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P. E. Fort, M. K. Losiewicz, T. W. Gardner; Alpha-Crystallin Neuroprotective Function in the Retina and its Regulation During Diabetes. Invest. Ophthalmol. Vis. Sci. 2010;51(13):3729.
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To investigate the role and regulation of the crystallin proteins in the retinal cells in relation with stress such as diabetic retinopathy. We previously showed that several proteins of the crystallin protein superfamily are expressed in the retina and over-expressed during diabetes. We also demonstrated that their regulation is sensitive to insulin but their basal function as well as the function of their upregulation during diabetes remained unclear.
iTRAQ method was used to characterize the proteome modifications in the retinas of diabetic mice and rats compared to their age-matched controls. Changes were confimed using western-blot and immunohistochemistry analysis, respectively, to determine levels of protein expression and cellular localization. Protein interactions were assessed using co-immunoprecipitation methods followed by western-blots analysis. Cell death was assessed using a Cell Death DNA fragmentation Elisa assay or a caspases-3/7 activity assay.
We demonstrated that the diabetes-induced crystallin upregulation was among the largest and the most conserved changes observed during diabetes since it is conserved across different models, strains and species. We showed that the interaction between alpha-crystallins and pro-apoptotic proteins observed in retinal tissue from non-diabetic animals was destabilized by diabetes and correlates with increased cell death. We also demonstrated that retinal neuronal cells over-expressing alpha-crystallin proteins were protected from diabetes related stress induced cell death. We showed that this was related to decreased caspases-dependant apoptosis.
This study shows for the first time that crystallin upregulation is one of the major and most conserved changes observed during diabetes. It also demonstrated that the normal function of alpha-crystallin in retinal cells is to prevent cell-death through interaction and inhibition of pro-apoptotic proteins. This study suggest that control of the function of alpha-crystallin could be a new therapeutic alternative for the prevention of retinal cell death during diabetes and potentially any retinal cell degeneration related disease.
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