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. 2013 Feb 15;304(4):E392-404.
doi: 10.1152/ajpendo.00398.2012. Epub 2012 Dec 11.

NOX2 deficiency attenuates markers of adiposopathy and brain injury induced by high-fat diet

Jennifer K Pepping  1 , Linnea R Freeman, Sunita Gupta, Jeffrey N Keller, Annadora J Bruce-Keller
Affiliations

NOX2 deficiency attenuates markers of adiposopathy and brain injury induced by high-fat diet

Jennifer K Pepping et al. Am J Physiol Endocrinol Metab. .

Abstract

The consumption of high-fat/calorie diets in modern societies is likely a major contributor to the obesity epidemic, which can increase the prevalence of cancer, cardiovascular disease, and neurological impairment. Obesity may precipitate decline via inflammatory and oxidative signaling, and one factor linking inflammation to oxidative stress is the proinflammatory, pro-oxidant enzyme NADPH oxidase. To reveal the role of NADPH oxidase in the metabolic and neurological consequences of obesity, the effects of high-fat diet were compared in wild-type C57Bl/6 (WT) mice and in mice deficient in the NAPDH oxidase subunit NOX2 (NOX2KO). While diet-induced weight gains in WT and NOX2KO mice were similar, NOX2KO mice had smaller visceral adipose deposits, attenuated visceral adipocyte hypertrophy, and diminished visceral adipose macrophage infiltration. Moreover, the detrimental effects of HFD on markers of adipocyte function and injury were attenuated in NOX2KO mice; NOX2KO mice had improved glucose regulation, and evaluation of NOX2 expression identified macrophages as the primary population of NOX2-positive cells in visceral adipose. Finally, brain injury was assessed using markers of cerebrovascular integrity, synaptic density, and reactive gliosis, and data show that high-fat diet disrupted marker expression in WT but not NOX2KO mice. Collectively, these data indicate that NOX2 is a significant contributor to the pathogenic effects of high-fat diet and reinforce a key role for visceral adipose inflammation in metabolic and neurological decline. Development of NOX-based therapies could accordingly preserve metabolic and neurological function in the context of metabolic syndrome.

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Figures

Fig. 1.
Fig. 1.
Effects of high-fat diet (HFD) on body weight and composition in WT and NAPDH oxidase subunit 2 knockout (NOX2KO) mice. Four-month old male C57Bl/6 (WT) mice and B6.129S-Cybbtm1Din/J (NOX2KO) mice were placed for 14 wk on HFD or the nutritionally matched low-fat control diet (CD) with 10 mice in each group. A: body weight in WT and NOX2KO mice over time following administration of CD or HFD. #Significant (P < 0.05) increase in body weight noted in NOX2KO vs. WT mice after 4 and 6 wk of diet. B: body fat as %total body weight in WT and NOX2KO mice over time following administration of CD or HFD. Significant (#P < 0.05, ##P < 0.01, respectively) increases in body fat in WT vs. NOX2KO mice after 11 and 14 wk of HFD. C: lean body mass as %total body weight in WT and NOX2KO mice over time following administration of CD or HFD. Significant (#P < 0.05, ##P < 0.01, respectively) decrease in lean body mass in WT vs. NOX2KO mice after 11 and 14 wk of HFD.
Fig. 2.
Fig. 2.
Effects of HFD on adipose depot weights in WT and NOX2KO mice. Subcutaneous inguinal and visceral epididymal fat pads were collected from WT and NOX2KO mice at the end of the 14-wk feeding trial and weighed. Data were collected from 10 mice in each group except for the NOX2KO/HFD group, which had 9 samples. A: inguinal fat depot weight in WT and NOX2KO mice following administration of CD or HFD. ***Significant (P < 0.001) increase in weight of inguinal fat depot in both WT and NOX2KO mice on HFD vs. CD. B: epididymal fat depots in WT and NOX2KO mice following administration of CD or HFD. *Significant (P < 0.05) increase in weight of epididymal fat depot in WT mice given HFD vs. WT mice on CD. #Significant (P < 0.05) increase in weight of epididymal fat depot in WT vs. NOX2KO mice on HFD.
Fig. 3.
Fig. 3.
Effects of HFD on adipocyte hypertrophy in WT and NOX2KO mice. Subcutaneous inguinal and visceral epididymal fat pads were collected from WT and NOX2KO mice at the end of the 14-wk feeding trial and processed for histological analyses of adipocyte size as described in materials and methods. Data were collected from 10 mice in each group except for the NOX2KO/HFD group, which had 9 samples. A: size of inguinal adipocytes in WT and NOX2KO mice after administration of CD or HFD. **Significant (P < 0.01) increase in adipocyte size in WT mice on HFD vs. WT mice on CD; ***significant (P < 0.001) increase in NOX2KO mice on HFD vs. NOX2KO mice on CD. B: size of epididymal adipocytes in WT and NOX2KO mice after administration of CD or HFD. ***Significant (P < 0.001) increase in adipocyte size in WT mice on HFD vs. WT mice on CD; ###significant decrease in adipocyte size in HFD-fed NOX2KO mice vs. HFD-fed WT mice. (C) Representative images of H&E stained adipocytes from which adipocyte size measures were based.
Fig. 4.
Fig. 4.
Effects of HFD on macrophage infiltration into adipose depots in WT and NOX2KO mice. Iba-1 expression was used to measure macrophage infiltration/activation in inguinal and epididymal fat. Data were collected from 10 mice in each group except for the NOX2KO/HFD group, which had 9 samples. A: HFD-induced Iba-1 expression in inguinal and epididymal adipose tissue in WT and NOX2KO mice as percentage of expression in CD mice. *Significant (P < 0.05) HFD-induced increase in Iba-1 expression in inguinal and epididymal fat in WT mice and inguinal fat in NOX2KO mice. B: representative Western blot images showing Iba-1 (17 kDa) expression in epididymal fat taken from WT and NOX2KO mice given CD or HFD. C: representative images of Iba-1 immunostaining showing macrophage activation and infiltration into epididymal fat taken from WT and NOX2KO mice given CD or HFD.
Fig. 5.
Fig. 5.
Effects of HFD on markers of adipocyte function and injury in WT and NOX2KO mice. Expression of (A) PPARγ, (B) adiponectin, (C) GADD153/CHOP, and (D) GRP78 were evaluated in tissue homogenates prepared from epididymal adipose depots. Data were collected from 10 mice in each group except for the NOX2KO/HFD group, which had 9 samples, and depict means ± SE expression in HFD mice presented as percentage values in CD mice (100% line on graph). Significant (*P < 0.05, **P < 0.01, ***P < 0.001, respectively) changes in expression in HFD mice vs. CD mice; #significant (P < 0.05) decreases in adiponectin expression in WT-HFD adipose vs. changes in expression in NOX2KO-HFD mice.
Fig. 6.
Fig. 6.
Expression of NOX2 in visceral adipose of WT mice. A: representative images of NOX2 Western blot showing that the antibody used for immunostaining did not recognize epitopes in NOX2KO tissue. B: representative images of NOX2 immunoreactivity (left, red), Iba-1 immunoreactivity (middle, black), and NOX2/Iba-1 double-labeled sections (right) of epididymal fat taken from WT mice given CD or HFD. Images reflect the predominant expression of NOX2 in Iba-1-positive macrophages, although NOX2-positive, Iba-1-negative cells can be observed (arrows in right), perhaps arising from preadipocytes or other stromovascular cells.
Fig. 7.
Fig. 7.
Effects of HFD on glucose regulation and tolerance in WT and NOX2KO mice. Mice were fasted for 4 h, after which an OGTT was performed to measure glucose regulation and insulin resistance. Data were collected from 10 mice in each group except for the NOX2KO/HFD group, which had 9 samples. A: fasting blood glucose levels measured in WT and NOX2KO mice after administration of HFD or CD. ***Significant (P < 0.001) increase in fasting blood glucose level in HFD-fed WT vs. CD-fed WT mice; ###significant decrease in fasting glucose in HFD-fed NOX2KO vs. HFD-fed WT mice. B: fasting insulin levels measured in WT and NOX2KO mice after administration of HFD or CD. ***Significant (P < 0.001) increase in fasting insulin in WT mice on HFD vs. WT mice on CD, and similarly an increase in NOX2KO mice on HFD vs. NOX2KO mice on CD. C: oral glucose tolerance, as indicated by area under the curve (AUC). ***Significantly (P < 0.001) larger AUC in WT mice on HFD vs. WT mice on CD; #significant decrease in glucose AUC in HFD-fed NOX2KO vs. HFD-fed WT mice. *Significantly (P < 0.05) larger AUC in NOX2KO mice on HFD vs. NOX2KO mice on CD. D: NEFA levels in mice 60 min after glucose (2 mg/kg) gavage. Significant (*P < 0.05 and **P < 0.001, respectively) increases in serum NEFA in HFD mice; ##significant decrease in NEFA following glucose load in HFD-fed NOX2KO vs. HFD-fed WT mice.
Fig. 8.
Fig. 8.
Effects of HFD on markers of brain injury in WT and NOX2KO mice. A: markers of cerebrovascular integrity (expression of tight junction proteins claudin-5, ZO-1, and occludin and matrix metalloproteinases MMP2 and MMP9). B: synaptic density [expression of postsynaptic marker protein synapse-associated protein-97 (SAP97), presynaptic protein synapsin 1, and phosphorylated synapsin 1] were evaluated in tissue homogenates prepared from the frontal cortex. C: reactive gliosis (expression of GFAP, Iba-1, iNOS, COX2). Data were collected from 10 mice in each group except for the NOX2KO/HFD group, which had 9 samples, and depict means ± SE expression in HFD mice presented as %values in CD mice (100% line) on graph. Significant (*P < 0.05, **P < 0.01, ***P < 0.001, respectively) changes in expression in WT HFD vs. WT CD mice.
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References

    1. Ahmed Z, Shaw G, Sharma VP, Yang C, McGowan E, Dickson DW. Actin-binding proteins coronin-1a and IBA-1 are effective microglial markers for immunohistochemistry. J Histochem Cytochem 55: 687–700, 2007 - PubMed
    1. Akiyama H, Barger S, Barnum S, Bradt B, Bauer J, Cole G, Cooper NR, Eikelenboom P, Emmerling M, Fiebich BL, Finch CE, Frautschy S, Griffin WS, Hampel H, Hull M, Landreth G, Lue L, Mrak R, Mackenzie IR, McGeer PL, O'Banion MK, Pachter J, Pasinetti G, Plata-Salaman C, Rogers J, Rydel R, Shen Y, Streit W, Strohmeyer R, Tooyoma I, Van Muiswinkel FL, Veerhuis R, Walker D, Webster S, Wegrzyniak B, Wenk G, Wyss-Coray T. Inflammation and Alzheimer's disease. Neurobiol Aging 21: 383–421, 2000 - PMC - PubMed
    1. Ali SS, Young JW, Wallace CK, Gresack J, Jeste DV, Geyer MA, Dugan LL, Risbrough VB. Initial evidence linking synaptic superoxide production with poor short-term memory in aged mice. Brain Res 1368: 65–70, 2011 - PMC - PubMed
    1. Appachi S, Kelly KR, Schauer PR, Kirwan JP, Hazen S, Gupta M, Kashyap SR. Reduced cardiovascular risk following bariatric surgeries is related to a partial recovery from “adiposopathy”. Obes Surg 12: 1928–1936, 2011 - PMC - PubMed
    1. Aprahamian TR, Sam F. Adiponectin in cardiovascular inflammation and obesity. Int J Inflam 2011: 376909, 2011 - PMC - PubMed

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