Differential Mechanisms of Nitric Oxide- and Peroxynitrite-Induced Cell Death

• Published in: Molecular pharmacology 2004-10, Vol.66 (4), p.1043-1053
• Main Authors: Meij, Johanna T A, Haselton, Carole L, Hillman, Kristin L, Muralikrishnan, Dhanasekaran, Ebadi, Manuchair, Yu, Lei
• Format: Article
• Language: English
• Subjects: Animals; Cell Death; Cell Survival - drug effects; Humans; Hybridomas; Mice; Nitric Oxide - pharmacology; Nitric Oxide Donors - antagonists & inhibitors; Nitric Oxide Donors - pharmacology; Peroxynitrous Acid - pharmacology; Rats; Time Factors; Tumor Cells, Cultured
• ISSN: 0026-895X; 1521-0111
• DOI: 10.1124/mol.104.001354

Nitric oxide (NO) contributes to cellular degeneration in various disorders, particularly in the nervous system. NO targets cell proteins such as soluble guanylyl cyclase, but its detrimental effects are generally attributed to its reaction product with superoxide, peroxynitrite. To understand the mechanisms of NO-induced cell stress, we studied the effects of the NO donors diethylenetriamine and spermine NONOate and the peroxynitrite donor 5-amino-3-(4-morpholinyl)-1,2,3-oxadiazolium chloride (SIN-1) in SH-SY5Y and NG108-15 neuroblastoma cells. All three compounds induced a dose- and time-dependent decrease in viable cells, which was not blocked by the soluble guanylyl cyclase inhibitor 1H-[1,2,4]oxadiazolo[4,3- a ]quinoxalin-1-one. The two NONOates were ∼15-fold more potent in SH-SY5Y than in NG108-15 cells, whereas the EC 50 values of SIN-1 in SH-SY5Y and NG108-15 cells were in the same order. This led us to conclude that the mechanisms of NO and peroxynitrite did not converge. This was supported by our other findings. NONOates induced DNA fragmentation and an increase in cellular caspase-3 activity that preceded the gradual decline in cell viability. In contrast, SIN-1 induced a transient decline in ATP levels and a delayed loss of cell viability with no significant increase in caspase-3 activity or DNA laddering. Moreover, post-treatment with insulin inhibited caspase-3 activation and loss of cell viability in NONOate- but not in SIN-1–exposed cells. These findings suggest that NO is a potent toxin independent of peroxynitrite formation.