-exercise). Importantly, this increase in PGC-1α Exon 1b expression was reduced compared to the response of low glycogen or cold water immersion alone, suggesting that the combination of prior low glycogen and CWI post-exercise impaired the response in gene expression versus these conditions individually. Data herein emphasise the influence of post-exercise cooling and low glycogen availability on Exon-specific control of total PGC-1 α gene expression and highlight the need for future research to assess Exon-specific regulation of PGC-1α.

Acute mechanical overload increases IGF-I and MMP-9 mRNA in 3D tissue-engineered skeletal muscle.

Skeletal muscle (SkM) is a tissue that responds to mechanical load following both physiological (exercise) or pathophysiological (bed rest) conditions. The heterogeneity of human samples and the experimental and ethical limitations of animal studies provide a rationale for the study of SkM plasticity in vitro. Many current in vitro approaches of mechanical loading of SkM disregard the three-dimensional (3D) structure in vivo. Tissue engineered 3D SkM, that displays highly aligned and differentiated myotubes, was used to investigate mechano-regulated gene transcription of genes implicated in hypertrophy/atrophy. Static loading (STL) and ramp loading (RPL) at 10 % strain for 60 min were used as mechano-stimulation with constructs sampled immediately for RNA extraction. STL increased IGF-I mRNA compared to both RPL and CON (control, p = 0.003 and 0.011 respectively) whilst MMP-9 mRNA increased in STL and RPL compared to CON (both p < 0.05). IGFBP-2 mRNA was differentially regulated in RPL and STL compared to CON (p = 0.057), whilst a reduction in IGFBP-5 mRNA was found for STL and RPL compared to CON (both p < 0.05). There was no effect in the expression of putative atrophic genes, myostatin, MuRF-1 and MAFBx (all p > 0.05). These data demonstrate a transcriptional signature associated with SkM hypertrophy within a tissue-engineered model that more greatly recapitulates the in vivo SkM structure compared previously published studies.

Laparoscopic fellowship training can deliver a competent laparoscopic surgeon and trainer.

AIM: The study investigated whether experience gained during a UK laparoscopic colorectal fellowship enabled the fellow subsequently to train consultant colleagues in laparoscopic surgery. METHOD: In one unit a newly appointed post-laparoscopic fellowship consultant (PFC) mentored his other two colleagues. Prospectively collected data regarding surgical outcome were compared with those of the year preceding the PFC appointment. RESULTS: In the preceding year 18.5% of 260 resections were attempted laparoscopically. This increased to 92.6% (of 270) in the year after (P < 0.0001). Respective conversion rates were 4.2% and 8.4% (P = 0.5524). In the first 6 months after PFC appointment, mentored consultants performed 23 supervised cases. In the second 6 months they carried out 58 procedures independently and trainees performed 38 supervised cases. There was no significant difference in anastomotic leakage and readmission and 30-day mortality rates between the pre- and post-PFC periods. CONCLUSION: A laparoscopic fellowship enables the PFC to mentor consultant colleagues safely and effectively.

Barriers, Attitudes, and Influences Towards Dietary Intake Amongst Elite Rugby Union Players.

Background: Dietary intakes can impact an athletes health and performance. Although evidence exists about what an athlete should eat, an athlete's nutritional intake is influenced by many factors. The limited research available suggests the main barriers preventing optimal nutritional intakes reported by athletes are lack of time, food accessibility, poor cooking skills, costs, taste, and time spent in "off-season." Although these factors have been shown to influence dietary intake they remain relatively unexplored in Rugby Union. This study aimed to describe the nutritional influences on dietary intake amongst Rugby Union player's. Methods: This was a qualitative study utilising in person individual interviews with all participants. Participants were Rugby Union players (n = 30) for either a Super Rugby franchise or one of their development squads in New Zealand. Participant's undertook recorded face to face interviews, which were later transcribed. A thematic approach was used to code the transcripts by the primary coder and the themes were subsequently evaluated by the research team. Results: Childhood upbringing, organisational skills, time and food security also emerged as barriers. Body composition and sport nutrition knowledge emerged as both barriers and enablers to nutritional intake. Influence on performance was an enabler to optimal dietary intake. Fully professional rugby players have access to dietitians, whereas development and semi-professional rugby players only have limited if any access, and they were more likely to seek nutritional information via social media. Conclusion: This study suggests a need for greater nutrition education at developmental levels with an emphasis on affordable food choices, meal planning and skills for interpreting online nutrition information.

DNA methylation across the genome in aged human skeletal muscle tissue and muscle-derived cells: the role of HOX genes and physical activity.

Skeletal muscle tissue demonstrates global hypermethylation with age. However, methylome changes across the time-course of differentiation in aged human muscle derived cells, and larger coverage arrays in aged muscle tissue have not been undertaken. Using 850K DNA methylation arrays we compared the methylomes of young (27 ± 4.4 years) and aged (83 ± 4 years) human skeletal muscle and that of young/aged heterogenous muscle-derived human primary cells (HDMCs) over several time points of differentiation (0, 72 h, 7, 10 days). Aged muscle tissue was hypermethylated compared with young tissue, enriched for; pathways-in-cancer (including; focal adhesion, MAPK signaling, PI3K-Akt-mTOR signaling, p53 signaling, Jak-STAT signaling, TGF-beta and notch signaling), rap1-signaling, axon-guidance and hippo-signalling. Aged cells also demonstrated a hypermethylated profile in pathways; axon-guidance, adherens-junction and calcium-signaling, particularly at later timepoints of myotube formation, corresponding with reduced morphological differentiation and reductions in MyoD/Myogenin gene expression compared with young cells. While young cells showed little alterations in DNA methylation during differentiation, aged cells demonstrated extensive and significantly altered DNA methylation, particularly at 7 days of differentiation and most notably in focal adhesion and PI3K-AKT signalling pathways. While the methylomes were vastly different between muscle tissue and HDMCs, we identified a small number of CpG sites showing a hypermethylated state with age, in both muscle tissue and cells on genes KIF15, DYRK2, FHL2, MRPS33, ABCA17P. Most notably, differential methylation analysis of chromosomal regions identified three locations containing enrichment of 6-8 CpGs in the HOX family of genes altered with age. With HOXD10, HOXD9, HOXD8, HOXA3, HOXC9, HOXB1, HOXB3, HOXC-AS2 and HOXC10 all hypermethylated in aged tissue. In aged cells the same HOX genes (and additionally HOXC-AS3) displayed the most variable methylation at 7 days of differentiation versus young cells, with HOXD8, HOXC9, HOXB1 and HOXC-AS3 hypermethylated and HOXC10 and HOXC-AS2 hypomethylated. We also determined that there was an inverse relationship between DNA methylation and gene expression for HOXB1, HOXA3 and HOXC-AS3. Finally, increased physical activity in young adults was associated with oppositely regulating HOXB1 and HOXA3 methylation compared with age. Overall, we demonstrate that a considerable number of HOX genes are differentially epigenetically regulated in aged human skeletal muscle and HDMCs and increased physical activity may help prevent age-related epigenetic changes in these HOX genes.

Methylome of human skeletal muscle after acute & chronic resistance exercise training, detraining & retraining.

DNA methylation is an important epigenetic modification that can regulate gene expression following environmental encounters without changes to the genetic code. Using Infinium MethylationEPIC BeadChip Arrays (850,000 CpG sites) we analysed for the first time, DNA isolated from untrained human skeletal muscle biopsies (vastus lateralis) at baseline (rest) and immediately following an acute (single) bout of resistance exercise. In the same participants, we also analysed the methylome following a period of muscle growth (hypertrophy) evoked via chronic (repeated bouts-3 sessions/wk) resistance exercise (RE) (training) over 7-weeks, followed by complete exercise cessation for 7-weeks returning muscle back to baseline levels (detraining), and finally followed by a subsequent 7-week period of RE-induced hypertrophy (retraining). These valuable methylome data sets described in the present manuscript and deposited in an open-access repository can now be shared and re-used to enable the identification of epigenetically regulated genes/networks that are modified after acute anabolic stimuli and hypertrophy, and further investigate the phenomenon of epigenetic memory in skeletal muscle.

Screening for occult gastrointestinal bleeding in hospital patients.

Stools have been tested for occult gastrointestinal bleeding in 278 outpatients and 170 hospital inpatients using the Haemoccult and Haemastix methods. Seventeen outpatients (6.1%) and 42 inpatients (24%) were positive with the Haemoccult technique. Thirty-three outpatients (11.9%) and 93 inpatients (54.7%) were positive with the Haemastix test. Following investigation of the Haemoccult-positive patients, only 2 cases (3.4%) were considered false positive. However, the false positive rate with Haemastix was 22.9% which is unacceptable in a screening test. Haemoccult may be useful as a screening test for asymptomatic general practice patients, but a test of greater sensitivity is needed for hospital patients.

The impact on work-related stress of mental health teams following team-based learning on clinical risk management.

Risk management is viewed as a systematic process based on multiprofessional and multi-agency decision-making. A learning pack was developed as part of a team-based learning project aiming to encourage and develop collaborative working practice. This brought different professionals and agencies working in mental health together to learn. There is little doubt that mental health practice is a source of stress for practitioners. Apart from the stress associated with managing 'risky' situations, risk management is also a relatively new concept. This can increase stress around ability to cope, both on an individual practitioner level and in teams. This article reports the impact that the learning pack had on team members' stress, specifically work-related stress. A range of scales were used to measure change in stress and results demonstrated reduced work-related pressure in a number of areas following the learning. The implications for team learning in relation to clinical risk management are discussed in light of the findings.

An approach to consensus building using the Delphi technique: developing a learning resource in mental health.

Health service research is best served through the collaboration of researchers and practitioners. This study utilizes a multi-method approach in developing a learning resource in clinical risk management in mental health. Phase one of the project is reported here; using a Delphi Technique to build the learning resource. Using the Delphi enabled a group of 'experts' to explore a complex issue and move towards consensus. In addition, the Delphi technique enabled the researchers to repeatedly access a dispersed group of 'experts' in an economic way. A three round Delphi was undertaken and results were analyzed (medians, means, standard deviations and thematic content analysis) to facilitate consensus building from a range of perspectives. The analytical model chosen is discussed in the context of prior use of the Delphi as a consensus building technique. The researchers conclude that analysis using medians and means to measure the level of agreement/disagreement with the content and structure of the learning pack, standard deviations to measure 'tightening' of the group view and Thematic Content Analysis of qualitative data was an effective approach. They argue that in the education and health fields, group views are particularly important and that this is a central tenet of the Delphi.

Anaesthesia in a child with Patau's syndrome.

A 5-year-old girl who had Patau's syndrome associated with double outlet right ventricle and pulmonary stenosis required general anaesthesia for the formation of a modified Blalock-Taussig shunt.