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Title: Impaired mitochondrial anti-oxidant defence in amyotrophic lateral sclerosis
Author: Wood-Allum, Clare Alison
ISNI:       0000 0001 3571 7552
Awarding Body: Sheffield University
Current Institution: University of Sheffield
Date of Award: 2006
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In this study a candidate-generating approach was first employed, using proteomics to identify mitochondrial proteins whose expression changed in the presence of mutant SODI in a wellvalidated model of SOD I-related familial amyotrophic lateral sclerosis (ALS). Proteins whose expression changed in a mutant-specific fashion had functions potentially relevant to the pathogenesis of ALS including roles in apoptosis, protein processing and anti-oxidant defence. I then validated selected protein changes of interest by Western blotting in mitochondrial preparations of further NSC34 cells and G93A transgenic mouse spinal cord. Peroxiredoxin 3 (Prx 3), a thioredoxin-dependent hydroperoxidase, was down-regulated in NSC34 cells expressing two different species of mutant SOD I and in 90 d G93A transgenic mouse whole spinal cord. Immunostaining for Prx 3 and the known mitochondrial matrix protein HSP60 in NSC34 cells revealed co-localizing staining patterns, confirming the expected mitochondrial localization of Prx 3. Prx 3 immunohistochemistry confirmed its expression within the mitochondria of human and murine spinal motor neurons, establishing potential relevance to the human disease. Q-PCR was then used to show a down regulation of Prx 3 mRNA in spinal motor neurons from patients with both sporadic and SOD I-related FALS. The small drug molecule ebselen was identified as a peroxiredoxin mimic and was shown selectively to protect motor neuronal cells expressing two different species of mutant human SOD I against apoptosis induced by withdrawal of serum. Finally, having postulated that ebselen may act by up-regulating anti-oxidant protein transcription as well as acting as an anti-oxidant in its own right, ebselen was shown to induce transcription of the anti-oxidant response element (ARE), an oxidative stress-sensitive promoter sequence common to many anti-oxidant response genes, including members of the peroxiredoxin family and the proteins which maintain them in their active, reduced state. Examination of the sequence 5' to the Prx 3 gene revealed a putative ARE response element.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available