Preclinical musculoskeletal junction testbed: co-culture of 3D tissue engineered skeletal muscle and bone constructs [Abstract]

Preclinical musculoskeletal junction testbed: co-culture of 3D tissue engineered skeletal muscle and bone constructs [Abstract

Physiological and pathophysiological concentrations of fatty acids induce lipid droplet accumulation and impair functional performance of tissue engineered skeletal muscle

Fatty acids (FA) exert physiological and pathophysiological effects leading to changes in skeletal muscle metabolism and function, however, in vitro models to investigate these changes are limited. These experiments sought to establish the effects of physiological and pathophysiological concentrations of exogenous FA upon the function of tissue engineered skeletal muscle (TESkM). Cultured initially for 14 days, C2C12 TESkM was exposed to FA‐free bovine serum albumin alone or conjugated to a FA mixture (oleic, palmitic, linoleic, and α‐linoleic acids [OPLA] [ratio 45:30:24:1%]) at different concentrations (200 or 800 µM) for an additional 4 days. Subsequently, TESkM morphology, functional capacity, gene expression and insulin signaling were analyzed. There was a dose response increase in the number and size of lipid droplets within the TESkM (p < .05). Exposure to exogenous FA increased the messenger RNA expression of genes involved in lipid storage (perilipin 2 [p < .05]) and metabolism (pyruvate dehydrogenase lipoamide kinase isozyme 4 [p < .01]) in a dose dependent manner. TESkM force production was reduced (tetanic and single twitch) (p < .05) and increases in transcription of type I slow twitch fiber isoform, myosin heavy chain 7, were observed when cultured with 200 µM OPLA compared to control (p < .01). Four days of OPLA exposure results in lipid accumulation in TESkM which in turn results in changes in muscle function and metabolism; thus, providing insight ito the functional and mechanistic changes of TESkM in response to exogenous FA.</div

Hypoxia Impairs Muscle Function and Reduces Myotube Size in Tissue Engineered Skeletal Muscle

Contemporary tissue engineered skeletal muscle models display a high degree of physiological accuracy compared with native tissue, and therefore may be excellent platforms to understand how various pathologies affect skeletal muscle. Chronic obstructive pulmonary disease (COPD) is a lung disease which causes tissue hypoxia and is characterized by muscle fiber atrophy and impaired muscle function. In the present study we exposed engineered skeletal muscle to varying levels of oxygen (O2; 21–1%) for 24 h in order to see if a COPD like muscle phenotype could be recreated in vitro, and if so, at what degree of hypoxia this occurred. Maximal contractile force was attenuated in hypoxia compared to 21% O2; with culture at 5% and 1% O2 causing the most pronounced effects with 62% and 56% decrements in force, respectively. Furthermore at these levels of O2, myotubes within the engineered muscles displayed significant atrophy which was not seen at higher O2 levels. At the molecular level we observed increases in mRNA expression of MuRF-1 only at 1% O2 whereas MAFbx expression was elevated at 10%, 5%, and 1% O2. In addition, p70S6 kinase phosphorylation (a downstream effector of mTORC1) was reduced when engineered muscle was cultured at 1% O2, with no significant changes seen above this O2 level. Overall, these data suggest that engineered muscle exposed to O2 levels of ≤5% adapts in a manner similar to that seen in COPD patients, and thus may provide a novel model for further understanding muscle wasting associated with tissue hypoxia

Neurodevelopmental outcome at 5 years of age after general anaesthesia or awake-regional anaesthesia in infancy (GAS)trial

Background: In laboratory animals, exposure to most general anaesthetics leads to neurotoxicity manifested by neuronal cell death and abnormal behaviour and cognition. Some large human cohort studies have shown an association between general anaesthesia at a young age and subsequent neurodevelopmental deficits, but these studies are prone to bias. Others have found no evidence for an association. We aimed to establish whether general anaesthesia in early infancy affects neurodevelopmental outcomes. Methods: In this international, assessor-masked, equivalence, randomised, controlled trial conducted at 28 hospitals in Australia, Italy, the USA, the UK, Canada, the Netherlands, and New Zealand, we recruited infants of less than 60 weeks' postmenstrual age who were born at more than 26 weeks

Resolvin E1 (R v

This is the peer reviewed version of the following article: BAKER, L. ...et al., 2018. Resolvin E1 (RvE1) attenuates LPS induced inflammation and subsequent atrophy in C2C12 myotubes. Journal of Cellular Biochemistry, 119 (7), pp.6094-6103, which has been published in final form at https://doi.org/10.1002/jcb.26807. This article may be used for non-commercial purposes in accordance with Wiley Terms and Conditions for Use of Self-Archived Versions.Resolution of inflammation is now known to be an active process which in part is instigated and controlled by specialised pro-resolving lipid mediators (SPM’s) derived from dietary omega-3 fatty acids. Resolvin E1 (RvE1) is one of these SPM’s derived from the omega-3 fatty acid eicosapentaenoic acid. Using both molecular and phenotypic functional measures we report that in a model of Lipopolysaccharide (LPS) induced inflammation, RvE1 attenuated mRNA gene expression levels of both interlukin-6 and monocyte chemoattractant protein-1 whilst having no effect on tumour necrosis factor-α or Interlukin-1β in C2C12 skeletal muscle myotubes. Findings at the molecular level were transferred into similar changes in extracellular protein levels of the corresponding genes with the greatest attenuation being noted in IL-6 protein concentrations. RvE1 instigated beneficial morphological changes through the prevention of endotoxin induced skeletal muscle atrophy, thus resulting in a rescue of endotoxin force losses in tissue engineered skeletal muscle. These findings demonstrate, in our model of endotoxin induced inflammation in skeletal muscle, that RvE1 has pro-resolving properties in this cell type. Our data provides rationale for further investigation into the mechanistic action of RvE1 in skeletal muscle, with the vision of having potential benefits for the prevention/resolution of in vivo skeletal muscle atrophy

The effect of chronic high insulin exposure upon metabolic and myogenic markers in C2C12 skeletal muscle cells and myotubes

This is the peer reviewed version of the following article: TURNER, M.C. ...et al., 2018. The effect of chronic high insulin exposure upon metabolic and myogenic markers in C2C12 skeletal muscle cells and myotubes.. Journal of Cellular Biochemistry, 119 (7), pp.5686-5695, which has been published in final form at https://doi.org/10.1002/jcb.26748. This article may be used for non-commercial purposes in accordance with Wiley Terms and Conditions for Use of Self-Archived Versions.Skeletal muscle is an insulin sensitive tissue and accounts for approximately 80% of post-prandial glucose disposal. This study describes the effects of insulin, delivered for 72 hours, to skeletal muscle myoblasts during differentiation or to skeletal muscle myotubes. After chronic treatment, cultures were acutely stimulated with insulin and analysed for total and phosphorylated Akt (Ser473), mRNA expression of metabolic and myogenic markers and insulin-stimulated glucose uptake. Skeletal muscle cells differentiated in the presence of insulin chronically, reduced acute insulin stimulated phosphorylation of Akt Ser473. In addition, there was a reduction in mRNA expression of Hexokinase II (HKII), GLUT4 and PGC-1α. Insulin-stimulated glucose uptake was attenuated when cells were differentiated in the presence of insulin. In contrast, myotubes exposed to chronic insulin showed no alterations in phosphorylation of Akt Ser473. Both HKII and GLUT4 mRNA expression were reduced by chronic exposure to insulin; while PGC-1α was not different between culture conditions and was increased by acute insulin stimulation. These data suggest that there are differential responses in insulin signalling, transcription and glucose uptake of skeletal muscle cells when cultured in either the presence of insulin during differentiation or in myotube cultures

PRISM (Polarized Radiation Imaging and Spectroscopy Mission): an extended white paper

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