Proceedings of The Physiological Society

University College Dublin (2009) Proc Physiol Soc 15, PC21

Poster Communications

Inhibition of skeletal muscle differentiation by tumour necrosis factor-α is reversed by the omega-3 polyunsaturated fatty acid eicosapentaenoic acid: a mechanism associated with inhibition of nuclear factor-κB and upregulation of peroxisome proliferator activated receptor γ.

P. Magee1, S. Pearson2, J. T. Allen1

1. Biomedical Sciences Research Institute, University of Salford, Salford, United Kingdom. 2. Centre for Rehabilitation & Human Performance Research, Institute for Health & Social Care Research, University of Salford, Salford, United Kingdom.

Eicosapentaenoic acid (EPA) is an omega-3 polyunsaturated fatty acid with anti-inflammatory and anti-cachetic (1, 2) properties which we reported to be protective against the damaging effects of TNF-α during skeletal muscle differentiation (3). Inflammatory cytokines such as TNF-α may contribute to muscle wasting through inhibition of myogenic differentiation via a nuclear factor-κB (NF-κB)-dependent pathway (4). Thus, we hypothesised that EPA may exert its actions downstream of TNF-α through NF-κB-mediated effects on gene transcription. C2C12 (or C2C12 stably transfected with an NF-κB reporter construct) myoblasts were differentiated by culture in growth medium containing 2% horse serum. EPA (50μM) was added at the start of differentiation and myotube formation allowed to progress for up to 48 hours in the presence or absence of TNF-α (20ng/ml). At various time-points, whole cell lysates were prepared for measurement of NF-κB activation by luminescence. In parallel, total RNA was extracted and cDNA synthesised for quantitative realtime PCR transcriptional analysis of peroxisome proliferator activated receptor (PPARγ), a downstream target for TNF-α. In response to TNF-α treatment, NF-κB activation was significantly (p<0.05) enhanced. However, EPA treatment significantly (p<0.05) attenuated this TNF-α-mediated activation of NF-κB at all time-points up to 24 hours after treatment. Furthermore, whereas TNF-α treatment alone downregulated PPARγ expression, normalised against β-actin, by 2-fold (p<0.05), EPA co-treatment reversed this inhibition, increasing PPARγ expression by 2-fold (p<0.05) at 12 hours. In summary, activation of NF-κB by TNF-α inhibits skeletal muscle differentiation but is blocked by administration of EPA and this is associated with upregulation of PPARγ expression. These data support a protective role for EPA, particularly where pathologically high levels of TNF-α may be present and support further work to elucidate its mechanisms of action.

Where applicable, experiments conform with Society ethical requirements