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ARS Home » Pacific West Area » Davis, California » Western Human Nutrition Research Center » Obesity and Metabolism Research » Research » Publications at this Location » Publication #316375
Title: Long-chain acylcarnitines activate cell stress and myokine release in C2C12 myotubes: calcium-dependent and independent effects

Author
item McCoin, Colin
item Knotts, Trina
item Ono-Moore, Kikumi
item Oort, Pieter
item ADAMS, SEAN - Arkansas Children'S Nutrition Research Center (ACNC)

Submitted to: American Journal of Physiology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 4/6/2015
Publication Date: 4/7/2015
Publication URL: http://ajpendo.physiology.org/content/early/2015/04/01/ajpendo.00602.2014
Citation: McCoin, C.S., Knotts, T.A., Ono-Moore, K.D., Oort, P.J., Adams, S.H. 2015. Long-chain acylcarnitines activate cell stress and myokine release in C2C12 myotubes: calcium-dependent and independent effects. American Journal of Physiology. 308 (11): E990-E1000. DOI: 10.1152/ajpendo.00602.2014.

Interpretive Summary: Long-chain fatty acylcarnitines (LCAC) are naturally-occurring metabolites found in the body’s tissues and blood, and patterns of LCAC are reflective of shifts in fat metabolism. In addition to marking changes in metabolism, more recently it has become appreciated that the metabolites themselves have bioactivities in certain cells, thereby eliciting inflammation and acting to limit the activity of the blood sugar-controlling hormone insulin, at least when LCAC are modestly elevated in concentration. It is possible that with especially high concentrations, as might be seen in the inherited disorders of fatty acid metabolism, LCAC have effects to elicit cell stress and could contribute to the pathology phenotypes of persons with these disorders. If true, nutritional and other strategies to limit accumulation of LCAC could be a viable means to prevent disease outcomes. In the current studies, the impact of LCAC on inflammatory, stress and death pathways in a skeletal muscle model were evaluated. Differentiated C2C12 myotubes treated with L-C14-, C16-, C18- and C18:1-carnitine displayed dose-dependent increases in IL-6 (a secreted myokine marker of cell stress) production with a concomitant rise in markers of cell permeability and death, which was not observed for shorter chain-lengths. L-C16-carnitine, used as a representative long-chain acylcarnitine at initial extracellular concentrations = 25 µM, increased IL-6 production 4.1-, 14.9- and 31.4- fold over vehicle at 25, 50 and 100 µM. Additionally, L-C16-carnitine activated cell stress-associated proteins JNK, ERK and p38 MAP kinases between 2.5 to 11-fold, and induced cell injury and death within 6 hours with modest activation of the apoptotic caspase-3 protein. L-C16-carnitine rapidly increased intracellular calcium, most clearly by 10 µM, implicating calcium as a potential mechanism for some activities of long-chain acylcarnitines. The intracellular calcium chelator, BAPTA-AM, blunted L-C16-carnitine-mediated IL-6 production by >65%. However, BAPTA-AM did not attenuate cell permeability and death responses, indicating that these outcomes are calcium-independent. Other data suggest that the effects of high experimental concentrations of long-chain acylcarnitines are through membrane disruption. Herein, a model is proposed in which acylcarnitine cell membrane interactions take place along a spectrum of cellular concentrations encountered in physiological-to-pathophysiological conditions, thus regulating function of membrane-based systems and impacting cell biology.

Technical Abstract: Acylcarnitines, important lipid biomarkers reflective of acyl-CoA status, are metabolites that possess bioactive and inflammatory properties. This study examined the potential for long-chain acylcarnitines to activate cellular inflammatory, stress and death pathways in a skeletal muscle model. Differentiated C2C12 myotubes treated with L-C14-, C16-, C18- and C18:1-carnitine displayed dose-dependent increases in IL-6 production with a concomitant rise in markers of cell permeability and death, which was not observed for shorter chain-lengths. L-C16-carnitine, used as a representative long-chain acylcarnitine at initial extracellular concentrations = 25 µM, increased IL-6 production 4.1-, 14.9- and 31.4- fold over vehicle at 25, 50 and 100 µM. Additionally, L-C16-carnitine activated JNK, ERK and p38 MAP kinases between 2.5 to 11-fold, and induced cell injury and death within 6 hours with modest activation of the apoptotic caspase-3 protein. L-C16-carnitine rapidly increased intracellular calcium, most clearly by 10 µM, implicating calcium as a potential mechanism for some activities of long-chain acylcarnitines. The intracellular calcium chelator, BAPTA-AM, blunted L-C16-carnitine-mediated IL-6 production by >65%. However, BAPTA-AM did not attenuate cell permeability and death responses, indicating that these outcomes are calcium-independent. The 16-carbon zwitterionic compound ASB-16 qualitatively mimicked the L-C16-carnitine-associated cell stress outcomes, suggesting that the effects of high experimental concentrations of long-chain acylcarnitines are through membrane disruption. Herein, a model is proposed in which acylcarnitine cell membrane interactions take place along a spectrum of cellular concentrations encountered in physiological-to-pathophysiological conditions, thus regulating function of membrane-based systems and impacting cell biology.