In vitro methods for Nrf2 pathway induction: Evaluation of the rotenone-induced activation of the Nrf2 pathway in a stem cell-derived neuronal model
Oxidative stress is one of the cellular responses determined at early stages of toxicity and can be induced by many different classes of chemicals. The nuclear factor (erythroid-derived 2)-like 2 (Nrf2) represents one of the key regulators of the Antioxidant-Response-Element-(ARE)-driven cellular defence mechanisms against oxidative stress. Nrf2 has been shown to have a cytoprotective, antioxidant and anti-inflammatory role in virtually all cell types in the body, both under physiological and disease conditions, as it regulates the expression of enzymes involved in detoxification processes. The SCR&Tox consortium, part of the SEURAT-1 research initiative, established to investigate Nrf2 signaling activation in neuronal models derived from hiPSCs. Analysis of Nrf2 signaling activation could serve as a base for the establishment of a horizontal assay to assess oxidative stress induction. Neuronal and glial cells undergo oxidative stress as an early response to different classes of chemicals, including inhibitors of mitochondrial respiration, such as rotenone, which is known to inhibit complex I of the mitochondrial respiratory chain. Moreover, human induced pluripotent stem cells (hiPSCs) are currently regarded as a powerful tool for drug and chemical screening and development of new in vitro testing strategies in the field of toxicology, including neurotoxicity and developmental neurotoxicity evaluation.
This technical report summarizes the strategy adopted by EURL-ECVAM to assess Nrf2 signalling activation in neuronal models derived from hiPSCs. In particular, as agreed within the SCR&Tox consortium, two hiPSC-neuronal models were used: (i) an Nrf2-Luciferase reporter neural stem cell model (called Clone 27 (Cl27), obtained from Phenocell, France), and (ii) a wild type IMR90-hiPSC-derived neuronal/glial model (obtained from I-Stem, France). The results obtained with the wild type (IMR90-hiPSC) neuronal model indicate that rotenone induces the activation of Nrf2 signaling, increases the number of astroglial cells and decreases the number of dopaminergic neurons. The obtained results suggest that hiPSC-derived mixed neuronal/glial culture could be a valuable in vitro model for the establishment of neuronal specific assays to evaluate Nrf2 pathway activation (biomarker of oxidative stress) using neuronal specific readouts that could be applied to in vitro neurotoxicity evaluation.
PISTOLLATO Francesca;
CASADO POBLADOR Juan;
PRICE Anna;
2016-11-09
Publications Office of the European Union
JRC103676
978-92-79-63194-8,
1831-9424,
EUR 28172 EN,
OP LA-NA-28172-EN-N,
https://publications.jrc.ec.europa.eu/repository/handle/JRC103676,
10.2787/36841,
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