Alphatrak glucometer

Tg(CAG-Z-miR-21-EGFP) [31 (link)] mice (backcrossed on a C57BL6/J background for >10 generations) were crossed with Ins1tm1(CreERT2)Thor [32 (link)] mice to generate Tg(βmiR-21) mice. Eight-week Tg(βmiR-21) mice and littermate controls (Cre+ and Cre-) were treated with 1 mg/day x 6 days intraperitoneal (IP) tamoxifen. IP glucose tolerance tests (IPGTTs) were performed 21-days post tamoxifen injection after overnight fast, using 2 g/kg body weight of glucose [28 (link)]. Tail vein glucose was determined (AlphaTRAK glucometer; Abbott) at 0, 10, 20, 30, 60, 90, and 120-min post injection. Insulin sensitivity was measured with IP insulin tolerance testing (IPITT) after a 2-h fast, using IP injection of 0.75 U/kg bodyweight of regular humulin-R insulin (Eli Lilly) [28 (link)]. Tail vein glucose was measured at 0, 10, 20, 30, and 60min post injection. Islets were isolated using collagenase 28 days after the initial tamoxifen injection [33 (link)].

To induce obesity, male C57BL/6J mice were fed a high-fat diet (HFD, D12492, 60% energy from fat; Research Diets Inc., New Brunswick, NJ, United States) for 12 weeks and then were injected intraperitoneally with lorcaserin (616202-92-7, Aladin, Shanghai, China) at a concentration of 2.5, 4 or 10 mg/kg once a day for 2 weeks based on previous researches (Burke et al., 2017 (link); Patel et al., 2020 (link)). Chronic injection of lorcaserin in diet-induced obesity (DIO) mice received intraperitoneal injections of lorcaserin at 2.5, 4 or 10 mg/kg as described above, whereas acute injection were only received a single intraperitoneal administration of lorcaserin. After fasting [16 h for intraperitoneal glucose tolerance test (IPGTT), 6 h for intraperitoneal insulin tolerance test (IPITT)], mice in acute and chronic treatment groups were all injected intraperitoneally with lorcaserin. 45 min later, 1.2 mg/kg glucose or 1 U/kg insulin were injected intraperitoneally into the mice, and blood was sampled from tail vein immediately prior to lorcaserin treatment, immediately prior to glucose or insulin injection, and 15, 30, 60, 90 and 120 min following glucose or insulin administration. Blood glucose was analyzed using an AlphaTRAK glucometer (Abbott Animal Health) and plasma insulin measurements with ELISA test kits (Ezassay Biotech Co., Ltd., Shenzhen City, China).

Body weights, fasting blood glucose (4 h fast), and glucose tolerance were assessed longitudinally; fasting blood glucose was measured using an AlphaTrak Glucometer (Abbott Laboratories, Abbott Park, IL, USA) and blood glucose was monitored over 2 h post-administration of a glucose bolus (1 g/kg body weight i.p.). Terminal glycated hemoglobin was measured by the Chemistry Core at the Michigan Diabetes Research Center. A Mouse Metabolic Phenotyping Center (MMPC; Vanderbilt University, Nashville, TN; University of Cincinnati, Cincinnati, OH, USA) determined fasting plasma insulin, triglycerides, and cholesterol and performed fast protein liquid chromatography (FPLC) analysis for cholesterol and triglyceride lipoprotein profiles. Plasma was diluted 1:200 and oxLDL was quantified via ELISA as per the manufacturer's instructions (#CSB-E07933 m, American Research Products, Waltham, MA, USA). Histomorphological assessment of epididymal white adipose tissue was determined in a blinded manner using MetaMorph Image Analysis Software v.7.7.0.8 (Molecular Devices, Sunnyvale, CA, USA) as previously described (Parlee et al., 2014 (link)).

Body weights were measured every 2 wk, from 12–24 wk. Four h fasting blood glucose (FBG) was measured from tail-blood using an AlphaTrak Glucometer (Abbott Laboratories, Abbott Park, IL). FBG was measured at 16, 20 and 24 wk. Terminal glycated hemoglobin (%HbA1c), and free plasma cholesterol and cholesterol esters were measured via ELISA, according to manufacturer’s protocols (Mouse HbA1c Assay Kit #80310, CrystalChem, Elk Grove Village, IL) (Cholesterol Assay Kit #ab65390, Abcam, Cambridge, MA). Total plasma cholesterol was measured by the Michigan Diabetes Research Center (University of Michigan, Ann Arbor, MI). Total plasma triglycerides were measured by the Mouse Metabolic Phenotyping Core (www.mmpc.org).