The nuclear factor erythroid 2-related factor 2 (Nrf2) signaling pathway, activated in human lung cells by cigarette smoke, regulates genes involved in the antioxidative stress response. Here, we evaluated the effect of cigarette whole smoke (3R4F reference cigarette) and whole aerosol from a commercially available tobacco heating product (THP) on cell viability and Nrf2 response in a 3D human airway model (EpiAirway™) transfected with a luciferase Nrf2 promoter.

EpiAirway™ tissues were exposed to smoke/aerosol generated under the Health Canada Intense regimen. Smoke/aerosol doses were controlled using dilution airflows of 0.5 to 6 L/min for 3R4F, and undiluted to 3 L/min for the THP. Eighteen hours post-exposure, luciferase activity and cell viability were measured. Post-exposure, smoke/aerosol deposition was also quantified using chemical analysis (e.g., glycerol, nicotine) and fluorescence of captured particulate matter in a dosimetry well within the exposure module.

Differential Nrf2 activation was observed following exposure to 3R4F smoke compared to the THP aerosol. A peak response of 1,484 ± 184-fold Nrf2 activation at 1.5 L/min (7.09 ± 1.58 µg nicotine) was observed for 3R4F while no meaningful response (<2 fold) was generated by the THP under comparable conditions. Undiluted aerosol (37.67 ± 2.44 µg nicotine) was required for the THP to induce a response: 445 ± 103-fold change. Cell viability remained >80% at these airflows. Moreover, the minimum exposure-correlated nicotine concentration required to induce a >2-fold increase in Nrf2 activation was significantly (p<0.001) lower for 3R4F than THP.

Collectively, these data show that changes in the Nrf2 promoter response may be useful in discriminating response to smoke/aerosol exposures from different tobacco product types.