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. 2020 Feb 13;10:2544. doi: 10.1038/s41598-020-59290-y

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© The Author(s) 2020

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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E-liquids stimulated the expression of inflammatory cytokines in HMEECs. (A) HMEECs were treated with PG/VG (1 to 5%), tobacco-flavored e-liquid (1 to 5%), and menthol-flavored e-liquid (1 and 2%) for 24 h. Quantitative real-time PCR was performed to evaluate inflammatory cytokine gene such as COX-2 and TNF-α expression levels. The expression levels of COX-2 and TNF-α significantly increased in e-liquid-concentration-dependent manner. (B) Cells were exposed to the average IC₅₀ values of each e-liquid for 24 h (PG/VG: 4.5%, Tobacco: 3.3%, and Menthol: 1.5%). Both flavored e-liquids significantly increased the mRNA levels of COX-2 and TNF-α on HMEECs. mRNA expression of COX-2 and TNF-α was significantly increased in menthol-flavored e-liquid-treated group compared to in PG/VG group. (C) Cells were treated with PG/VG (4.5%), tobacco-flavored e-liquid (3.3%), and menthol-flavored e-liquid (1.5%) for 24 h. The results showed that the levels of COX-2 and TNF-α protein levels were increased by treatment with e-liquids. Densitometric analysis of western blot is shown as a graph. Similar to their mRNA profiles, COX-2 and TNF- α protein expression was significantly higher following tobacco- and menthol-flavored e-liquid exposure compared to the control group. Western blot data were quantified and normalized to β-actin levels. All data were obtained from three independent experiments and the error bars indicate the mean ± SD. *p < 0.05, **p < 0.01, and ***p < 0.001 compared to the corresponding control.