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High-throughput Screening of ToxCast™ Phase I Chemicals in a Mouse Embryonic Stem Cell (mESC) Assay Reveals Disruption of Potential Toxicity Pathways
Citation:
CHANDLER, K. J., E. S. HUNTER, S. C. JEFFAY, H. P. NICHOLS, M. Hoopes, M. BARRIER, N. SIPES, N. KLEINSTREUER, A. Singh, AND T. Knudson. High-throughput Screening of ToxCast™ Phase I Chemicals in a Mouse Embryonic Stem Cell (mESC) Assay Reveals Disruption of Potential Toxicity Pathways. Presented at Society of Toxicology (SOT) annual meeting, Washington, DC, March 06 - 10, 2011.
Impact/Purpose:
Little information is available regarding the potential for many commercial chemicals to induce developmental toxicity. The mESC Adherent Cell Differentiation and Cytoxicity (ACDC) assay is a high-throughput screen used to close this data gap.
Description:
Little information is available regarding the potential for many commercial chemicals to induce developmental toxicity. The mESC Adherent Cell Differentiation and Cytoxicity (ACDC) assay is a high-throughput screen used to close this data gap. Thus, ToxCast™ Phase I chemicals were evaluated using the ACDC assay in a J1 mESC assay at 0.0125-12.5uM. Differentiation was measured by In-Cell Western™ of Myosin Heavy Chain (MHC), a cardiomyocyte marker, and cytotoxicity using DRAQ5/Sapphire700. AC50 and AC20 values were computed by curve-fitting methods. Fifty-six of 309 chemicals caused a 50% change in cytotoxiclty and/or differentiation endpoints in the concentration range tested. Comparison with 548 ToxCast™ assays revealed significant correlations between the inhibition of cardiomyocyte differentiation and perturbation of developmentally regulated genes such as PAX6 (p≤(O.15) and BMPR2 (p≤ O.003), as well as multiple components of the oxidative stress pathway (ABCG2, HIF1a, NRF2, MTF2, p53) (p ≤O.15), while strong associations between general cell-based cytotoxicity assays were shown with the ES cell cytotoxicity endpoint. In addition, a strong multivariate model suggests ABCG2, a xenobiotic transporter, along with assays that evaluate DNA texture, cell loss, mitotic arrest and steatosis, predict decreased ES cell differentiation (BA=0.84). Taken together, this work demonstrated the activity of chemicals in ES cells and identified potential disruption of developmental and oxidative stress signaling pathways by a focused group of pesticides. [This work does not necessarily reflect Agency policy.]