Development of autoimmune diabetes in NOD mice is associated with the formation of peroxynitrite in pancreatic islet β-cells

Peroxynitrite (ONOO^-) is a highly reactive oxidant species produced by the reaction of the free radicals superoxide (O₂^.-) and nitric oxide (NO^.). Here we report a marked increase in nitrotyrosine (NT), a marker of peroxynitrite, in islet cells from NOD mice developing spontaneous autoimmune diabetes. By using specific antibodies and immunohistochemical methods, we found that NT-positive cells were significantly more frequent in islets from acutely diabetic NOD mice (22 ± 6%) than in islets from normoglycemic NOD mice (7 ± 1%) and control BALB/c mice (2 ± 1%). The NT⁺ cells in islets were identified to be macrophages and also β-cells. Most of the β-cells in islets from acutely diabetic NOD mice were NT⁺ (73 ± 8%), whereas significantly fewer β-cells were NT⁺ in islets from normoglycemic NOD mice (18 ± 4%) and BALB/c mice (5 ± 1%). Also, the percentage of β-cells in islets from NOD mice (normoglycemic and diabetic) correlated inversely with the frequency of NT⁺ β-cells. This study demonstrates for the first time that peroxynitrite, a reaction product of superoxide and nitric oxide, is formed in pancreatic islet β-cells of NOD mice developing autoimmune diabetes. This suggests that both oxygen and nitrogen free radicals contribute to β-cell destruction in IDDM via peroxynitrite formation in the islet β-cells.