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Defining a Model for Mitochondrial Function in mESC Differentiation
Citation:
Royland, J., S. Warren, S. Jeffay, M. Hoopes, H. Nichols, AND S. Hunter. Defining a Model for Mitochondrial Function in mESC Differentiation. Presented at International Society for Stem Cell Research, Vancouver, CANADA, June 18 - 21, 2014.
Impact/Purpose:
Will be presented at the 12th International society for Stem Cell Research, Vancouver, Canada June 18-21, 2014
Description:
Defining a Model for Mitochondrial Function in mESC DifferentiationDefining a Model for Mitochondrial Function in mESC Differentiation Differentiating embryonic stem cells (ESCs) undergo mitochondrial maturation leading to a switch from a system dependent upon glycolysis to a reliance on oxidative phosphorylation. This switch acts as a signal to the cell to decrease the rate of proliferation and initiate differentiation into downstream cell lineages, a critical step in embryogenesis and a potential target for disruption by xenobiotic compounds. We investigated the sensitivity of this metabolic shift to interruption of oxidative phosphorylation at various points of the mitochondrial electron transport chain in a differentiating mouse ESC model system. The impacts of exposures to chemical inhibitors of electron transport complexes were assessed using in-cell Western analysis of the differentiation biomarker, Goosecoid (GSC), a transcription factor highly expressed during embryonic gastrulation. Additional fluorescent bioassays were used to monitor cell number, mitochondria number and cell death. The effect on cell proliferation by inhibitor exposure varied depending on the complex tested. We found inhibitors of complex I led to a 91% decrease in cell number, which was > complex V (69% decrease) ≈ complex IV (66% decrease) > complex III (25% decrease) ≈ complex II (21% decrease). The pattern of altered differentiation as measured by GSC was similar (complex I inhibitor, 97%; complexes IV and V, ~50%; and little effect in complexes II and III, 12 and <1%, respectively). In contrast, inhibitors of electron transport at complexes I, IV and V showed increased number of mitochondria/cell in the surviving cell population. Mitochondria/cell decreased following inhibition of complexes II and III (~56%). These data will be combined with other measures of mitochondrial function to help define a mitochondrial adverse outcome pathway, to expand the application of the mESC developmental model and to aid in developing in silico models for toxicity testing. (This abstract does not necessarily reflect USEPA policy).