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Project Number: CSS 12.01 Adverse Outcome Pathway Discovery and Development (AOP-DD)Task Number: Task 1.1d: Putative AOP development related to Fatty Liver Disease (steatosis) Product number: Key Product 1 (AOP knowledgebase development)
Citation:
Angrish, M., J. Rooney, C. McQueen, AND B. Chorley. Project Number: CSS 12.01 Adverse Outcome Pathway Discovery and Development (AOP-DD)Task Number: Task 1.1d: Putative AOP development related to Fatty Liver Disease (steatosis) Product number: Key Product 1 (AOP knowledgebase development). Society of Toxicology Annual Meeting, Baltimore, MA, March 12 - 16, 2017.
Impact/Purpose:
This work describes the development of a targeted adverse outcome pathway based toxicity test for hepatic steatosis. This targeted approach to toxicity testing not only identifies chemical hazards, but also helps to elucidate the mechanism causing that adverse outcome.
Description:
Expansion of the Steatosis Adverse Outcome Pathway to Mechanistic Toxicity TestsMichelle M. Angrish†, John P. Rooney‡, Charlene A. McQueen†, and Brian N. Chorley†*†National Health and Environmental Effects Research Laboratory, ORD, US EPA, RTP, NC27709, United States,‡ORISE Participant, US EPA, Research Triangle Park, NCAbstractCurrently, risk assessors utilizes a number of liver endpoints, including steatosis, in rodent toxicology studies to assess the safety of exogenous chemical exposures. Critically, there is often a biological disconnect between rodent physiologic responses and the human condition. Furthermore, while liver function tests help to screen for and evaluate liver damage, they are not predictive of progressive disease. For these reasons and others including cost and throughput of chemical assessment, in vitro based human cell models are being developed to better predict adverse human health outcomes. Here, we utilized a networked Adverse Outcome Pathway to assess key biological events that can lead to the initiation and progression of steatosis. Assays for lipid uptake, lipid efflux, fatty acid oxidation, lipid accumulation, total nuclei, cytotoxicity, and gene expression were applied in HepaRG cells. Six compounds were screened using the assays and include: T0901317 (a liver X receptor [LXR] agonist), cyclosporin A (an immunosuppressant used in organ transplant), 22(r)-hydroxycholesterol (an endogenous LXR agonist), wyeth-14,643 (a peroxisome proliferator activated receptor [PPAR] α agonist), troglitazone (a PPARγ agonist), and amiodarone (an antiarrhythmic medication). Effects were considered significant if cell viability was ≥70% and the |log2 fold change| of test samples was ≥3 times the baseline median absolute deviation of DMSO controls. Only T0901317 and cyclosporin A dose-dependently increased hepatocyte lipids. T0901317 decreased lipid efflux, increased triglyceride synthesis and lipid efflux genes Dgat1, Dgat2, and ApoB100, and decreased lipid synthesis genes Fasn, Pparg, Acaca, and Srebf. Cyclosporin A increased fatty acid uptake and efflux. 22(R)-hydroxycholesterol and wyeth-14,643 increased fatty acid uptake and Cd36 mRNA. Troglitazone and amiodarone decreased fatty acid uptake and Cd36. These results suggest T0901317 post-transcriptionally altered lipid efflux pathways, whereas cyclosporin A caused steatosis by altering lipid efflux and uptake. This work is an important step towards developing an in vitro test to screen chemicals for potential to cause steatosis. This abstract does not necessarily reflect official Agency policy.