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Yanli Zhu, Lihong Zhang, Yo Sasaki, Jeffrey Milbrandt, Jeffrey M. Gidday; Protection Of Retinal Ganglion Cell Soma And Axons From Glaucomatous Injury In Mice By Cytoplasmic Overexpression Of Nmnat1. Invest. Ophthalmol. Vis. Sci. 2011;52(14):3082.
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© ARVO (1962-2015); The Authors (2016-present)
The NAD biosynthetic enzyme Nmnat is largely responsible for the profound slowing of peripheral nerve axonal degeneration in mice carrying a mutation for the nuclear protein Wlds (Araki T, Science 2004; MacDonald JM, Neuron 2006, Sasaki Y, J Neurosci 2006 and 2009). Moreover, cytoplasmic overexpression of Nmnat-1 affords robust axonal protection in a model of sciatic nerve transection (Sasaki Y, J Neurosci 2009). However, when glaucoma is induced in Wlds Tg rats (Beirowski B, Eur J Neurosci 2008) and mice (Wang AL, J Neurosci Res 2006), only retinal ganglion cell (RGC) axons are protected, not RGC soma. The present study assessed whether cytoplasmic Nmnat-1 overexpression affords protection of both RGC soma and axons against glaucomatous injury.
Experimental primary open-angle glaucoma was induced by episcleral vein ligation in littermate-matched adult male wild-type (WT; C57Bl/6) and cyt-Nmnat-1 transgenic (Tg) mice in which neurons overexpress Nmnat-1 in the cytoplasm; immunoblotting confirmed a robust expression of Nmnat-1 protein in both retina and optic nerve (ON) of these Tg mice. After three wks of sustained IOP elevation (by TonoLab tonometer, in mmHg, vs. baseline: WT=18.4±0.9 vs. 9.8±0.8; Tg=17.3±0.6 vs. 9.9±0.9), flat-mounted retinae from experimental eyes and fellow contralateral control eyes from WT and Tg groups were immunostained for NeuN to quantify across all quadrants surviving RGC soma (Buckingham BP 2008); in cross sections of the post-laminar ON, axon neurofilaments were immunostained with SMI-32 and astrocytes with GFAP. Images collected by confocal fluorescence microscopy, analyses by blinded observer, and statistics by nonparametric ANOVA.
In WT mice (n=6), 34% (p<0.05 vs. fellow eye) of RGC soma in the peripheral retina were lost, but in Tg mice (n=6), only a 2% soma loss occurred (p<0.05 vs. WT). In WT ON, reductions in both SMI32+ axon density and axonal fluorescence intensity were clearly evident, with a concomitant increase in GFAP staining in regions of axonal loss. In contrast, in Tg ON, SMI32+ axon integrity was well preserved, with only mild changes in GFAP expression.
Cytoplasmic overexpression of Nmnat-1 exerts a robust protective effect on both RGC soma and axons in experimental glaucoma, and underscores the need for further mechanistic investigations to determine if therapeutic approaches to stabilize this protein in RGC cytoplasm could be a viable approach for treating glaucoma.
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