Differential Gene Expression Using RNA Sequencing Profiling in a Reconstituted Airway Epithelium Exposed to Conventional Cigarette Smoke or Electronic Cigarette Aerosols

Abstract

The use of electronic cigarettes (e-cigarettes) potentially offers a safer alternative to conventional tobacco products. The advance in molecular biology and computational sciences offers new perspective to assess adverse biological responses for product risk assessment by combining omics screens with knowledge-based biological pathways. Our aim was to compare transcriptomic perturbations in MucilAirÔ, a commercially available lung epithelial tissue, after short repeated exposure to cigarette smoke (3R4F) and e-cigarette (Vype ePen) aerosols. We performed deep RNA sequencing and secreted inflammatory cytokine profiling postexposure. One hundred twenty-three genes were differentially expressed at fold change (FC) >1.5 and p-false discovery rate (pFDR) <0.1 for 3R4F exposure and 0 genes for Vype ePen aerosol exposure. When a relaxed filter pFDR <0.5 and FC >1.5 was applied, 29 genes were identified with e-cigarette aerosol exposure and used for validation of potential candidates by quantitative reverse transcription polymerase chain reaction (qRT-PCR). Gene enrichment analysis was conducted and predicted a response to 3R4F smoke exposure in biological processes involving inflammation and oxidative stress pathways. No enrichment could be performed for Vype ePen aerosol exposure due to the lack of regulated gene candidates at those exposure conditions even after qRT-PCR validation. Of a panel of 33 cytokines screened, 8 were upregulated (FC >1.5 p < 0.05) following 3R4F smoke exposure, which was in agreement with our enrichment analysis. In conclusion, aerosol from the tested e-cigarette caused limited perturbations in gene and inflammatory cytokine expression compared to conventional cigarette smoke, as assessed using next-generation sequencing-based systems biology approaches in 3D commercially available reconstituted lung epithelial tissues.

A lower impact of an acute exposure to electronic cigarette aerosols than to cigarette smoke in human organotypic buccal and small airway cultures was demonstrated using systems toxicology assessment

“…We also had previously reported that the pattern of gene expression changes was comparable between cultures from two donors following CS exposure [35]. Furthermore, Banerjee et al reported that a similar trend of inflammatory mediator secretion was detected in cultures from six different donors following CS exposure [50]. These studies thus support that the results of the present study, despite using data from one donor, can still provide valuable insights into the underlying mechanisms associated with CS and EC aerosol exposures.…”

Reduced biological effect of e-cigarette aerosol compared to cigarette smoke evaluated in vitro using normalized nicotine dose and RNA-seq-based toxicogenomics

“…We observed CYP1A1 induction between 1348- and 876-fold at pFDR < 0.01 at 24 hrs and 48 hrs, respectively. We also previously reported CYP1A1 induction by cigarette smoke in MucilAir™ in shorter exposure runs 24 . Finally, for the e-cigarette aerosol treatment, only FGF14 and EGR1 were significant in the qPCR at p < 0.05, FC > [1.5] at the 1/7 aerosol dilution (Supplementary Table S21 , Fig.…”

Liquid‐liquid phase separation and fibrillation of the prion protein modulated by a high‐affinity DNA aptamer

“…We were able to follow fusion and fission events characteristic of protein liquid droplets (not shown). 41 We suggest that 1% (v/v) dimethyl sulfoxide (DMSO), which is the SYPRO orange vehicle, can result in this non-equilibrium state depicted by vacuolized droplets possessing varying densities. To provide insights into the structure of the rPrP 90-231 phase-separated state, we added SYPRO orange, a dye that only exhibits fluorescence when bound to hydrophobic surfaces exposed to solvent.…”