Abstract
Background and Objectives: Exhaled nitric oxide (NO) is decreased by smoking while oxides of nitrogen such as nitrites/nitrates (NOx) are increased. It was hypothesised that in vitro cigarette smoke extract (CSE) would either inhibit NO generation by increasing the NO synthase inhibitor, NG, NG-dimethyl-L-arginine (ADMA) or increase NOx levels via an oxidation pathway, which in turn could be inhibited by the antioxidant N-acetylcysteine NAC. Methods: Transformed airway cells (A549) were cultured with control medium, 1.0% CSE in culture medium, or 0.8 mM NAC with 1.0% CSE. Baseline L-arginine, NOx and ADMA levels were measured in the media. Conditioned media were then sampled at 1hour, 6 hours, 24 hours, 48 hours and 72 hours after incubation. Results: CSE induced significantly higher NOx levels (mean (SD) peak increase of 135.8 (126.6)% after incubation for 6 hours (p x which was partially reversed by NAC pre-treatment. ADMA levels were also increased after CSE exposure, suggesting that it activates the NO pathway via oxidative-stress while inhibition probably occurs via both ADMA and NOS.
Highlights
Smoking is known to decrease nitric oxide (NO) production but the mechanism is unknown [1]
Cell numbers increased after 24 hours incubation in all three groups, but there was slower growth in the cells exposed to cigarette smoke extract (CSE) conditioned media when compared with control cells, an effect which was reversed by NAC pre-treatment
ADMA has been recognised as an endogenous analogue of L-arginine, and it competitively inhibits nitric oxide synthase (NOS) [4]
Summary
Smoking is known to decrease nitric oxide (NO) production but the mechanism is unknown [1]. It has been established that these analogues inhibit the activity of nitric oxide synthase (NOS), presumably by competitive antagonism at the binding site for L-arginine [2,3,4]. These molecules are capable of decreasing NOS-related nitric oxide (NO) production, and have been proved to participate in a variety of physiological and pathological processes [5,6]. It was hypothesised that in vitro cigarette smoke extract (CSE) would either inhibit NO generation by increasing the NO synthase inhibitor, NG, NG-dimethyl-L-arginine (ADMA) or increase NOx levels via an oxidation pathway, which in turn could be inhibited by the antioxidant N-acetylcysteine NAC. ADMA levels were increased after CSE exposure, suggesting that it activates the NO pathway via oxidative-stress while inhibition probably occurs via both ADMA and NOS
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