Echomri

Body composition was measured by MRI using an EchoMRI (Echo Medical Systems). Indirect calorimetry was performed in groups of 7–8 mice using a Comprehensive Laboratory Animal Monitoring System (Columbus Instruments) with free access to food and tap water. Mice were individually housed at room temperature, and a standard 12-hour light/dark cycle was maintained throughout the measurements. Mice were acclimated to the cages for a period of 48 hours before the start of 4 days of measurements at 20-minute intervals. Food intake was assessed by real-time feed weight measurements. Oxygen consumption and carbon dioxide production were measured, and based on this respirometry, EE as well as CHO and FA oxidation were calculated as previously described (82 (link)).

Body composition was measured with Echo-MRI (Echo Medical Systems). After body composition measure mice were individually placed in the Oxymax Comprehensive Lab Animal Monitoring System (Columbus Instruments). Oxygen consumption (VO2), carbon dioxide production (VCO₂) and ambulatory activity were measured over the course of 2 days. Respiratory exchange ratio (RER, or Respiratory quotient) and heat production were calculated but not reported because the assumptions are not met in abnormal metabolic states [35 (link)–37 (link)]. VO2 and VCO2 were normalized over grams of body weight. Food intake was calculated from the difference between the food allocated to every mouse at the start of the recording and the amount of food that was left at the end of the recording. All experiments were performed at the IBP Phenotyping Core Facility, University of Minnesota. Data were analyzed with unpaired t-test.

Experiments were performed with six different batches of male C57Bl/6J mice (Charles River, Maastricht, the Netherlands). The batches differed in duration (4–34 weeks) and type of HFD (45% or 60% energy derived from lard fat; D12451 or D12492, Research Diet Services, Wijk bij Duurstede, the Netherlands) (electronic supplementary material [ESM] Table 1). Body weight was measured and lean and fat mass was assessed by MRI-based body composition analysis (Echo MRI, Echo Medical Systems, Houston, TX, USA). At the end of the diet intervention, mice were killed, perfused and organs were dissected for further analysis. All experiments were approved by the animal ethics committee of Leiden University Medical Center.

After the mice were sacrificed, their body composition was determined by whole-body nuclear magnetic resonance imaging (EchoMRI, Echo Medical Systems, Waco, TX).