A comparative study of two in-vitro methodologies used to assess E-cigarette cytotoxicity

Though widely perceived as harmless, limited information is available on E-cigarette (EC) cytotoxicity. Numerous factors impede EC research, including cell model employed and method of exposure. The present study aims to compare two in vitro methodologies to explore the effects of these choices. Bronchial epithelial cells BEAS-2B were plated in 24 well plates at a density of 3x105 cells/mL/well. Aqueous extracts of a brand anonymised strawberry flavoured EC (ECE) and tobacco cigarette (CSE) were obtained using standard methods and applied to the epithelial cell cultures. Additionally, a multi-cellular human airways model consisting of bronchial epithelial cells and human pulmonary fibroblasts co-cultured at air-liquid interface (ALI) was exposed to the same strawberry flavoured EC vapour (ECV) and whole cigarette smoke (WCS) for 7 mins according to ISO standard using an in-house built smoking machine. 24h post exposure, XTT cell viability analysis showed that, in submerged cultures, ECE and CSE significantly decreased cell viability to 43.60 – 3.65% (p < 0.0001) and 46.27% – 11.37% (p < 0.0001) respectively, compared to untreated cells. In contrast, while exposure of ALI cultures to WCS significantly reduced cell viability to 60.63 – 4.20% (p < 0.0001) compared to cells exposed to air only, ECV exposure had no significant effect on cell viability (103.6 – 15.62% control). These results indicate that the choice of cell model and EC exposure system can significantly impact upon EC cytotoxicity. It also highlights the urgent need for standard testing protocols for EC safety assessment.