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. 2017 May 1;312(5):E381-E393.
doi: 10.1152/ajpendo.00408.2016. Epub 2017 Feb 21.

Loss of Macrophage Fatty Acid Oxidation Does Not Potentiate Systemic Metabolic Dysfunction

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Free PMC article

Loss of Macrophage Fatty Acid Oxidation Does Not Potentiate Systemic Metabolic Dysfunction

Elsie Gonzalez-Hurtado et al. Am J Physiol Endocrinol Metab. .
Free PMC article

Abstract

Fatty acid oxidation in macrophages has been suggested to play a causative role in high-fat diet-induced metabolic dysfunction, particularly in the etiology of adipose-driven insulin resistance. To understand the contribution of macrophage fatty acid oxidation directly to metabolic dysfunction in high-fat diet-induced obesity, we generated mice with a myeloid-specific knockout of carnitine palmitoyltransferase II (CPT2 Mϕ-KO), an obligate step in mitochondrial long-chain fatty acid oxidation. While fatty acid oxidation was clearly induced upon IL-4 stimulation, fatty acid oxidation-deficient CPT2 Mϕ-KO bone marrow-derived macrophages displayed canonical markers of M2 polarization following IL-4 stimulation in vitro. In addition, loss of macrophage fatty acid oxidation in vivo did not alter the progression of high-fat diet-induced obesity, inflammation, macrophage polarization, oxidative stress, or glucose intolerance. These data suggest that although IL-4-stimulated alternatively activated macrophages upregulate fatty acid oxidation, fatty acid oxidation is dispensable for macrophage polarization and high-fat diet-induced metabolic dysfunction. Macrophage fatty acid oxidation likely plays a correlative, rather than causative, role in systemic metabolic dysfunction.

Keywords: adipose tissue; fatty acid; inflammation; macrophage; obesity.

Figures

Fig. 1.
Fig. 1.
Generation of mice with a macrophage-specific loss of mitochondrial fatty acid β-oxidation. A: mRNA for carnitine palmitoyltransferase (CPT) II (Cpt2) in bone marrow-derived macrophages (BMDM) from control [wild-type (WT)] mice and BMDM from mice with a specific knockout of Cpt2 [CPT2 Mϕ-KO (KO), n = 3]. B: Western blot for Cpt2 in control and CPT2 Mϕ-KO BMDM. C: Western blot of mitochondrial proteins in control and CPT2 Mϕ-KO BMDM in the presence and absence of IL-4 stimulation (n = 2). D: oxidation of [1-14C]oleic acid to 14CO2 in control and CPT2 Mϕ-KO BMDM (n = 6). E: oxidation of [U-14C]glucose to 14CO2 in control and CPT2 Mϕ-KO BMDM (n = 6). F: incorporation of [3H]acetate into lipid in control and CPT2 Mϕ-KO BMDM (n = 6). G: [1,2-3H]2-deoxyglucose uptake in control and CPT2 Mϕ-KO BMDM (n = 6). H: steady-state metabolite concentrations in IL-4- or LPS-IFNγ-stimulated control and CPT2 Mϕ-KO BMDM (n = 5–6). Values are means ± SE. *P < 0.05, ***P < 0.001 for genotype comparison. #P < 0.05, ###P < 0.001 for treatment comparison.
Fig. 2.
Fig. 2.
Macrophage mitochondrial fatty acid β-oxidation is not required for alternative activation in vitro. A–D: quantitative RT-PCR analysis of M1 and M2 macrophage markers, fatty acid synthesis and transcriptional regulator genes, fatty acid oxidation genes, and oxidative stress genes in control and CPT2 Mϕ-KO BMDM in the presence and absence of IL-4 or LPS-IFNγ. Values are means ± SE (n = 6). *P < 0.05, **P < 0.01, ***P < 0.001.
Fig. 3.
Fig. 3.
Loss of macrophage fatty acid oxidation does not affect body composition. A: body weight of control and CPT2 Mϕ-KO male mice fed a low-fat diet (LFD) or a high-fat diet (HFD, n = 10–14). B: EchoMRI measurement of body composition of control and CPT2 Mϕ-KO mice fed a low- or high-fat diet (n = 10–14). C: inguinal white adipose tissue (iWAT), gonadal white adipose tissue (gWAT), and liver wet weight of control and CPT2 Mϕ-KO male mice fed a low- or high-fat diet (n = 10–14). D: serum metabolites [β-hydroxybutyrate (β-HB), triacylglycerol (TAG), nonesterified fatty acid (NEFA), cholesterol, and glycerol] in control and CPT2 Mϕ-KO male mice fed a high-fat diet (n = 7). E: hematoxylin-eosin-stained sections of gWAT from control and CPT2 Mϕ-KO male mice fed a low- or high-fat diet. Scale bar = 200 μm. F: hematoxylin-eosin-stained sections of gWAT from liver of control and CPT2 Mϕ-KO male mice fed a low- or high-fat diet. Scale bar = 200 μm. Values are means ± SE.
Fig. 4.
Fig. 4.
Macrophage fatty acid β-oxidation is dispensable for polarization in vivo. A–C: quantitative RT-PCR analysis of macrophage markers in gWAT(n = 6), liver (n = 6), and spleen (n = 6) of control and CPT2 Mϕ-KO male mice fed a low- or high-fat diet. Values are means ± SE. #P < 0.05; ##P < 0.01; ###P < 0.001 for treatment comparison.
Fig. 5.
Fig. 5.
Loss of macrophage fatty acid β-oxidation does not affect polarization of adipose tissue macrophages from high-fat diet-fed mice. A and B: total adipose cell number and percentage of adipose tissue macrophages in gWAT of control and CPT2 Mϕ-KO male mice fed a high-fat diet. CF: percentage of CD206+, CD301+, TNFα+, and IFNγ+ adipose tissue macrophages in control and CPT2 Mϕ-KO male mice fed a high-fat diet. Values are means ± SE (n = 5).
Fig. 6.
Fig. 6.
Loss of macrophage fatty acid oxidation does not potentiate oxidative stress. A: quantitative RT-PCR analysis of oxidative stress genes in gWAT of control and CPT2 Mϕ-KO male mice fed a low- or high-fat diet (n = 6). B: thiobarbituric acid-reactive substances assay of gWAT, liver, and serum of control and CPT2 Mϕ-KO male mice fed a high-fat diet (n = 5). MDA, malondialdehyde. Values are means ± SE. #P < 0.05, ###P < 0.001 for treatment comparison.
Fig. 7.
Fig. 7.
Loss of macrophage fatty acid oxidation does not alter progression of high-fat diet-induced insulin resistance. A and B: intraperitoneal glucose tolerance test (ipGTT) and intraperitoneal insulin tolerance test (ipITT), including area under the curve and area above the curve, respectively, for control and CPT2 Mϕ-KO male mice fed low-fat (n = 10–13) and high-fat (n = 8–10) diets. Values are means ± SE.
Fig. 8.
Fig. 8.
Loss of macrophage fatty acid oxidation does not induce a mitochondrial DNA stress response. Quantitative RT-PCR analysis of interferon-stimulated genes in control and CPT2 Mϕ-KO BMDM in the presence and absence of polyinosinic:polycytidylic acid [poly(I:C)]. Values are means ± SE (n = 6). ***P < 0.001.
Fig. 9.
Fig. 9.
Loss of macrophage fatty acid β-oxidation does not result in free fatty acid-induced inflammation. Quantitative RT-PCR analysis of inflammatory response genes in control and CPT2 Mϕ-KO BMDM in the presence and absence of 100 μM FFA. Values are means ± SE (n = 6).

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