Zdrazilova 2021 MitoFit ace-sce

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Zdrazilova 2021 MitoFit ace-sce

Publications in the MiPMap
Zdrazilova L, Hansikova H, Gnaiger E (2021) Comparable respiratory activity in attached and suspended human fibroblasts. MitoFit Preprints 2021.7. doi:10.26124/mitofit:2021-0007

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Comparable respiratory activity in attached and suspended human fibroblasts


Zdrazilova Lucie, Hansikova Hana, Gnaiger Erich (2021-09-21) MitoFit Prep

Abstract: Version 1 (v1) 2021-09-21 doi:10.26124/mitofit:2021-0007

Measurement of oxygen consumption of cultured cells is widely used for diagnosis of mitochondrial diseases, drug testing, biotechnology and toxicology. Fibroblasts are cultured in monolayers but physiological measurements are carried out in suspended or attached cells. We address the question whether respiration differs in attached and suspended cells using multiwell respirometry (Agilent Seahorse XF24) and high-resolution respirometry (Oroboros O2k), respectively. Respiration of human dermal fibroblasts measured in culture medium was baseline-corrected for residual oxygen consumption and expressed as oxygen flow per cell.

No differences were observed in ROUTINE respiration of living cells and LEAK respiration obtained after inhibition of ATP synthase by oligomycin. Multiple steps of uncoupler titrations in the O2k allowed for evaluation of maximum electron transfer capacity, which was higher than respiration obtained in the XF24 due to a limitation to two uncoupler titrations.

Quantitative evaluation of respiration measured in different platforms revealed that short-term suspension of fibroblasts did not affect respiratory activity and coupling control. Consistent results obtained with different platforms provide a test for reproducibility and allow for building an extended respirometric database.

Keywords: attached cells; ace, suspended cells; sce, fibroblasts, high-resolution respirometry; HRR, Oroboros O2k, Seahorse Bioanalyzer XF24, ROUTINE respiration; R, LEAK respiration; L, electron transfer capacity; E, residual oxygen consumption; Rox, bioenergetic cluster analysis; BCA Bioblast editor: Gnaiger E O2k-Network Lab: AT Innsbruck Oroboros, CZ Prague Zeman J

ORCID: ORCID.png Zdrazilova Lucie, ORCID.png Hanskova Hana, ORCID.png Gnaiger Erich

Data availability

Original files are available Open Access at Zenodo repository: 10.5281/zenodo.5518059

References

LinkReferenceViewYear
Gnaiger E (2021) Bioenergetic cluster analysis – mitochondrial respiratory control in human fibroblasts. MitoFit Preprints 2021.8. doi:10.26124/mitofit:2021-0008
MitoFit pdf
Bioenergetic cluster analysis – mitochondrial respiratory control in human fibroblasts
2021-09-21

Support

Template NextGen-O2k.jpg
This work was partially funded by the European Union’s Horizon 2020 research and innovation programme under grant agreement No. 859770, NextGen-O2k project, and by Institutional project GAUK110119. Contribution to COST Action CA15203 MitoEAGLE with financial support of Short-Term Scientific missions (LD).

Cited by

Gnaiger E (2021) Bioenergetic cluster analysis – mitochondrial respiratory control in human fibroblasts. MitoFit Preprints 2021.8.


Gnaiger E (2021) Bioenergetic cluster analysis – mitochondrial respiratory control in human fibroblasts. MitoFit Preprints 2021.8. - »Bioblast link«


Labels: MiParea: Respiration, Instruments;methods 


Organism: Human  Tissue;cell: Fibroblast  Preparation: Intact cells 

Regulation: Coupling efficiency;uncoupling  Coupling state: LEAK, ROUTINE, ET  Pathway: ROX  HRR: Oxygraph-2k 

SUIT-003, MitoEAGLEPublication, MitoFit 2021 BCA