Flavored e-cigarette liquids reduce proliferation and viability in the CALU3 airway epithelial cell line

Am J Physiol Lung Cell Mol Physiol. 2017 Jul 1;313(1):L52-L66. doi: 10.1152/ajplung.00392.2016. Epub 2017 Apr 20.


E-cigarettes are generally thought of as a safer smoking alternative to traditional cigarettes. However, little is known about the effects of e-cigarette liquids (e-liquids) on the lung. Since over 7,000 unique flavors have been identified for purchase in the United States, our goal was to conduct a screen that would test whether different flavored e-liquids exhibited different toxicant profiles. We tested the effects of 13 different flavored e-liquids [with nicotine and propylene glycol/vegetable glycerin (PG/VG) serving as controls] on a lung epithelial cell line (CALU3). Using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay as an indicator of cell proliferation/viability, we demonstrated a dose-dependent decrease of MTT metabolism by all flavors tested. However, a group of four flavors consistently showed significantly greater toxicity compared with the PG/VG control, indicating the potential for some flavors to elicit more harmful effects than others. We also tested the aerosolized "vapor" from select e-liquids on cells and found similar dose-dependent trends, suggesting that direct e-liquid exposures are a justifiable first-pass screening approach for determining relative e-liquid toxicity. We then identified individual chemical constituents for all 13 flavors using gas chromatography-mass spectrometry. These data revealed that beyond nicotine and PG/VG, the 13 flavored e-liquids have diverse chemical constituents. Since all of the flavors exhibited some degree of toxicity and a diverse array of chemical constituents with little inhalation toxicity available, we conclude that flavored e-liquids should be extensively tested on a case-by-case basis to determine the potential for toxicity in the lung and elsewhere.

Keywords: COPD; aerosols; cancer; nicotine; tobacco.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Aerosols
  • Cell Death / drug effects
  • Cell Line
  • Cell Proliferation / drug effects
  • Cell Survival / drug effects
  • Cinnamomum aromaticum / chemistry
  • Electronic Nicotine Delivery Systems*
  • Epithelial Cells / cytology*
  • Epithelial Cells / drug effects
  • Epithelial Cells / metabolism
  • Gas Chromatography-Mass Spectrometry
  • Humans
  • Inhibitory Concentration 50
  • Lung / cytology*
  • Menthol / pharmacology
  • Nicotine / pharmacology


  • Aerosols
  • Menthol
  • Nicotine