Echomri 900

A nuclear echo magnetic resonance imaging whole-body composition analyzer (Echo MRI 900; EchoMedical Systems, Houston, TX, USA) was used^{35 (link)} to repeatedly assess body fat and lean mass changes in conscious mice during the study at T0, T16, T18 and T23 (Fig. 2).

The EchoMRI-900 quantitative nuclear magnetic resonance (qMR) system (Echo Medical Systems, Houston, TX) was used to determine fat mass and lean mass in conscious mice. Blood samples were collected via tail vein or cardiac puncture performed on terminally anesthetized mice. Blood glucose, triglycerides, cholesterol and free fatty acid, and plasma leptin and insulin were measured as described previously (51 (link)). Plasma β-hydroxybutyrate was measured using a colorimetric assay (Cayman Chemical Company, Ann Arbor, MI). Plasma alanine aminotransferase (ALT) activity was measured using kits on a Daytona autoanalyzer (Randox). Glucose and insulin tolerance tests were carried out on mice as described elsewhere (51 (link)). The respiratory exchange ratio (RER) and O₂ consumption were determined by open-circuit indirect calorimetry (Columbus Instruments).

Mice were injected daily intraperitoneally (ip) for 10 days, then twice-daily for a total of 21 days with the Jak2 inhibitor NVP-BSK805 [0.03 mg in 0.1 mL dimethyl sulfoxide (DMSO)] or its vehicle. The peripheral dose was chosen based on the doses used for icv administration. Another group of mice underwent anesthesia and cannula implantation in the brain lateral ventricle, as detailed in Andre et al. (2017) (link), allowing administration of NVP-BSK805 (3.12 μg/μL in 1 μL DMSO, icv) or vehicle once a week for 3 weeks. Animals were free-fed and all injections were performed during the light phase. FI and BW were recorded daily. Assessment of body fat and lean mass in conscious mice was carried out using a nuclear echo magnetic resonance imaging whole-body composition analyzer, which gives information on the total quantity of fat and lean mass in the body by using NMR-MRI-based technology (Echo MRI 900; Echo Medical Systems, Houston, TX, United States), as done previously (Andre et al., 2017 (link)), before and after 3 weeks of treatment. Feed efficiency, intended as efficiency of conversion of ingested food into fat mass, was calculated as the ratio between fat mass gain and cumulative caloric intake over the period of the study.

Body weight was measured on a weekly basis from baseline to week 10. Total body composition [two compartments: fat mass (FM) and fat-free mass (FFM)] was measured at baseline (time 0: “T0”), after 5 weeks (time 1: “T1”), and at week 10 (time 2: “T2”), at the end of the study, through Echo-MRI (EchoMRI^® 900, 3-in-1 composition analyzer Echo Medical Systems, Houston, TX, United States) (Wall et al., 2015 (link)). This type of analyzers deliver precise body composition measurements of fat, lean, free water, and total water masses in lived animals.

Body compositions of WT and Creb3l4 KO mice were determined using non-invasive quantitative magnetic resonance relaxometry on an EchoMRI-900 (Echo Medical Systems, Houston, TX, USA) at the Phenogenomic Research Center, Woo Jung BSC, Inc., Korea. Scans were then performed by placing animals in a thin-walled plastic cylinder (3-mm thick, 4.5-cm inner diameter, based on mouse body weight), which was inserted into a mouse cylindrical sensory antenna, the A100, to limit movement. All quantitative magnetic resonance measurements were made during the light phase (0700–1900 hours). The accumulation factor used was set for extra-high precision ( × 3), resulting in a scan time of approximately 2.5 min.

DIO-F344 rats (42 weeks old, HFD for 37 weeks, baseline BW 480 ± 18 g) were randomized based on body fat mass measured by an EchoMRI-900 (Echo Medical Systems), BW and food intake, and were administered drugs once daily for 4 weeks (N = 6–7 per group, total N = 41). After the repeated doses, blood samples were collected from the tail vein at the indicated time points after a single further dosing and plasma total GLP-1 levels were measured as described above. Ffar1^-/- and wild-type mice (23 weeks old, baseline BW 45 ± 5 g for wild and 44 ± 5 g for Ffar1-/-) fed HFD for 16 weeks were randomized based on their BW and food intake, and were administered drugs once daily for 3 days (N = 5–6 per group, total N = 17 in each genotype). Blood samples were collected from the tail vein before and 1 h after the 4^th dosing and plasma total GLP-1 and GIP levels were measured as described above.