Effects of different diets used in diet-induced obesity models on insulin resistance and vascular dysfunction in C57BL/6 mice

Abstract

The aim of the present study was to compare different diets used to induce obesity in a head-to-head manner with a focus on insulin resistance and vascular dysfunction. Male C57BL/6J mice were put on standard chow diet (SCD), normal-fat diet (NFD), cafeteria diet (CAF) or high-fat diet (HFD) for 12 weeks starting at the age of 6 weeks. Both CAF and HFD led to obesity (weight gain of 179% and 194%, respectively), glucose intolerance and insulin resistance to a comparable extent. In aortas containing perivascular adipose tissue (PVAT), acetylcholine-induced vasodilation was best in the NFD group and worst in the CAF group. Reduced phosphorylation of endothelial nitric oxide synthase at serine 1177 was observed in both CAF and HFD groups. Plasma coagulation activity was highest in the HFD group and lowest in the SCD group. Even the NFD group had significantly higher coagulation activity than the SCD group. In conclusions, CAF and HFD are both reliable mouse diets in inducing visceral obesity, glucose intolerance and insulin resistance. CAF is more effective than HFD in causing PVAT dysfunction and vascular dysfunction, whereas hypercoagulability was mostly evident in the HFD group. Coagulation activity was higher in NFD than NCD group.

Figure 1

Development of the bodyweight. Male C57BL/6J mice were put on standard chow diet (SCD), cafeteria diet (CAF), normal-fat diet (NFD) or high-fat diet (HFD) at the age of 6 weeks for another 12 weeks. Food intake per day per mouse in gram (A) and changes in food consumption per week per mouse in gram over the feeding period (B) are pictured. ANOVA followed by Fisher’s protected least significant difference test as post hoc method was performed to compare all four groups. Panel C shows the development of the bodyweight over time. Panel D shows the bodyweight after 12 weeks of experimental feeding. Significances marked with ^*,# and ⁺refer to comparisons to the SCD, CAF and NFD group, respectively. ^*,#P < 0.05; ^***,###,+++P < 0.001; ns, not significant. n = 15 (SCD and CAF) or 20 (NFD and HFD), respectively.

Figure 2

Weight of fat mass. Male C57BL/6J mice were put on experimental diets at the age of 6 weeks for another 12 weeks. The weights of epididymal (A), lumbar (B) and mesenteric fat pads (C) at the end of the experiment are shown. Significances marked with ^{*, #} and ⁺refer to comparisons to the SCD, CAF and NFD group, respectively *P < 0.05; **P < 0.01; ^***###,+++P < 0.001; ns, not significant. n = 15 (SCD and CAF) or 20 (NFD and HFD), respectively.

Figure 3

Glucose intolerance. Male C57BL/6J mice were put on experimental diets at the age of 6 weeks for another 12 weeks. Panel A shows the fasting glucose levels in the 12th week of feeding. n = 15 (SCD and CAF) or 20 (NFD and HFD), respectively. Panel B shows the plasma insulin concentration in the 12th week of feeding. n = 6 (SCD, NFD and HFD) or 5 (CAF), respectively. Panel C shows the results of GTT at the end of the experiment after overnight fasting. n = 5 (SCD, CAF and HFD) and 9 (NFD), respectively. Panels E shows the results of ITT (1 IU/kg body weight insulin was injected intraperitoneally) in the 12^th week of experimental feeding after overnight fasting. n = 10 (SCD and NFD) and 9 (CAF and HFD), respectively. The area under the curve (AUC) of GTT and ITT are shown in panels D and F, respectively. Significances marked with ^*,# and ⁺refer to comparisons to the SCD, CAF and NFD group, respectively. ^*,#,+P < 0.05, ^**,##P < 0.01, ^***,###,+++P < 0.001, ns, not significant.

Figure 4

Vascular function. Male C57BL/6J mice were put on experimental diets at the age of 6 weeks for another 12 weeks. Vasodilation was induced by acetylcholine in norepinephrine-pre-contracted aorta in myograph experiments, without PVAT (A), with PVAT (B–E), or in the presence of NO synthase inhibitor L-NAME (E). *P < 0.05, **P < 0.01, ***P < 0.001. n = 9 (SCD and CAF) and 15 (NFD and HFD), respectively.

Figure 5

Phosphorylation of eNOS. Male C57BL/6J mice were put on experimental diets at the age of 6 weeks for another 12 weeks. Phosphorylation of eNOS at Ser1177 (A) and of Akt at Ser473 (B) in aortic PVAT was studied by Western blot analyses. The blots shown are representative of four independent experiments with similar results. Significances marked with ^*,# and ⁺refer to comparisons to the SCD, CAF and NFD group, respectively. ^*,#P < 0.05, ^**,##,++P < 0.01, ***P < 0.001, ns, not significant. n = 12.

Figure 6

Coagulation. Male C57BL/6J mice were put on experimental diets at the age of 6 weeks for another 12 weeks. Prothrombin time (PT) (A) and activated partial thromboplastin time (APTT) (B) were measured with citrate plasma on a KC4 Delta Amelung Coagulometer. Significances marked with ^*,# and ⁺refer to comparisons to the SCD, CAF and NFD group, respectively. *P < 0.05, ***P < 0.001, ns, not significant. n = 10 (SCD and CAF), 9 (NFD) and 11 (HFD), respectively.

Figure 7

Activity of coagulation factors. Male C57BL/6J mice were put on experimental diets at the age of 6 weeks for another 12 weeks. The activity of coagulation factors was determined using deficient plasma (shorter coagulation time means higher activity) on a KC4 Delta Amelung Coagulometer. Shorter coagulation time indicates higher activity. Significances marked with ^*,# and ⁺refer to comparisons to the SCD, CAF and NFD group, respectively. ^*,#,+P < 0.05, **P < 0.01, ***P < 0.001, ns, not significant. n = 9 (SCD, NFD and CAF) and 11 (HFD), respectively.