The Role of Glutathione in Nitric Oxide Induced Cytoprotection

• Published in: Japanese Journal of Pharmacology 1997, Vol.75 (suppl), p.96-96
• Main Authors: Ponte, Jose F., Huot, Anne E.
• Format: Article
• Language: English
• ISSN: 0021-5198; 1347-3506
• DOI: 10.1016/S0021-5198(19)41778-2

Glutathione (GSH) is an essential tripeptide important in providing cells with their reducing milieu and protection against oxidative damage. We previously reported that priming L1210 cells with 200 μM Sodium Nitroprusside (SNP) for 24 hours significantly inhibited apoptosis seen in unprimed cells after exposure to 1000 μM SNP for 12 hours. The current investigation assessed the role of GSH in nitric oxide (NO) induced cytoprotection. Unprimed cells exposed to 1000 μM SNP had a 60% reduction, and cells exposed to 200 μM SNP had a 69% increase, in total GSH compared to control cells. Priming cells with 200 μM SNP prior to exposure with 1000 μM SNP not only inhibited this reduction, but increased total GSH levels 138% above levels of control cells. This suggests that priming prepares the cell for a quicker and heightened response to oxidative damage. The level of GSH returns to that of control cells after 48 hours. The correlation of this with NO induced cytoprotection was assessed by priming cells for 24 hours and delaying the addition of 1000 μM SNP for 12 or 24 hours. We assessed viability with the MTT assay and apoptosis with the DNA fragmentation ELISA assay. There is a direct correlation between GSH concentration and cytoprotection. When the lethal insult is delayed for 24 hours post priming, the GSH falls from 138% to 50% over the control and cytoprotection was ablated 47% as measured with the MTT assay and 69% with the DNA fragmentation assay. To determine if GSH was essential to cytoprotection, we primed with both 200 μM SNP and 50 μM L-Buthionine-Sulfoximine (BSO), an inhibitor of GSH synthesis which completely depletes GSH, for 24 hours and then exposed the cells to 1000 μM SNP. Despite the fact that GSH is depleted, these cells are still resistant to induction of cell death by lethal dose NO. Further, the duration of protection is similar to that of cells primed with only 200 μM SNP. Thus, while GSH may be important in NO induced cytoprotection it is not sufficient, suggesting that NO may induce multiple cytoprotective response modifications.