AAAR 35th Annual Conference October 17 - October 21, 2016 Oregon Convention Center Portland, Oregon, USA
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Banana Pudding Flavored E-liquid Alters Cell Proliferation and Ca2+ Signaling in Lung Epithelia
TEMPERANCE ROWELL, Robert Tarran, University of North Carolina at Chapel Hill
Abstract Number: 104 Working Group: Electronic Cigarettes - Health Effects
Abstract E-cigarettes are popular and available in over 7,000 flavors, yet little is known about their effects on pulmonary epithelia. We purchased flavored e-liquids from the Vapor Girl (http://www.thevaporgirl.com/) to characterize their biological effects on human bronchial epithelial cultures (HBECs) and CALU3 lung epithelial cells. We screened 13 e-liquid flavors for effects on cell proliferation/viability and ability to alter cell signaling (e.g. Ca$^(2+)). Ca$^(2+) signaling regulates cell division, mucus secretion, ciliary beat frequency, and Cl$^(-)/fluid secretion in airway epithelia. Altered Ca$^(2+) signaling could change cell homeostasis, thus we measured changes in cytosolic Ca$^(2+) and other pathway components (i.e. STIM1 puncta, kinase phosphorylation, IP$_3 generation).
Cells were plated on glass coverslips or in multiwell plates and exposed to e-liquids diluted into media either acutely (10 min) or over 24h. Cell proliferation/viability was measured using the MTT assay. Cytosolic Ca$^(2+) changes were measured using Fura-2-AM. Relative kinase phosphorylation was measured using a Phospho-Kinase Array and IP$_3 generation was measured using a competitive ELISA. All assays were done with HBECs and/or CALU3s except STIM1 puncta visualization, which used transiently transfected STIM1-mCherry HEK293 cells.
Several flavors inhibited cell proliferation in a dose-dependent manner, including Banana Pudding (Southern Style) (BPSS). BPSS also elicited an acute cytosolic Ca$^(2+) signal involving both the endoplasmic reticulum (ER) and store-operated Ca$^(2+) entry (SOCE), formed STIM1 puncta, and altered phosphorylation of ERK1/2, which regulates STIM1 phosphorylation.
Our data showed that BPSS-flavored e-liquid altered ER/SOCE Ca$^(2+)-signaling mechanisms in airway epithelia, which could have biological consequences to Ca$^(2+)-mediated HBEC innate defenses. This suggested that other flavors may alter cell signaling and other important lung epithelial functions. Investigations into the effects of flavored e-cigarette aerosol on Ca$^(2+)-dependent aspects of innate defense are ongoing.
This study was funded by UNC TCORS NIH (P50-HL-120100-01) and the Marsico Lung Institute Tissue Core provided HBECs.