Nitric oxide-induced cytotoxicity: involvement of cellular resistance to oxidative stress and the role of glutathione in protection.
A series of experiments were designed to examine the potential cytotoxicity of nitric oxide (NO), or reactive species derived from NO, in HA1 fibroblasts and H2O2-resistant variants of this cell line, designated OC14 cells. A 1-h exposure at 37 degrees C to a 1.7 mM bolus dose of NO, prepared in N2-gassed medium, significantly reduced clonogenic survival in the HA1 fibroblasts line to 60% of control cells treated with N2-gassed medium alone. The OC14 cells were found to be completely resistant (100% survival) to NO-mediated injury in comparable experiments. A second set of experiments was designed to determine the role of the intracellular antioxidant, glutathione, in protection against NO-mediated injury. Depletion of total glutathione resulted in a significant reduction in HA1 and OC14 clonogenic survival to 8% and 50% when compared with respective control cells. The effect of total glutathione depletion on NO-initiated toxicity in HA1 cells was dose- and cell-density dependent and was observed to occur within 5 min of exposure to NO. Further evidence of cytotoxicity was demonstrated by loss of trypan blue dye exclusion properties in glutathione-depleted HA1 cells after NO exposure. Other experiments demonstrated that nitrate and nitrite exposure produced no cytotoxicity in glutathione-depleted HA1 cells and that coincubation of NO-saturated medium with oxyhemoglobin inhibited NO-induced cytotoxicity in glutathione-depleted HA1 cells.(ABSTRACT TRUNCATED AT 250 WORDS)
Walker
M W
MW
University of Virginia Health Sciences Center, Department of Pediatrics, Charlottesville 22908.
Kinter
M T
MT
Roberts
R J
RJ
Spitz
D R
DR
D000818:Animals; D019328:Buthionine Sulfoximine; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects* / Q000502:physiology; D003114:Colony-Forming Units Assay; D006224:Cricetinae; D005978:Glutathione / Q000378:metabolism*; D008717:Methionine Sulfoximine / Q000031:analogs & derivatives / Q000494:pharmacology; D008954:Models, Biological; D009569:Nitric Oxide / Q000633:toxicity*; D018384:Oxidative Stress / Q000502:physiology*
D008717:Methionine Sulfoximine; D009569:Nitric Oxide; D019328:Buthionine Sulfoximine; D005978:Glutathione