Protection against cigarette smoke-induced damage to intact transformed rabbit corneal cells by N-acetyl-L-cysteine

Abstract

In order to assess cigarette smoke-induced oxidative damage to intact cells, an assay was developed to measure cell detachment and protection. Due to the complex nature of cigarette smoke, which contains molecules that can interfere with conventional spectrophotometric and fluorometric biochemical assays, transformed rabbit corneal cells were radiolabeled with tritiated thymidine and then subjected to direct stream smoke. As a result, cell damage in response to the smoke from only two cigarettes could be measured in a time-dependent manner. When cells were prelabeled with N-acetyl-L-cysteine (NAC), a substrate for glutathione synthesis, a significant reduction in damage was measured. Additionally, when buthionine sulfoximine (BSO), an inhibitor of glutathione synthesis, was incubated with cells, a reduction in the effectiveness of NAC was observed, although NAC still retained some activity. Furthermore, vitamin E conferred no protection to cells in this system nor was NAC active in a separate assay that appears to favor peroxyl radical generation. From these results we conclude that cigarette smoke damage can easily be determined at the cellular level with this technique and that NAC acted to prevent this damage in two ways: first, as glutathione precursor and, secondly, as an antioxidant capable of scavenging non-peroxyl radicals.