Elevated fasting insulin concentrations associate with impaired insulin signaling in skeletal muscle of healthy subjects independent of obesity.

BACKGROUND: Insulin signaling is impaired in the skeletal muscle of obese subjects but whether defects in skeletal muscle insulin signaling also characterize insulin resistance of non-obese individuals is unknown. The detection of insulin signaling defects in muscle biopsies is hampered by the variation of the contaminating non-muscle elements such as blood, connective tissue, fat, and blood vessel structures. Freeze-drying and macroscopic purification of the muscle fibers prior to the analysis might offer a possibility to minimize the analytical variation due to these contaminants.

METHODS: In the present study we first determined whether insulin signaling could be reliably assessed in freeze-dried muscle specimens, which are free of non-muscle contaminants, and then applied this method to the study of insulin signaling in weight-matched insulin-sensitive and insulin-resistant non-diabetic men.

RESULTS: In rat muscle, increases in tyrosine phosphorylation of insulin receptor (IR) and activity of the insulin receptor substrate-1 (IRS-1)-associated phosphatidylinositol (PI) 3-kinase activity by insulin were similar or higher in freeze-dried and purified muscle than wet muscle. Prior to freeze-drying and purification, biopsies of human vastus lateralis muscle contained between 1% and 40% non-muscle contaminants (11+/-3%, mean+/-SEM, n=19). In freeze-dried biopsies of human vastus lateralis muscle taken before and after 30 min of hyperinsulinemia (serum free insulin 61+/-1 mU/l) in 13 non-diabetic men, insulin increased IR tyrosine phosphorylation 1.4-fold (p<0.05) and IRS-1-associated PI 3-kinase activity 1.7-fold (p<0.005). Insulin-stimulated PI 3-kinase activity was significantly inversely correlated with the fasting serum insulin concentration (r=-0.57, p<0.05). When divided according to the median fasting serum insulin concentration, the men with high fasting insulin [HI, n=7, age 44+/-3 years, body mass index (BMI) 25+/-1 kg/m(2)] as compared to the men with low fasting insulin [LI, n=6, age 45+/-3 years (NS), BMI 24+/-1 kg/m(2) (NS)] had lower rates of whole-body glucose uptake (3.4+/-0.4 vs 5.5+/-0.3 mg/kg min, p<0.005), higher fasting plasma glucose concentrations (5.9+/-0.2 vs 5.2+/-0.1 mmol/l, p<0.05), higher fasting serum triglycerides (1.4+/-0.2 vs 0.9+/-0.1 mmol/l, p<0.05) and lower high-density lipoprotein (HDL) cholesterol concentrations (1.3+/-0.1 vs 1.7+/-0.1 mmol/l, p<0.05). Insulin-stimulated IR tyrosine phosphorylation (p<0.05) and IRS-1-associated PI 3-kinase activity (p<0.05) were significantly lower in the HI than the LI group.

CONCLUSIONS: Taken together these data demonstrate that early insulin signaling events can be reliably assessed in freeze-dried human skeletal muscle, and that in vivo insulin resistance and its accompanying features are associated with defects in early insulin signaling events in human skeletal muscle independent of body weight.