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Extracellular microRNAs are sensitive biomarkers of chemical-mediated toxicity and other mechanisms of action
Citation:
Nelson, G., J. Harrill, AND B. Chorley. Extracellular microRNAs are sensitive biomarkers of chemical-mediated toxicity and other mechanisms of action. Society of Toxicology 62nd Annual Meeting and ToxExpo 2023, Nashville, TN, March 19 - 23, 2023. https://doi.org/10.23645/epacomptox.22714315
Impact/Purpose:
Poster presented to the Society of Toxicology 62nd Annual Meeting and ToxExpo March 2023. This study develops microRNA biomarkers for development of high throughput chemical screening. These efforts will support NAM development.
Description:
Previous in vivo and in vitro chemical exposure studies have indicated that dose responsive microRNAs (miRNAs) are linked to the known mechanisms of action (MoA) of chemical perturbagens. The plausibility of using extracellular miRNA profiles as non-destructive biomarkers of chemical exposure is currently being investigated. We had previously identified the presence of 181 extracellular miRNAs of human hepatic HepaRG cells by small RNA sequencing, and a panel of 65 were chosen for direct measurement in media by Abcam Fireplex assay based on highest level of presence. Cells were treated with a set of 40 chemicals representing 14 MoAs. After nine days in culture, cells were dosed over a 4- or 8-point range with half-log spacing. MiRNA and lactate dehydrogenase (LDH) release into the media were measured after 24h of exposure. MicroRNA profiling identified three basic patterns: 1) response of most measured miRNAs, 2) a partial response of miRNAs (either higher or lower compared to vehicle control), and 3) no or little change. A response of many measured miRNAs usually correlated with cellular toxicity as indicated by significant LDH release. In contrast, the Cyp3a4 inhibitors amiodarone, ketoconazole and itraconazole demonstrated a reduction of release into the media compared to control for most of the measured miRNAs. Many chemicals/doses not resulting in overt toxicity demonstrated a partial pattern of released miRNAs into the media, some of which were consistent within chemical MoA. Benchmark dose (BMD) analysis identified the potential dose-responsive miRNA biomarkers common within a MoA group. Unsupervised hierarchical clustering of both miRNA measurements and BMD responses revealed some clustering of chemicals by MoA for several groups, including HMGCR inhibitors (statins), HDAC inhibitors, PPAR-gamma activators, unfolded protein response and heat shock response. BMD estimates identified examples of altered extracellular miRNA release that were more sensitive than measurements of toxicity. This included 19 miRNAs that demonstrated BMDs more than one-half below the BMD for LDH release when HepaRG were exposed to statins. Future analyses will examine paired intracellular miRNA and mRNA profiles to further link findings with known mechanistic pathways of toxicity. In summary, extracellular miRNAs may be useful as accessible early indicators of toxicity and may inform chemical MoA. Further work to define these profiles could be invaluable for screening of chemicals and mixtures of unknown effect by a simple and non-destructive sampling of the media. This abstract does not necessarily reflect EPA policy.
URLs/Downloads:
DOI: Extracellular microRNAs are sensitive biomarkers of chemical-mediated toxicity and other mechanisms of action
230228_SOT_MIRNA_NELSON FINAL TO STICS.PDF (PDF, NA pp, 759.797 KB, about PDF)