The down-/up-regulation of mitochondrial proteins and their effects on neuronal viability following in vitro ischaemia-related insults

Jonathan Teoh

    Research output: ThesisDoctoral Thesis

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    Abstract

    [Truncated abstract] Therapeutic agents to directly modulate ischaemic neuronal death are presently unavailable. One approach to developing such therapeutics is based on the identification of proteins that are differently expressed as the result of 'preconditioning', a natural adaptive response utilised by neural cells to counter damaging insults. In this context, our laboratory has identified over 40 different proteins that are either up- or down-regulated in rat primary cortical neuronal cultures following preconditioning. The current study focused on characterising the roles played by seven mitochondria-related proteins in a number of in vitro neuronal injury models of ischaemic death, to determine if they exert neuroprotective or neurodamaging effects. The approach taken was to examine the effects of up- and down-regulating each protein in cortical neuronal cultures prior to their exposure to injury. These proteins included citrate synthase (CS), prohibitin (PHB), voltage-dependent anion channel 1 (VDAC1), heat shock protein 60 (HSP60), glucose-regulated protein 78 (GRP78), glucose-regulated protein 75 (GRP75) and nucleoside diphosphate kinase A (NDPKA). As a means of down-regulating proteins, the RNA interference [RNAi] technique was first established using cortical neuronal cultures. Validation of the RNAi technique was established using a green fluorescent protein (GFP) short interfering RNA (siRNA) construct to down-regulate adenoviral-mediated GFP expression. Subsequently, using the RNAi technique, it was demonstrated that the down-regulation of specific mitochondrial proteins did not result in any significant increases or decreases in neuronal survival following oxygen-glucose deprivation, hydrogen peroxide oxidative stress or L-glutamic acid excitotoxicity. Interestingly, down-regulation of the anti-apoptotic protein, Bcl-XL (B-cell lymphoma extra large) only exacerbated cell death in the hydrogen peroxide injury model...
    Original languageEnglish
    QualificationDoctor of Philosophy
    Publication statusUnpublished - 2013

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