Abstract
Chloropicrin (CP) is a common agricultural fumigant historically used as a chemical warfare agent and is a concern for potential use in warfare and terrorist applications. Our inability to effectively treat CP-induced injuries makes it essential to better understand CP toxicity. We set out to elucidate variables that must be understood to achieve optimal exposure conditions for in vitro investigations given that such models are important for the study of CP injury and potential therapeutics. To this end, we evaluated the effects of volatility, cell seeding density, and serum concentration of cell culture medium on CP toxicity in an immortalized human corneal epithelial cell line. We found that even with very dilute solutions, CP remained highly volatile, so much so that a 0.0019% CP solution resulted in 90% cell death at time 0, but was nearly nontoxic 45 min later. Not surprisingly, the CP-induced IL-8 response was shown to vary with cell viability in this experiment. After exposure with 0.00115% CP, cells that were 12% confluent experienced over 40% more cell death than cells exposed at 87% confluency. Exposure with the same CP dose in medium containing concentrations of fetal bovine serum (FBS) ranging from 0.1% to 15% exhibited a 17% difference in cell viability. Given that chemical toxicity can be significantly influenced by volatility, cell density, and serum content of cell culture medium, these phenomena should be explored during the development and optimization of toxicant exposure models.
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The views expressed in this manuscript are those of the author(s) and do not reflect the official policy of the Department of Army, Department of Defense, or the U.S. Government. This research was supported in part by an appointment to the Postgraduate Research Participation Program at the U.S. Army Medical Research Institute of Chemical Defense administered by the Oak Ridge Institute for Science and Education through an interagency agreement between the US Department of Energy and USAMRDC.
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This research was supported by an interagency agreement (AOD18016-001-00000) between NIH/NIAID and the USAMRICD.
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ALR: was the principal investigator for the study. He was responsible for the overall study, participated in the execution of all experiments and is responsible for the final draft of the manuscript. BTC: designed all studies, participated in the execution of all experiments, analyzed all data and drafted the majority of the manuscript. AFM: assisted in study design, participated in execution of all experiments, and reviewed the final draft of the manuscript.
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Chalmers, B.T., Merriman, A.F. & Ruff, A.L. Considerations for the optimization of in vitro models of chloropicrin toxicity. Arch Toxicol 97, 255–261 (2023). https://doi.org/10.1007/s00204-022-03400-4
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DOI: https://doi.org/10.1007/s00204-022-03400-4