Mouse albumin

Six-week-old male C57BL/6 mice were obtained from Daehan Biolink (EumSeong, Korea). TGase2-null (Tgm2^−/−) mice (C57BL/6) [9 (link)] were kindly provided by Dr. Soo-Yul Kim (National Cancer Center, Goyang, Korea). All animal experiments conformed to the Guide for the Care and Use of Laboratory Animals from the National Institutes of Health and were approved by the Institutional Animal Care and Use Ethics Committee of Kangwon National University. Mice were maintained under pathogen-free conditions in a temperature-controlled room with a 12-h light/dark cycle. Diabetic mice were generated by a single daily intraperitoneal injection for 5 consecutive days of streptozotocin (50 mg/kg body weight; MilliporeSigma, Burlington, MA, USA) [34 (link)]. Mice with fasting blood glucose levels ≥19 mmol/L, polyuria, and glucosuria were considered diabetic.
Six weeks after streptozotocin injections, diabetic mice (n = 8) were treated with 10 mg/kg midazolam (Bukwang, Ansan, Korea) by subcutaneous injection to the nape of the neck every 3 days for 6 weeks (Figure 1a). Albumin and creatinine concentrations were determined using mouse albumin (Abcam, Cambridge, UK) and creatine (BioVision, Milpitas, CA, USA) ELISA kits.

Urine was collected from the bladder after sacrifice and stored at –80^°C. Mouse albumin (Abcam) and mouse NGAL (R&D Systems) ELISAs were performed according to the manufacturer’s protocols. Creatinine was measured using the Creatinine Parameter Assay Kit (R&D Systems). Albumin and NGAL levels were normalized to urinary creatinine levels.