Introduction
Exercise training is associated with enhanced glucose tolerance and insulin action in healthy and insulin resistant people. The molecular mechanism for enhanced glucose uptake with exercise training may be partly related to increased expression and activity of an array of key proteins known to regulate glucose metabolism in skeletal muscle. We determined whether habitual exercise training is associated with an altered expression of proteins involved in insulin signaling.
Methods
Eighteen healthy young male volunteers participated in the study. The study groups consisted of 11 habitual runners (trained) and 6 sedentary (untrained) controls. Informed consent was obtained from each subject. The Ethical Committee at Karolinska Institutet approved the study protocol. After local anesthesia (Mepivakain chloride 5 mg/ml), an incision (5 mm long/10 mm deep) was made in the skin and fascia, and a muscle biopsy was obtained from the vastus lateralis portion of the quadriceps femoris by means of a Weil-Blakesley´s conchotome. Muscle tissue was immediately frozen and stored in liquid nitrogen until further analysis.
Skeletal muscle biopsies (60-70 mg) were homogenized in ice-cold buffer (50 mM Tris HCl, 0.1% Trition X 100, 1 mM EDTA, 1 mM EGTA, 50 mM NaF, 5 mM Na2P2O7, 10 mM glycerophosphate, 1 mM Na3VO4, 1 µM microcystin, 0.1% b-mercaptoethanol). Aliquots of lysates (20 µg) were mixed with Laemmli sample buffer, and proteins were separated by SDS-PAGE and analyzed by Western blot with appropriate antibody.
Results
Insulin receptor (IR), insulin receptor substrate (IRS) 1 and IRS-2 expression was decreased 44% (P<0.05), 57% (P<0.001) and 77% (P<0.001) respectively, in trained vs. untrained muscle. Training did not alter expression of the downstream signaling target, Akt kinase. ERK 1/2 MAP kinase expression was 190% greater (P<0.05), whereas p38 MAP kinase expression was 32% lower (P<0.05) in trained vs. untrained muscle. GLUT4 protein expression was 2-fold higher (P<0.05) and the GLUT4 vesicle-associated protein, the insulin-regulated aminopeptidase (IRAP), was increased 4.7-fold (P<0.05) in trained muscle.
Conclusions
Here we provide molecular evidence for differential effects of habitual exercise on the expression of an array of key signaling proteins in skeletal muscle. Regular exercise-training of moderate intensity is associated with a striking down regulation of early components of the insulin signaling cascade, concomitant with a profound up-regulation of GLUT4 and IRAP in skeletal muscle. Furthermore, different MAP kinase signaling pathways appear to play specialized roles in modulating exercise-adaptations on gene expression since ERK 1/2 MAP kinase expression was increased in trained muscle, while p38 MAP kinase expression was decreased.
References