α,β-Unsaturated Aldehydes in Cigarette Smoke Release Inflammatory Mediators from Human Macrophages

Abstract

Smoking cigarettes is the major risk factor for chronic obstructive pulmonary disease (COPD). COPD is a condition associated with chronic pulmonary inflammation, characterized by macrophage activation, neutrophil recruitment, and cell injury. Many substances contained in cigarette smoke, including reactive oxygen species (ROS), have been proposed to be responsible for the inflammatory process of COPD. However, this issue remains unsettled. By gas chromatography/mass spectrometry (GC/MS) we show that acrolein and crotonaldehyde, two alpha,beta-unsaturated aldehydes, are contained in aqueous cigarette smoke extract (CSE) at micromolar concentrations and mimic CSE in evoking the release of the neutrophil chemoattractant IL-8 and of the pleiotropic inflammatory cytokine TNF-alpha from the human macrophagic cell line U937. In addition, acrolein (10-30 microM) released IL-8 also from cultured human alveolar macrophages and THP-1 macrophagic cells. 4-hydroxy-2-nonenal (30-100 microM), an endogenous alpha,beta-unsaturated aldehyde that is abundant in lungs of patients with COPD, stimulated the release of IL-8 from U937 cells, whereas the saturated aldehyde, acetaldehyde, was ineffective. CSE-evoked IL-8 release was remarkably (> 80%) inhibited by N-acetyl-cysteine (0.1-3 mM) or glutathione monoethyl ester (1-3 mM). Both compounds, by forming covalent adducts (Michael adducts), completely removed unsaturated aldehydes from CSE. Our data demonstrate that alpha,beta-unsaturated aldehydes are major mediators of cigarette smoke-induced macrophage activation, and suggest that they might contribute to pulmonary inflammation associated with cigarette smoke.

Cigarette smoke and α,β‐unsaturated aldehydes elicit VEGF release through the p38 MAPK pathway in human airway smooth muscle cells and lung fibroblasts

“…Interestingly, augmented VEGF release in response to cigarette smoke does not appear to be common to all pulmonary cell types as our results indicate that epithelial (SAEC) cultures exposed to CSE do not up-regulate VEGF release, a finding consistent with previous reports (St-Laurent et al, 2009;Thaikoottathil et al, 2009). Because acrolein or crotonaldehyde are two a,b-unsaturated aldehydes contained in CSE at concentrations (Facchinetti et al, 2007) capable of eliciting VEGF release, and the effect of CSE was counteracted by the a,b-unsaturated aldehyde scavenger NAC, we conclude that acrolein, crotonaldehyde and possibly other electrophilic compounds, are, at least in part, the smoke components involved in stimulation of VEGF release.…”

“…This is consistent with the observation that, in induced sputum, VEGF levels are higher in healthy smokers as well as in patients with chronic bronchitis when compared with non-smokers (Kanazawa et al, 2003;Kanazawa, 2007;Rovina et al, 2007). Notably, elevation of VEGF levels in the sputum of asymptomatic smokers and smokers with bronchitis type of COPD correlates with smoking pack years and with IL-8 levels (Rovina et al, 2007), a chemokine up-regulated by direct cigarette smoke exposure (Facchinetti et al, 2007). Moreover, VEGF expression is increased in pulmonary muscular arteries of smokers with normal lung function when compared with non-smokers (Santos et al, 2003;Kranenburg et al, 2005).…”

“…We observed that 4-HNE was capable of activating, at sub-toxic concentrations, VEGF release from ASMC cultures. Interestingly, 4-HNE also stimulated the secretion of IL-8 in macrophages (Facchinetti et al, 2007), lung fibroblasts and epithelial cells (Moretto et al, 2009) as well as eliciting neutrophil recruitment (Schaur et al, 1994). Thus, VEGF might be triggered in chronic cigarette smokers or COPD patients not only by smoke-inhaled acrolein and crotonaldehyde, but also by endogenously formed unsaturated aldehydes such as 4-HNE.…”

“…Cigarette smoke extract was freshly generated under standardized conditions as previously described (Sato et al, 1999;Walters et al, 2005;Facchinetti et al, 2007). Briefly, aqueous CSE was obtained by bubbling smoke generated from the combustion of four cigarettes (Marlboro Red, 12 mg tar, 0.9 mg nicotine each) through 50 mL of DMEM w/o phenol red and subsequently filtered through a 0.2 mm pore filter (Millipore).…”

“…Culpitt et al, 2003;Walters et al, 2005;Demirjian et al, 2006;Yang et al, 2006) as well as from lung fibroblasts (Sato et al, 1999;Numanami et al, 2003;Li et al, 2007) and airway epithelial cells (Mio et al, 1997;Masubuchi et al, 1998;Kode et al, 2006). Recently we provided evidence that the a,b-unsaturated aldehydes acrolein and crotonaldehyde are the main components of smoke responsible for IL-8 release from macrophages (Facchinetti et al, 2007) as well as from airway epithelial cells and fibroblasts through the stimulation of both extracellular signal-regulated kinase type 1 and 2 (ERK1/2) and mitogen-activated protein kinase (p38 MAPK) intracellular pathways (Moretto et al, 2009).…”