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Metabolism


Metabolism is the sum of biochemical processes that either produce or consume energy. Metabolic dysfunction can be the cause as well as the consequence of many human diseases. Ncardia offers a range of assays to monitor several indicators of the metabolic profile on iPSC-derived cell models. This provides an adaptable platform to assess the impact of candidate compounds on cell metabolism with relevant human in vitro models.

Assay
Benefits
  • Monitor key indicators of the metabolic profile
  • Extensive menu of assays available
  • Multiplexing options
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Applications

Abnormal cellular metabolism is involved in the development of atherosclerosis, cardiomyopathies and neurodegenerative disorders, among other human diseases. Our scientists have developed and optimized several assays to evaluate the most relevant indicators for metabolic performance, including mitochondrial function, glycolytic activity, oxidative stress and more:

  • Oxygen Consumption Rate (OCR) assay
    Evaluation of the oxidative phosphorylation capabilities of cells as an indicator of the mitochondrial function.
  • Extracellular Acidification Rate (ECAR) assay
    Evaluation of the glycolytic activity in cells by recording lactate production.
  • Intracellular Oxygen Concentration assay
    Quantification of real-time cellular oxygenation under hypoxic conditions and upon drug treatment to investigate the interplay between cellular oxygenation and metabolism.
  • TMRE, MitoTracker and JC-1 assays
    Monitoring of the mitochondrial membrane potential in whole cells as an indicator of mitochondrial function.
  • MitoSOX assay
    Assessing the superoxide concentration inside the mitochondria as an indicator of reactive oxygen species toxicity.
  • CellROX assay
    Novel fluorogenic probe to evaluate oxidative stress in live cells.

Pairing OCR and the ECAR metabolic assays for a comprehensive characterization of metabolic activity in Ncardia’s human iPSC-derived cardiomyocytes.

The combination of these two metabolic assays with physiologically relevant models provides a good indication of drug-induced changes on cellular metabolism to determine drug efficacy and safety. Here we tested the effect of two known compounds, oligomycin and carbonyl cyanide-4 (trifluoromethoxy) phenylhydrazone (FCCP).

Oligomycin inhibits ATP synthase resulting in reduced mitochondrial respiration (OCR) and increase glycolysis (ECAR). Carbonyl cyanide-4 phenylhydrazone (FCCP) collapses the proton gradient and disrupts mitochondrial membrane potential increasing oxygen consumption (OCR) with no or little effect on glycolysis (ECAR).

Assessing mitochondrial depolarization in Ncyte CNS neurons to quantify cytotoxic damage of compounds.

Mitochondrial membrane potential controls respiratory rate, ATP production and generation of reactive oxygen species, which are essential processes for cell homeostasis. Our scientists use confocal microscopy and JC-1 dye as a sensitive readout of mitochondrial depolarization and disruption after treatment with the therapeutic compounds of interest.

Assessment of oxidative stress levels in Ncyte Cardiomyocytes to measure changes in cardiac metabolism during ischemia/reperfusion.

During myocardial infarction, oxygen supply to cardiac tissue is reduced and it is rapidly restored after reperfusion treatment, inducing changes in cell metabolism. Our scientists can model ischemia/reperfusion on human iPSC-derived cardiomyocytes by using an atmospheric control unit (ACU) and measure oxygen consumption in real time to study cells’ metabolism under different environmental conditions.

Cell oxygenation traces from cardiomyocytes with oxygen consumption (light red) and from cardiomyocytes in hypoxia (dark red). The black trace represents the oxygen level inside the chamber.