Tissue engineered skeletal muscle allows investigation of the cellular and molecular mechanisms that regulate skeletal muscle pathology. The fabricated model must resemble characteristics of in vivo tissue and incorporate cost-effective and high content primary human tissue. Current models are limited by low throughput due to the complexities associated with recruiting tissue donors, donor specific variations, as well as cellular senescence associated with passaging. This research presents a method using fused deposition modelling (FDM) and laser sintering (LS) 3D printing to generate reproducible and scalable tissue engineered primary human muscle, possessing aligned mature myotubes reminiscent of in vivo tissue. Many existing models are bespoke causing variability when translated between laboratories. To this end, a scalable model has been developed (25-500 μL construct volumes) allowing fabrication of mature primary human skeletal muscle. This research provides a strategy to overcome limited biopsy cell numbers, enabling high throughput screening of functional human tissue.

Patients with chronic kidney disease (CKD) exhibit reduced exercise capacity, poor physical function and symptoms of fatigue. The mechanisms that contribute to this are not clearly defined but may involve reductions in mitochondrial function, mass and biogenesis. Here we report on the effect of non‐dialysis dependent CKD (NDD‐CKD) on mitochondrial mass and basal expression of transcription factors involved in mitochondrial biogenesis compared to a healthy control cohort (HC). In addition, we sought to investigate the effect of a 12‐week exercise‐training programme on these aspects of mitochondrial dysfunction in a NDD‐CKD cohort.For the comparison between NDD‐CKD and HC populations, skeletal muscle biopsies were collected from the vastus lateralis (VL) of n=16 non‐dialysis dependent CKD patient’s stage 3b‐5 (NDD‐CKD) and n=16 healthy controls matched for age, gender and physical activity (HC). To investigate the effect of exercise training, VL biopsies were collected from n=17 NDD‐CKD patients before and after a 12‐week exercise intervention that was comprised of aerobic exercise (AE) or a combination of aerobic exercise and resistance training (CE). Mitochondrial mass was analysed by citrate synthase activity and mitochondrial protein content by Porin expression, whilst the expression of transcription factors involved in mitochondrial biogenesis were quantified by real‐time qPCR. NDD‐CKD patients exhibited a significant reduction in mitochondrial mass when compared to HC, coupled to a reduction in PGC‐1α, NRF‐1, Nrf2, TFam, mfn2 and SOD1/2 gene expression. 12‐weeks of exercise training resulted in a significant increase in PGC‐1α expression in both groups, with no further changes seen across indicators of mitochondrial biogenesis. No significant changes in mitochondrial mass were observed in response to either exercise programme. NDD‐CKD patients exhibit reduced skeletal muscle mitochondrial mass and gene expression of transcription factors involved in mitochondrial biogenesis compared to HC. These reductions were not restored following 12‐weeks of exercise training implying exercise resistance in this cohort. The reasons for this lack of improvement are currently unknown and require further investigation, as reversing the dysregulation of these processes in NDD‐CKD may provide a therapeutic opportunity to improve muscle fatigue and dysfunction in this population.

Single bouts of exercise do not cause compensatory changes in appetite, food intake or appetite regulatory hormones on the day that exercise is performed. It remains possible that such changes occur over an extended period or in response to a higher level of energy expenditure. This study sought to test this possibility by examining appetite, food intake and appetite regulatory hormones (acylated ghrelin, total peptide-YY, leptin and insulin) over two days, with acute bouts of exercise performed on each morning. Within a controlled laboratory setting, 15 healthy males completed two, 2-day long (09:00-16:00) experimental trials (exercise and control) in a randomised order. On the exercise trial participants performed 60 min of continuous moderate-high intensity treadmill running (day one: 70.1 ± 2.5% VO2peak, day two: 70.0 ± 3.2% VO2max (mean ± SD)) at the beginning of days one and two. Across each day appetite perceptions were assessed using visual analogue scales and appetite regulatory hormones were measured from venous blood samples. Ad libitum energy and macronutrient intakes were determined from meals provided two and six hours into each day and from a snack bag provided in-between trial days. Exercise elicited a high level of energy expenditure (total = 7566 ± 635 kJ across the two days) but did not produce compensatory changes in appetite or energy intake over two days (control: 29,217 ± 4006 kJ; exercise: 28,532 ± 3899 kJ, P > 0.050). Two-way repeated measures ANOVA did not reveal any main effects for acylated ghrelin or leptin (all P > 0.050). However a significant main effect of trial (P = 0.029) for PYY indicated higher concentrations on the exercise vs. control trial. These findings suggest that across a two day period, high volume exercise does not stimulate compensatory appetite regulatory changes.

Resolution of inflammation is now known to be an active process which in part is instigated and controlled by specialized pro‐resolving lipid mediators (SPM's) derived from dietary omega‐3 fatty acids. Resolvin E1 (R_vE₁) 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, R_vE₁ attenuated mRNA levels of both interlukin‐6 and monocyte chemoattractant protein‐1 whilst having no effect on tumor 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. R_vE₁ instigated beneficial morphological changes through the prevention of LPS induced skeletal muscle atrophy, in tandem with attenuation of the LPS induced reduction in contractile force in tissue engineered skeletal muscle. These findings demonstrate, in our model of endotoxin induced inflammation in skeletal muscle, that R_vE₁ has pro‐resolving properties in this cell type. Our data provides rationale for further investigation into the mechanistic action of R_vE₁ in skeletal muscle, with the vision of having potential benefits for the prevention/resolution of in‐vivo skeletal muscle atrophy.

Skeletal muscle wasting is a common complication of chronic kidney disease (CKD), characterized by the loss of muscle mass, strength and function, which significantly increases the risk of morbidity and mortality in this population. Numerous complications associated with declining renal function and lifestyle activate catabolic pathways and impair muscle regeneration, resulting in substantial protein wasting. Evidence suggests that increasing skeletal muscle mass improves outcomes in CKD, making this a clinically important research focus. Despite extensive research, the pathogenesis of skeletal muscle wasting is not completely understood. It is widely recognized that microRNAs (miRNAs), a family of short non-coding RNAs, are pivotal in the regulation of skeletal muscle homoeostasis, with significant roles in regulating muscle growth, regeneration and metabolism. The abnormal expression of miRNAs in skeletal muscle during disease has been well described in cellular and animal models of muscle atrophy, and in recent years, the involvement of miRNAs in the regulation of muscle atrophy in CKD has been demonstrated. As this exciting field evolves, there is emerging evidence for the involvement of miRNAs in a beneficial crosstalk system between skeletal muscle and other organs that may potentially limit the progression of CKD. In this article, we describe the pathophysiological mechanisms of muscle wasting and explore the contribution of miRNAs to the development of muscle wasting in CKD. We also discuss advances in our understanding of miRNAs in muscle–organ crosstalk and summarize miRNA-based therapeutics currently in clinical trials.