Chronic low-level JUUL aerosol exposure causes pulmonary immunologic, transcriptomic, and proteomic changes.

E-cigarettes currently divide public opinion, with some considering them a useful tool for smoking cessation and while others are concerned with potentially adverse health consequences. However, it may take decades to fully understand the effects of e-cigarette use in humans given their relative newness on the market. This highlights the need for comprehensive preclinical studies investigating the effects of e-cigarette exposure on health outcomes. Here, we investigated the impact of chronic, low-level JUUL aerosol exposure on multiple lung outcomes. JUUL is a brand of e-cigarettes popular with youth and young adults. To replicate human exposures, 8- to 12-week-old male and female C57BL/6J mice were exposed to commercially available JUUL products (containing 59 mg/ml nicotine). Mice were exposed to room air, PG/VG, or JUUL daily for 4 weeks. After the exposure period, inflammatory markers were assessed via qRT-PCR, multiplex cytokine assays, and differential cell count. Proteomic and transcriptomic analyses were also performed on samples isolated from the lavage of the lungs; this included unbiased analysis of proteins contained within extracellular vesicles (EVs). Mice exposed to JUUL aerosols for 4 weeks had significantly increased neutrophil and lymphocyte populations in the BAL and some changes in cytokine mRNA expression. However, BAL cytokines did not change. Proteomic and transcriptomic analysis revealed significant changes in numerous biological pathways including neutrophil degranulation, PPAR signaling, and xenobiotic metabolism. Thus, e-cigarettes are not inert and can cause significant cellular and molecular changes in the lungs.

Differential impact of JUUL flavors on pulmonary immune modulation and oxidative stress responses in male and female mice.

JUUL is a popular e-cigarette brand that manufactures e-liquids in a variety of flavors, such as mango and mint. Despite their popularity, the pulmonary effects of flavored JUUL e-liquids that are aerosolized and subsequently inhaled are not known. Therefore, the purpose of this study was to evaluate if acute exposure to JUUL e-cigarette aerosols in three popular flavors elicits an immunomodulatory or oxidative stress response in mice. We first developed a preclinical model that mimics human use patterns of e-cigarettes using 1 puff/min or 4 puffs/min exposure regimes. Based on cotinine levels, these exposures were representative of light/occasional and moderate JUUL users. We then exposed C57BL/6 mice to JUUL e-cigarette aerosols in mango, mint, and Virginia tobacco flavors containing 5% nicotine for 3 days, and assessed the inflammatory and oxidative stress response in the lungs and blood. In response to the 1 puff/min regime (light/occasional user), there were minimal changes in BAL cell composition or lung mRNA expression. However, at 4 puffs/min (moderate user), mint-flavored JUUL significantly increased lung neutrophils, while mango-flavored JUUL significantly increased Tnfα and Il13 mRNA in the lungs. Both the 1- and 4 puffs/min regimes significantly increased oxidative stress markers in the blood, indicating systemic effects. Thus, JUUL products are not inert; even short-term inhalation of flavored JUUL e-cigarette aerosols differentially causes immune modulation and oxidative stress responses.

Sex-specific alterations in pulmonary metabolic, xenobiotic and lipid signalling pathways after e-cigarette aerosol exposure during adolescence in mice.

BACKGROUND: E-cigarette use is now prevalent among adolescents and young adults, raising concerns over potential adverse long-term health effects. Although it is hypothesised that e-cigarettes promote inflammation, studies have yielded conflicting evidence. Our previous work showed that JUUL, a popular e-cigarette brand, elicited minimal lung inflammation but induced significant molecular changes in adult C57BL/6 mice. METHODS: Now, we have profiled immunological and proteomic changes in the lungs of adolescent male and female BALB/c and C57BL/6 mice exposed to a flavoured JUUL aerosol containing 18 mg/mL of nicotine for 14 consecutive days. We evaluated changes in the immune composition by flow cytometry, gene expression levels by reverse transcription-quantitative PCR and assessed the proteomic profile of the lungs and bronchoalveolar lavage (BAL) by tandem mass tag-labelled mass spectroscopy. RESULTS: While there were few significant changes in the immune composition of the lungs, proteomic analysis revealed that JUUL exposure caused significant sex-dependent and strain-dependent differences in lung and BAL proteins that are implicated in metabolic pathways, including those related to lipids and atherosclerosis, as well as pathways related to immune function and response to xenobiotics. Notably, these changes were more pronounced in male mice. CONCLUSIONS: These findings raise the possibility that vaping dysregulates numerous biological responses in lungs that may affect disease risk, disproportionally impacting males and raising significant concerns for the future health of male youth who currently vape.