Preserved endothelial function in human obesity in the absence of insulin resistance

Abstract

Background: Insulin resistance (IR) is frequently associated with endothelial dysfunction and has been proposed to play a major role in cardiovascular disease (CVD). On the other hand, obesity has long been related to IR and increased CVD. However it is not known if IR is a necessary condition for endothelial dysfunction in human obesity, allowing for preserved endothelial function in obese people when absent. Therefore, the purpose of the study was to assess the relationship between IR and endothelial dysfunction in human obesity and the mechanisms involved.

Methods: Twenty non-insulin resistant morbid obese (NIR-MO), 32 insulin resistant morbid obese (IR-MO), and 12 healthy subjects were included. Serum concentrations of glucose, insulin, interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-α), resistin and adiponectin were determined. IR was evaluated by HOMA-index. Endothelium-dependent relaxation to bradykinin (BK) in mesenteric microvessels was assessed in wire myograph.

Results: Serum IL-6, and TNF-α levels were elevated only in IR-MO patients while resistin was elevated and adiponectin reduced in all MO individuals. Mesenteric arteries from IR-MO, but not from NIR-MO subjects displayed blunted relaxation to BK. Vasodilatation was improved in IR-MO arteries by the superoxide scavenger, superoxide dismutase (SOD) or the mitochondrial-targeted SOD mimetic, mito-TEMPO. NADPH oxidase inhibitors (apocynin and VAS2870) and the nitric oxide synthase (NOS) cofactor, tetrahydrobiopterin failed to modify BK-induced vasodilatations. Superoxide generation was higher in vessels from IR-MO subjects and reduced by mito-TEMPO. Blockade of TNF-α with infliximab, but not inhibition of inducible NOS or cyclooxygenase, improved endothelial relaxation and decreased superoxide formation.

Conclusions: Endothelial dysfunction is observed in human morbid obesity only when insulin resistance is present. Mechanisms involved include augmented mitochondrial superoxide generation, and increased systemic inflammation mediated by TNF-α. These findings may explain the different vascular risk of healthy vs unhealthy obesity.

Figure 1

Insulin resistance is related to endothelial dysfunction in morbid obesity. (A) Vasodilation to bradykinin (BK) in mesenteric arterial segments derived from control subjects, non-insulin resistant (NIR-MO) and insulin resistant morbid obese (IR-MO) subjects. Data are expressed as mean±SEM of the percentage of the contraction elicited by K⁺. N= the number of subjects/ n= the number of vascular segments. ^*p< 0.001 vs NIR-MO and ^¶p< 0.001 vs control subjects by two-factors ANOVA. (B) Negative correlation between Homeostasis Model Assessment of Insulin Resistance (HOMA-IR) score and pD₂ values for BK. Each point represents the averaged pD₂ values of segments from one single subject. (C) Effect of preincubation with the superoxide dismutase (SOD; 100 U/ml) on the relaxant responses to BK in mesenteric arteries from IR-MO. ^*p< 0.05 vs IR-MO subjects by two-factors ANOVA.

Figure 2

Sources of superoxide anions in mesenteric arteries of morbid obese subjects with insulin resistance (IR-MO). Effects of preincubation with the NADPH oxidase inhibitors, apocynin (100 μmol/l) (A); or VAS2870 (10 μmol/l) (B); the NOS cofactor, tetrahydrobiopterin (BH₄; 10 μmol/l) (C); and the SOD mimetic targeted to the mitochondria, mito-TEMPO (5 μmol/l) (D) on the relaxant response to bradykinin in isolated mesenteric microvessels from IR-MO subjects. Data are expressed as mean±SEM of the percentage of the contraction elicited by K⁺. N= the number of subjects/n= the number of vascular segments. ^*p< 0.001 vs IR-MO by two-factors ANOVA.

Figure 3

Detection of superoxide generation in vessel segments from morbid obese subjects without insulin resistance (NIR-MO) and with insulin resistance (IR-MO) and the effect of mitochondria antioxidant, mito-TEMPO and the anti-TNF-α, infliximab on superoxide generation in IR-MO microarteries. Representative photomicrographs (A) and quantitative analysis (B) of the staining (red signal) with the superoxide-detecting fluorescent probe, dihydroethidium (DHE), in mesenteric microvessels derived from control, NIR-MO and IR-MO subjects treated or not with the superoxide scavenger targeted to the mitochondria, mito-TEMPO (5 μmol/l), and the anti-TNF-α, infliximab (100 μmol/l) on superoxide generation in IR-MO vessels. Original magnification is (20X). ^*p< 0.001 vs control subjects; p < 0.01 vs NIR-MO subjects; ^† p< 0.05 vs IR-MO subjects respectively. Each column represents the mean±SEM of 4 to 6 experiments. Panel (C) shows the correlation between the percentages of DHE positive nuclei and pD₂ values for BK.

Figure 4

Role of inflammation on endothelial dysfunction in morbid obese subjects with insulin resistance (IR-MO). Effects of preincubation with the iNOS inhibitor, 1400W (10 μmol/l) (A), the COX inhibitor, indomethacin (10 μmol/l) (B), and the anti-TNF-α, infliximab (100 μmol/l) (C) on the relaxant response to bradykinin (BK) in isolated mesenteric microvessels from IR-MO. Data are expressed as mean±SEM of the percentage of the contraction elicited by K⁺. N= the number of subjects/n= the number of vascular segments. ^*p< 0.05 vs IR-MO subjects by two-factors ANOVA.