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The effect of simvastatin treatment and cholesterol depletion on the redox status of cultured astrocytes

thesis
posted on 22.02.2017, 02:27 authored by Cane, Joannah
Cholesterol is an essential lipid molecule in many organisms. In addition to uptake from circulating lipoproteins, cells can synthesise cholesterol from acetyl-CoA via the mevalonate pathway. Unlike circulating cholesterol which may have endogenous or dietary origins, brain cholesterol is exclusively produced in situ, predominantly by glial cells. Ubiquinone or Coenzyme Q (CoQ) is also synthesised via the mevalonate pathway. Best known for its role as an electron-transferring agent in the mitochondrial electron transport chain (ETC), CoQ is also known to have anti-oxidative properties as a scavenger of radicals and enhancing other antioxidant defenses. Drugs called statins inhibit the mevalonate pathway and are used to treat elevated blood cholesterol. Primarily, statin treatment aims to reduce hepatic and blood cholesterol, however lipophilic statins are able to enter the brain and can disrupt brain-cholesterol homeostasis in addition to altering endogenous antioxidant capacity by decreasing CoQ levels. Further investigations into the conditions under which statins may be detrimental, and when they can be used advantageously are therefore needed. The initial aim of this thesis was to further explore the effect of statins on the brain; specifically how simvastatin treatment and cellular cholesterol levels interact with endogenous antioxidant systems in cultured astrocytes. These investigations extended into examination of the potential for interactions between cellular cholesterol and cellular iron uptake. Simvastatin treatment was not found to influence cellular defences against oxidative stress as predicted. However, simvastatin treatment resulted in a significant decline in total glutathione (GSH) levels. This result is the opposite of the effect on GSH that was expected and was not reflected in the measures of total antioxidant capacity. The simvastatin-induced decrease in GSH was not found to result in a decrease in cell viability when exposed to peroxides and also did not result in decreased specific detoxification rates of extracellular peroxides. It is suggested that the observed decrease does not represent a loss of cytosolic GSH but is likely to represent lost mitochondrial GSH which detoxifies free-radical leakage from the ETC. This result, combined with observed increases in cellular lactate dehydrogenase (LDH) suggests that statio treated astrocytes may be shifting to anaerobic metabolism, negating the requirement for mitochondrial GSH and allowing cellular resources to be redirected to replenishing essential cholesterol. Oxidative stress is tightly linked to the availability of redox-active trace metals, including iron. Astrocytes are pivotal in brain iron metabolism, capable of accumulating iron from transferrin (Tf) and non-transferrin bound iron (NTBI). Decreased cellular cholesterol may protect astrocytes from oxidative damage by decreasing intracellular iron, as uptake of iron from Tf is a cholesterol dependant process. As predicted, cholesterol depletion lead to decreased cellular accumulation of iron from Tf but not from NTBI. Contrary to predictions, simvastatin co-incubation with NBTI lead to a significant increase in the amount of iron accumulation in the cells, whereas the presence of the statio did not affect cellular iron uptake from Tf. This result suggests that beneficial effects of statio treatment resulting from a decrease in cellular iron uptake by the Tf-TfR pathway may be overridden by increased uptake of NTBI. In conclusion, this thesis provides valuable insights into the effect of simvastatin treatment on cellular redox status, including that statio treatment may ultimately lead to a metabolic shift to anaerobic energy production in astrocytes. In addition, cholesterol disruptions have varying effects on astrocytes' uptake of iron which may be influencing their susceptibility to oxidative damage.

History

Principal supervisor

Alfons Lawen

Year of Award

2015

Department, School or Centre

Biochemistry and Molecular Biology

Campus location

Australia

Course

Doctor of Philosophy

Degree Type

DOCTORATE

Faculty

Faculty of Medicine Nursing and Health Sciences