Biological role of glutathione in nitric oxide-induced toxicity in cell culture and animal models

Glutathione (GSH) plays an important role in cellular defense response in many in vitro and in vivo models. Here we investigated its role in NO-induced toxicity in cell culture and mouse models. Wild-type (TK6) and p53-null (NH32) human lymphoblastoid cells were treated with NO at a steady-state concentration of 0.6 μM, similar to the level estimated to occur in inflamed tissues. In both cell types, GSH was depleted by this exposure in a dose- and time-dependent manner. Contrary to expectations, prior depletion of GSH by treatment with l-buthionine-SR-sulfoximine did not potentiate NO-induced cell killing or DNA deamination in TK6 cells. In activated RAW264.7 murine macrophages producing NO, intracellular GSH content did not change, although γ-glutamate-cysteine ligase was upregulated. NO overproduction in RcsX lymphoma-bearing SJL mice resulted in significantly elevated GSH levels in various organs. Administration of the NO synthase inhibitor N-methylarginine abolished the increase in GSH in these animals. Collectively, these data indicate a multifaceted and complex involvement of GSH in responses of cells and tissues to toxic levels of NO. NO treatment effectively depleted GSH levels in human lymphoblastoid cells, but this alteration was not a critical initiating factor for NO-mediated toxicity. Murine macrophages maintained GSH homeostasis when exposed to endogenously produced NO. In RcsX lymphoma-bearing mice, upregulation of de novo synthesis of GSH appeared to be a response to the toxic effects of NO.