Challenges in developing high-throughput assays to evaluate inhalation toxicity of volatile organic compounds and aerosols

Inhalation is one of the three primary modes of chemical exposure, but key challenges in developing reproducible exposure systems with robust analytical dosimetry methods have limited the use of in vitro models for the assessment of inhaled chemicals. Direct cell-toxicant interaction at ALI mimics realistic human exposures, but volatile organic compounds (VOCs) and aerosols possess distinct transport and deposition mechanisms which complicate the reliability and reproducibility of ALI exposure technology unless these are addressed. To meet screening demands for a diverse list of TSCA-nominated inhalable compounds, we utilized fluorescent tracers and Computational Fluid Dynamics (CFD) modeling to develop an Aerosol-compatible Cell Culture Exposure System (ACCES) which achieves serial dilution of aerosols and volatile compounds at ALI. This design incorporates six exposure concentrations with four technical replicates per dose within a standard 24-well plate and allows real-time sampling of VOC concentrations via gas chromatography (GC)-coupled detection methods. However, this same real-time sampling approach cannot be utilized to quantify aerosol concentrations without impacting the performance of the ACCES. As an alternative approach, we aerosolized two fluorescent tracers (sodium fluorescein and rhodamine 6G) and developed methods to directly quantify deposition and cellular uptake in human bronchial epithelial cells. Both fluorescent tracers revealed that cell-free alternatives (filters, cell culture media, etc.) may not accurately predict cell deposition at ALI and should be used with caution. Deposition studies in two other commercial ALI exposure systems, the VITROCELL Cloud and MedTec CelTox, confirmed these results. Overall, this presentation focuses on the need for different approaches to evaluate toxicity for both volatile organic compounds and aerosols; and highlights efforts to improve dosimetry methods by directly quantifying cell deposition.

Impact/Purpose

Inhalation is one of the three primary modes of chemical exposure, but key challenges in developing reproducible exposure systems with robust analytical dosimetry methods have limited the use of in vitro models for the assessment of inhaled chemicals. Overall, this presentation focuses on the need for different approaches to evaluate toxicity for both volatile organic compounds and aerosols; and highlights efforts to improve dosimetry methods by directly quantifying cell deposition.

Citation

Murray, J. Challenges in developing high-throughput assays to evaluate inhalation toxicity of volatile organic compounds and aerosols. Society of Toxicology, Salt Lake City, UT, March 10 - 14, 2024.