Blood glucose monitoring system

The mice in each group fasted for 12 h before the blood glucose test. Blood samples (10 μL) were drawn from the tail vein and the glucose concentrations were measured using the blood glucose monitoring system (Roche). Serum insulin concentrations were measured according to the mouse insulin enzyme-linked immunoassay (ELISA) kit instructions with the standard preparation (R&D Systems, Suzhou, China). In brief, blood samples (approximately 800 μL) were taken from the orbit and centrifuged at 3,000 rpm for 15 min at 4 °C. The supernatants were then collected. The sample (10 μL) with the diluent (40 μL) was added to the pre-coated plate, and 100 μL HRP-conjugate reagent was then mixed in each well and incubated for 60 min at 37 °C in the dark. After washing, chromogen was added to each well, which was then incubated at 37 °C in the dark for 15 min. Finally, the stop solution was added and the resulting color was recorded at 450 nm using a microplate absorbance reader (CORONA SH-1000 Lab, Japan).

Six-week-old male C57BL/6J mice (from the Animal Center of Xi'an Jiaotong University, Xi'an, China) were allowed to acclimatize to their environment for 1 week. All experimental procedures were carried out in accordance with the guidelines of Xi'an Jiaotong University on Animal Care. All chemicals and reagents were purchased from Sigma (Oakville, ON, Canada), unless otherwise stated. The mice were randomly divided into 2 groups as follows: i) the control group (Cont, n=10); ii) the STZ group (n=10), where the animals were rendered hyperglycemic by an intraperitoneal injection of STZ, dissolved in citrate buffer (0.1 M at pH 4.2), at 35 mg/kg body weight for 5 consecutive days. The mice in the control group was treated with citrate buffer only. All the mice were sacrificed by cardiac exsanguination under light ether anesthesia 8 weeks after the STZ injection. The fasting blood glucose (FBG) levels were measured using a blood glucose monitoring system (Roche Diagnostics, Manheim, Germany), and the body weight of the mice was recorded every 2 weeks during the experimental period.