Cigarette Smoke Extract Suppresses Human Dendritic Cell Function Leading to Preferential Induction of Th-2 Priming

Abstract

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.

Nicotine and oxidative cigarette smoke constituents induce immune-modulatory and pro-inflammatory dendritic cell responses

“…Aqueous CSE was prepared from Kentucky research cigarettes 1RF4 as recently described (Vassallo et al, 2005). The nicotine levels in the CSE preparations were measured in the institutional clinical laboratory using liquid chromatography-tandem mass spectrometry: the mean nicotine content in 4 separate samples of 100% CSE was 21,460±2778 ng/ml.…”

“…The final concentration of CSE used in the experiments described was 1-2% -final nicotine concentration in vitro of 1% CSE was equal to 214.6±27.8 ng/ml. The concentrations of <3% were chosen because of preliminary viability studies that demonstrated a lack of nonspecific toxicity and >95% viability compared with controls, as determined by the XTT assay and AnnexinV/propridium iodide staining with these CSE preparations (Vassallo et al, 2005).…”

“…Monocytes were isolated using depleting antibody cocktails (StemCell laboratories) and monocyte-derived DCs were generated with GM-CSF and IL-4 as previously described (Vassallo et al, 2005). Maturation was induced by 18-hour culture with 100 ng/ml LPS [from E. coli; Sigma] or 1µg/ml soluble recombinant human CD40L [Axxora Platform].…”

“…To determine whether cigarette smoke induces neutrophilic chemokine production from DCs, human monocyte-derived DCs were incubated with CSE, in the presence or absence of either 100ng/ml LPS, or 1µg/ ml recombinant human CD40L as maturational agents. In these experiments, a CSE concentration of 1 -2% was selected because prior studies demonstrated non-specific toxicity (cell death) in DCs exposed to CSE levels >3% (Vassallo et al, 2005). Following 18 hours of incubation, CXCL8/IL-8 levels produced by immature or maturing DCs were measured in the supernatants using ELISA.…”

“…In prior studies, we demonstrated that cigarette smoke constituents suppress Th-1 stimulatory functions of DCs while preserving the capacity of DCs to generate interleukin-6 and interleukin-10 upon maturation with lipopolysaccharide (LPS) (Vassallo et al, 2005). The current study was designed to determine mechanisms by which cigarette smoke constituents activate DCs to produce soluble factors that promote local inflammation and immune-suppression in the airways.…”