Browsing School, Graduate by Subject "idebenone"
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Effects of Idebenone on the Mitochondrial Respiration of Neurons, Astrocytes, and MicrogliaNeuroinflammation and mitochondrial bioenergetic dysfunction are present in most neurodegenerative diseases. Microglia, the resident immune cells of the brain, release cytotoxic factors such as nitric oxide (NO), superoxide, and cytokines following proinflammatory activation. NO mediates mitochondrial respiratory inhibition in microglia and neighboring cells by competing with oxygen at complex IV of the electron transport chain. Additional modes of inhibition by NO include post-translational modifications to upstream complexes I and II. Idebenone is a clinically safe prodrug that, in its reduced form idebenol, can act in place of ubiquinol and donate electrons directly to complex III. This dissertation tested the overarching hypothesis that idebenone can support mitochondrial respiration in cells with sufficient quinone-reduction capacity to convert idebenone to idebenol, despite complex I and II impairment by NO. In astrocytes but not neurons, idebenone and two related quinones could rescue maximal oxygen consumption rate (OCR) when a complex I inhibitor was present. This difference between astrocytes and neurons was due to a disparity in cellular quinone-reduction capacity mediated by the expression of NADPH:quinone oxidoreductase 1 (NQO1). Astrocytes were sensitive to respiratory impairment by an NO donor or co-cultured proinflammatory microglia at a physiologically-relevant oxygen level and idebenone was able to partially reverse this impairment. Interestingly, microglia upregulated their quinone-reduction capacity following proinflammatory stimulation and idebenone was also able to partially reverse respiratory impairment in microglia following activation. Surprisingly, in contrast to astrocytes, NQO1 was not responsible for idebenone reduction in activated microglia. Biochemical isolation of the responsible enzyme identified inducible nitric oxide synthase (iNOS) among the few candidates common to three distinct fractionation approaches. Assays performed with recombinant iNOS revealed a novel idebenone reduction activity with exciting implications for future studies. This dissertation’s findings suggest that insufficient quinone-reduction capacity in diseased target cells may be a mechanistic reason for the failure of idebenone in clinical trials. These results support new strategic approaches for the use of idebenone and similar drugs to overcome mitochondrial bioenergetic dysfunction.
Mitochondrial Functional Changes Contribute to Proinflammatory Microglial ActivationMicroglia are the innate immune cells of the central nervous system. They have many important physiological functions, from surveying the surrounding environment to synaptic pruning. However, following brain injury or during neurodegenerative diseases, microglia become 'activated,' migrate towards the site of injury, and produce inflammatory factors. Under neuropathological conditions, microglia may become excessively activated, a condition characterized by prolonged release of damaging proinflammatory factors. Mitochondrial function and dysfunction are increasingly being implicated in the process of microglial activation. This dissertation tested the overarching hypothesis that mitochondrial dysfunction stimulates proinflammatory microglial activation. Specifically, we found that in response to lipopolysaccharide (LPS) and interferon-γ (IFN-γ) in vitro, microglial mitochondria undergo structural remodeling and become significantly shortened. We utilized the putative mitochondrial fragmentation inhibitor mdivi-1 which successfully attenuated production of proinflammatory factors in vitro and in vivo. Surprisingly, mdivi-1 did not impair inflammation through modulation of mitochondrial structure, but instead impaired acetylation of the proinflammatory transcription factor NF-κB. We then set out to determine the importance of mitochondrial bioenergetic changes in microglial activation. We found that the combination of LPS/IFN-γ induced a significant impairment in mitochondrial oxygen consumption. This impairment was mediated by selective degradation of mitochondrial Complex I subunits at atmospheric 21% O2 and by nitric oxide at 3% O2, an oxygen tension thought to be more physiologically relevant to that present in the brain. Finally, we utilized the exogenous electron donor idebenone in an attempt to bypass this respiratory impairment. Indeed, we found that idebenone acutely reversed respiratory inhibition, and that this respiratory bypass was able to attenuate microglial production of proinflammatory factors. Together, the findings in this dissertation link mitochondrial structural and bioenergetic dysfunction to proinflammatory microglial activation, and move the field towards successfully finding a target for reversing damaging neuroinflammation following brain injury or in neurodegenerative disease.