Topical Area: Aging and Chronic Disease
Chronic inflammation is a hallmark of skeletal muscle (SkM) with advancing age and is thought to augment age-related SkM atrophy (sarcopenia). Arginine, a conditionally essential amino acid, is necessary for protein synthesis. Intriguingly, arginine has been identified as being important for attenuating nuclear factor kappa B (NF-κB) activity in non-muscle cells. Our objective was to determine the age-related effects of arginine and inflammation on markers of protein synthesis and breakdown in human myotubes.
Methods : Muscle progenitor cells (MPCs) were obtained from SkM biopsy tissue of young (YNG, n= 5) and old (OLD, n = 5) women. MPCs were differentiated for 5 days (myotubes) followed by treatment with a pro-inflammatory cytokine, TNFα, for 6h, 24h, 48h, or 96h. Inflammatory activity and arginine uptake were determined using immunoblotting and radioactive isotopes. Protein synthesis was assessed with puromycin incorporation. MURF1 and MAFBX1 gene expression were used as indicators of proteolysis.
Prior to TNFα treatment, OLD (vs. YNG) myotubes had greater arginine uptake (p = 0.02), but there were no age-related differences in basal p65 NF-κB activity, puromycin incorporation, or MAFBX1 and MURF1 expression (p > 0.05). Treatment with TNFα induced NF-κB activity in both YNG and OLD myotubes at 6 and 24h of treatment (p < 0.0001), with no additional increase in activity from 24-96h. CAT2, an arginine transporter, was transiently induced at 6h of TNFα treatment (p < 0.01), with no age-related difference in response (p > 0.05). 96h of TNFα increased arginine uptake in OLD (p < 0.05), but not YNG myotubes (p >0.05). 6h TNFα had no effect (p > 0.05), but 96h of TNFα reduced puromycin incorporation in both YNG and OLD myotubes (p < 0.05). Compared to no treatment control, 96h TNFα unexpectedly decreased MAFBX1 expression in both YNG and OLD myotubes (p < 0.001). Intriguingly, in the presence of TNFα, arginine increased MURF1 expression in YNG and OLD myotubes compared to no treatment control (p < 0.05).
Conclusions : Unexpectedly, arginine amplified markers of proteolysis, and did not affect protein synthesis. Future studies will investigate regulatory transcription factors involved in protein breakdown and a crude marker of protein balance (e.g. myotube diameter).
Funding Sources : President’s Council for Cornell Women