Diabetologia

, Volume 60, Issue 11, pp 2262–2273 | Cite as

VLDL and apolipoprotein CIII induce ER stress and inflammation and attenuate insulin signalling via Toll-like receptor 2 in mouse skeletal muscle cells

  • Gaia Botteri
  • Marta Montori
  • Anna Gumà
  • Javier Pizarro
  • Lídia Cedó
  • Joan Carles Escolà-Gil
  • Diana Li
  • Emma Barroso
  • Xavier Palomer
  • Alison B. Kohan
  • Manuel Vázquez-Carrera
Article

Abstract

Aim/hypothesis

Here, our aim was to examine whether VLDL and apolipoprotein (apo) CIII induce endoplasmic reticulum (ER) stress, inflammation and insulin resistance in skeletal muscle.

Methods

Studies were conducted in mouse C2C12 myotubes, isolated skeletal muscle and skeletal muscle from transgenic mice overexpressing apoCIII.

Results

C2C12 myotubes exposed to VLDL showed increased levels of ER stress and inflammatory markers whereas peroxisome proliferator-activated receptor γ co-activator 1α (PGC-1α) and AMP-activated protein kinase (AMPK) levels were reduced and the insulin signalling pathway was attenuated. The effects of VLDL were also observed in isolated skeletal muscle incubated with VLDL. The changes caused by VLDL were dependent on extracellular signal-regulated kinase (ERK) 1/2 since they were prevented by the ERK1/2 inhibitor U0126 or by knockdown of this kinase by siRNA transfection. ApoCIII mimicked the effects of VLDL and its effects were also blocked by ERK1/2 inhibition, suggesting that this apolipoprotein was responsible for the effects of VLDL. Skeletal muscle from transgenic mice overexpressing apoCIII showed increased levels of some ER stress and inflammatory markers and increased phosphorylated ERK1/2 levels, whereas PGC-1α levels were reduced, confirming apoCIII effects in vivo. Finally, incubation of myotubes with a neutralising antibody against Toll-like receptor 2 abolished the effects of apoCIII on ER stress, inflammation and insulin resistance, indicating that the effects of apoCIII were mediated by this receptor.

Conclusions/interpretation

These results imply that elevated VLDL in diabetic states can contribute to the exacerbation of insulin resistance by activating ERK1/2 through Toll-like receptor 2.

Keywords

AMPK apoCIII ERK1/2 TLR2 VLDL 

Abbreviations

ACC

Acetyl-CoA carboxylase

AMPK

AMP-activated protein kinase

Apo

Apolipoprotein

apoCIII Tg

Transgenic mice overexpressing human apoCIII

BiP

Binding immunoglobulin protein

CPT-1

Carnitine palmitoyltransferase 1

CHOP

CCAAT-enhancer-binding protein homologous protein

eIF2α

Εukaryotic initiation factor 2α

EMSA

Electrophoretic mobility shift assay

ER

Endoplasmic reticulum

ERK

Extracellular signal-regulated kinase

FAO

Fatty acid oxidation

GRP78

Glucose-regulated protein 78

IκB

Inhibitor of κB

ΙΚΚ-β

IκΒ kinase β

IRβ

Insulin receptor β-subunit

IRE-1α

Inositol-requiring 1 transmembrane kinase/endonuclease-1α

MAPK

Mitogen-activated protein kinase

MCAD

Medium chain acyl-CoA dehydrogenase

MCP-1

Monocyte chemoattractant protein 1

MEK

MAPK–ERK

NRF1

Nuclear respiratory factor 1

NRF2

Nuclear factor-E2-related factor 2

OXPHOS

Oxidative phosphorylation

PERK

Eukaryotic translation initiation factor-2α kinase 3

PGC-1α

Peroxisome proliferator-activated receptor γ co-activator 1α

PPAR

Peroxisome proliferator-activated receptor

SOCS

Suppressor of cytokine signalling 3

STAT3

Signal transducer and activator of transcription 3

TLR

Toll-like receptor

TRB3

Tribbles 3

UPR

Unfolded protein response

XBP1

X-box binding protein-1

Notes

Acknowledgements

We thank the University of Barcelona’s Language Advisory Service for revising the manuscript.

Data availability

Data are available on request from the authors.

Funding

This study was partly supported by funds from the Spanish Ministerio de Economía y Competitividad (SAF2012-30708 and SAF2015-64146-R to MVC), the Generalitat de Catalunya (2014SGR-0013 to MVC), NIH NIDDK (DK101663 to ABK), USDA NIFA (11874590 to ABK) and USDA NIFA Hatch Formula Funds (2015-31200-06009 to ABK), an Instituto de Salud Carlos III grant (PI16-00139 to JCE-G) and European Union ERDF funds. CIBER de Diabetes y Enfermedades Metabólicas Asociadas (CIBERDEM) is an Instituto de Salud Carlos III project (Grant CB07/08/0003 to MVC). GB was supported by an FPI grant from the Spanish Ministerio de Economía y Competitividad.

Duality of interest

The authors declare that there is no duality of interest associated with this manuscript.

Contribution statement

All authors processed the samples, analysed and prepared the data and were involved in drafting the article. GB, AG, JCEG, XP and ABK contributed to data interpretation and revised the article. MVC designed the experiments, interpreted the data and was primarily responsible for writing the manuscript. All authors approved the final version of the manuscript. MVC is the guarantor of this work.

Supplementary material

125_2017_4401_MOESM1_ESM.pdf (758 kb)
ESM(PDF 758 kb)

References

  1. 1.
    Xiao C, Dash S, Morgantini C, Hegele RA, Lewis GF (2016) Pharmacological targeting of the atherogenic dyslipidemia complex: the next frontier in CVD prevention beyond lowering LDL cholesterol. Diabetes 65:1767–1778CrossRefPubMedGoogle Scholar
  2. 2.
    Adiels M, Olofsson SO, Taskinen MR, Borén J (2008) Overproduction of very low-density lipoproteins is the hallmark of the dyslipidemia in the metabolic syndrome. Arterioscler Thromb Vasc Biol 28:1225–1236CrossRefPubMedGoogle Scholar
  3. 3.
    Hiukka A, Fruchart-Najib J, Leinonen E, Hilden H, Fruchart JC, Taskinen MR (2005) Alterations of lipids and apolipoprotein CIII in very low density lipoprotein subspecies in type 2 diabetes. Diabetologia 48:1207–1215CrossRefPubMedGoogle Scholar
  4. 4.
    Campos H, Perlov D, Khoo C, Sacks FM (2001) Distinct patterns of lipoproteins with apoB defined by presence of apoE or apoC-III in hypercholesterolemia and hypertriglyceridemia. J Lipid Res 42:1239–1249PubMedGoogle Scholar
  5. 5.
    Aalto-Setälä K, Fisher EA, Chen X et al (1992) Mechanism of hypertriglyceridemia in human apolipoprotein (apo) CIII transgenic mice. Diminished very low density lipoprotein fractional catabolic rate associated with increased apo CIII and reduced apo E on the particles. J Clin Invest 90:1889–1900CrossRefPubMedPubMedCentralGoogle Scholar
  6. 6.
    Kawakami A, Aikawa M, Alcaide P, Luscinskas FW, Libby P, Sacks FM (2006) Apolipoprotein CIII induces expression of vascular cell adhesion molecule-1 in vascular endothelial cells and increases adhesion of monocytic cells. Circulation 114:681–687CrossRefPubMedGoogle Scholar
  7. 7.
    Lee HY, Birkenfeld AL, Jornayvaz FR et al (2011) Apolipoprotein CIII overexpressing mice are predisposed to diet-induced hepatic steatosis and hepatic insulin resistance. Hepatology 54:1650–1660CrossRefPubMedPubMedCentralGoogle Scholar
  8. 8.
    Pollin TI, Damcott CM, Shen H et al (2008) A null mutation in human APOC3 confers a favorable plasma lipid profile and apparent cardioprotection. Science 332:1702–1705CrossRefGoogle Scholar
  9. 9.
    DeFronzo RA, Gunnarsson R, Björkman O, Olsson M, Wahren J (1985) Effects of insulin on peripheral and splanchnic glucose metabolism in noninsulin-dependent (type II) diabetes mellitus. J Clin Invest 76:149–155CrossRefPubMedPubMedCentralGoogle Scholar
  10. 10.
    Abdul-Ghani MA, DeFronzo RA (2010) Pathogenesis of insulin resistance in skeletal muscle. J Biomed Biotechnol 2010:476279CrossRefPubMedPubMedCentralGoogle Scholar
  11. 11.
    Schenk S, Saberi M, Olefsky JM (2008) Insulin sensitivity: modulation by nutrients and inflammation. J Clin Invest 118:2992–3002CrossRefPubMedPubMedCentralGoogle Scholar
  12. 12.
    Salvadó L, Palomer X, Barroso E, Vázquez-Carrera M (2015) Targeting endoplasmic reticulum stress in insulin resistance. Trends Endocrinol Metab 26:438–448CrossRefPubMedGoogle Scholar
  13. 13.
    Könner AC, Brüning JC (2011) Toll-like receptors: linking inflammation to metabolism. Trends Endocrinol Metab 22:16–23CrossRefPubMedGoogle Scholar
  14. 14.
    Nguyen A, Tao H, Metrione M, Hajri T (2014) Very low density lipoprotein receptor (VLDLR) expression is a determinant factor in adipose tissue inflammation and adipocyte-macrophage interaction. J Biol Chem 289:1688–1703CrossRefPubMedGoogle Scholar
  15. 15.
    Salvadó L, Barroso E, Gómez-Foix AM et al (2014) PPARβ/δ prevents endoplasmic reticulum stress-associated inflammation and insulin resistance in skeletal muscle cells through an AMPK-dependent mechanism. Diabetologia 57:2126–2135CrossRefPubMedGoogle Scholar
  16. 16.
    Alkhateeb H, Chabowski A, Bonen A (2006) Viability of the isolated soleus muscle during long-term incubation. Appl Physiol Nutr Metab 31:467–476CrossRefPubMedGoogle Scholar
  17. 17.
    Koh HJ, Toyoda T, Didesch MM et al (2013) Tribbles 3 mediates endoplasmic reticulum stress-induced insulin resistance in skeletal muscle. Nat Commun 4:1871CrossRefPubMedPubMedCentralGoogle Scholar
  18. 18.
    Howard JK, Flier JS (2006) Attenuation of leptin and insulin signaling by SOCS proteins. Trends Endocrinol Metab 17:365–371CrossRefPubMedGoogle Scholar
  19. 19.
    Handschin C, Spiegelman BM (2006) Peroxisome proliferator-activated receptor gamma coactivator 1 coactivators, energy homeostasis, and metabolism. Endocr Rev 27:728–735CrossRefPubMedGoogle Scholar
  20. 20.
    Patti ME, Butte AJ, Crunkhorn S et al (2003) Coordinated reduction of genes of oxidative metabolism in humans with insulin resistance and diabetes: potential role of PGC1 and NRF1. Proc Natl Acad Sci U S A 100:8466–8471CrossRefPubMedPubMedCentralGoogle Scholar
  21. 21.
    Mootha VK, Lindgren CM, Eriksson KF et al (2003) PGC-1α-responsive genes involved in oxidative phosphorylation are coordinately downregulated in human diabetes. Nat Genet 34:267–273CrossRefPubMedGoogle Scholar
  22. 22.
    Miura S, Kai Y, Ono M, Ezaki O (2003) Overexpression of peroxisome proliferator-activated receptor γ coactivator-1α down-regulates GLUT4 mRNA in skeletal muscles. J Biol Chem 278:31385–31390CrossRefPubMedGoogle Scholar
  23. 23.
    Vega RB, Huss JM, Kelly DP (2000) The coactivator PGC-1 cooperates with peroxisome proliferator-activated receptor α in transcriptional control of nuclear genes encoding mitochondrial fatty acid oxidation enzymes. Mol Cell Biol 20:1868–1876CrossRefPubMedPubMedCentralGoogle Scholar
  24. 24.
    Wu Z, Puigserver P, Andersson U et al (1999) Mechanisms controlling mitochondrial biogenesis and respiration through the thermogenic coactivator PGC-1. Cell 98:115–124CrossRefPubMedGoogle Scholar
  25. 25.
    Zhang BB, Zhou G, Li C (2009) AMPK: an emerging drug target for diabetes and the metabolic syndrome. Cell Metab 9:407–416CrossRefPubMedGoogle Scholar
  26. 26.
    Zhou L, Zhang J, Fang Q et al (2009) Autophagy-mediated insulin receptor down-regulation contributes to endoplasmic reticulum stress-induced insulin resistance. Mol Pharmacol 76:596–603CrossRefPubMedPubMedCentralGoogle Scholar
  27. 27.
    Wenz T, Rossi SG, Rotundo RL, Spiegelman BM, Moraes CT (2009) Increased muscle PGC-1α expression protects from sarcopenia and metabolic disease during aging. Proc Natl Acad Sci U S A 106:20405–20410CrossRefPubMedPubMedCentralGoogle Scholar
  28. 28.
    Chung S, Lapoint K, Martinez K et al (2006) Preadipocytes mediate lipopolysaccharide-induced inflammation and insulin resistance in primary cultures of newly differentiated human adipocytes. Endocrinology 147:5340–5351CrossRefPubMedGoogle Scholar
  29. 29.
    Coll T, Jové M, Rodríguez-Calvo R et al (2006) Palmitate-mediated downregulation of peroxisome proliferator-activated receptor-gamma coactivator 1α in skeletal muscle cells involves MEK1/2 and nuclear factor-κB activation. Diabetes 55:2779–2787CrossRefPubMedGoogle Scholar
  30. 30.
    Hwang SL, Jeong YT, Li X et al (2013) Inhibitory cross-talk between the AMPK and ERK pathways mediates endoplasmic reticulum stress-induced insulin resistance in skeletal muscle. Br J Pharmacol 169:69–81CrossRefPubMedPubMedCentralGoogle Scholar
  31. 31.
    Reaven GM, Mondon CE, Chen YD, Breslow JL (1994) Hypertriglyceridemic mice transgenic for the human apolipoprotein C-III gene are neither insulin resistant nor hyperinsulinemic. J Lipid Res 35:820–824PubMedGoogle Scholar
  32. 32.
    Salerno AG, Silva TR, Amaral ME et al (2007) Overexpression of apolipoprotein CIII increases and CETP reverses diet-induced obesity in transgenic mice. Int J Obes 31:1586–1595CrossRefGoogle Scholar
  33. 33.
    Chen M, Breslow JL, Li W, Leff T (1994) Transcriptional regulation of the apoC-III gene by insulin in diabetic mice: correlation with changes in plasma triglyceride levels. J Lipid Res 35:1918–1924PubMedGoogle Scholar
  34. 34.
    Altomonte J, Cong L, Harbaran S et al (2004) Foxo1 mediates insulin action on apoC-III and triglyceride metabolism. J Clin Invest 114:1493–1503CrossRefPubMedPubMedCentralGoogle Scholar
  35. 35.
    Caron S, Verrijken A, Mertens I et al (2011) Transcriptional activation of apolipoprotein CIII expression by glucose may contribute to diabetic dyslipidemia. Arterioscler Thromb Vasc Biol 31:513–519CrossRefPubMedGoogle Scholar
  36. 36.
    Atzmon G, Rincon M, Schechter CB et al (2006) Lipoprotein genotype and conserved pathway for exceptional longevity in humans. PLoS Biol 4:e113CrossRefPubMedPubMedCentralGoogle Scholar
  37. 37.
    Jørgensen AB, Frikke-Schmidt R, Nordestgaard BG, Tybjærg-Hansen A (2014) Loss-of-function mutations in APOC3 and risk of ischemic vascular disease. N Engl J Med 371:32–41CrossRefPubMedGoogle Scholar
  38. 38.
    Åvall K, Ali Y, Leibiger IB et al (2015) Apolipoprotein CIII links islet insulin resistance to β-cell failure in diabetes. Proc Natl Acad Sci U S A A112:E2611–E2619CrossRefGoogle Scholar
  39. 39.
    Digenio A, Dunbar RL, Alexander VJ et al (2016) Antisense-mediated lowering of plasma apolipoprotein C-III by volanesorsen improves dyslipidemia and insulin sensitivity in type 2 diabetes. Diabetes Care 39:1408–1415CrossRefPubMedGoogle Scholar
  40. 40.
    Ozaki KI, Awazu M, Tamiya M et al (2016) Targeting the ERK signaling pathway as a potential treatment for insulin resistance and type 2 diabetes. Am J Physiol Endocrinol Metab 310:E643–E651PubMedGoogle Scholar
  41. 41.
    Bost F, Aouadi M, Caron L et al (2005) The extracellular signal-regulated kinase isoform ERK1 is specifically required for in vitro and in vivo adipogenesis. Diabetes 54:402–411CrossRefPubMedGoogle Scholar
  42. 42.
    Jager J, Corcelle V, Grémeaux T et al (2011) Deficiency in the extracellular signal-regulated kinase 1 (ERK1) protects leptin-deficient mice from insulin resistance without affecting obesity. Diabetologia 54:180–189CrossRefPubMedGoogle Scholar
  43. 43.
    Dong Y, Zhang M, Wang S et al (2010) Activation of AMP-activated protein kinase inhibits oxidized LDL-triggered endoplasmic reticulum stress in vivo. Diabetes 59:1386–1396CrossRefPubMedPubMedCentralGoogle Scholar
  44. 44.
    Dong Y, Zhang M, Liang B et al (2010) Reduction of AMP-activated protein kinase α2 increases endoplasmic reticulum stress and atherosclerosis in vivo. Circulation 121:792–803CrossRefPubMedPubMedCentralGoogle Scholar
  45. 45.
    Cantó C, Auwerx J (2009) PGC-1α, SIRT1 and AMPK, an energy sensing network that controls energy expenditure. Curr Opin Lipidol 20:98–105CrossRefPubMedPubMedCentralGoogle Scholar
  46. 46.
    Rubartelli A, Lotze MT (2007) Inside, outside, upside down: damage-associated molecular-pattern molecules (DAMPs) and redox. Trends Immunol 28:429–436CrossRefPubMedGoogle Scholar
  47. 47.
    Senn JJ (2006) Toll-like receptor-2 is essential for the development of palmitate-induced insulin resistance in myotubes. J Biol Chem 281:26865–26875CrossRefPubMedGoogle Scholar
  48. 48.
    Ninomiya-Tsuji J, Kishimoto K, Hiyama A, Inoue J, Cao Z, Matsumoto K (1999) The kinase TAK1 can activate the NIK-IκB as well as the MAP kinase cascade in the IL-1 signalling pathway. Nature 398:252–256CrossRefPubMedGoogle Scholar
  49. 49.
    Kuo LH, Tsai PJ, Jiang MJ et al (2011) Toll-like receptor 2 deficiency improves insulin sensitivity and hepatic insulin signalling in the mouse. Diabetologia 54:168–179CrossRefPubMedGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany 2017

Authors and Affiliations

  • Gaia Botteri
    • 1
    • 2
    • 3
  • Marta Montori
    • 1
    • 2
    • 3
  • Anna Gumà
    • 2
    • 4
  • Javier Pizarro
    • 1
    • 2
    • 3
  • Lídia Cedó
    • 2
    • 5
  • Joan Carles Escolà-Gil
    • 2
    • 5
    • 6
  • Diana Li
    • 7
  • Emma Barroso
    • 1
    • 2
    • 3
  • Xavier Palomer
    • 1
    • 2
    • 3
  • Alison B. Kohan
    • 7
  • Manuel Vázquez-Carrera
    • 1
    • 2
    • 3
  1. 1.Pharmacology Unit, Department of Pharmacology, Toxicology and Therapeutic Chemistry, Faculty of Pharmacy and Food Sciences, Institut de Biomedicina de la Universidad de Barcelona (IBUB)University of BarcelonaBarcelonaSpain
  2. 2.Centro de Investigación Biomédica en Red de Diabetes y Enfermedades Metabólicas Asociadas (CIBERDEM)Instituto de Salud Carlos IIIBarcelonaSpain
  3. 3.Institut de Recerca Sant Joan de Déu (IR-SJD)BarcelonaSpain
  4. 4.Department of Biochemistry and Molecular Biology and IBUBUniversity of BarcelonaBarcelonaSpain
  5. 5.Institut d’Investigacions Biomèdiques (IIB) Sant PauBarcelonaSpain
  6. 6.Department of Biochemistry and Molecular BiologyAutonomous University of BarcelonaBarcelonaSpain
  7. 7.Department of Nutritional SciencesUniversity of ConnecticutStorrsUSA

Personalised recommendations