Characterization and cytotoxicity assessment of nargile smoke using dynamic exposure

Objectives: Nargile (waterpipe) smoking has gained popularity in the Middle East and throughout the world. In this research, a new dynamic methodology was conceived. This methodology was deployed for direct in vitro assessment of cytotoxicity, genotoxicity, and apoptotic potential of smoke generated from a single nargile session. Materials and methods: A549 cells were deployed in a designed system to assess the cytotoxicity of generated smoke. The smoke was characterized using Gas chromatography-mass spectrometry (GC-MS) profiling for major organic compounds, whereas the remaining chemical and physical parameters were tabulated from published data. The cytoxicity of smoke generated from five commercial flavored tobacco products was assessed using the (3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulphophenyl)-2-H-tetrazolium) (MTS) assay. The genotoxicity was also measured using the comet assay, while apopoptosis was evaluated using Annexin V/propidium iodide staining.Results: The data indicated acute cytotoxicity emanating from smoke products in all tested tobacco flavors. Significant loss of viability and mitochondrial activity was observed 40 min post smoke exposure (Double-Apple flavored), while DNA damage onset was reported as early as 20 min of exposure. Microscopical analysis showed a systematic increase in cell rounding post exposure indicating cellular loss of adhesion and potential membrane damages. Finally, the Annexin V/propidium iodide cellular staining showed signs of late apoptosis or necrosis in exposed cells. Conclusions: The presented data clearly indicated significant in vitro cytotoxicity, genotoxicity and apoptosis/necrosis associated with a 60-min single session of nargile smoking.