4.6. Body Composition and Energy Expenditure Measurement
Body composition (lean and fat mass) was evaluated in each mouse using magnetic resonance (EchoMRI; Echo Medical Systems, Houston, TX, USA). Energy expenditure was measured by indirect calorimetry in an Oxymax Lab Animal Monitoring System (CLAMS; Columbus Instruments, Columbus, OH, USA). Animals were individually housed in Plexiglas cages with an open flow system connected to CLAMS for 24 h. Prior to the test, animals were acclimatized for 24 h, fasted for 12 h in the light period and fed during the dark period. Throughout the test, O2 consumption (VO2, mL/kg/h or mL/kg lean mass/h) and CO2 production (VCO2, mL/kg/h) were measured sequentially during 90 s. Multiple linear regression was performed to evaluate the slopes of change of VO2 (mL/h) per unit change of body weight or lean mass. The respiratory exchange ratio (RER) was calculated as the average ratio of produced CO2 to O2 inhaled (VCO2/VO2).