Cardoso 2022 ESCI Bari

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Luiza Cardoso
Cardoso LHD, Donnelly C, Komlódi T, Gnaiger E (2022) Bioenergetic fingerprinting with coupling and pathway control of coenzyme Q redox state and respiration in permeabilized HEK 293T cells. 56th Annual Scientific Meeting of the European Society for Clinical Investigation.

Link: 56th Annual Scientific Meeting of the European Society for Clinical Investigation

Cardoso Luiza HD, Donnelly Chris, Komlodi Timea, Gnaiger Erich (2022)

Event: ESCI 2022 Bari IT

Multiple mitochondrial electron transfer (ET) pathways converge at the Q-junction and reduce coenzyme Q, which is oxidized by downstream Complexes III and IV. Diagnostic fingerprinting is challenged with targeting specific segments of pathway and coupling control of oxidative phosphorylation (OXPHOS) [1].

We characterised the Q redox state in pathway and coupling control using the Oroboros NextGen-O2k with the electrochemical Q-Module [2], monitoring simultaneously respiration and the reduced Q-fraction in permeabilized HEK 293T cells. Multiple combinations of substrates (pyruvate, malate, succinate) and inhibitors (rotenone, malonate) were used to interrogate the NADH-pathway (N), succinate-pathway (S), or their combination (NS). Coupling control was analysed in the S-pathway (succinate and rotenone), varying from LEAK respiration to OXPHOS-, and ET-capacity.

The reduced Q-fraction increased proportionally with increasing respiration, from N-, S-, and NS-pathway control. This reflects the variable ET push upstream of the Q-junction. In coupling control, the reduced Q-fraction decreased with an increase of respiration, opposite to pathway control. This is caused by the pull effect of coupling control downstream of the Q-junction: Stimulation of respiration by ADP increases the pull and Q becomes more oxidized.

Combined measurements of respiration and the Q-redox state provide unique diagnostic fingerprinting approaches for in-depth diagnostics of mitochondrial function in health and disease.

Keywords: Coenzyme Q, Q-junction, Q-redox state, Electron transfer system, ETS-reactive Q-pool, Mitochondrial coenzyme Q, Supercomplexed Q, Free Q-pool according to the fluid-state model, Qfree, Cyclic voltammetry, CV, High-resolution respirometry, HRR, Isolated mitochondria, imt, Mouse heart mitochondria, Mouse brain mitochondria, Oxygen consumption, SUIT protocols, Coupling control, Pathway control, NS-pathway, Additivity Bioblast editor: Plangger M O2k-Network Lab: AT Innsbruck Oroboros, CH Lausanne Place N, AT Innsbruck Gnaiger E, AT Innsbruck MitoFit


Affiliations and support

Luiza H.D. Cardoso1, Chris Donnelly1,2, Timea Komlódi1, Erich Gnaiger1*
  1. Oroboros Instruments, Innsbruck, Austria
  2. Institute of Sport Sciences, University of Lausanne, Switzerland
This work was part of the NextGen-O2k project, with funding from the European Union’s Horizon 2020 research and innovation programme under grant agreement nº 859770. Chris Donnelly was supported by the Swiss National Science Foundation under grant agreement nº 194964.

References

  1. Gnaiger E (2020) Mitochondrial pathways and respiratory control. An introduction to OXPHOS analysis. 5th ed. Bioenerg Commun 2020.2. https://doi.org/10.26124/bec:2020-0002
  2. Komlódi T, Cardoso LHD, Doerrier C, Moore AL, Rich PR, Gnaiger E (2021) Coupling and pathway control of coenzyme Q redox state and respiration in isolated mitochondria. Bioenerg Commun 2021.3. https://doi.org/10.26124/bec:2021-0003

List of abbreviations, terms and definitions - MitoPedia

» MitoPedia: Terms and abbreviations


Labels: MiParea: Respiration 


Organism: Human  Tissue;cell: HEK  Preparation: Permeabilized cells 


Coupling state: LEAK, OXPHOS, ET  Pathway: N, S, NS  HRR: Oxygraph-2k