ABSTRACT
RATIONALE Tobacco smoking, via nicotine receptor (α7 nAChR), increases risk for the susceptibility to infection from SARS-CoV-2 via increased expression of ACE2 in the lung. Given the modifiable nature and high risk of electronic cigarettes, with potential to deliver toxic concentrations of nicotine, we investigated if electronic cigarettes are also a danger in relation to COVID-19. We investigated the cytotoxicity of electronic cigarette exposure and whether flavored vaping has the potential to increase susceptibility to SARS-CoV-2 infection in large and small airway epithelial cells through the upregulation of ACE2. METHODS We exposed bronchial epithelial (BEAS-2B) and primary human small airway epithelial (SAEC) cells to e-cigarette aerosol condensates produced from propylene glycol/vegetable glycerin (PG/VG) or locally bought [Juicius Maximus, watermelon flavor (WM)] e-liquid (± added nicotine), and cigarette smoke extract (CSE). Aerosols were produced from a Drag2 e-cigarette device (Vapoo) operated at 60W, with a 0.4Ω U2 dual coil (Vapoo), and condensates were collected in flasks over dry ice with the aid of a peristaltic pump, then stored at -80°C. We investigated if e-cigarette exposure, alike cigarette smoke, increases the expression of ACE2 in lung epithelial cells. Cytotoxicity (CCK-8), membrane integrity by lactate dehydrogenase (LDH) release, and ACE2 protein expression (immunofluorescence) were measured for both 4- and 24-hour treatments;ACE2 gene expression was measured using qPCR for 4- hour treatment only. RESULTSFor both BEAS-2B cells and SAECs, nicotine-free condensates and higher concentrations of nicotine-containing condensates were highly cytotoxic (PG/VG, JM ± nicotine(60mg);∗∗∗p<0.0001, ∗∗∗p<0.001). Higher LDH release was seen in BEAS-2B cells treated for 24 hours with higher concentrations of nicotine-containing condensates (PG/VG, JM + nicotine(18 or 60mg): ∗∗∗∗p<0.0001, ∗∗p<0.01) indicating that the membrane integrity was disturbed. Cells appear to be under significant distress. Small increases in LDH were also seen with 4 hours treatment (∗∗∗∗p<0.0001, ∗∗∗p<0.001, ∗∗p<0.01∗p<0.05). ACE2 protein expression was observably increased in all treatments compared to media controls, particularly for 24 hours exposures. ACE2 gene expression was significantly increased in cells exposed to WM + 60mg/mL nicotine (∗∗p<0.1) and cigarette smoke extract (∗∗∗∗p<0.0001) compared with media controls. CONCLUSIONS Our study confirms that electronic cigarettes condensates are highly cytotoxic. Data also suggests that vaping can result in an increase in lung ACE2 expression as we have shown for tobacco smoking. The health care community needs to stay alert;vaping and smoking are avoidable risk factors for SARS-CoV-2 infection and should be avoided during and post- the COVID-19 pandemic.