Abstract
Aims/hypothesis
Saturated fatty acids (SFAs) such as palmitate activate inflammatory pathways and elicit an endoplasmic reticulum (ER) stress response in macrophages, thereby contributing to the development of insulin resistance linked to the metabolic syndrome. This study addressed the question of whether or not mitochondrial fatty acid β-oxidation (FAO) affects macrophage responses to SFA.
Methods
We modulated the activity of carnitine palmitoyl transferase 1A (CPT1A) in macrophage-differentiated THP-1 monocytic cells using genetic or pharmacological approaches, treated the cells with palmitate and analysed the proinflammatory and ER stress signatures.
Results
To inhibit FAO, we created THP-1 cells with a stable knockdown (KD) of CPT1A and differentiated them to macrophages. Consequently, in CPT1A-silenced cells FAO was reduced. CPT1A KD in THP-1 macrophages increased proinflammatory signalling, cytokine expression and ER stress responses after palmitate treatment. In addition, in human primary macrophages CPT1A KD elevated palmitate-induced inflammatory gene expression. Pharmacological inhibition of FAO with etomoxir recapitulated the CPT1A KD phenotype. Conversely, overexpression of a malonyl-CoA-insensitive CPT1A M593S mutant reduced inflammatory and ER stress responses to palmitate in THP-1 macrophages. Macrophages with a CPT1A KD accumulated diacylglycerols and triacylglycerols after palmitate treatment, while ceramide accumulation remained unaltered. Moreover, lipidomic analysis of ER phospholipids revealed increased palmitate incorporation into phosphatidylethanolamine and phosphatidylserine classes associated with the CPT1A KD.
Conclusions/interpretation
Our data indicate that FAO attenuates inflammatory and ER stress responses in SFA-exposed macrophages, suggesting an anti-inflammatory impact of drugs that activate FAO.
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Abbreviations
- AMPK:
-
AMP-dependent protein kinase
- CPT1:
-
Carnitine palmitoyl transferase 1
- CPT1A:
-
Liver isoform of carnitine palmitoyl transferase 1
- CV:
-
Control virus
- DAG:
-
Diacylglycerol
- DFPU:
-
Difluorophenylurethane
- eIF2α:
-
Eukaryotic initiation factor 2α
- ER:
-
Endoplasmic reticulum
- FAO:
-
Fatty acid β-oxidation
- IRE1α:
-
Inositol requiring enzyme 1α
- IκB:
-
Inhibitor of κB
- KD:
-
Knockdown
- LC–MS/MS:
-
Liquid chromatography–tandem mass spectrometry
- MAPK:
-
Mitogen-activated protein kinase
- NLS:
-
Neutral loss scan
- OCR:
-
Oxygen consumption rate
- PBA:
-
Phenylbutyric acid
- PC:
-
Phosphatidylcholine
- PE:
-
Phosphatidylethanolamine
- PEMT:
-
Phosphatidylethanolamine-methyltransferase
- PERK:
-
Protein kinase RNA-like endoplasmic reticulum kinase
- PIS:
-
Precursor ion scan
- PKC:
-
Protein kinase C
- PMA:
-
Phorbol 12-myristate 13-acetate
- PS:
-
Phosphatidylserine
- SFA:
-
Saturated fatty acid
- shRNA:
-
Small hairpin RNA
- siRNA:
-
Small interfering RNA
- sXBP1:
-
Spliced X-box binding protein 1
- TAG:
-
Triacylglycerol
- TLR:
-
Toll-like receptor
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Acknowledgements
We thank I. Elschner (Institute of Biochemistry I, Goethe University Frankfurt, Frankfurt, Germany) for technical assistance. We also thank D. Kauhanen and K. Tarasov (Zora Biosciences Oy, Espoo, Finland) for assistance with the shotgun lipidomics analyses.
Funding
This work was supported by grants from Deutsche Forschungsgemeinschaft (grant no. BR999) and by the Else Kröner Fresenius Foundation (Translational Research Innovation—Pharma [TRIP]). Mass spectrometric measurements of DAGs were supported by a grant (RCM) from the US National Institutes of Health (Lipid Maps, GM006938).
Duality of interest
The authors declare that there is no duality of interest associated with this manuscript.
Contribution statement
DN acquired data, drafted and revised the manuscript. SL acquired data and edited the manuscript. TJL and RCM contributed to DAG analysis and revised the manuscript. KE contributed to the shotgun lipidomics analysis and revised the manuscript. NF and GG contributed to ceramide analysis and revised the manuscript. BB analysed data and revised the manuscript. All of the authors approved the final version of the manuscript. BB is responsible for the integrity of the work as a whole.
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Dmitry Namgaladze and Sebastian Lips contributed equally to this study.
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Namgaladze, D., Lips, S., Leiker, T.J. et al. Inhibition of macrophage fatty acid β-oxidation exacerbates palmitate-induced inflammatory and endoplasmic reticulum stress responses. Diabetologia 57, 1067–1077 (2014). https://doi.org/10.1007/s00125-014-3173-4
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DOI: https://doi.org/10.1007/s00125-014-3173-4