What are the effects of e-cigarettes and their constituents on SARS-CoV-2 infection?

In a recent study posted to the bioRxiv* preprint server, researchers evaluated whether e-cigarette (EC) aerosols promoted severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection.

Study: New Insights into How JUUL™ Electronic Cigarette Aerosols and Aerosol Constituents Affect SARS-CoV-2 Infection of Human Bronchial Epithelial Cells. Image Credit: Hazem.m.kamal/Shutterstock
Study: New Insights into How JUUL™ Electronic Cigarette Aerosols and Aerosol Constituents Affect SARS-CoV-2 Infection of Human Bronchial Epithelial Cells. Image Credit: Hazem.m.kamal/Shutterstock

This news article was a review of a preliminary scientific report that had not undergone peer-review at the time of publication. Since its initial publication, the scientific report has now been peer reviewed and accepted for publication in a Scientific Journal. Links to the preliminary and peer-reviewed reports are available in the Sources section at the bottom of this article. View Sources

Background

Smoking can cause lung diseases, including cancer. Although there is an intense interest in exploring the association between tobacco/nicotine use and the coronavirus disease 2019 (COVID-19), the relationship is often contradictory and poorly defined. For instance, patient-derived data indicate that smoking is protective against COVID-19, while many studies, including meta-analyses, have found smoking as a risk factor for COVID-19 progression.

Further, elevated levels of angiotensin-converting enzyme 2 (ACE2) receptors have been detected in the respiratory tract biopsies from smokers, underscoring the increased susceptibility of smokers. ECs are nicotine delivery devices that heat e-liquids to generate aerosols containing multiple chemical substances. EC aerosols could have adverse effects on the respiratory system.

The study and findings

The present study tested whether EC aerosols enhanced SARS-CoV-2 infection in the human bronchial epithelial cells. BEAS-2B cells were used to examine the effects of ‘JUUL Virginia Tobacco’ fluids, aerosols, and constituents of JUUL products on ACE2 levels.

Cells were treated in a submerged culture with 0.5% of 5% JUUL Virginia Tobacco fluid (containing 0.3 mg/ml nicotine after dilution), 0.03 or 0.3 mg/ml of nicotine, 0.5% propylene glycol/vegetable glycerol (PG/VG), or a mix of 0.5% PG/VG with 0.03 or 0.3 mg/ml nicotine. Micrographs of cells labeled with ACE2 antibodies revealed elevated ACE2 expression in those exposed to 0.3 mg/ml of nicotine, further confirmed by western blotting analysis.

The increase in ACE2 levels by nicotine occurred in a dose-dependent manner. ACE2 levels were increased in JUUL-treated cells, but the increase was insignificant relative to controls. Similarly, BEAS-2B cells were cultured at an air-liquid interface (ALI) in the Cultex exposure system or a Vitrocell cloud chamber.

Cells were exposed to one puff of aerosols generated from phosphate-buffered saline (PBS) or 0.3 or 0.03 mg/ml nicotine in PBS. ACE2 expression was higher in nicotine-treated cells. In contrast, cells were exposed to ten puffs of humified clean air or aerosols from JUUL Virginia Tobacco EC. Additionally, the JUUL device was used with other refillable pods consisting of lab-made fluids (of PG/VG or a mix of PG/VG with 6 mg/ml or 60 mg/ml of nicotine).

ACE2 expression increased in cells exposed to PG/VG or JUUL aerosols, albeit insignificant; aerosols from lab-made fluids significantly increased ACE2 expression. Next, submerged treatments were performed to examine the effects of nicotine, PG/VG, or JUUL aerosols on transmembrane protease, serine 2 (TMPRSS2) levels, and activity. TMPRSS2 expression increased in cells exposed to 0.3 mg/ml of nicotine.

Cells exposed to lab-made fluids containing nicotine exhibited a dose-dependent increase in TMPRSS2 levels. TMPRSS2 activity, measured by the cleavage of a specific substrate, increased when cells were treated with JUUL fluids. Similar experiments were repeated at ALI using the cloud chamber or the exposure system. In the cloud chamber, nicotine exposure did not affect TMRPSS2 levels, but enzymatic activity was significantly higher with 0.3 mg/ml of nicotine.

In the Cultex system, TMPRSS2 levels were not significantly different from those in controls (exposed to clear air); however, enzymatic activity was significantly reduced in PG/VG-exposed cells and increased in cells exposed to PG/VG-nicotine mix but remained unchanged in cells exposed to JUUL aerosols.

Last, the authors infected how JUUL fluids/aerosols or individual constituents affect SARS-CoV-2 infection of BEAS-2B cells. SARS-CoV-2 pseudoviruses were generated, which expressed SARS-CoV-2 spike and contained ZsGreen, a reported plasmid to identify infected cells. Twenty-four hours after submerged treatment, pseudoviruses were added to the cultures and incubated for 24 hours.

Fluorescence was measured using flow cytometry and microscopy. Viral infection increased significantly in JUUL fluid-treated cells. In PG/VG-treated cells, viral infection was marginally elevated, nicotine treatment induced a dose-dependent increase in viral infection, and infection was higher when nicotine was used with PG/VG.

Similarly, pseudovirus infection was carried out at ALI. Nicotine (0.3 mg/ml) exposure slightly enhanced viral infection. Enhanced infection was observed in cells exposed to JUUL aerosols and the PG/VG-nicotine mix.

Conclusions

The study demonstrated that JUUL fluids and aerosols increased ACE2 expression, the activity of TMRPSS2, and the infection of BEAS-2b cells. The effect of ECs on viral infection relied on exposure type and chemical formulations. In submerged cultures and cloud chamber exposures, nicotine enhanced ACE2 expression and pseudovirus infection.

Contrastingly, in the Cultex system, nicotine and PG/VG were associated with enhanced ACE2 expression and infection. Although TMPRSS2 levels and activity were higher in submerged cultures, only enzymatic activity was elevated in ALI exposures. In summary, the findings suggested that JUUL aerosols modulated cellular infection machinery that enhanced infection by SARS-CoV-2 pseudo-particles.

This news article was a review of a preliminary scientific report that had not undergone peer-review at the time of publication. Since its initial publication, the scientific report has now been peer reviewed and accepted for publication in a Scientific Journal. Links to the preliminary and peer-reviewed reports are available in the Sources section at the bottom of this article. View Sources

Journal references:

Article Revisions

  • May 15 2023 - The preprint preliminary research paper that this article was based upon was accepted for publication in a peer-reviewed Scientific Journal. This article was edited accordingly to include a link to the final peer-reviewed paper, now shown in the sources section.
Tarun Sai Lomte

Written by

Tarun Sai Lomte

Tarun is a writer based in Hyderabad, India. He has a Master’s degree in Biotechnology from the University of Hyderabad and is enthusiastic about scientific research. He enjoys reading research papers and literature reviews and is passionate about writing.

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