INT281878
From wiki-pain
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Sentences Mentioned In
Key: | Protein | Mutation | Event | Anatomy | Negation | Speculation | Pain term | Disease term |
Transduction with AdAcon increases aconitase expression and activity in primary midbrain cultures | |||||||||||||||
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In summary, since catalase can only attenuate cell death when m-aconitase is overexpressed, this suggests 1) that the excess H2O2 from m-aconitase inactivation is being released from astrocytes and injuring neighboring neurons and 2) that removal of astrocyte-derived extracellular H2O2 can prevent death. | |||||||||||||||
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The exacerbation of Fe2+ and H2O2 production in m-aconitase overexpressing cells suggests its role as their source. | |||||||||||||||
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Overexpression of m-aconitase was achieved by transducing cells with an adenoviral construct expressing m-aconitase cDNA and a GFP reporter (AdAcon) or with an adenoviral construct only expressing GFP (AdGFP) to control for viral-mediated effects. | |||||||||||||||
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More importantly, neurons from AdAcon transduced cultures revealed greater damage than AdGFP transduced cells at 500 µM PQ2+ supporting that m-aconitase overexpression exacerbates neuronal death upon oxidative inactivation. | |||||||||||||||
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M-aconitase was overexpressed in mature primary midbrain cultures using an adenoviral vector. | |||||||||||||||
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Overexpression of m-aconitase resulted in a concentration-dependent decrease of cell viability compared to GFP-only expressing cells (Fig. 5). | |||||||||||||||
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Although astrocytes were the predominant cell type to overexpress m-aconitase, the neuronal population was more susceptible to death compared to astrocytes. | |||||||||||||||
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To confirm the role of m-aconitase versus other cellular proteins in the production of Fe2+ and H2O2, we specifically overexpressed m-aconitase. | |||||||||||||||
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We hypothesized that increasing the levels of m-aconitase would increase the amount of target enzyme available for oxidative inactivation leading to exacerbation of H2O2 production, Fe2+ formation and cell death. | |||||||||||||||
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Astrocytic overexpression of m-aconitase results in neuronal death | |||||||||||||||
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Fenton reaction) in cells expressing basal levels of m-aconitase, extracellular H2O2 was also mediating cell death in m-aconitase overexpressing cultures. | |||||||||||||||
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However, in m-aconitase overexpressing cells, either catalase or HBED was sufficient to significantly inhibit cell death (Fig. 7). | |||||||||||||||
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To confirm the role of m-aconitase as a source of Fenton reagents and death, we overexpressed m-aconitase using an adenoviral construct thereby increasing the target available for inactivation by ROS. | |||||||||||||||
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To confirm the mechanism of H2O2 and Fe2+ in the death of cells overexpressing m-aconitase, we asked whether cell death could be prevented by their pharmacological removal using catalase and HBED respectively. | |||||||||||||||
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Co-localization of GFP fluorescence with MAP2 and GFAP performed 24 hrs after transduction with AdAcon, revealed the expression of m-aconitase largely in GFAP positive cells, suggesting that m-aconitase was predominately overexpressed in astrocytes (Fig. 8a,b). | |||||||||||||||
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generating compound PQ2+ on aconitase activity, H2O2 production, mitochondrial Fe2+ accumulation and cell death in primary midbrain cultures. | |||||||||||||||
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generating compound PQ2+ on aconitase activity, H2O2 production, mitochondrial Fe2+ accumulation and cell death in primary midbrain cultures. | |||||||||||||||
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Oxidative inactivation of m-aconitase overexpressing cultures resulted in exacerbation of H2O2 production, Fe2+ accumulation and increased neuronal death. | |||||||||||||||
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We proceeded to test whether increasing m-aconitase expression would exacerbate the release of Fenton reactants following oxidative inactivation. | |||||||||||||||
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General Comments
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