Multi-parametric metabolic assessment of cells under stress conditions

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dc.check.embargoformatE-thesis on CORA onlyen
dc.check.opt-outNot applicableen
dc.check.reasonThis thesis is due for publication or the author is actively seeking to publish this materialen
dc.contributor.advisorPapkovsky, Dmitri B.en
dc.contributor.authorWaters, Alicia H. C.
dc.contributor.funderEuropean Commissionen
dc.date.accessioned2015-11-19T10:13:12Z
dc.date.issued2015
dc.date.submitted2015
dc.description.abstractGlycolysis, glutaminolysis, the Krebs cycle and oxidative phosphorylation are the main metabolic pathways. Exposing cells to key metabolic substrates (glucose, glutamine and pyruvate); investigation of the contribution of substrates in stress conditions such as uncoupling and hypoxia was conducted. Glycolysis, O2 consumption, O2 and ATP levels and hypoxia inducible factor (HIF) signalling in PC12 cells were investigated. Upon uncoupling with FCCP mitochondria were depolarised similarly in all cases, but a strong increase in respiration was only seen in the cells fed on glutamine with either glucose or pyruvate. Inhibition of glutaminolysis reversed the glutamine dependant effect. Differential regulation of the respiratory response to FCCP by metabolic environment suggests mitochondrial uncoupling has a potential for substrate-specific inhibition of cell function. At reduced O2 availability (4 % and 0 % O2), cell bioenergetics and local oxygenation varied depending on the substrate composition. Results indicate that both supply and utilisation of key metabolic substrates can affect the pattern of HIF-1/2α accumulation by differentially regulating iO2¬, ATP levels and Akt/Erk/AMPK pathways. Inhibition of key metabolic pathways can modulate HIF regulatory pathways, metabolic responses and survival of cancer cells in hypoxia. Hypoxia leads to transcriptional activation, by HIF, of pyruvate dehydrogenase (PDH) kinase which phosphorylates and inhibits PDH, a mitochondrial enzyme that converts pyruvate into acetyl-CoA. The levels of PDH (total and phosphorylated), PDH kinase and HIF-1α were analysed in HCT116 and HCT116 SCO2-/- (deficient in complex IV of the respiratory chain) grown under 20.9 % and 3 % O2. Data indicate that regulation of PDH can occur in a manner independent of the HIF-1/PDH kinase 1 axis, mitochondrial respiration and the demand for acetyl-CoA. Collectively these results can be applied to many diseases; reduced nutrient supply and O2 during ischemia/stroke, hypoglycaemia in diabetes mellitus and cancer associated changes in uncoupling protein expression levels.en
dc.description.sponsorshipEuropean Commission (EU FP7 Marie Curie Initial Training Network CHEBANA Program 264772)en
dc.description.statusNot peer revieweden
dc.description.versionAccepted Version
dc.format.mimetypeapplication/pdfen
dc.identifier.citationWaters, A. H. C. 2015. Multi-parametric metabolic assessment of cells under stress conditions. PhD Thesis, University College Cork.en
dc.identifier.endpage183
dc.identifier.urihttps://hdl.handle.net/10468/2075
dc.language.isoenen
dc.publisherUniversity College Corken
dc.rights© 2015, Alicia H. C. Waters.en
dc.rights.urihttp://creativecommons.org/licenses/by-nc-nd/3.0/en
dc.subjectBioenergticsen
dc.subjectMetabolismen
dc.subjectUncouplingen
dc.subjectMitochondriaen
dc.subjectHypoxia inducible factoren
dc.subjectHypoxiaen
dc.thesis.opt-outfalse
dc.titleMulti-parametric metabolic assessment of cells under stress conditionsen
dc.typeDoctoral thesisen
dc.type.qualificationlevelDoctoralen
dc.type.qualificationnamePhD (Science)en
ucc.workflow.supervisord.papkovsky@ucc.ie
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