Protein thiol oxidation in the rat lung following e-cigarette exposure.

E-cigarette (e-cig) aerosols are complex mixtures of various chemicals including humectants (propylene glycol (PG) and vegetable glycerin (VG)), nicotine, and various flavoring additives. Emerging research is beginning to challenge the "relatively safe" perception of e-cigarettes. Recent studies suggest e-cig aerosols provoke oxidative stress; however, details of the underlying molecular mechanisms remain unclear. Here we used a redox proteomics assay of thiol total oxidation to identify signatures of site-specific protein thiol modifications in Sprague-Dawley rat lungs following in vivo e-cig aerosol exposures. Histologic evaluation of rat lungs exposed acutely to e-cig aerosols revealed mild perturbations in lung structure. Bronchoalveolar lavage (BAL) fluid analysis demonstrated no significant change in cell count or differential. Conversely, total lung glutathione decreased significantly in rats exposed to e-cig aerosol compared to air controls. Redox proteomics quantified the levels of total oxidation for 6682 cysteine sites representing 2865 proteins. Protein thiol oxidation and alterations by e-cig exposure induced perturbations of protein quality control, inflammatory responses and redox homeostasis. Perturbations of protein quality control were confirmed with semi-quantification of total lung polyubiquitination and 20S proteasome activity. Our study highlights the importance of redox control in the pulmonary response to e-cig exposure and the utility of thiol-based redox proteomics as a tool for elucidating the molecular mechanisms underlying this response.