Cigarette smoke irreversibly modifies glutathione in airway epithelial cells

Abstract

In patients with chronic obstructive pulmonary disease (COPD), an imbalance between oxidants and antioxidants is acknowledged to result in disease development and progression. Cigarette smoke (CS) is known to deplete total glutathione (GSH + GSSG) in the airways. We hypothesized that components in the gaseous phase of CS may irreversibly react with GSH to form GSH derivatives that cannot be reduced (GSX), thereby causing this depletion. To understand this phenomenon, we investigated the effect of CS on GSH metabolism and identified the actual GSX compounds. CS and H(2)O(2) (control) deplete reduced GSH in solution [Delta = -54.1 +/- 1.7 microM (P < 0.01) and -39.8 +/- 0.9 microM (P < 0.01), respectively]. However, a significant decrease of GSH + GSSG was observed after CS (Delta = -75.1 +/- 7.6 microM, P < 0.01), but not after H(2)O(2). Exposure of A549 cells and primary bronchial epithelial cells to CS decreased free sulfhydryl (-SH) groups (Delta = -64.2 +/- 14.6 microM/mg protein, P < 0.05) and irreversibly modified GSH + GSSG (Delta = -17.7 +/- 1.9 microM, P < 0.01) compared with nonexposed cells or H(2)O(2) control. Mass spectrometry (MS) showed that GSH was modified to glutathione-aldehyde derivatives. Further MS identification showed that GSH was bound to acrolein and crotonaldehyde and another, yet to be identified, structure. Our data show that CS does not oxidize GSH to GSSG but, rather, reacts to nonreducible glutathione-aldehyde derivatives, thereby depleting the total available GSH pool.

Intracellular Metabolism of α,β-Unsaturated Carbonyl Compounds, Acrolein, Crotonaldehyde and Methyl Vinyl Ketone, Active Toxicants in Cigarette Smoke: Participation of Glutathione Conjugation Ability and Aldehyde–Ketone Sensitive Reductase Activity

“…4,22,23) In addition, it has been reported that a substantial amount of GSH in epithelial cells exposed to the gaseous phase of cigarette smoke is irreversibly modified to the GSH-ACR adduct (m/z 364) and GSH-CA adduct (m/z 378) by direct infusion mass spectra of the cell lysate, but the GSH-MVK adduct (m/z 378) and GSH-CA adduct are not separated. 7) Furthermore, it has been reported that the GSH-ACR and GSH-CA adducts can be detected in airway epithelial cells exposed to cigarette smoke, although the rela- tive yields were limited.…”

“…[2][3][4] Among them, the α,β-unsaturated aldehydes acrolein (ACR) and crotonaldehyde (CA), abundantly present in cigarette smoke, have been often reported to strongly react with protein and peptide thiol groups. [5][6][7][8][9][10] In particular, glutathione (GSH), a tripeptide containing a thiol group, is an important antioxidant and essential cofactor for antioxidant enzyme glutathione peroxidase. ACR and CA can chemically or enzymatically react with GSH to form their corresponding Michael adducts.…”

“…Colombo et al 12) demonstrated by MS analyses that exposure of human gingival fibroblasts to cigarette smoke rapidly depletes intracellular GSH, and also that the total GSH consumption is due to the release of GSH-ACR and GSH-CA adducts from the cells. Furthermore, Van der Toorn et al 7) demonstrated by directinfusion MS combined with enzymatic assays that a substantial amount of GSH in epithelial cells is irreversibly modified to GSH-ACR and GSH-CA derivatives, thereby depleting the total available GSH pool. Consequently, α,β-unsaturated aldehydes as well as oxidants are thought to mediate oxidative stress, which has been implicated in the pathogenesis of smoking-related diseases.…”

Carbonyl Compounds in the Gas Phase of Cigarette Mainstream Smoke and Their Pharmacological Properties

“…[12][13][14] In contrast to ROS, carbonyl compounds, which are relatively stable, extremely hydrophilic, and chemically reactive, pass through the airway epithelial barrier, enter the systemic circulation, and increase systemic oxidative stress and oxidative damage, contributing to the development of chronic diseases such as atherosclerosis, diabetes, and chronic obstructive pulmonary disease (COPD). [15][16][17] In addition, carbonyl compounds can induce carbonylation of proteins, which is an irreversible oxidative damage often associated with loss of protein function and accumulation of damaged or unfolded proteins. 18) Cigarette smoke may induce endogenous ROS production via continuous activation of reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX) in inflammatory cells including macrophages, neutrophils, and epithelial cells, leading to chronic inflammatory conditions such as COPD.…”

Pathogenic Mechanisms in Emphysema: From Protease Anti-Protease Imbalance to Apoptosis

“…The gaseous phase of cigarette smoke contains many reactive oxidants such as superoxide anion, nitric oxides and peroxynitrites, as reviewed recently (MacNee b2005; Lin & Thomas 2010). Oxidants and free radicals inhaled in tobacco smoke, can damage airway epithelial cells, and impair antioxidants, such as glutathione to non-reducible glutathione-aldehyde derivatives (van Der Toorn et al, 2007). Oxidants from tobacco smoke may also inactivate antiproteases, predisposing to a protease-antiprotease imbalance from the increased numbers of neutrophils and macrophages in smokers' lungs.…”