Thirdhand smoke: Chemical dynamics, cytotoxicity, and genotoxicity in outdoor and indoor environments

•Chemical dynamics, toxicity, and aging of THS were studied indoors and outdoors.•Chemical dynamics varied indoors vs outdoors and with the type of fabric.•In the indoor environment, TSNA increased during aging.•THS residues from relatively few cigarettes were both cytotoxic and genotoxic.•Protein enhanced the efficiency of extraction of cytotoxic chemicals from THS.

We tested the toxicity of thirdhand smoke (THS) using two controlled laboratory exposure scenarios and low levels of THS. One exposure modeled THS in a car parked outdoors, while the second modeled THS in a room without sunlight. The fabrics were exposed to cigarette smoke and then extracted in culture medium. Concentrations of nicotine, nicotine related alkaloids, and tobacco specific nitrosamines (TSNAs) were determined in fresh and aged extracts. The concentration of TSNAs increased with aging in the indoor experiment. THS extracts were used for cytotoxicity testing using mouse neural stem cells (mNSC), human dermal fibroblasts (hDF) and human palatal mesenchyme cells (hPM). Extracts from the car experiment inhibited mNSC proliferation in a live cell imaging assay and induced single strand DNA breaks in mNSC and hDF. In the indoor experiment, THS extracts made with medium containing serum proteins were significantly more toxic than extracts made with basal medium, and mNSC and hPM were more sensitive than hDF. These data indicate that: (1) aging of THS chemical differs on different fabrics and differs with and without sunlight; (2) very few cigarettes are sufficient to produce a toxic THS residue; and (3) protein enhances the efficiency of extraction of cytotoxic chemicals.

Graphical abstractGraphical overview of the assays and results. Two THS generation experiments were conducted and THS constituents were quantified using LC-MS/MS. Aging of THS chemicals, which caused the chemical composition to change, differed in indoor and outdoor environments. THS extracts were tested for cytotoxicity using the MTT assay, a live cell imaging assay, and the comet assay. THS extracts from the outdoor (car) experiment were genotoxic to mouse neural stem cells (mNSC) and human dermal fibroblasts (hDF). Extracts from the indoor experiments were highly cytotoxic to mNSC when extracted with serum proteins.Download full-size image