Early responses of insulin signaling to high-carbohydrate and high-fat overfeeding

Abstract

Background: Early molecular changes of nutritionally-induced insulin resistance are still enigmatic. It is also unclear if acute overnutrition alone can alter insulin signaling in humans or if the macronutrient composition of the diet can modulate such effects.

Methods: To investigate the molecular correlates of metabolic adaptation to either high-carbohydrate (HC) or high-fat (HF) overfeeding, we conducted overfeeding studies in 21 healthy lean (BMI < 25) individuals (10 women, 11 men), age 20-45, with normal glucose metabolism and no family history of diabetes. Subjects were studied first following a 5-day eucaloric (EC) diet (30% fat, 50% CHO, 20% protein) and then in a counter balanced manner after 5 days of 40% overfeeding of both a HC (20% fat, 60% CHO) diet and a HF (50% fat, 30% CHO) diet. At the end of each diet phase, in vivo insulin sensitivity was assessed using the hyperinsulinemic-euglycemic clamp technique. Ex vivo insulin action was measured from skeletal muscle tissue samples obtained 15 minutes after insulin infusion was initiated.

Results: Overall there was no change in whole-body insulin sensitivity as measured by glucose disposal rate (GDR, EC: 12.1 ± 4.7; HC: 10.9 ± 2.7; HF: 10.8 ± 3.4). Assessment of skeletal muscle insulin signaling demonstrated increased tyrosine phosphorylation of IRS-1 (p < 0.001) and increased IRS-1-associated phosphatidylinositol 3 (PI 3)-kinase activity (p < 0.001) following HC overfeeding. In contrast, HF overfeeding increased skeletal muscle serine phosophorylation of IRS-1 (p < 0.001) and increased total expression of p85α (P < 0.001).

Conclusion: We conclude that acute bouts of overnutrition lead to changes at the cellular level before whole-body insulin sensitivity is altered. On a signaling level, HC overfeeding resulted in changes compatible with increased insulin sensitivity. In contrast, molecular changes in HF overfeeding were compatible with a reduced insulin sensitivity.

Figure 1

Phosphorylation of IRS-1. Tyrosine (A) and Serine (B) phosphorylation of IRS-1 in skeletal muscle following Eucaloric feeding (EC), high-carbohydrate overfeeding (HC), and high-fat overfeeding (HF). HC overfeeding increased tyrosine phosphorylation of IRS-1 compared to EC feeding. HF overfeeding increased serine phosphorylation of IRS-1 compared to EC feeding.

Figure 2

Representative blots. Representative blots from ex vivo skeletal muscle tissue following Eucaloric feeding (EC), high-carbohydrate overfeeding (HC), and high-fat overfeeding (HF).

Figure 3

PI 3-kinase activity and subunit expression. IRS-1-associated PI 3-kinase activity (A), IRS-1-associated p110 protein expression (B), and Total p85α protein expression (C) in skeletal muscle following Eucaloric feeding (EC), high-carbohydrate overfeeding (HC), and high-fat overfeeding (HF). HC overfeeding increased IRS-1 associated PI 3-kinase activity and IRS-1 associated p110 expression compared to EC feeding. HF overfeeding increased total p85α expression compared to EC feeding.

Figure 4

mTOR and S6K1 activity. Phosphorylation of mTOR (A) and S6K1 (B) in skeletal muscle following Eucaloric feeding (EC), high-carbohydrate overfeeding (HC), and high-fat overfeeding (HF). Both HC and HF overfeeding increased phosphorylation of mTOR and S6K1 compared to EC feeding.