The main aim of this thesis was to examine the hypothesis that the A/J strain variant of H55N substitution affects citrate synthase (CS) enzyme activity and metabolic health in mice fed a high fat diet (HFD). C57BL/6J (B6) mice and congenic B6.A-(rs3676616-D10Utsw1)/KjnB6 (B6.A) mice, a strain which carries the A/J allele of Cs on the B6 strain background, were fed a HFD (45% kcal from fat) for 12 weeks. CS activity, but not that of ß-hydroxyacyl-coenzyme dehydrogenase was lower in the gastrocnemius muscle of B6.A mice compared to B6 mice (P< 0.001). During HFD feeding the glucose tolerance of mice decreased progressively and to a greater extent in B6.A females compared to B6 females, with males showing a similar trend. Interestingly, after 12 weeks of HFD feeding only B6.A males showed increases (P< 0.05) in their resting metabolic rate; moreover; core body temperature were also increased (P< 0.05) for congenic B6.A of both sexes by the end of the study. However, body weight and fat gain did not differ between B6.A and B6 mice. The second aim of the thesis was to test the hypothesis that low CS activity promotes palmitate-induced lipotoxicity in muscle cells. After 18 hours of incubation in 0.8 mM palmitate, C2C12 muscle cells with a ~50% reduction in CS activity showed low (P< 0.001) viability, increased (P< 0.001) levels of cleaved Caspase-3, high levels of AMP-activated protein kinase and acetyl-CoA carboxylase phosphorylation (P< 0.05), low levels of protein kinase B phosphorylation, high mitogen-activated protein kinases activation (P< 0.001) compared to the control shRNA cells. This was coupled with higher levels of mitochondrial proteins (P< 0.05), which are involved in oxidative phosphorylation. C2C12 cells with reduced CS activity also showed high reactive oxygen species production (P< 0.05), low intracellular ATP levels (P< 0.05), and lower basal mitochondrial respiration (P< 0.001). In summary, the A/J strain variant of H55N is associated with low CS enzyme activity and impaired metabolic health when fed HFD. Palmitate has a lipotoxic effect on Cs shRNA transfected cells and can lead to cell death.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:704533 |
| Date | January 2016 |
| Creators | Alhindi, Yosra |
| Publisher | University of Aberdeen |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://digitool.abdn.ac.uk:80/webclient/DeliveryManager?pid=231391 |
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