Alterations to redox chemistry of A549 adenocarcinomic human alveolar basal epithelial cells upon exposure to e-cigarette vapors

Stefanie Rigg, Purdue University

Abstract

Cigarettes are known to cause harm to nearly every organ in the body. Due to the highly addictive nature of nicotine, many people have begun to look for healthier alternatives to cigarettes, such as the electronic nicotine delivery system (ENDS). While there has been significant research on the effects of cigarette smoke on the body, there is little known about the effects of ENDS. It was recently shown that ENDS vapor exposure altered the redox physiology of lung epithelial cells and induced pro-inflammatory cytokine secretion in mouse bronchial alveolar lavage fluid. Based on these findings, I hypothesized that exposure to ENDS vapor would affect cell viability in adenocarcinoma human alveolar basal epithelial (A549) cells similar to cigarette smoke. To test this, ENDS vapors extract using an e-juice containing 6mg/20 mL nicotine (eCSE) and 0 mg/20 mL nicotine (NFeCSE), cigarette smoke extract (CSE), pure nicotine, and glycerol were used as test reagents on A549 cells. Cells exposed to eCSE and NFeCSE demonstrated a decrease in viability, while cells exposed to CSE had no change compared to the PBS control. Similarly, a decrease in cellular proliferation, as measured by BrdU incorporation, was observed in cells exposed to both eCSE and NFeCSE, while cells exposed to CSE had no change in proliferation compared to the PBS control. In order to elucidate the mechanism behind reduced viability with ENDS exposure, cells were examined for the activation of apoptotic pathways and cellular oxidative stress, as well as activation of the Akt cellular proliferation signaling pathway. Apoptosis was measured by quantifying caspase-3 activation in cells following exposure to the test reagents. Cells exposed to NFeCSE displayed an increase in caspase-3 activation, while cells exposed to eCSE and CSE displayed no change compared to the PBS control. Glutathione (GSSG and GSH) levels of cells treated with eCSE and NFeCSE both exhibited elevated levels of the oxidized form, glutathione disulfide (GSSG), indicative of oxidative stress. Activation of Akt signaling was elevated in eCSE, NFeCSE, CSE, and glycerol treated cells, suggesting that the combination of glycerol and nicotine, which are both known to activate the Akt pathway, could be influencing pro-survival signaling cascades. Thus, A549 cell exposure to eCSE results in caspase-3 independent cytotoxicity due to activation of Akt pro-survival signaling cascades with concurrent oxidative stress. This is in contrast to CSE exposed A549 cells, which undergo Akt-mediated cellular proliferation, with no indication of oxidative stress. These results indicate that ENDS vapor has a differential effect on A549 cells compared to CSE and questions the advertisement of ENDS as a healthy alternative to cigarettes. ^

Degree

M.S.

Advisors

Lindsay Gielda, Purdue University, Radmila Sarac, Purdue University.

Subject Area

Biology|Cellular biology

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