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Manik Goel, Stacey Peek, Rachel Lamb, Susan M Culican; Phr1 disruption is neuroprotective for retinal ganglion cells in a mouse glaucoma model. Invest. Ophthalmol. Vis. Sci. 2014;55(13):2423. doi: https://doi.org/.
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Loss of the E3-ubiquitin ligase Phr1 has been demonstrated to increase retinal ganglion survival (RGC) survival following optic nerve crush injury. The purpose of our study was to determine whether Phr1 disruption is neuroprotective and preserves RGC survival in vivo in a mouse glaucoma model.
Induction of glaucoma: Elevation of intraocular pressure (IOP) was induced unilaterally by injection of polystyrene microbeads to the anterior chamber of the right eye of each animal in adult C57Bl/6 mice (control, n= 11) and Phr1 retinal knockout mice maintained in the C57Bl/6 background (Phr1-rko, n= 8). IOP measurement: IOP measurements were made using the hand held TonoLab tonometer on anesthetized mice. IOP was measured twice prior to bead injection, and at days 1 and 3, and weeks 1, 2, 3 and 4 after injection. RGC counts: 4 weeks after the injection, mice were sacrificed and retinas were dissected. Retinas were stained with anti-Brn3a antibody and the density of Brn3a+ RGCs was determined by counting the RGC number in 24 regions of the retina, 3 peripheral and 3 central in each of 4 quadrants and dividing by the total area measured (cells/mm2).
When compared to pre-injection IOP, the injected right eyes in both control and Phr1-rko showed a statistically significant elevation of IOP by 7 days post-injection. IOP increase was maximal at 2 weeks post-injection (control= 20.5 mm Hg, p<0.001; Phr1-rko=19.8 mm Hg, p=0.002) and remained elevated for 4 weeks after the injection. At 4 weeks after bead injection, the density of RGCs remaining in the injected eye was significantly decreased relative to the un-injected eye in control mice (2951±307 vs. 3483±294 cells/mm2, p=0.01) but not in the Phr1-rko mice (3203±174 vs. 3213±194 cells/mm2, p=0.74). Thus, although both strains of mice showed elevation of IOP in response to intracameral microbead injection, a loss of RGCs was observed from control eyes but Phr1-rkos were protected against IOP induced RGC loss and maintained significantly higher RGC counts as a percentage of the unmanipulated eye (85.4% vs 99.7%, p=0.009).
We have shown that RGCs deficient in the E3 ubiquitin ligase Phr1 have improved survival after IOP elevation than wild-type controls. Our results support further investigation of Phr1 as a potential clinically relevant therapeutic target for RGC neuroprotection in glaucoma and other optic neuropathies.
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