Vascular inflammation, insulin resistance, and reduced nitric oxide production precede the onset of peripheral insulin resistance

Abstract

Objective: Obesity causes inflammation and insulin resistance in the vasculature as well as in tissues involved in glucose metabolism such as liver, muscle, and adipose tissue. To investigate the relative susceptibility of vascular tissue to these effects, we determined the time course over which inflammation and insulin resistance develops in various tissues of mice with diet-induced obesity (DIO) and compared these tissue-based responses to changes in circulating inflammatory markers.

Methods and results: Adult male C57BL/6 mice were fed either a control low-fat diet (LF; 10% saturated fat) or a high-fat diet (HF, 60% saturated fat) for durations ranging between 1 to 14 weeks. Cellular inflammation and insulin resistance were assessed by measuring phospho-IkappaBalpha and insulin-induced phosphorylation of Akt, respectively, in extracts of thoracic aorta, liver, skeletal muscle, and visceral fat. As expected, HF feeding induced rapid increases of body weight, fat mass, and fasting insulin levels compared to controls, each of which achieved statistical significance within 4 weeks. Whereas plasma markers of inflammation became elevated relatively late in the course of DIO (eg, serum amyloid A [SAA], by Week 14), levels of phospho-IkappaBalpha in aortic lysates were elevated by 2-fold within the first week. The early onset of vascular inflammation was accompanied by biochemical evidence of both endothelial dysfunction (reduced nitric oxide production; induction of intracellular adhesion molecule-1 and vascular cell adhesion molecule-1) and insulin resistance (impaired insulin-induced phosphorylation of Akt and eNOS). Although inflammation and insulin resistance were also detected in skeletal muscle and liver of HF-fed animals, these responses were observed much later (between 4 and 8 weeks of HF feeding), and they were not detected in visceral adipose tissue until 14 weeks.

Conclusions: During obesity induced by HF feeding, inflammation and insulin resistance develop in the vasculature well before these responses are detected in muscle, liver, or adipose tissue. This observation suggests that the vasculature is more susceptible than other tissues to the deleterious effects of nutrient overload.

Figures

Figure 1. Time course of the effect HF feeding on vascular inflammation and basal NO levels

A. Levels of phospho-IκBα in lysates of thoracic aorta after low fat (L) or high fat (H) feeding, with a representative Western blot for each time point. B. Fold increase in VCAM mRNA levels as measured by quantitative PCR. C. Fold increase in ICAM protein. D. Levels of NO in lysates of thoracic aorta. *P<0.05 vs. LF controls.

Figure 2. Time course of the effect of HF feeding on vascular insulin signaling

A. pAkt levels in lysates of thoracic aorta as measured by ELISA. For each time point, pAkt levels were normalized to the low fat (L), saline vehicle condition. B. peNOS levels in lysates of thoracic aorta as measured by Western blot analysis, quantified using densitometry, and the ratio of peNOS/total eNOS was calculated. *P<0.05 vs. low fat controls (arrow indicates 140 kD molecular weight).

Figure 3. Time course of the effect of HF feeding on liver, muscle, and adipose tissue phospho-IκBα

Levels of phospho-IκBα were determined in lysates of liver (A), skeletal (quadriceps) muscle (B), and mesenteric adipose tissue (C) by Western blot in mice fed a LF (L) vs. HF (H) diet for periods ranging from 1-14 wk. Data are expressed as fold increase over the LF-fed, vehicle condition. *P

Figure 4. Time course of the effect of HF feeding on insulin signaling in liver, muscle, and adipose tissue

After consuming a LF (L) or HF (H) diet for periods of 1-14 wk, mice received either saline or insulin. Fifteen minutes later, animals were sacrificed and tissues harvested for analysis of pAkt levels in lysates of liver (A), skeletal (quadriceps) muscle (B), and mesenteric adipose tissue (C). *P