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Long-chain acylcarnitines activate cell stress and myokine release in C2C12 myotubes: calcium-dependent and -independent effects

McCoin, CS;Knotts, TA;Ono-Moore, KD;Oort, PJ;Adams, SH;

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 M, increased IL-6 production 4.1-, 14.9-, and 31.4-fold over vehicle at 25, 50, and 100 M. Additionally, l-C16 carnitine activated c-Jun NH2-terminal kinase, extracellular signal-regulated kinase, and p38 mitogen-activated protein kinase between 2.5- and 11-fold and induced cell injury and death within 6 h with modest activation of the apoptotic caspase-3 protein. l-C16 carnitine rapidly increased intracellular calcium, most clearly by 10 M, 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 amidosulfobetaine-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.