Airway Exposure to E-Cigarette Vapors Impairs Autophagy and Induces Aggresome Formation

Electronic cigarettes (e-cigarettes) are proposed to be a safer alternative to tobacco cigarettes. Hence, we evaluated if e-cigarette vapors (eCV) impair cellular proteostasis similar to cigarette smoke exposure.

First, we evaluated the impact of eCV exposure (2.5 or 7.5 mg) on Beas2b cells that showed significant increase in accumulation of total polyubiquitinated proteins (Ub, insoluble fractions) with time-dependent decrease in proteasomal activities from 1 h (p < 0.05), 3 h (p < 0.001) to 6 h (p < 0.001) of eCV exposure compared to room air control. We verified that even minimal eCV exposure (1 h) induces valosin-containing protein (VCP; p < 0.001), sequestosome-1/p62 (aberrant autophagy marker; p < 0.05), and aggresome formation (total poly-Ub-accumulation; p < 0.001) using immunoblotting (IB), fluorescence microscopy, and immunoprecipitation (IP). The inhibition of protein synthesis by 6 h of cycloheximide (50 μg/ml) treatment significantly (p < 0.01) alleviates eCV-induced (1 h) aggresome bodies. We also observed that eCV (1 h)-induced protein aggregation can activate oxidative stress, apoptosis (caspase-3/7), and senescence (p < 0.01) compared to room air controls. We verified using an autophagy inducer carbamazepine (20 μM, 6 h) or cysteamine (250 μM; 6 h, antioxidant) that eCV-induced changes in oxidative stress, poly-ub-accumulation, proteasomal activity, autophagy, apoptosis, and/or senescence could be controlled by autophagy induction. We further confirmed the role of acute eCV exposure on autophagy impairment in murine lungs (C57BL/6 and CD1) by IB (Ub, p62, VCP) and IP (VCP, p62), similar to in-vitro experiments.

In this study, we report for the first time that eCV exposure induces proteostasis/autophagy impairment leading to oxidative stress, apoptosis, and senescence that can be ameliorated by an autophagy inducer.

eCV-induced autophagy impairment and aggresome formation suggest their potential role in chronic obstructive pulmonary disease-emphysema pathogenesis. Antioxid. Redox Signal. 00, 000-000.