Cigarette smoke dampens antiviral signaling in small airway epithelial cells by disrupting TLR3 cleavage

Abstract

Cigarette smokers and people exposed to second-hand smoke are at an increased risk for pulmonary viral infections, and yet the mechanism responsible for this heightened susceptibility is not understood. To understand the effect of cigarette smoke on susceptibility to viral infection, we used an air-liquid interface culture system and exposed primary human small airway epithelial cells (SAEC) to whole cigarette smoke, followed by treatment with the viral mimetic polyinosinic polycytidylic acid (poly I:C) or influenza A virus (IAV). We found that prior smoke exposure strongly inhibited production of proinflammatory (interleukin-6 and interleukin-8) and antiviral [interferon-γ-induced protein 10 (IP-10) and interferons] mediators in SAECs in response to poly I:C and IAV infection. Impaired antiviral responses corresponded to increased infection with IAV. This was associated with a decrease in phosphorylation of the key antiviral transcription factor interferon response factor 3 (IRF3). Here, we found that cigarette smoke exposure inhibited activation of Toll-like receptor 3 (TLR3) by impairing TLR3 cleavage, which was required for downstream phosphorylation of IRF3 and production of IP-10. These results identify a novel mechanism by which cigarette smoke exposure impairs antiviral responses in lung epithelial cells, which may contribute to increased susceptibility to respiratory infections.

Cigarette smoke increases susceptibility to infection in lung epithelial cells by upregulating caveolin-dependent endocytosis

“…We have previously shown that cigarette smoke impaired the anti-viral responses in primary human SAEC following treatment with poly I:C [4]. Poly I:C is recognized by the pattern recognition receptor (PRR) toll-like receptor 3 (TLR3).…”

“…Small airway epithelial cells were cultured at the air-liquid interface and exposed to whole cigarette smoke for 60 min as previously described [4]. Following smoke exposure, media in the basal compartment was replaced to remove any residual smoke components.…”

“…Levels of interferon gamma inducible protein 10 (IP-10), interleukin 6 (IL-6), and interleukin 8 (IL-8) in the culture supernatant were determined by ELISA. Interferon activity in the culture supernatants was determined using an ISRE reporter cell line as described previously [4].…”

“…Virus infection was performed essentially as described [4]. Briefly, SAEC were exposed to air or cigarette smoke for 60 min as described above and then rested for 5 hours before infection with Influenza A/WSN/1933 H1N1 (IAV WSN) virus at the indicated MOI for 1 hour.…”

“…Respiratory infections can trigger acute exacerbations of COPD symptoms that are responsible for the bulk of the COPDrelated morbidity and mortality. We have previously shown that cigarette smoke impairs TLR3 cleavage in primary human small airway epithelial cells resulting in impaired antiviral responses [4]. Similarly, smoke-exposed mice have increased infectivity with influenza A virus [5].…”