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K. Rodgers, D. Wang, Y. Ben, J. Qu, C. L. Grosskreutz; Autophagy in Experimental Glaucoma. Invest. Ophthalmol. Vis. Sci. 2010;51(13):2117.
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It has been observed that autophagy may be involved in several chronic neurodegenerative diseases including Alzheimer’s, Huntington’s and Parkinson’s disease. Whether autophagy is neuroprotective or detrimental in neuronal diseases is the subject of significant debate. The role of autophagy in glaucoma has yet to be investigated, and is the subject of this study. LC3-II is present on activated autophagosomes, and is thus widely used as a marker of autophagy. In this study, we investigated LC3-II expression in experimental glaucoma.
Brown Norway Rats were injected unilaterally (OS) with saline in the episcleral vein to induce glaucoma. Intraocular pressure (IOP) was measured three times per week. The area under the pressure-time curve (experimental-control), or AUC, was used to assess the IOP exposure. After sustained IOP elevation, retinal protein was extracted for immunoblot analysis or eyes were fixed and sectioned for immunohistochemistry. Western blots were run with a 1:1000 concentration of LC3-II antibody (n=7). A two-tailed student t-test was used for western blot statistics. LC3-II immunohistochemistry was performed using a 1:100 antibody concentration (n=3). Densitometry was performed to quantitate protein expression and alpha tubulin was used as a loading control. Optic nerve damage was assessed using a stereologically informed grading scheme (1=no damage, 5=severe damage).
Following the induction of experimental glaucoma, the peak IOP was 35.66 ± 5.12 mmHg with an average AUC of 211.67± 82.29. Western blot analysis of LC3-II shows a 0.69 ± 0.35 (mean ± S.D.) ratio of glaucomatous to control eye (n=7, p= 0.039) with an average optic nerve grade of 3.2 ± 0.9. Immunohistochemistry demonstrated decreased LC3-II immunoreactivity in the glaucomatous retinal ganglion cell layer in two of three animals investigated.
Based on these studies, we find that the autophagy marker LC3-II is decreased in experimental glaucoma. These findings are somewhat surprising as autophagy has been reported to be activated following crush injury to the optic nerve. Since autophagy can remove toxic protein buildup in tissues, and is thought to confer protection to cells under stress conditions, we speculate that failure of this system may render retinal ganglion cells more susceptible to death under conditions of elevated IOP.
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