OXIDATIVE STRESS MEASUREMENT BY IN-VIVO ELECTRON-SPIN-RESONANCE SPECTROSCOPY IN RATS WITH STREPTOZOTOCIN-INDUCED DIABETES

Enhanced oxidative stress in diabetic patients may contribute to the p athogenesis of diabetic angiopathy. We have recently developed a metho d to determine the electron spin resonance (ESR, electron paramagnetic resonance; EPR) of reactive oxygen species and free radicals in vivo, using the nitroxide derivative, carbamoyl-PROXYL as a probe. In this study, diabetes was induced in Wistar rats by streptozotocin (STZ) inj ection (65 mg/kg. body weight, intravenously). Two, 4, and a weeks lat er, the animals received carbamoyl-PROXYL (300 nmol/g, intravenously), and ESR was measured at the upper abdominal level at a frequency of 3 00 MHz. The intensity of the carbamoyl-PROXYL ESR signal decreased gra dually after the injection, and the spin clearance fate was determined over the first 5 min. At all time points, the spin clearance rate was significantly greater in the diabetic rats than in control rats. More over, the spin clearance rate in the diabetic rats was significantly c orrelated with urinary malondialdehyde (MDA) levels, which serve as a marker for lipid peroxidation. Daily treatment with 4 units neutral pr otamin Hagedorn (NPH) insulin for 4 weeks reduced the spin clearance r ate in the diabetic rats. Simultaneous injection of carbamoyl-PROXYL a nd superoxide dismutase reduced the spin clearance rate in the diabeti c rats in a dose-dependent manner. Injection of the antioxidant a-toco pherol (40 mg/kg, intraperitoneally) for 2 weeks restored the spin cle arance rate in the diabetic rats without concomitant glycaemic restora tion. These results suggest that a diabetic state enhances the generat ion of free radicals in vivo, and that both glycaemic control and anti oxidant treatment can reduce this oxidative stress. Non-invasive in vi vo ESR measurement may be useful for evaluating oxidative stress in di abetes.