Metabolic cage

Glucose tolerance tests (GTTs) were performed on male and female Mkp-2^+/+ and Mkp-2^−/− mice fed either chow or HFD for 16 to 24 weeks. Mice were fasted overnight for 16 h followed by intraperitoneally (i.p.) injection of glucose (2 g/kg). Blood glucose levels were measured at time points 0, 15, 30, 60, 90, and 120 min. For insulin tolerance tests (ITTs), mice were fasted for 5 h and were injected (i.p.) with 0.75 mU/g human insulin (Humlin R; Elly and Company, Indianapolis, IN, USA). The blood glucose levels were measured at time points 0, 15, 30, 60, 90, and 120 minutes. Plasma insulin levels were measured using ultra-sensitivity mouse insulin Elisa kit (Crystal Chem, Elk Grove Village, IL, USA; Catalog #90080).
Conscious male chow-fed Mkp-2^+/+ and Mkp-2^−/− mice aged between 8 to 12 weeks old were individually housed under controlled temperature (23 °C) and lighting (12 h light, 12 h dark cycle, lights on at 0700 h) with free access to food and water. Mice were adapted for one week and kept in metabolic cages (Promethion, Sable Systems, Las Vegas, NV, USA) for one week, and the food and water intake, energy expenditure, respiratory exchange ratio (RER), and physical activity were measured. Data were analyzed using regression analysis using ANCOVA.

Rats in each group were housed in metabolic cages (Sable Systems International, America), respectively, and 24-h urine was collected to calculate the total amount of urine. Three mL of urine was centrifuged at 3000 rpm for 10 min at 4°C, and then, the supernatant was collected. Urine albumin level was measured with an automatic biochemical analyzer (Olympus corporation, Japan). Peripheral blood was collected from rats in each group via the tail veins and then was centrifuged at 3000 r/min for 20 min. After that, the serum in the upper layer was drawn to determine the level of the blood urea nitrogen (BUN), serum creatinine (SCr), and albumin (ALB).

8–11-week-old male Sprague–Dawley (SD) rats were purchased from Charles River Laboratories (Wilmington, MA). Rats were cohoused (2 rats/ cage) and maintained on a 12-h light/dark cycle with ad libitum access to chow (Teklad Diet #2018), high fat diet (HF, Research Diets D42151), or high fat diet with oligofructose (HF-OFS, Research Diets D19112708; BENEO, Orafti P95), which is macronutrient and calorie matched to the HF diet with the addition of 10% OFS (Supplementary Table 1). Male rats were chosen, as they readily develop obesity when placed on a HF diet and do not have an estrus cycle, which can influence food intake [30 (link)–32 (link)]. Body composition was measured by quantitative magnetic resonance imaging using EchoMRI-1100 (EchoMRI, Houston, Texas), and rats were placed in metabolic cages (Sable Systems International, Las Vegas, NV, USA) for up to 10 days to record food intake and metabolism.