title: Cu/Zn superoxide dismutase (SOD1) mutations associated with familial amyotrophic lateral sclerosis (ALS) affect cellular free radical release in the presence of oxidative stress.
abstract: BACKGROUND
  The exact molecular mechanisms by which mutations in Cu/Zn superoxide dismutase (SOD1) cause motor neuron injury remain incompletely understood, though a body of evidence suggests that the mutant protein exerts a cell-specific toxic gain of function. The role of nitric oxide (NO) in SOD1-related motor neuron injury has been particularly controversial. Theoretically, there are arguments to suggest that NO may exert an important role in motor neuron injury, but there is relatively little direct experimental support for this hypothesis.
  
  
  OBJECTIVE
  The present study aimed to examine further the potential role for NO in motor neuron injury caused by mutant SOD1.
  
  
  METHODS
  We have generated a cellular model of familial amyotrophic lateral sclerosis (ALS) by stably transfecting NSC34 cells with one of three mutant forms of SOD1 (G93A, G37R, I113T). In the presence of mutant SOD1, NSC34 cells show increased cell death following oxidative stress induced by serum withdrawal. This model of motor neuron death involves cellular release of superoxide and NO radicals, which were directly measured in real time using microelectrode biosensors.
  
  
  RESULTS
  The expression of both normal and mutant SOD1 decreased the measured extracellular superoxide release, but had divergent effects on the measured release of NO. Normal SOD1 increased the measured NO release, whereas cells expressing mutant SOD1 released less NO. Co-administration of two different nitric oxide synthase inhibitors (L-NAME and L-N-methyl arginine) did show some neuroprotective effect, but this was only partial, and the effect was more marked using nuclear integrity as a measure of cell viability, rather than MTT conversion. Cells expressing mutant SOD1 were, however, more sensitive to toxicity induced by extrinsic exposure to NO, using a slow-release NO donor.
  
  
  CONCLUSIONS
  NO is likely to contribute to motor neuron injury, but this does not fully account for all the cellular toxic effects of mutant SOD1.
authors.affiliation: Academic Neurology Unit, University of Sheffield, UK.
authors.forename: Mark R
authors.initials: MR
authors.lastname: Cookson
authors.forename: Fiona M
authors.initials: FM
authors.lastname: Menzies
authors.forename: Philip
authors.initials: P
authors.lastname: Manning
authors.forename: Christopher J
authors.initials: CJ
authors.lastname: Eggett
authors.forename: Denise A
authors.initials: DA
authors.lastname: Figlewicz
authors.forename: Calum J
authors.initials: CJ
authors.lastname: McNeil
authors.forename: Pamela J
authors.initials: PJ
authors.lastname: Shaw
mesh_terms: D000690:Amyotrophic Lateral Sclerosis / Q000201:enzymology / Q000235:genetics; D000704:Analysis of Variance; D000818:Animals; D017304:Annexin A5 / Q000378:metabolism; D017209:Apoptosis / Q000187:drug effects; D015153:Blotting, Western; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D016895:Culture Media, Serum-Free / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005609:Free Radicals / Q000378:metabolism*; D006801:Humans; D007150:Immunohistochemistry; D009154:Mutation; D019331:NG-Nitroarginine Methyl Ester / Q000494:pharmacology; D009569:Nitric Oxide / Q000378:metabolism; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors / Q000378:metabolism; D018384:Oxidative Stress / Q000502:physiology*; D013482:Superoxide Dismutase / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D000072105:Superoxide Dismutase-1; D013481:Superoxides / Q000378:metabolism; D013997:Time Factors; D014162:Transfection
chemical_list: D017304:Annexin A5; D016895:Culture Media, Serum-Free; D004791:Enzyme Inhibitors; D005609:Free Radicals; C000606290:SOD1 protein, human; D013481:Superoxides; D009569:Nitric Oxide; D019001:Nitric Oxide Synthase; C109415:SOD1 G93A protein; D013482:Superoxide Dismutase; D000072105:Superoxide Dismutase-1; D019331:NG-Nitroarginine Methyl Ester