All ETDs from UAB

Advisory Committee Chair

Jianhua Zhang

Advisory Committee Members

Shannon Bailey

Steven Carroll

Helen Kim

Aimee Landar

Andy West

Document Type

Dissertation

Date of Award

2012

Degree Name by School

Doctor of Philosophy (PhD) Heersink School of Medicine

Abstract

Neurodegenerative disease encompasses a wide range of conditions and pathologies that can manifest at any age depending on the etiology. A major factor in both early onset and age-related neurodegeneration is mitochondrial dysfunction. To investigate how mitochondrial bioenergetics is affected by cellular stress, we used an in vitro culture system to examine mitochondrial function in response to oxidative stress. We also studied an in vivo model of neuronal ceroid lipofuscinosis to determine the impact of deficient autophagy-lysosomal activity on mitochondrial morphology, composition and function. In vitro we found that retinoic acid-induced differentiation of dopaminergic neuroblastoma SH-SY5Y cells exhibited increased mitochondrial membrane potential, and no change in mitochondrial number. The differentiated cells also exhibited greater stimulation of mitochondrial respiration with uncoupling and an increased bioenergetic reserve capacity, which was suppressed by the inhibitor of glycolysis, 2-deoxy-D-glucose (2-DG). Differentiated cells were substantially more resistant to cytotoxicity and mitochondrial dysfunction induced by the reactive lipid species 4-hydroxynonenal (HNE) or the reactive oxygen species generator 2,3-dimethoxy-1,4-naphthoquinone (DMNQ). In vivo, deficiency in lysosomal protease Cathepsin D (CD) resulted in increased autophagosomes, lysosomal storage of autofluorescent lipofuscin pigment, decreased turnover of lysosomal substrates and neurodegeneration. Our prior studies demonstrated that CD knockout (CD-/-) mouse at postnatal day 25 (P25) accumulated endogenous α- synuclein and ubiquitinated protein, and had increased parkin mRNA, in the brain. Because autophagy and Parkin are important for mitochondrial quality control, we determined whether brains of CD-/- mice exhibited changes in mitochondrial composition, mitophagy signaling, mitochondrial morphology or function. We found a significant decrease in fission protein Drp1 in both P1 and P21 CD-/- brains and an increase in cytochrome oxidase subunit IV in P21 brain. However total mtDNA levels and average mitochondrial areas were similar between CD wild-type (CD+/+) and CD-/- cortex. Interestingly, PGC1-α mRNA was decreased and complex IV activity was decreased in CD-/- cortex, suggesting that in response to decreased autophagic flux downstream of CD deficiency, mitochondrial homeostasis is altered. Importantly the above changes occur before signs of neurodegeneration or accumulation of α- synuclein. Together, this thesis provided new information regarding factors important for mitochondrial quality control.

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