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Jorgelina M. Calandria, Nicolas G. Bazan; Ataxin-1 Poly-Q-induced Proteotoxic Stress And Apoptosis Are Attenuated By Docosahexaenoic Acid-derived Neuroprotectin D1 In Human Retinal Pigment Epithelial Cells. Invest. Ophthalmol. Vis. Sci. 2012;53(14):1137.
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Poly-glutamine-dependent misfolding induces toxicity and consequently contributes to degeneration in the central nervous system. In Spinocerebellar ataxia 1 (SCA1), Ataxin-1 mutation causes the accumulation and partial loss of function of the protein leading to cytotoxicity. Ataxin-1 interacts with transcription factors and spliceosome components. In addition to an involvement of the cerebellum in SCA1, the retina also undergoes degeneration (J. Neurol. Neurosurg. Psychiatry 77:1293, 2006), thus providing the basis for applying this model to retinal studies. We hypothesize that Neuroprotectin D1 (NPD1), a lipid mediator derived from DHA, promotes the survival of the RPE cells undergoing proteotoxic stress by increasing the turn-over of the poly-Q proteins.
Ataxin-1 containing 82 glutamines-tract was expressed in ARPE-19 cells. Apoptosis was assessed using activated caspase-3 immunostaining, TUNEL and Hoechst assay. Akt phosphorylation status on NPD1 treated cells was determined by western blot. Localization and accumulation of Ataxin-1 was determined by immunocytochemistry.
Ataxin-1 82Q expression in RPE cells induced apoptosis in a time-dependent manner. This was evidenced by the increase in the percentage of TUNEL and Hoechst positive cells when compared with the controls expressing wild-type protein.Caspase-3 also showed the same pattern of activation. NPD1 (100 nM) decreased apoptosis induced by Ataxin-1 82Q. The expression of the mutant Ataxin-1 also induced accumulation of the Serine-776 phosphorylated form in the nucleus. ARPE-19 cells expressing Ataxin-1 82Q in the presence of NPD1 presented lower levels of phosphorylated Ataxin-1 signal in the nucleus. The translocation into the nucleus and phosphorylation of Serine-473 of Akt, the main kinase that phosphorylates Ataxin-1, was increased differentially when the 82Q form was expressed even in the presence of NPD1, suggesting that NPD1 does not prevent the phosphorylation of Ataxin-1 and may be acting on its dephosphorylation instead.
These results demonstrate that: 1) the RPE cellular model depicts similar features to the changes triggered by the mutation in Purkinje cells, laying the groundwork for unveiling new pathways and therapeutic approaches in other misfolded protein-induced pathologies in the retina and 2) NPD1 rescues the deleterious effects of proteotoxic stress caused by extended polyQ tract Ataxin-1 expression in RPE cells due to the increase in turnover. These findings may lead to new therapeutic applications for DHA and NPD1 for preventing and attenuating the initiation of retinal degenerations.
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