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Diabetes 51:1076-1082, 2002
© 2002 by the American Diabetes Association, Inc.

Acute Hyperglycemia Induces Nitrotyrosine Formation and Apoptosis in Perfused Heart From Rat

Antonio Ceriello1, Lisa Quagliaro2, Michele D’Amico3, Clara Di Filippo3, Raffaele Marfella4, Francesco Nappo4, Liberato Berrino3, Francesco Rossi3, and Dario Giugliano4

1 Department of Pathology and Medicine, Experimental and Clinical, University of Udine, Udine, Italy
2 Morpurgo-Hofmann Research Laboratory on Aging, Udine, Italy
3 Department of Experimental Medicine, Sezione di Farmacologia L. Donatelli, Second University of Naples, Naples, Italy
4 Department of Geriatric and Metabolic Diseases, Second University of Naples, Naples, Italy

This study investigated coronary perfusion pressure, nitric oxide (NO) and superoxide production, nitrotyrosine (NT) formation, and cardiac cell apoptosis in isolated hearts perfused with high glucose concentration. Coronary perfusion pressure; NO and superoxide anion generation; immunostaining for NT, inducible NO synthase (iNOS), and the constitutive type of NO synthase (NOS) eNOS; iNOS and eNOS mRNA expression by Western blot and RT-PCR; and apoptosis of cardiac cells were studied in hearts perfused for 2 h with solutions containing D-glucose at a concentration of 11.1 mmol/l (control), D-glucose at the concentration of 33.3 mmol/l (high glucose), or D-glucose (33.3 mmol/l) plus glutathione (0.3 mmol/l). Perfusion of isolated hearts in conditions of high glucose concentration caused a significant increase of coronary perfusion pressure (P < 0.001) and an increase of both NO and superoxide generation. However, superoxide production was 300% higher than baseline, whereas NO production was 40% higher (P < 0.001 for both). This effect was accompanied by the formation of NT, and an increase of iNOS expression. eNOS remained unchanged. At the end of the experiments, cardiac cell apoptosis was evident in hearts perfused with high glucose. The effects of high glucose were significantly prevented by glutathione. This study demonstrates that high glucose for 2 h is enough to increase iNOS gene expression and NO release in working rat hearts. Upregulation of iNOS and raised NO generation are accompanied by a marked concomitant increase of superoxide production, a condition favoring the production of peroxynitrite, a powerful pro-oxidant that can mediate the toxic effects of high glucose on heart by itself and/or via the formation of nitrotyrosine, as suggested by the detection of cell apoptosis.


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