Dvi 1650

At the end of the experiments, the fasting plasma glucose (FPG), 24 h urinary albumin excretion, hemoglobin A1c (HbA_1C), urinary creatinine (Ucr), serum creatinine (Scr), Ccr = Ucr/ScrxV (V:ml/min, urine per minute), and the ratio of kidney weight to bodyweight and serum AGEs was measured for each rat. Scr and Ucr were measured using an automatic biochemical analysis instrument (DVI 1650; Bayer AG, Leverkusen, Germany). HbA_1C was measured by high-pressure liquid phase methods (ADAMS A1c HA-8180; Arkray, Inc., Kyoto, Japan). The level of AGEs in serum was measured by the enzyme-linked immunosorbent assay (ELISA) method (19 (link)) (Hitachi 850; Hitachi, Tokyo, Japan). To collect urine samples, rats were placed in individual metabolic cages for 24 h prior to sacrificing. ELISA (Nephrat II; Exocell, Philadelphia, PA, USA) was used to determine 24 h urinary albumin excretion levels.

After 24 weeks, the rats underwent the analysis of renal function. The serum fasting plasma glucose (FPG) level, serum creatinine and urinary creatinine were detected by an automatic biochemical analysis instrument (DVI 1650; Bayer, Leverkusen, Germany). The level of glycosylated hemoglobin A_1c (HbA_1c) was measured using the high performance liquid chromatography method (HA-8160 HbA_1c Analyzer; Menarini Diagnostics, Florence, Italy). To measure advanced glycation end products (AGEs) in serum, a fluorescence spectrophotometer (Hitachi F-2500; Hitachi, Tokyo, Japan) was used to determine specific fluorescence by measuring 440 nm emissions after excitation at 370 nm [10 (link)]. To collect urine samples, the rats had free access to tap water but were deprived of food for 24 h in individual cages before being killed. The urine samples were centrifuged (1500 × g, 15 min) and supernatants were stored in poly-ethylene containers at –20°C. Urinary albumin concentrations in fresh urine samples were determined by enzyme-linked immunosorbent assay (ELISA, Nephrat II; Exocell, Philadelphia, PA), and this value was multiplied by urine volume to get the total 24-h urine albumin excretion (mg).