Degree Name

Master of Science (MS)

Semester of Degree Completion


Thesis Director

Britto P. Nathan

Thesis Committee Member

Michael A. Menze


Mitochondrial dysfunction, such as reductions in ATP production via oxidative phosphorylation, is a recognized pathological symptom of neurodegenerative diseases including Alzheimer's disease (AD). One of the known risk factors for the development of AD is the inheritance of a specific genotype of a lipid transporting protein known as apolipoprotein E (apoE). How apoE contributes to the development and progression of AD is poorly understood. The purpose of this study was to determine if a relationship exists between mitochondrial dysfunction and genetic predisposition to AD via apoE. Mitochondrial bioenergetics and enzymatic activity were investigated in an AD mouse model (apoE KO) and in wild-type (WT) mice. Since AD develops with age, mitochondrial function was studied in both young (4-6 months old) and old (12-22 months old) mice. Additionally, estrogen therapy was utilized as a possible therapeutic to prevent mitochondrial dysfunction.

We isolated mitochondria located in nerve cell terminals (synaptosomes) using Percoll gradient centrifugation from WT (C57BL/6J) and apoE KO (Apoetn1Unc) mice. Mitochondria were isolated from the following three treatment groups: mice possessing ovaries injected with vehicle control solution (Sham), ovariectomized mice with vehicle control injection (OVX), and ovariectomized mice with 50 ng/g injection of 17β-estradiol (OVX + E2). Mitochondrial function was evaluated based on oxygen consumption of permeabilized and non-permeabilized synaptosomes and was measured using the OROBOROS Oxygraph-2k at 37°C. Succinate dehydrogenase (SDH) and citrate synthase (CS) activity was also assayed to help elucidate the bioenergetic profile of the isolated synaptosomes.

Our results revealed no significant differences in oxygen consumption from intact and permeabilized synaptosomes between Sham and OVX young WT and apoE KO mice. Additionally, no significant differences were detected in SDH and CS activity between young WT and apoE KO mice. However, estrogen treatment dramatically increased oxygen consumption and enzymatic activity in apoE KO for all respiration parameters, whereas, a minimal effect was observed in WT mice. No significant differences in oxygen consumption from permeabilized and intact synaptosomes were detected between young and old OVX and OVX + E2 treated mice, but old Sham mice had significantly higher oxygen consumption rates compared to young Sham mice for most all permeabilized parameters in both WT and apoE KO mice. Estrogen treatment had no effect on mitochondrial respiration in old mice, but estrogen treated old apoE KO mice had significantly higher SDH activity compared to its Sham.

Increases in respiration and enzymatic activity prompted investigation of mitochondrial quantity via Western blot. We used the voltage dependent-anion-channel (VDAC) as a mitochondrial quantity marker. The results revealed that estrogen treatment increased mitochondrial quantity in estrogen treated young apoE KO and old Sham mice. We speculate that increases in mitochondrial quantity in young apoE KO mice and old Sham mice were due to estrogen and reactive oxygen species (ROS) induced mitochondrial biogenesis, respectively. Our data suggests that predisposition of AD via apoE inheritance is not an underlying source of mitochondrial abnormalities; however, the effectiveness of estrogen as a neuro-therapeutic may be dependent upon an individual's apoE genotype, as well as, their age.

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