(1) Objective: The aim of this study was to clarify the role of adipokines in the regulation of glucose metabolism in middle-aged obese subjects with impaired glucose tolerance in response to a long-term exercise and dietary intervention. (2) Methods: Skeletal muscle, plasma and serum samples were examined in 22 subjects from an exercise-diet intervention study aiming to prevent type 2 diabetes. The subjects were further divided into two subgroups (non-responders n = 9 and responders n = 13) based on their achievement in losing at least 3 kg. (3) Results: The two-year exercise-diet intervention reduced leptin levels and increased adiponectin levels in responders; the changes in leptin levels were significantly associated with changes in their weights (r = 0.662, p < 0.01). In responders, insulin sensitivity (Bennett and McAuley index) increased and was associated with changes in maximal oxygen uptake (VO₂peak) (r = 0.831, p < 0.010 and r = 0.890, p < 0.01). In addition, the VO₂peak and oxidative capacity of skeletal muscle improved in responders, but not in non-responders. However, there were no changes between the two groups in expressions of the glucose transporter protein-4 (GLUT-4) gene or of AMP-activated protein kinase (AMPK)-alpha 1 or AMPK-alpha 2 proteins. (4) Conclusions: The exercise-diet intervention decreased serum leptin and increased serum adiponectin concentrations, improved glucose control without affecting GLUT-4 gene expression in the skeletal muscle in responders.