Background: The effect of a weight-loss intervention on the masses of lean tissues and organs in humans is not well known. Objective: We studied the effects of a diet and exercise weight-loss intervention on skeletal muscle (SM) mass and selected organs over 2 y using MRI in overweight adults with type 2 diabetes. Design: Participants were 53 women and 39 men [mean 6 SD: age 58 6 7 y; body mass index (BMI; in kg/m 2 ) 32 6 3] enrolled in the Look AHEAD (Action for Health in Diabetes) trial and randomly assigned to an intensive lifestyle intervention (ILI) or diabetes support and education (DSE) on whom 2 y of data were collected. MRI-derived measurements of SM, heart, liver, kidney, spleen, and pancreas were acquired. Results: Adjusted for baseline weight, height, age, sex, and ethnicity, the ILI group weighed (mean 6 SE) 6.6 6 0.7 kg less after 1 y and 5.2 6 0.7 kg less after 2 y, whereas the DSE group did not change significantly (20.4 6 0.6 and 21.0 6 0.7 kg after 1 and 2 y, respectively; P-interaction , 0.001). Total SM decreased in both groups during year 1 (21.4 6 0.2 kg; P , 0.001) with appendicular SM regained during year 2. Liver and spleen masses decreased in the ILI group (20.12 6 0.02 and 20.006 6 0.003 kg, respectively) but were unchanged in the DSE group (0.00 6 0.02 and 0.004 6 0.003 kg, respectively). Kidney mass decreased by 0.013 6 0.003 kg (P , 0.001) over 2 y in both groups. Conclusions: Decreases in liver (in Caucasians but not African Americans) and spleen were detected after a 6.2-kg weight reduction compared with a control group. SM and kidney mass decreased in both groups. Appendicular SM was regained during the second year whereas trunk SM was not. No evidence of a disproportionate loss of high-metabolic rate organs (heart, liver, kidney, spleen) compared with SM was found.Am J Clin Nutr 2017;105:78-84.