Proceedings of The Physiological Society

University of Manchester (2010) Proc Physiol Soc 19, C110

Oral Communications

Inhibition of murine skeletal muscle differentiation by TNF-related weak inducer of apoptosis (TWEAK) is reversed by the omega-3 polyunsaturated fatty acid eicosapentaenoic acid (EPA).

P. Magee1, J. Allen1, S. Pearson2

1. Biomedical Sciences Research Institute, University of Salford, Manchester, United Kingdom. 2. Centre for Health, Sport and Rehabilitation Sciences Research, University of Salford, Manchester, United Kingdom.


  • Fig.1. EPA protects against the deleterious effects of TWEAK on differentiated myotubes. Myotubes differentiated for 96h were incubated for a further 72h in DM in the presence of several EPA (50

TWEAK is a member of the TNF superfamily that has been implicated in several pathological responses including inflammation, and has been shown to modulate myogenic differentiation (1). TWEAK has been suggested to exert these effects indirectly via TNF-stimulation possibly through NF-κB pathway activation (2). Eicosapentaenoic acid (EPA) is an omega-3 polyunsaturated fatty acid with anti-inflammatory and anti-cachetic properties which the authors have previously reported to be protective against the damaging effects of TNF-α during skeletal muscle differentiation (3). We hypothesised that EPA may have a similar anti-inflammatory activity against TWEAK-induced skeletal muscle damage. C2C12 myoblasts were differentiated by culture in growth medium containing 2% horse serum (DM). Cells in DM were pre-treated or co-treated with EPA (50μM) and myotube formation allowed to progress for up to 72 hours in the presence or absence of TWEAK (10ng/ml). In parallel, the same experiments were performed on fully differentiated myotubes. Myogenesis was evaluated morphologically by a myogenic index, by myotube metrics and by myosin heavy chain (MyHC) expression. In parallel, total RNA was extracted and cDNA synthesised for quantitative realtime PCR transcriptional analysis of TNF-α expression, normalised to β-actin. EPA co-treatment attenuated the inhibitory effects of TWEAK on myoblast differentiation (Fig1. Myotube images A and B), significantly increasing myogenic fusion (p<0.05) and myotube diameters (p<0.05) in comparison to TWEAK treated cultures. EPA co-treatment also ameliorated TWEAK-induced loss of MyHC from fully differentiated myotubes , again resulting in a significant increase in myotube diameter (p<0.05) and the fraction of nuclei residing in myotubes (p<0.05), compared to TWEAK-treated cultures. TWEAK upregulated TNF-α expression. However, EPA treatment reversed this, significantly decreasing TNF-α transcription levels (p<0.05), compared with TWEAK alone. In summary, EPA treatment protects against TWEAK-induced inhibition of myotube differentiation, EPA protects mature myotubes against TWEAK-induced atrophy, and EPA prevents TWEAK-induced increases in TNF expression that could account for, at least in part, the TWEAK actions. These data encourage a possible therapeutic role for EPA, particularly in inflammatory conditions where pathologically high levels of TNF-α and TWEAK may be present and support further work to elucidate its mechanisms of action.

Where applicable, experiments conform with Society ethical requirements