Irisin is assumed to be a relevant link between muscle and weight maintenance as well as to mediate exercise benefits on health. The aim of this study was to assess the possible associations between irisin levels and glucose homeostasis in obese subjects with metabolic syndrome (MetS) following an energy-restricted treatment. Ninety-six adults with excessive body weight and MetS features underwent a hypocaloric dietary pattern for 8 weeks, within the RESMENA randomized controlled trial (www.clinicaltrials.gov; NCT01087086). After the intervention, dietary restriction significantly reduced body weight and evidenced a dietary-induced decrease in circulating levels of irisin in parallel with improvements on glucose homeostasis markers. Interestingly, participants with higher irisin values at baseline (above the median) showed a greater reduction on glucose (P=0.022) and insulin (P=0.021) concentrations as well as on the homeostasis model assessment index (P=0.008) and triglycerides (P=0.006) after the dietary intervention, compared with those presenting low-irisin baseline values (below the median). Interestingly, a positive correlation between irisin and carbohydrate intake was found at the end of the experimental period. In conclusion, irisin appears to be involved in glucose metabolism regulation after a dietary-induced weight loss.