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Use of cellular ethanol metabolism for screening the effects of volatile agents
Citation:
Madden, M., B. Winters, M. Wallace, A. Ghio, L. Dailey, AND J. Pleil. Use of cellular ethanol metabolism for screening the effects of volatile agents. International Association of Breath Research, Longborough, England, UK, September 08 - 11, 2019.
Impact/Purpose:
This presentation will illustrate improved methodology related to high through-put screening of the potential toxicity of gaseous chemicals.
Description:
Background Many agents with insufficient toxicity data are gas phase and cannot be examined using standard cell culture, with media impeding the delivery of the agent to cells. We are developing a system using cells that can tolerate a lack of fluid medium on the apical side for potential screening of gaseous agents. Changes in the conversion of an exogenously added substrate to a gaseous metabolite in vitro can be utilized to monitor and screen biological responses induced by an agent of interest (i.e., “probe molecule approach” to toxicity testing). Methods BEAS-2B human airway epithelial cell line cultures were incubated with or without ethanol (1-2 %) in a flow-through system and the production of acetaldehyde (C2) and other gaseous carbonyls were collected on the outflow line with dinitrophenylhydrazine packed cartridges. Samples were analyzed by HPLC-UV. Results Results showed increased amounts of C2 with ethanol exposure (1 and 2% for 2 and 6 hr, respectively) compared to vehicle controls. Control blanks showed negligible C2 background. Cell viability was >95% by trypan blue exclusion. In separate studies where the cells were allowed to desiccate, carbonyls in the C4-C6 range were observed, suggesting these carbonyls are possible gaseous biomarkers of cell death. Conclusions The use of BEAS-2B cells and this flow-through gas system shows promise for 1) being able to expose cells to a volatile substance (ethanol) and preserve cell viability for several hours, and 2) capture a gas phase metabolite for use in probe molecule approaches for screening the induction of toxicity by a gas phase agent. Further optimization of the system is underway. The use of in vitro metabolism for chemical screening may reflect in vivo metabolism responses that can be captured in exhaled breath. [This abstract may not reflect official US EPA policy.]