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dc.contributor.advisorPearson, Andree
dc.contributor.advisorHampton, Mark
dc.contributor.authorBrinsden, Mark
dc.date.available2015-11-10T22:05:39Z
dc.date.copyright2015
dc.identifier.citationBrinsden, M. (2015). Measuring Mitochondrial Dysfunction in Humans (Thesis, Bachelor of Biomedical Sciences with Honours). University of Otago. Retrieved from http://hdl.handle.net/10523/6056en
dc.identifier.urihttp://hdl.handle.net/10523/6056
dc.description.abstractMitochondria are essential organelles found in almost every cell in the human body. They host a number of important metabolic pathways and carry out essential biological functions such as ATP synthesis and regulating cell death. There is a slow decline in mitochondrial function associated with ageing and mitochondrial dysfunction is proposed to act causally in a number of diseases. Previously it has been difficult to measure the health of human mitochondria as tests have required tissue from invasive muscle biopsies. The Seahorse XF Analyser is a recent technological advance that enables researchers to test mitochondrial function in small numbers of live cells. Recently, using the Seahorse analyser, peripheral blood cells such as platelets, monocytes and lymphocytes have been shown to display individually distinct bioenergetic profiles. During circulation, these cells are exposed to metabolic or environmental stressors throughout the body, potentially allowing them to act as biomarkers of bioenergetic health and ageing. Different cell preparations were trialled to purify and isolate platelets, monocytes and lymphocytes from freshly drawn whole blood. These protocols succeeded in preparing platelet and T-lymphocyte samples for XF analysis, however inconsistent results indicated that the protocols need further development. The Seahorse XF analyser was used to measure bioenergetic function in human platelets and T-lymphocytes from healthy donors ranging from 21 to 56 years of age. Each cell type required optimisation experiments to determine the optimal inhibitor and substrate concentrations to generate a meaningful bioenergetic profile. Similarly, seeding densities were determined to ensure oxygen consumption values that were suitable to the instruments sensitivity. These cell types have elastic metabolic phenotypes, and appeared sensitive to metabolic switching during early stages of the XF assay. Platelets were particularly difficult to work with because of their inclination to cease using oxidative phosphorylation and switch metabolism to using purely glycolytic pathways. Platelet susceptibility for metabolic switching is undetermined at this point. Lymphocyte optimization experiments also indicated possible premature activation during the assay. Further work is needed to fine tune this protocol to ensure consistent and uniform measurements before accurate BHI values are calculated for donors of different ages.
dc.format.mimetypeapplication/pdf
dc.language.isoen
dc.publisherUniversity of Otago
dc.rightsAll items in OUR Archive are provided for private study and research purposes and are protected by copyright with all rights reserved unless otherwise indicated.
dc.subjectSeahorse
dc.subjectMitochondrial
dc.subjectBHI
dc.subjectBioenergetic
dc.subjectPlatelets
dc.subjectPBMCs
dc.titleMeasuring Mitochondrial Dysfunction in Humans
dc.typeThesis
dc.date.updated2015-11-10T06:34:50Z
dc.language.rfc3066en
thesis.degree.disciplineCFFR - Centre for Free Radical Research
thesis.degree.nameBachelor of Biomedical Sciences with Honours
thesis.degree.grantorUniversity of Otago
thesis.degree.levelHonours
otago.openaccessOpen
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