Dendritic cells (DC) are key regulators of immune responses. In the current study, we hypothesized that cigarette smoke-induced aberrance in DC function is an important mechanism by which smokers develop cancer, infection, and allergy—diseases common in smokers. We demonstrate that cigarette smoke extract (CSE) inhibits DC-mediated priming of T cells, specifically inhibiting the secretion of IFN-γ whereas enhancing the production of IL-4 in the MLR. Conditioning with CSE did not effect cytokine (IL-10, IL-6, or IL-12) production from immature DCs, but significantly inhibited IL-12p70 release by LPS-matured DCs. In contrast, IL-10 secretion by LPS-activated CSE-conditioned DCs was enhanced when compared with control DCs. CSE also induced cyclooxygenase-2 protein levels in maturing DCs and significantly augmented endogenous PGE2 release. Conditioning of DCs with CSE also suppressed LPS-mediated induction of CD40, CD80, and CD86, and suppressed maturation-associated CCR7 expression. Although CSE has been reported to induce apoptosis of fibroblasts and epithelial cells, the immunomodulatory effects observed with CSE were not due to diminished DC viability. The effects of CSE on DC function were not exclusively mediated by nicotine, because equivalent, or even higher concentrations of nicotine than those found in CSE, failed to suppress DC-induced T cell priming. These data provide evidence that soluble components extracted from cigarette smoke suppress key DC functions and favor the development of Th-2 immunity.

IL-12p70, a heterodimer composed of p35 and p40 subunits, is a key polarizing cytokine produced by maturing dendritic cells (DCs). We report that cigarette smoke extract (CSE), an extract of soluble cigarette smoke components, suppresses both p35 and p40 production by LPS or CD40L-matured DCs. Suppression of IL-12p70 production from maturing DCs was not observed in the presence of nicotine concentrations achievable in CSE or in the circulation of smokers. The suppressed IL-12p70 protein production by CSE-conditioned DCs was restored by pretreatment of DCs or CSE with the antioxidants N-acetylcysteine and catalase. Inhibition of DC IL-12p70 by CSE required activation of ERK-dependent pathways, since inhibition of ERK abrogated the suppressive effect of CSE on IL-12 secretion. Oxidative stress and sustained ERK phosphorylation by CSE enhanced nuclear levels of the p40 transcriptional repressor c-fos in both immature and maturing DCs. Suppression of the p40 subunit by CSE also resulted in diminished production of IL-23 protein by maturing DCs. Using a murine model of chronic cigarette smoke exposure, we observed that systemic and lung DCs from mice “smokers” produced significantly less IL-12p70 and p40 protein upon maturation. This inhibitory effect was selective, since production of TNF-α during DC maturation was enhanced in the smokers. These data imply that oxidative stress generated by cigarette smoke exposure suppresses the generation of key cytokines by maturing DCs through the activation of ERK-dependent pathways. Some of the cigarette smoke-induced inhibitory effects on DC function may be mitigated by antioxidants.

Chronic airway inflammation is a cardinal feature of chronic obstructive pulmonary disease (COPD), a destructive cigarette smoke-induced lung disease. Although it is apparent that dendritic cells (DCs) are an important constituent of the chronic inflammatory cell influx found in airways of COPD patients, the functional roles of DCs in the pathogenesis of smoking-induced emphysema are unknown. We postulated that DCs activated by cigarette smoke constituents directly participate in the chronic inflammation that characterizes COPD airways. Concordant with this hypothesis, we observed that incubation of DCs with cigarette smoke extract (CSE), and chronic exposure of mice to cigarette smoke, both augmented the generation of neutrophilic chemokines by immature and lipopolysaccharide (LPS) or CD40L-matured DCs. The generation of interleukin-8 (CXCL8/IL-8) by human DCs conditioned with CSE was suppressed by the antioxidant n-acetyl cysteine (NAC), implying the involvement of oxidant sensitive pathways as a primary mechanism involved in the enhanced CXCL8/IL-8 generation. Cigarette smoke extract and nicotine also augment the production of secreted prostaglandin E 2 and intracellular cyclooxygenase-2 (COX-2) in maturing DCs. Whereas NAC suppressed production of CXCL8 by CSEconditioned DCs, it augmented production of PGE 2 and cellular COX-2 levels in maturing DCs. These studies indicate that the stimulation of DCs by cigarette smoke-induced oxidative stress and nicotine promote the generation of pro-inflammatory responses that promote chronic inflammation in smokers. Certain pharmacologic strategies such as anti-oxidant therapy may be only partially effective in mitigating cigarette smoke-induced pro-inflammatory DC-mediated responses in smokers.

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Little is known about the role of dendritic cells [DCs] in emphysema, a disease characterized by prominent neutrophilic airway inflammation. We hypothesized that lung DCs participate in the pathogenesis of emphysema by producing chemotactic factors that recruit neutrophils to the lung. Cigarette smoke extract (CSE 0.5–2%) generated from 1RF4 Kentucky research cigarettes dose‐dependently induced interleukin‐8 [IL‐8] production from both immature and maturing DCs. Although relatively high nicotine concentrations induced IL‐8 production in vitro, nicotine concentrations equivalent to those obtained in CSE or in the blood of smokers failed to induce IL‐8 release from DCs. The antioxidant n‐acetyl cysteine [NAC] completely suppressed IL‐8 generation by CSE‐activated DCs. Using a cigarette smoking chamber, we observed that cigarette smoking enhanced the production of the murine IL‐8 homologs keratinocyte‐derived chemokine [KC] and macrophage inflammatory protein‐2 [MIP‐2] by both LPS and CD40‐matured lung DCs. In parallel to the observation that oxidative stress is important in smoking‐induced chemokine generation by DCs, we observed an increase in endogenous DC heme‐oxygenase‐1 [HO‐1] levels in human DCs activated by CSE and also in DCs harvested from mice exposed to cigarette smoke. Generation of IL‐8 by maturing DCs activated by CSE was abrogated when induction of cellular HO‐1 was inhibited by zinc protoporhyrin, suggesting that HO‐1‐dependent pathways are necessary for cigarette smoke‐induced IL‐8 production by DCs. These studies indicate a role for DCs in the pathogenesis of neutrophilic airway inflammation observed in smokers with emphysema. Funded by a Parker B Francis Fellowship grant and a Flight Attendant Medical Research Institute award to RV.

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Dendritic cells (DC) are key regulators of immunity. Chronic obstructive pulmonary disease [COPD], a destructive lung disease that affects smokers, is associated with chronic airway inflammation. Although increased DC numbers are found in COPD airways, their functional roles in the pathogenesis of COPD are not well‐defined. We postulated that DCs activated by cigarette smoke constituents directly promote chronic airway inflammation. Concordant with this hypothesis, we observed that incubation of DCs with cigarette smoke extract [CSE], and chronic exposure of mice to cigarette smoke, both augmented the generation of neutrophilic chemokines by immature and lipopolysaccharide [LPS] or CD40L‐matured DCs. The generation of IL‐8 by human DCs conditioned with CSE was completely suppressed by the anti‐oxidant n‐acetyl cysteine [NAC]. Cigarette smoke extract and nicotine augmented the production of secreted prostaglandin‐E2 and intracellular cyclo‐oxygenase‐2 in maturing DCs. Whereas the production of IL‐8 by CSE‐conditioned DCs was suppressed by anti‐oxidants, PGE2 production was augmented by anti‐oxidants, and suppressed by inhibitors of nicotinic signaling. These studies indicate that certain pharmacologic strategies such as anti‐oxidant therapy may be only partially effective in mitigating pro‐inflammatory DC‐mediated responses in smokers. Supported by FARMRI research grant to RV.