Clara Cell 10-kDa Protein Gene Transfection Inhibits NF-κB Activity in Airway Epithelial Cells

Abstract

BackgroundClara cell 10-kDa protein (CC10) is a multifunctional protein with anti-inflammatory and immunomodulatory effects. Induction of CC10 expression by gene transfection may possess potential therapeutic effect. Nuclear factor κB (NF-κB) plays a key role in the inflammatory processes of airway diseases.Method/ResultsTo investigate potential therapeutic effect of CC10 gene transfection in controlling airway inflammation and the underlying intracellular mechanisms, in this study, we constructed CC10 plasmid and transfected it into bronchial epithelial cell line BEAS-2B cells and CC10 knockout mice. In BEAS-2B cells, CC10's effect on interleukin (IL)-1β induced IL-8 expression was explored by means of RT-PCR and ELISA and its effect on NF-κB classical signaling pathway was studied by luciferase reporter, western blot, and immunoprecipitation assay. The effect of endogenous CC10 on IL-1β evoked IL-8 expression was studied by means of nasal explant culture. In mice, CC10's effect on IL-1β induced IL-8 and nuclear p65 expression was examined by immunohistochemistry. First, we found that the CC10 gene transfer could inhibit IL-1β induced IL-8 expression in BEAS-2B cells. Furthermore, we found that CC10 repressed IL-1β induced NF-κB activation by inhibiting the phosphorylation of IκB-α but not IκB kinase-α/β in BEAS-2B cells. Nevertheless, we did not observe a direct interaction between CC10 and p65 subunit in BEAS-2B cells. In nasal explant culture, we found that IL-1β induced IL-8 expression was inversely correlated with CC10 levels in human sinonasal mucosa. In vivo study revealed that CC10 gene transfer could attenuate the increase of IL-8 and nuclear p65 staining in nasal epithelial cells in CC10 knockout mice evoked by IL-1β administration.ConclusionThese results indicate that CC10 gene transfer may inhibit airway inflammation through suppressing the activation of NF-κB, which may provide us a new consideration in the therapy of airway inflammation.

Low plasma CC16 levels in smokers are associated with a higher risk for chronic bronchitis

“…Secondly, even after adjustment in multivariable models for age, sex, baseline BMI, pack years of smoking and baseline current smoking status, the observed effect not only persists but is strengthened. These observed associations are consistent with published results wherein CC16 has been associated with anti-inflammatory activity in lungs [2,13] and with regression of bronchial dysplasia in former smokers [14]. Taken together, these studies support an integral role for this protein in lung health.…”

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Club cell protein (CC16 in humans) plays a crucial role in maintaining homeostasis of the airway epithelium [1] and has anti-inflammatory activity in the lung [2]. Serum CC16 levels are low in cigarette smokers [3] and increase following smoking cessation [4], but patients with chronic obstructive pulmonary disease (COPD) show progressive decrease in CC16 with disease severity [5].

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Club Cell Protein 16 (CC16) Augmentation: A Potential Disease-modifying Approach for Chronic Obstructive Pulmonary Disease (COPD)

“…CC16 −/− nasal epithelial cells produce greater amounts of IL-8 when activated with IL-1β than cells from WT mice both in vitro and in vivo 113 . Moreover, plasmid-mediated over-expression of CC16 in the nasal epithelium of CC16 −/− mice reduces their increased release of IL-8 when IL-1β is administered 113 .…”

“…CC16 −/− nasal epithelial cells produce greater amounts of IL-8 when activated with IL-1β than cells from WT mice both in vitro and in vivo 113 . Moreover, plasmid-mediated over-expression of CC16 in the nasal epithelium of CC16 −/− mice reduces their increased release of IL-8 when IL-1β is administered 113 . Gamez et al reported that epithelial cells isolated from COPD airways produce more IL-8 compared with cells isolated from healthy smoker and non-smoker airways, and this increased release of IL-8 was reduced by adding rCC16 to the COPD cell cultures 114 .…”

“…CC16 inhibits activation of NFκB in lung cancer cells 138 , uterine endometrial cells 139 and airway epithelial cells 113; 48 in vitro . Plasmid-mediated over-expression of CC16 in nasal epithelial cells inhibits NFκB activation by blocking the phosphorylation of the inhibitor of NFκB subunit-α (IκB-α) thereby reducing degradation of this inhibitor 113 .…”