Korean Diabetes Journal 2004;28(4):273-283.
Published online August 1, 2004.
Increased ROS Production by High Glucose in Cultured Mouse Insulinoma Cell Line.
Jin Hwa Kim, Jung In Kim, Dong Hyun Choi, Young Uk Seo, Young Dae Kim, Jong Chan Oh, Beom Ju Lee, Keo Woon Park, Sang Yong Kim, Hak Yoen Bae, Byoung Rai Lee
1Division of Endocrinology, Department of Internal Medicine, College of Medicine, Chosun University, Gwangju, Korea.
2Department of Biochemistry, College of Medicine, Chosun University, Gwangju, Korea.
Abstract
BACKGROUND
To investigate the effects of high glucose on oxidative stress of islet beta cells, the effects of high glucose on antioxidant enzymes, the production of free reactive substances and paraquat-induced cytotoxicity were examined in cultured mouse insulinoma cells (MIN6N8a). METHODS: The MIN6N8a cell line (Obtained from Diabetic research center, Univer sity of Calgary, Canada) was maintained in RPMI1640 medium supplemented with 10% fetal bovine serum, at 37degrees C under an atmosphere of 5% CO2 and 100 % humidity. The MIN6N8a cells were cultured in high glucose (22.4 mM) and normogl-ucose (5.6 mM) containing RPMI1640 media, for 1-6 days, and the superoxide dismutase (SOD), catalase and glutathione peroxidase (GSHPx) activities in the MIN6N8a cells assayed. The levels of reactive oxygen species in the MIN6N8a cells was determined using dihydroethidium (DHE). Paraquatinduced cytotoxicity was determined using the 3-[4,5-dimethylthiazol-2-yl]-2, 5diphenyl tetraz olium bromide (MTT) method. RESULTS: No difference was observed catalase in the catalase and GSHPx activities in MIN6N8a cells between the high glucose (22.4 mM) and normoglucose (5.6 mM) groups. The CuZn-SOD activity of MIN6N8a cells was decreased by 32% in the high glucose (25.4 mM) medium compared to normoglucose (5.6 mM) medium, while the Mn-SOD activity was increased by 24% in high glucose group. The paraquat induced cytotoxicity of MIN6N8a cells was potentiated by high glucose. and the amount of DHE oxidation increased. CONCLUSION: The Oxidative stress in MIN6N8a cells was increased by high glucose as a resulted of the decreased CuZn-SOD activity and increased production of reactive oxygen species. Increased oxidative stress in MIN6N8a cells by high glucose may play some roles in the pathogenesis of diabetes.
Key Words: CuZn-SOD, MIN6N8a cells, High glucose
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