SUPEROXIDE, NITRIC-OXIDE, PEROXYNITRITE AND CYTOKINE COMBINATIONS ALL-CAUSE FUNCTIONAL IMPAIRMENT AND MORPHOLOGICAL-CHANGES IN RAT ISLETS OF LANGERHANS AND INSULIN-SECRETING CELL-LINES, BUT DICTATE CELL-DEATH BY DIFFERENT MECHANISMS

We have shown that nitric oxide treatment for 30-90 min causes inhibit ion of insulin secretion, DNA damage and disturbs sub-cellular organiz ation in rat and human islets of Langerhans and HIT-T15 cells. Here ra t islets and beta-cell lines were treated with various free radical ge nerating systems S-nitrosoglutathione (nitric oxide), xanthine oxidase plus hypoxanthine (reactive oxygen species), 3-morpholinosydnonimine (nitric oxide, superoxide, peroxynitrite, hydrogen peroxide) and perox ynitrite and their effects over 4 h to 3 days compared with those of t he cytokine combination interleukin-1 beta, tumour necrosis factor-alp ha and interferon-gamma. End points examined were de novo protein synt hesis, cellular reducing capacity, morphological changes and apoptosis by acridine orange cytochemistry, DNA gel electrophoresis and electro n microscopy. Treatment (24-72 h) with nitric oxide, superoxide, perox ynitrite or combined cytokines differentially decreased redox function and inhibited protein synthesis in rat islets of Langerhans and in in sulin-containing cell lines; cytokine effects were arginine and nitric oxide dependent, Peroxynitrite gave rare apoptosis in HIT-T15 cells a nd superoxide gave none in any cell type, but caused the most beta cel l-specific damage in islets. S-nitrosoglutathione was the most effecti ve agent at causing DNA laddering or chromatin margination characteris tic of apoptotic cell death in insulin-containing cells, Cytokine-indu ced apoptosis was observed specifically in islet beta cells, combined cytokine effects on islet function and death most resembled those of t he mixed radical donor SIN-1.