BTEX in vitro exposure tool using human lung cells: trips and gains

Abstract

Cytotoxicity of benzene, toluene, ethylbenzene and xylenes (BTEX) to human lung cells was explored using three different exposure methods: Method 1 - in normal 96-well plates using DMSO as a carrier vehicle, we exposed (a) human lung carcinoma A549 cells, (b) A549 cells over-expressed with cytochrome P450 2E1 cells, and (c) normal lung fibroblast LL-24 cells to benzene, toluene, ethylbenzene and xylene individually and in a mixture which models car exhaust gases for between 1-88 h. We found that the order of the BTEX potency is benzene<toluene<ethylbenzene=m-xylene with acute BTEX toxicity to A549≈LL-24>CYP2E1 over-expressed A549 cells. A significant difference was found between inter-assay responses for all 24h exposures (P<0.005) suggesting a poor assay repeatability. No sign of potency increase was found from 6 to 72 h exposures. Method 2 - Using sealed vials to expose A549 cells to benzene, toluene and ethylbenzene, we observed a twenty-fold increase in their cytotoxicity, but also with no time-course effect. Method 3 - Using air exposed hanging-drop cell culture, we were able to see both an increase of demonstration of toxicity and a time-course effect from 1 to 12h exposure. We conclude that exposing cells in sealed and unsealed media using DMSO as a carrier vehicle was not suitable for BTEX exposure studies. Hanging-drop air exposure has more potential. It should be noted that if there are any changes in their exposure matrixes, its exposure mass distribution in cells could differ.

Keywords: A549; BTEX; Hanging drop; In-vitro air exposure; Media exposure.