Sub-ohm vaping increases the levels of carbonyls, is cytotoxic, and alters gene expression in human bronchial epithelial
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RESEARCH
Sub‑ohm vaping increases the levels of carbonyls, is cytotoxic, and alters gene expression in human bronchial epithelial cells exposed at the air–liquid interface Alexandra Noël* , Ekhtear Hossain, Zakia Perveen, Hasan Zaman and Arthur L. Penn
Abstract Background: Exposure to electronic-cigarette (e-cig) aerosols induces potentially fatal e-cig or vaping-associated lung injury (EVALI). The cellular and molecular mechanisms underlying these effects, however, are unknown. We used an air–liquid interface (ALI) in vitro model to determine the influence of two design characteristics of third-generation tank-style e-cig devices—resistance and voltage—on (1) e-cig aerosol composition and (2) cellular toxicity. Methods: Human bronchial epithelial cells (H292) were exposed to either butter-flavored or cinnamon-flavored e-cig aerosols at the ALI in a Vitrocell exposure system connected to a third-generation e-cig device. Exposures were conducted following a standard vaping topography profile for 2 h per day, for 1 or 3 consecutive days. 24 h after ALI exposures cellular and molecular outcomes were assessed. Results: We found that butter-flavored e-cig aerosol produced under ‘sub-ohm’ conditions ( 1 Ω and voltage > 4.5 V), contains lower carbonyl levels ( ± 1.5 was considered significant.
Extracellular NO measurements
Statistical analysis
The NO concentrations in the H292 cell culture media were determined by the Griess reagent assay (catalog #30,100, Biotum, Fremont, CA). 50 µL of the cell culture medium was removed from the basal side of the Transwell and incubated with an equal volume of the Griess reagent at room temperature for 15 min. Sodium nitrite was used to generate a standard curve. We followed the manufacturers’ instructions. The absorbance was determined with a Tecan Infinite 2000 spectrometer (Tecan Group Ltd, Mannedorf, Switzerland) at a wavelength of 490 nm. For each sample, the cell medium absorbance was normalized to the total cell count. Absorbance values for the air control groups were set at 100%. Samples were run in triplicate.
All statistical analyses were performed using Microsoft Excel or GraphPad Prism 7 software. Data are expressed as means ± standard error of the mean (SEM). Data are also expressed as percent change relative to the respective air control group, set at 100%. Statistically significant differences between groups were analyzed using either a Student-t test or a one-way analysis of variance (ANOVA) followed by a Tukey’s post-hoc test, when testing 3 or more groups. Statistical significance was achieved with a p-value 100%) when compared to
Noël et al. Respir Res
(2020) 21:305
Page 10 of 20
Fig. 5. 1 Day of butter-flavored e-cig aerosol exposure under sub-ohm conditions-induced oxidative stress-mediated lung cell injury is alleviated by n-acetyl cysteine (NAC) pre-treatment. H292 cells were pre-treated with or without NAC at a concentration of 5 mM for 12 h before exposure to butter-flavored e-cig aerosol for 1 day at the air–liquid interface (ALI).
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