Hong, CT; (2014) Neuroprotection in Parkinson's Disease: Controlling Mitochondria-Dependent Apoptosis. Doctoral thesis , UCL (University College London).

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Abstract

Objective: To investigate the neuroprotective approach of controlling mitochondria-dependent apoptosis in Parkinson’s disease cellular models. Background: Parkinson’s disease is a neurodegenerative disease and there is evidence indicating that mitochondria-dependent apoptosis is related to dopaminergic neuron loss. pUL37x1, an immediate early protein expressed during cytomegalovirus infection, modulates mitochondria-dependent apoptosis by inactivating Bcl-2-associated X protein (Bax), and meclizine is believed to increase glycolysis and hyperpolarize mitochondria, which inhibits apoptosis. Methods: The neuroprotective and anti-apoptotic effects of pUL37x1 over-expression and meclizine were investigated in SH-SY5Y, a neuroblastoma cell line and primary rat cortical culture cells. Cell death was induced by either staurosporine or 6-hydroxydopamine and measured by lactate dehydrogenase release and propidium iodide binding assay. Apoptotic markers were measured by the release of cytochrome c and the activation of caspase-3. Mitochondrial membrane potential was measured by Tetramethylrhodamine, methyl ester fluorescence obtained by confocal microscope. Extracellular acidification rate (ECAR), a glycolytic activity parameter and oxygen consumption rate (OCR) were measured by XF analyser. Statistics was performed by either ANOVA with Dunnett’s post-hoc analysis or two-tailed Student’s t-test. Results: Both pUL37x1 over-expression and meclizine significantly protected against toxin-induced cell death in SH-SY5Y and rat primary cortical culture cells. Both approaches also down-regulated apoptosis. In terms of meclizine, the protection resulted from glycolysis-related mitochondrial hyperpolarization. Hyperpolarization and protection would decline following glycolytic inhibition. pUL37x1 over-expression prevented apoptotic cell death by two means: Bax-dependent and glycolysis-dependent mechanisms. First, pUL37x1 over-expression led to Bax mitochondria-translocation, and in contrast to control, Bax silencing did not provide more protection on pUL37x1 over-expressing cells. Second, pUL37x1 over-expression increased cellular glycolysis and hyperpolarized mitochondria, and glycolytic inhibitors attenuated the protection, indicating a glycolysis-dependent protective mechanism. Conclusions: The success of neuroprotection by pUL37x1 and meclizine in Parkinson’s disease cellular models not only confirms the significance of controlling mitochondria-dependent apoptosis, but also indicates two novel approaches to neuroprotection.

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