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Chendong Pan, Lihua Guo, Evan Berger, Anna Demetriades; Neurotrophin-4 Promotes Retinal Ganglion Cell Survival Following Neuronal Injury In Thy1-YFP Mice. Invest. Ophthalmol. Vis. Sci. 2013;54(15):779.
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Neurotrophin-4 is a neurotrophic factor that regulates the survival, differentiation and mature function of neurons. This study aims to determine if an AAV vector expressing NT4 prevents retinal ganglion cell loss following optic nerve crush injury in Thy1-yellow fluorescent protein (Thy1-YFP) mice.
Thy1-YFP mice, which express yellow fluorescent protein as a marker of viable retinal neurons, were imaged before and after optic nerve crush injury. Two weeks prior to optic nerve crush injury, mice were treated with a 1μl intravitreal injection containing 1x10e9 vector genomes of either AAV.NT4 or AAV.Null in one eye with no treatment in the contralateral eye. Mice were sacrificed at two weeks following crush injury and retinal flatmounts prepared. Fluorescent spots were counted manually in the same retinal area of each animal. ELISA was performed to quantify NT4 levels in the retina and optic nerve of injected eyes.
ELISA confirmed that NT4 expression was significantly increased in the retina and optic nerve of eyes injected with AAV.NT4 compared to control eyes (n=5). In vivo imaging demonstrated less fluorescent retinal neuron loss in AAV.NT4 treated eyes compared to both AAV.Null treated eyes and uninjected control eyes. At two weeks, the number of fluorescent cells (mean±SEM) in untreated eyes without optic nerve crush injury was 3213±95 cells/mm2 (n=8). Eyes that received intravitreal AAV.NT4 (1598±45 cells/mm2 )(n=10) showed more remaining fluorescent cells than eyes that received intravitreal AAV.Null (1167±124 cells/mm2 )(n=6) or intravitreal PBS (1112±106 cells/mm2)(n=9) or no injection (1017±82 cells/mm2)(n=5). Thus, the number of of fluorescent cells remaining in optic nerve crushed eyes that received AAV.NT4 was over 150% greater than optic nerve crushed eyes that received no injection (p<0.01).
Our results indicate that increased NT4 expression in the retina and optic nerve following intravitreal injection of AAV.NT4 results in protection of retinal neurons in Thy1-YFP mice following optic nerve crush injury. In Thy1-YFP mice, fluorescent cells predominantly represent retinal ganglion cells; therefore, these findings suggest that intravitreal AAV.NT4 provides a neuroprotective effect on retinal ganglion cells after optic nerve injury.
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