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. 2015 Feb 4;10(2):e0116861.
doi: 10.1371/journal.pone.0116861. eCollection 2015.

Exposure to Electronic Cigarettes Impairs Pulmonary Anti-Bacterial and Anti-Viral Defenses in a Mouse Model

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Free PMC article

Exposure to Electronic Cigarettes Impairs Pulmonary Anti-Bacterial and Anti-Viral Defenses in a Mouse Model

Thomas E Sussan et al. PLoS One. .
Free PMC article

Abstract

Electronic cigarettes (E-cigs) have experienced sharp increases in popularity over the past five years due to many factors, including aggressive marketing, increased restrictions on conventional cigarettes, and a perception that E-cigs are healthy alternatives to cigarettes. Despite this perception, studies on health effects in humans are extremely limited and in vivo animal models have not been generated. Presently, we determined that E-cig vapor contains 7 x 10(11) free radicals per puff. To determine whether E-cig exposure impacts pulmonary responses in mice, we developed an inhalation chamber for E-cig exposure. Mice that were exposed to E-cig vapor contained serum cotinine concentrations that are comparable to human E-cig users. E-cig exposure for 2 weeks produced a significant increase in oxidative stress and moderate macrophage-mediated inflammation. Since, COPD patients are susceptible to bacterial and viral infections, we tested effects of E-cigs on immune response. Mice that were exposed to E-cig vapor showed significantly impaired pulmonary bacterial clearance, compared to air-exposed mice, following an intranasal infection with Streptococcus pneumonia. This defective bacterial clearance was partially due to reduced phagocytosis by alveolar macrophages from E-cig exposed mice. In response to Influenza A virus infection, E-cig exposed mice displayed increased lung viral titers and enhanced virus-induced illness and mortality. In summary, this study reports a murine model of E-cig exposure and demonstrates that E-cig exposure elicits impaired pulmonary anti-microbial defenses. Hence, E-cig exposure as an alternative to cigarette smoking must be rigorously tested in users for their effects on immune response and susceptibility to bacterial and viral infections.

Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.

Figures

Figure 1
Figure 1. Schematic of our E-cig exposure model.
Figure 2
Figure 2. E-cig-induced pulmonary response.
(A) EPR spectra of E-cig TPM. The presented spectrum is a result of subtraction of a Cambridge filter pad EPR signal before and after collection of TPM. (B) Lipid peroxidation was measured by thiobarbituric acid reactive substances (TBARS) in lung homogenates from C57BL/6 mice that were exposed to air or E-cig vapor for 1.5 h, twice per day, for 2 wks. (C) Inflammatory cells were quantified in the BAL at 24h after the final exposure. (D) Cytokines were analyzed in cell-free BAL fluid from air and E-cig exposed mice at 24h after the final exposure. N = 10 mice per group. *p<0.05 by Student’s two-tailed t-test.
Figure 3
Figure 3. E-cig exposure reduces pulmonary bacterial clearance in mice infected with S. pneumoniae.
Mice were exposed to air or E-cig for 2 wks, then infected intranasally with 1x105 colony forming units (CFU) of S. pneumoniae. Inflammation (A) and bacterial CFUs (B) were determined in BAL at 24h after infection (N = 10 mice per group). (C) In a separate group of mice, bacterial CFUs were quantified in lung homogenates (N = 10 mice per group). (D) Alveolar macrophages from air or E-cig exposed mice were harvested and infected with S. pneumoniae at multiplicities of infection of 10 and 20. Bacterial CFUs were quantified in cell-free culture media at 4 h after infection. (E) Intracellular (internalized) and extracellular bacteria (cell-free culture media) were quantified at 1 h after ex vivo infection of alveolar macrophages with an MOI of 20. (F) Alveolar macrophages were harvested at 2h after final exposure, stained with antibodies against CD36 and MARCO, and analyzed by flow cytometry. (G) Bacterial clearance was measured in alveolar macrophages from mice exposed to air or traditional E-cig vapor. *p<0.05 by Student’s two-tailed t-test.
Figure 4
Figure 4. E-cig exposure impairs viral clearance and causes significant morbidity and mortality in mice following influenza virus infection.
Mice were exposed to air or E-cig for 2 wks, then infected intranasally with either TCID50 102 (A-B) or TCID50 103 (C) of H1N1 virus. (A) Viral titer was determined by TCID50 assay in lung homogenates at 4 days after infection (N = 5 mice per group). (B-C) Mice were weighed daily after infection with either TCID50 102 (B) or 103 (C), and values are presented as percent of starting weight (N = 10 mice per group). For mice that died during the experiments, body weights were included in the analysis up until the day of death. (D) Mortality curves in response to intranasal infection with TCID50 102 or 103 H1N1 (N = 10 mice per group). *p<0.05 by Student’s two-tailed t-test.
Figure 5
Figure 5. Influenza-induced inflammation is altered by E-cig exposure.
Mice were exposed to air or E-cig for 2 wks, then infected intranasally with TCID50 102 of H1N1. BAL was collected at day 4 (N = 5) and day 8 (N = 4) after infection, followed by quantification of inflammatory cells (A) and cytokines (B). *p<0.05 by Student’s two-tailed t-test.

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