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Protective Effect of Unsaturated Fatty Acids on Palmitic Acid-Induced Toxicity in Skeletal Muscle Cells is not Mediated by PPARδ Activation

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Lipids

Abstract

Unsaturated free fatty acids (FFA) are able to prevent deleterious effects of saturated FFA in skeletal muscle cells although the mechanisms involved are still not completely understood. FFA act as endogenous ligands of peroxisome proliferator-activated receptors (PPAR), transcription factors regulating the expression of genes involved in lipid metabolism. The aim of this study was to determine whether activation of PPARδ, the most common PPAR subtype in skeletal muscle, plays a role in mediating the protective effect of unsaturated FFA on saturated FFA-induced damage in skeletal muscle cells and to examine an impact on mitochondrial respiration. Mouse C2C12 myotubes were treated for 24 h with different concentrations of saturated FFA (palmitic acid), unsaturated FFA (oleic, linoleic and α-linolenic acid), and their combinations. PPARδ agonist GW501516 and antagonist GSK0660 were also used. Both mono- and polyunsaturated FFA, but not GW501516, prevented palmitic acid-induced cell death. Mono- and polyunsaturated FFA proved to be effective activators of PPARδ compared to saturated palmitic acid; however, in combination with palmitic acid their effect on PPARδ activation was blocked and stayed at the levels observed for palmitic acid alone. Unsaturated FFA at moderate physiological concentrations as well as GW501516, but not palmitic acid, mildly uncoupled mitochondrial respiration. Our results indicate that although unsaturated FFA are effective activators of PPARδ, their protective effect on palmitic acid-induced toxicity is not mediated by PPARδ activation and subsequent induction of lipid regulatory genes in skeletal muscle cells. Other mechanisms, such as mitochondrial uncoupling, may underlie their effect.

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Abbreviations

FFA:

Free fatty acids

PPAR:

Peroxisome proliferator-activated receptors

PDK4:

Pyruvate dehydrogenase kinase 4

UCP3:

Uncoupling protein 3

CPT1:

Carnitine palmitoyltransferase 1

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Acknowledgments

This work was supported by the research project of Grant Agency of Charles University GAUK 661912 and grants PRVOUK-31 and UNCE 204015.

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Authors and Affiliations

  1. Department of Nutrition and Centre for Research on Diabetes, Metabolism and Nutrition, Third Faculty of Medicine, Charles University in Prague, Prague, Czech Republic

    Jana Tumova, Michal Andel & Jan Trnka

  2. Department of Sport Medicine, Third Faculty of Medicine, Charles University in Prague, Prague, Czech Republic

    Lucia Malisova

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  1. Jana Tumova
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Correspondence to Jan Trnka.

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11745_2015_4058_MOESM1_ESM.eps

The mRNA expression of PPARδ. a, b) Myotubes were treated with 1 μM PPARδ agonist GW501516 (a) or 1 μM PPARδ antagonist GSK0660 (b) for 24 h. c, d, e) Myotubes were treated with 100 μM (c), 300 μM (d) and 600 μM (e) total concentration of palmitic acid (PAM), unsaturated FFA (OLA, LNA, ALA) or with combinations: PAM + unsaturated FFA in a ratio 1:1 or PAM + 1 μM GW501516 for 24 h. The mRNA level relative to the internal control GAPDH was determined by RT-PCR analysis and fold change of expression was calculated using the ΔΔCt method. Results are presented as mean ± SEM (n = 3). * p < 0.05 compared to BSA control (C-BSA) (EPS 788 kb).

11745_2015_4058_MOESM2_ESM.eps

Mitochondrial respiration (oxygen consumption rates, OCR) in myotubes measured under different respiratory conditions. a, b) Myotubes were treated with 100 μM (c), 300 μM (d) and 600 μM (e) total concentration of palmitic acid (PAM), unsaturated FFA (OLA, LNA, ALA) or with combinations: PAM + unsaturated FFA in a ratio 1:1 or PAM + 1 μM GW501516 for 24 h. c) Myotubes were treated with 1 μM PPARδ agonist GW501516 for 24 h. Basal and maximal respiration were assessed by a stress test as described in the Methods section. Results are presented as mean ± SEM (n ≥ 3). * p < 0.05 compared to control (C) (EPS 950 kb).

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Tumova, J., Malisova, L., Andel, M. et al. Protective Effect of Unsaturated Fatty Acids on Palmitic Acid-Induced Toxicity in Skeletal Muscle Cells is not Mediated by PPARδ Activation. Lipids 50, 955–964 (2015). https://doi.org/10.1007/s11745-015-4058-0

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