Role of vascular oxidative stress in obesity and metabolic syndrome

Abstract

Obesity is associated with vascular diseases that are often attributed to vascular oxidative stress. We tested the hypothesis that vascular oxidative stress could induce obesity. We previously developed mice that overexpress p22phox in vascular smooth muscle, tg(sm/p22phox), which have increased vascular ROS production. At baseline, tg(sm/p22phox) mice have a modest increase in body weight. With high-fat feeding, tg(sm/p22phox) mice developed exaggerated obesity and increased fat mass. Body weight increased from 32.16 ± 2.34 g to 43.03 ± 1.44 g in tg(sm/p22phox) mice (vs. 30.81 ± 0.71 g to 37.89 ± 1.16 g in the WT mice). This was associated with development of glucose intolerance, reduced HDL cholesterol, and increased levels of leptin and MCP-1. Tg(sm/p22phox) mice displayed impaired spontaneous activity and increased mitochondrial ROS production and mitochondrial dysfunction in skeletal muscle. In mice with vascular smooth muscle-targeted deletion of p22phox (p22phox(loxp/loxp)/tg(smmhc/cre) mice), high-fat feeding did not induce weight gain or leptin resistance. These mice also had reduced T-cell infiltration of perivascular fat. In conclusion, these data indicate that vascular oxidative stress induces obesity and metabolic syndrome, accompanied by and likely due to exercise intolerance, vascular inflammation, and augmented adipogenesis. These data indicate that vascular ROS may play a causal role in the development of obesity and metabolic syndrome.

Figure 1

Augmented obesity in high-fat diet–fed tg^sm/p22phox mice. A: Representative mice from WT and tg^sm/p22phox groups fed with high-fat diet for 6 weeks. B: Body weight gain in WT and tg^sm/p22phox mice fed with control or high-fat diet for 6 weeks. C: White fat mass. D: Liver weight in WT and tg^sm/p22phox mice fed with control or high-fat diet for 6 weeks. Data are presented as mean ± SEM; n = 10–14 for A–D.

Figure 2

Changes in water intake, food intake, and energy intake in WT and tg^sm/p22phox mice fed with control or high-fat diet for 6 weeks. A: Water intake was measured weekly, and there were no significant changes among the four different groups except for weeks 3–5. B: Weekly food intake was decreased in WT mice after high-fat feeding for 2 weeks. C: Energy intake was calculated into kilocalories from grams of food ingested as described in Research Design and Methods. Data are presented as mean ± SEM; n = 7–11 for A–C.

Figure 3

NMR analysis of body weight, fat mass, and muscle mass in WT and tg^sm/p22phox mice fed with control or high-fat diet for 6 weeks. A: Body weight was measured weekly. High-fat diet feeding induced an exaggerated body weight gain in tg^sm/p22phox mice. B: Total fat mass was measured weekly and found to be substantially more increased by high-fat diet feeding in tg^sm/p22phox mice. C: Total muscle mass was monitored weekly and found not to be different either at baseline or at 6 weeks after high-fat diet feeding between WT and tg^sm/p22phox mice. Data are presented as mean ± SEM; n = 5 for A–C.

Figure 4

Leptin resistance and dyslipidemia in high-fat diet–fed tg^sm/p22phox mice. A: Plasma leptin levels were measured weekly as described in Research Design and Methods. A remarkable increase in plasma leptin levels was observed in high-fat diet–fed tg^sm/p22phox mice, while it did not occur in the WT mice fed with a high-fat diet. These data implicate a leptin resistance phenotype. B: Total cholesterol levels were increased in both WT and tg^sm/p22phox mice fed with high-fat diet. C: High-fat diet feeding induced a significant reduction in HDL cholesterol in tg^sm/p22phox mice. D: Plasma MCP-1 levels at 6 weeks of high-fat feeding were markedly increased in tg^sm/p22phox mice. Data are presented as mean ± SEM; n = 7–11 for A–C, n = 6–7 for D). E: Plasma MCP-1 levels were positively correlated with plasma leptin levels (n = 27 of 4 groups).

Figure 5

Insulin resistance in high-fat diet–fed tg^sm/p22phox mice. A: Fasting glucose levels were measured weekly over 6 weeks. Changes from baseline were presented. B: Weekly circulating insulin levels were determined by ELISA. Insulin levels were elevated in a time-dependent manner in high-fat diet–fed tg^sm/p22phox mice. Data are presented as mean ± SEM; n = 7–11 for A and B.

Figure 6

Impaired glucose tolerance in high-fat diet–fed tg^sm/p22phox mice. Intraperitoneal glucose tolerance test was performed at weeks 3 and 5. A: Glucose intolerance was observed in high-fat diet–fed tg^sm/p22phox mice at week 3. B: Glucose intolerance was aggravated by high-fat feeding at week 5 in tg^sm/p22phox mice. Data are presented as mean ± SEM; n = 7–11 for A and B.

Figure 7

Decreased spontaneous activity accompanied by mitochondrial dysfunction in skeletal muscle of high-fat–fed tg^sm/p22phox mice. A: Spontaneous activity was monitored over 8 weeks of high-fat diet feeding and progressively declined in the tg^sm/p22phox mice while remaining constant in the WT mice. B: Mitochondrial fraction from skeletal muscle was prepared as described in Research Design and Methods and subjected to superoxide detection using electron spin resonance. Mitochondrial superoxide production from high-fat diet–fed tg^sm/p22phox mice was increased more than threefold compared with WT controls fed high-fat diet. C: Calcium-induced swelling of skeletal muscle mitochondria was significantly augmented in high-fat diet–fed tg^sm/p22phox mice compared with WT controls fed high-fat diet. n = 11–13. Data are presented as mean ± SEM.

Figure 8

Prevention of obesity induction in p22phox knockout mice. p22phox^loxp/loxp crossed with mice expressing Cre recombinase driven by the tamoxifen inducible smooth muscle myosin heavy chain promoter, Tg^smmhc/cre. A: Expression of p22phox was decreased upon tamoxifen introduction. B: High-fat feeding for 6 weeks failed to induce body weight gain in p22phox knockout mice. n = 6. C: Leptin level was attenuated in high-fat diet–fed p22phox knockout mice, while it was increased in vehicle corn oil–treated mice with high-fat diet feeding or cre-negative mice. n = 5–6. Data are presented as mean ± SEM.

Figure 9

Effect of high-fat feeding on mesenteric fat leukocytes in presence and absence of VSM p22phox. Mice were fed either a control diet or a high-fat diet for 6 weeks. Mice fed a high-fat diet were p22phox^loxp/loxp, and half had the VSMC-specific Cre transgene induced by tamoxifen injection (gray bars). As control, p22phox^loxp/loxp Cre-negative mice were also fed a high-fat diet and were treated with tamoxifen (black bars). The mesenteric vasculature with all adjacent fat was removed en bloc, digested, and subjected to FACS analysis for measurement of total leukocytes and T-cell subtypes. A: Populations of leukocytes were analyzed by FACS. B: T-cell subtypes were also analyzed by FACS. Data are presented as mean ± SEM; n = 4 for A and B.