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Two distinct mechanisms of nitric oxide-mediated neuronal cell death show thiol dependency
1Stokes Research Institute, Children's Hospital of Philadelphia, University of Pennsylvania 19104, USA.
Abstract:
To better understand the mechanism(s) underlying nitric oxide (. NO)-mediated toxicity, in the presence and absence of concomitant oxidant exposure, postmitotic terminally differentiated NT2N cells, which are incapable of producing. NO, were exposed to PAPA-NONOate (PAPA/NO) and 3-morpholinosydnonimine (SIN-1). Exposure to SIN-1, which generated peroxynitrite in the range of 25-750 nM/min, produced a concentration- and time-dependent delayed cell death. In contrast, a critical threshold concentration (>440 nM/min) was required for. NO to produce significant cell injury. Examination of cells by electron microscopy shows a largely necrotic injury after peroxynitrite exposure but mainly apoptotic-like morphology after. NO exposure. Cellular levels of reduced thiols correlated with cell death, and pretreatment with N-acetylcysteine (NAC) fully protected from cell death in either PAPA/NO or SIN-1 exposure. NAC given within the first 3 h posttreatment further delayed cell death and increased the intracellular thiol level in SIN-1 but not. NO-exposed cells. Cell injury from. NO was independent of cGMP, caspases, and superoxide or peroxynitrite formation. Overall, exposure of non-. NO-producing cells to. NO or peroxynitrite results in delayed cell death, which, although occurring by different mechanisms, appears to be mediated by the loss of intracellular redox balance.
Insights
Nitric oxide (NO) and peroxynitrite cause delayed cell death through redox imbalance. N-acetylcysteine (NAC) protects cells, highlighting the role of reduced thiols in NO-mediated toxicity.
Area of Science:
- Neuroscience
- Toxicology
- Cell Biology
Background:
- Nitric oxide (NO) is a signaling molecule with complex roles in cellular function and toxicity.
- Understanding NO-mediated toxicity is crucial, especially when combined with oxidant exposure.
- Terminally differentiated NT2N cells offer a model to study NO toxicity without endogenous NO production.
Purpose of the Study:
- To elucidate the mechanisms of nitric oxide (NO)-mediated cell toxicity.
- To compare the toxic effects of NO and peroxynitrite on neuronal cells.
- To investigate the role of intracellular thiols and potential protective agents.
Main Methods:
- Exposure of postmitotic NT2N cells to PAPA-NONOate (NO donor) and SIN-1 (peroxynitrite generator).
- Assessment of cell death morphology (necrosis vs. apoptosis) using electron microscopy.
- Quantification of cellular reduced thiols and evaluation of N-acetylcysteine (NAC) protection.
Main Results:
- Peroxynitrite induced concentration- and time-dependent delayed cell death with necrotic morphology.
- NO required a critical threshold concentration to cause significant cell injury, primarily apoptotic-like.
- N-acetylcysteine (NAC) provided full protection against both NO and peroxynitrite, indicating redox balance disruption.
Conclusions:
- Both NO and peroxynitrite induce delayed cell death in neuronal cells via distinct mechanisms.
- Loss of intracellular redox balance, indicated by reduced thiol levels, is a key mediator of NO and peroxynitrite toxicity.
- NAC demonstrates significant protective potential against NO- and peroxynitrite-induced cell death.