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M. M. Patel, X. Ding, D. Shen, A. A. Herzlich, M. Annamalai, J. Tuo, C.-C. Chan, S. Kaushal; Celastrol Decreases Endoplasmic Reticulum (ER) Stress and Clinical Progression in the Cx3cr1-/-/Ccl2-/- Murine Model of Age-Related Macular Degeneration. Invest. Ophthalmol. Vis. Sci. 2008;49(13):3767.
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Glycoprotein accumulation in age-related macular degeneration (AMD)-associated drusen deposits, alterations of chaperone proteins in proteomic analyses in human AMD and in the Cx3cr1-/-/Ccl2-/- double knockout (DKO) murine model of AMD, as well as protein misfolding in AMD-related inherited maculopathies suggest a role for endoplasmic reticulum (ER) stress in AMD pathogenesis. This study sought to determine the role of ER stress in the DKO model of AMD and to assess the ability of celastrol, a plant-derived triterpine that upregulates heat shock proteins (hsp), to induce a cytoprotective effect on the retina and RPE of DKO mice.
DKO and age-matched wild-type (WT) mice were treated three times weekly by intraperitoneal injection for four weeks with celastrol or placebo (3:1 DMSO:PBS). Fundoscopy was conducted prior to and after treatment. Eyes from celastrol or placebo-treated mice, and from additional DKO and age-matched WT mice were harvested for frozen section, protein extraction, or mRNA isolation. Analyses of protein expression of hsp70, ER stress markers p-PERK, p-eIF2α, and p-IRE1α, and ER stress-induced apoptosis marker CHOP were performed by immunohistochemistry and western blotting. RT-PCR for CHOP was also performed on eyes from DKO and WT mice. In addition, in vitro studies were performed to assess the effect of celastrol treatment on hsp40, hsp70, and hsp90 protein expression.
Both WT and DKO mice express p-PERK, p-eIF2α, p-IRE1α, and CHOP in all layers of the retina. Compared to WT, DKO mice exhibit diffusely increased p-PERK, p-eIF2α, p-IRE1α, and CHOP protein expression in the retina. No significant difference in CHOP mRNA levels was detected. In vitro studies demonstrated increased hsp70 and hsp90 protein expression after administration of celastrol. Treatment of DKO mice with celastrol, as compared to treatment with placebo, reduced protein expression of ER stress markers p-PERK and p-IRE1α and decreased clinical progression of AMD lesions, as determined by number and size of subretinal and retinal lesions.
These findings demonstrate that the Cx3cr1-/-/Ccl2-/- murine model of AMD exhibits increased ER stress and responds to cytoprotective strategies such as celastrol, implicating a role for cellular stress reduction in the pathogenesis and treatment of AMD.
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