Evaluation of in vitro Mass Balance Distributions Models for Direct Liquid Application Exposures at the Air-Liquid Interface (ALI)
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It is critical to report internal or cellular dose for New Approach Methods (NAMs) following chemical exposures to facilitate evidence integration for risk assessment and in vitro to in vivo extrapolation (IVIVE). However, analytical methods to directly measure cellular uptake in cell cultures grown at the air-liquid interface (ALI) for inhalation toxicity screening can be time-intensive and costly. In silico dosimetry tools are an attractive alternative to predict cellular dose, but they have been developed for traditional 2D cell culture systems which do not possess apical and basolateral compartments. We aimed to determine whether two mass balance models (IV-MBM DP v1.0, IV-MBM EQP v2.0) could predict cellular uptake of a positively-charged fluorescent tracer (Rhodamine 6G) following direct liquid application to human bronchial epithelial cells grown at the ALI. Both mass balance models are unique in how they calculate chemical partitioning and distribution within in vitro models and include predictions for partitioning into cells, plastic, and media components. Empirical data were obtained by extracting and quantifying rhodamine in apical solutions, basolateral media, 16HBE cells, and cell culture inserts across a variety of apical dosing solutions. Cellular uptake of rhodamine showed little correspondence to in silico predictions which suggests that current in vitro mass balance models are insufficient to predict internal doses for direct application exposures of ALI models. Until in silico dosimetry models are modified and validated for ALI cell systems, direct analytical measurements of cellular uptake are recommended. Abstract does not reflect views or policies of the U.S. EPA.