EpiAirway Nrf2 – oxidative stress model: Practical application of in vitro systems with clinical relevance
The nuclear factor erythroid 2-related factor 2 (Nrf2) signaling pathway regulates genes that contribute to the antioxidative stress response. This pathway is involved in respiratory diseases, including chronic obstructive pulmonary disease (COPD). Cigarette smoke has been shown to activate this pathway in lung. Here, we evaluated the effects of whole smoke/aerosol from two marketed combustible (nonmenthol and menthol) cigarettes (CC), two smoke-free alternative products: glo™ (heated tobacco product, HTP), and Vuse Alto® (electronic nicotine delivery system, ENDS) on cell viability and Nrf2 response in a 3D human airway model (EpiAirway™) transfected with a luciferase Nrf2 promoter. EpiAirway™ tissues were exposed at the air liquid interface to whole smoke or aerosol generated on a Vitrocell® VC10®. Whole smoke/aerosol doses were controlled using dilution airflows of 0.5 to 8 L/min for CC, and undiluted to 4 or 5 L/min for HTP or ENDS, respectively. Eighteen hours post-exposure, luciferase activity and cell viability were measured. Relative luciferase activity was expressed as fold change over the air exposed control. Post-exposure, whole smoke/aerosol deposition was quantified using chemical analysis (e.g., glycerol, nicotine, carbonyls).
Differential Nrf2 activation was observed following exposure to CC whole smoke compared to the tested HTP and the ENDS. Maximum Nrf2 fold increase occurred at the undiluted dose for the HTP and ENDS versus 3 L/min for the CC. Moreover, the minimum exposure-correlated nicotine concentration required to induce a > 2-fold increase (threshold response) in Nrf2 activation was > 30× and > 100× lower for CC than the HTPs and ENDS, respectively.
These data indicate that the tested HTP and ENDS induce significantly lower oxidative stress than the CC. Additionally, the 3D Nrf2 EpiAirway™ in vitro model can be used to assess and discriminate responses of a biomarker (oxidative stress) relevant to smoking-related disease pathways (e.g., respiratory and cardiovascular disease).