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Abstract

Attenuation by Metallothionein of Early Cardiac Cell Death via Suppression of Mitochondrial Oxidative Stress Results in a Prevention of Diabetic Cardiomyopathy Lu Cai, Yuehui Wang, Guihua Zhou, Teresa Chen, Ye Song, Xiaokun Li, Y. James Kang Diabetes was induced by streptozotocin (STZ) in both cardiac-specific, metallothionein (MT)-overexpressing transgenic mice and wild-type (WT) controls. However, cardiac apoptosis along with a release of mitochondrial cytochrome c , as well as a depletion of mitochondrial reduced glutathione, was observed in the WT, but not in the cardiac-specific, MT-overexpressing transgenic (MT-TG) diabetic mice 7 to 21 days after STZ treatment. Morphological abnormalities, revealed by electron microscope examination, and fibrosis, shown by Sirius-red staining, were also observed in the WT, but not in the MT-TG diabetic mice 6 months after STZ treatment. These results suggest that cardiac apoptosis is a major early cellular event in diabetic hearts and that its attenuation results in a prevention of diabetic cardiomyopathy. We aimed to test whether attenuation of early-phase cardiac cell death can prevent diabetic cardiomyopathy. Our previous study showed that cardiac apoptosis as a major early cellular response to diabetes is induced by hyperglycemia-derived oxidative stress that activates a mitochondrial cytochrome c -mediated caspase-3 activation pathway. Metallothionein (MT) as a potent antioxidant prevents the development of diabetic cardiomyopathy. Diabetes was induced by a single dose of streptozotocin (STZ) (150 mg/kg) in cardiac-specific, metallothionein-overexpressing transgenic (MT-TG) mice and wild-type (WT) controls. On days 7, 14, and 21 after STZ treatment, cardiac apoptosis was examined by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assay and caspase-3 activation. Cardiomyopathy was evaluated by cardiac ultrastructure and fibrosis in the diabetic mice 6 months after STZ treatment. A significant reduction in diabetes-induced increases in TUNEL-positive cells, caspase-3 activation, and cytochrome c release from mitochondria was observed in the MT-TG mice as compared to WT mice. Cardiac protein nitration (3-nitrotyrosine [3-NT]) and lipid peroxidation were significantly increased, and there was an increase in mitochondrial oxidized glutathione and a decrease in mitochondrial reduced glutathione in the WT, but not in the MT-TG, diabetic mice. Double staining for cardiomyocytes with alpha sarcomeric actin and caspase-3 or 3-NT confirmed the cardiomyocyte-specific effects. A significant prevention of diabetic cardiomyopathy and enhanced animal survival were observed in the MT-TG diabetic mice as compared to WT diabetic mice. These results suggest that attenuation of early-phase cardiac cell death by MT results in a significant prevention of the development of diabetic cardiomyopathy. This process is mediated by MT suppression of mitochondrial oxidative stress.