Adipocyte-Specific Deletion of Manganese Superoxide Dismutase Protects From Diet-Induced Obesity Through Increased Mitochondrial Uncoupling and Biogenesis
Article Figures & Tables
Figures
- Figure 1
MnSOD deletion in adipocytes increased superoxide generation but did not enhance lipid peroxidation. Representative Western blots and the corresponding densitometry of MnSOD protein expression normalized by SDHA expression in isolated adipocytes from iWAT (A) and eWAT (B). MnSOD protein expression normalized GAPDH protein expression in whole interscapular BAT homogenates (C). D: Superoxide levels in isolated adipocytes from iWAT of WT or AdSod2 KO mice fed an NCD or HFD for 21 weeks. E: Representative Western blot of 4-HNE and GAPDH protein expression in iWAT. F: Densitometry of 4-HNE normalized by GAPDH. G: Relative mRNA expression of major antioxidant enzymes in iWAT. n = 3–6 mice per group. Data are mean ± SEM. *P < 0.05, **P < 0.005 vs. WT under the same feeding condition; #P < 0.05, ##P < 0.005 vs. NCD within the same genotype.
- Figure 2
MnSOD deficiency in adipocytes reduced diet-induced weight gain and adiposity. Body weights (A), weight gain (B), iWAT weight/body weight (BW) × 103 (C), eWAT weight/BW × 103 (D), BAT weight/BW × 103 (E) in WT or AdSod2 KO mice maintained on NCD or HFD for 21 weeks. F: Representative images of iWAT and eWAT sections stained with hematoxylin and eosin. G and H: Distribution of adipocyte diameter in iWAT and eWAT, respectively. I and J: Adipocyte number per area in iWAT and eWAT, respectively. n = 12–18 mice per group for A–E. Ten images per animal were used to quantify adipocyte diameter and number for five to six mice per group. Scale bar = 40 μm. Data are mean ± SEM. *P < 0.05, **P < 0.005 vs. WT under the same feeding condition; #P < 0.05, ##P < 0.005 vs. NCD within the same genotype.
- Figure 3
Deletion of MnSOD in adipocytes increased metabolic rate and EE when mice are maintained on an HFD. VO2 (A) and VCO2 (B) at night averaged over a 72-h period. Calculated RER (C) and EE (D). Food intake (E) and ambulatory activity (F) in WT and AdSod2 KO mice fed NCD or HFD for 12 weeks. n = 6–9 mice per group. Data are mean ± SEM. *P < 0.05 vs. WT under the same feeding condition; ##P < 0.005 vs. NCD within the same genotype.
- Figure 4
Increased metabolic rate and FA utilization in HFD-fed AdSod2 KO mice at thermoneutrality. WT and AdSod2 KO mice fed NCD or HFD for 8–9 weeks were housed in metabolic cages for 72 h, and their VO2, VCO2, and RER were recorded at 23°C, 15°C, and 30°C. A–C: Changes in VO2, VCO2, and RER, respectively, over 72 h. D–F: Average night data for VO2, VCO2, and RER, respectively. n = 4 mice per group. Data are mean ± SEM. *P < 0.05, **P < 0.005 vs. WT under the same feeding condition; ##P < 0.005 vs. NCD within the same genotype.
- Figure 5
MnSOD deficiency triggered an HFD-dependent increase in FA oxidation, mitochondrial respiration, and mitochondrial biogenesis. WT and AdSod2 KO mice were maintained on an HFD for 21 weeks. A: Palmitate oxidation in iWAT, eWAT, BAT, and hindlimb muscle. B: Fold change in mRNA expression of FA oxidation and mitochondrial biogenesis genes in iWAT. C and D: OCR and mtDNA/nuclear DNA ratios in iWAT. E and F: Electron micrographs and quantification of mitochondrial density in iWAT. n = 8–9 mice per group in A, and n = 6 mice per group in B. OCR, mtDNA, and mitochondrial density were quantified in three different sections of iWAT from three to four mice per group. Data are mean ± SEM. *P < 0.05, **P < 0.005 vs. WT. cpm, counts per minute.
- Figure 6
Lack of MnSOD increased superoxide generation, uncoupled respiration, and enhanced mitochondrial content in cultured adipocytes. APs were isolated from iWAT of WT or AdSod2 KO mice fed NCD or HFD for 12 weeks and differentiated into adipocytes. A: Representative Western blots of MnSOD and tubulin protein expression before differentiation (day 0) and 6 and 12 days postdifferentiation. Representative images of Oil Red O staining of neutral lipids (B) and the corresponding quantification at 6 days postdifferentiation (C). Superoxide levels (D), OCRs (E), and mtDNA/nuclear DNA ratios (F) at 12 days postdifferentiation. G: Fold change in Ucp1 and Cidea mRNA in iWAT. n = 3 independent experiments (each included five to six mice per group). Scale bar = 40 μm in B. Data are mean ± SEM. *P < 0.05 vs. WT under the same feeding condition; #P < 0.05 vs. NCD within the same genotype. A.U., arbitrary units; DIFF, differentiated; UND, undifferentiated.
- Figure 7
AdSod2 KO mice are protected from diet-induced hepatic steatosis, adipose inflammation, and glucose and insulin intolerance. A and B: Representative images of liver sections stained with Oil Red O and liver triglyceride content in WT and AdSod2 KO mice fed NCD and HFD for 21 weeks. C and D: Macrophage (F4/80) staining and quantification in eWAT. E: Fold change in mRNA expression of pro- and anti-inflammatory genes in eWAT. F–H: Body weights, GTTs, and areas under the curve (AUCs) for GTT in WT and AdSod2 KO mice fed NCD or HFD for 6 weeks. I and J: Insulin tolerance tests (ITTs) and AUCs for ITT in WT and AdSod2 KO mice fed NCD or HFD for 21 weeks. n = 4–5 mice per group for A–D, n = 6 mice per group for E, and n = 8–17 mice per group for F–J. Scale bars = 20 μm in A and 400 μm in C. Data are mean ± SEM. *P < 0.05, **P < 0.005 vs. WT under the same feeding condition; #P < 0.05, ##P < 0.005 vs. NCD within the same genotype.
Tables
- Table 1
Serum metabolites, hormones, and lipids in WT and AdSod2 KO mice fed NCD or HFD
NCD HFD WT AdSod2 KO WT AdSod2 KO Glucose (mg/dL) 109.9 ± 8.1 99.3 ± 10.8 146 ± 6.3## 129.8 ± 5.9# T0 insulin (ng/mL) 0.34 ± 0.06 0.3 ± 0.06 0.54 ± 0.06 0.21 ± 0.03** T30 insulin (ng/mL) 0.48 ± 0.09 0.47 ± 0.1 0.47 ± 0.08 0.35 ± 0.05 Leptin (ng/mL) 5 ± 0.5 5.4 ± 0.8 14.6 ± 4.3 4.1 ± 0.9* Adiponectin (ng/mL) 7,443.1 ± 505.1 7,242.1 ± 504.8 6,890 ± 413.8 7,183 ± 579.6 FFA (mmol/L) 0.46 ± 0.03 0.63 ± 0.07 0.46 ± 0.06 0.28 ± 0.03* Triglycerides (mg/dL) 104.1 ± 8.1 92.2 ± 10.6 74.9 ± 16.8 63.3 ± 16.1 Cholesterol (mg/dL) 236.9 ± 27.7 230.7 ± 10.8 195.8 ± 31.6 236.8 ± 19.1 Data are mean ± SEM. Serum metabolites, hormones, and lipids were measured after a 6-h fast. T0 insulin and T30 insulin correspond to insulin measured before glucose injection and 30 min after glucose injection, respectively, during GTTs. All other metabolites and hormones were determined by blood samples extracted before glucose injection before GTTs. n = 5 mice per genotype for NCD and n = 8 mice per genotype for HFD.
*P < 0.05 vs. WT mice under the same feeding condition.
**P < 0.005 vs. WT mice under the same feeding condition.
#P < 0.05 vs. NCD within the same genotype.
##P < 0.005 vs. NCD within the same genotype.
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