Interplay between proteasome function and inflammatory responses in e-cig vapor condensate-challenged lung epithelial cells

Exposure to cigarettes and other nicotine-based products results in persistent inflammation in the lung. In recent years, electronic cigarettes (e-cigs) have become extremely popular among adults and youth alike. E-cigarette vapor-induced oxidative stress promotes protein breakdown, DNA damage and cell death, culminating in a variety of respiratory diseases. The proteasome, a multi-catalytic protease, superintends protein degradation within the cell. When cells are stimulated with inflammatory cytokines such as IFN-γ and TNF-α, the constitutive catalytic proteasome subunits are replaced by the inducible subunits-low-molecular mass polypeptide (LMP)2 (β1i), multi-catalytic endopeptidase complex-like (MECL)1 (β2i), and LMP7 (β5i), which are required for the production of certain MHC class I-restricted T-cell epitopes. In this study, we used human alveolar epithelial cells (A549) and exposed them to filtered air or (1%) tobacco-flavored (TF) electronic cigarette vapor condensate (ECVC) ± nicotine (6 mg/ml) (TF-ECVC ± N) for 24 h. We observed an increase in the levels of IFN-γ, TNF-α, and inducible proteasome subunits (LMP7/PSMB8, LMP2/PSMB9, MECL1/PSMB10), and a reduced expression of constitutive proteasome subunits (β1/PSMB6 and β2/PSMB7) in challenged A549 cells. Interestingly, knockdown of the inducible proteasome subunit LMP7 reversed ECVC-induced expression of NADPH oxidase and immunoproteasome subunits in A549 cells. In addition, pre-exposure to an LMP7 inhibitor (ONX-0914) abrogated the mRNA expression of several NOX subunits and rescued the excessive production/release of inflammatory cytokines/chemokines (IL-6, IL-8, CCL2, and CCL5) in ECVC-challenged cells. Our findings suggest an important role of LMP7 in regulating the expression of inflammatory mediators during ECVC exposure. Overall, our results provide evidence for proteasome-dependent ROS-mediated inflammation in ECVC-challenged cells.