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Raghu R Krishnamoorthy, Dorota L Stankowska; Brn3b Mediated Axonal Regeneration of the Optic Nerve in a Rodent Model of Glaucoma. Invest. Ophthalmol. Vis. Sci. 2014;55(13):2183.
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The purpose of this study was to determine if adeno-associated virus-2 (AAV-2) mediated overexpression of transcription factor Brn3b in retinal ganglion cells could promote axonal regeneration and improvement in visual acuity after IOP-mediated optic nerve injury in glaucomatous rats.
The Morrison’s rat model of glaucoma was used to elevate IOP in one eye of Brown Norway rats, while the corresponding contralateral eye served as control. One week after IOP elevation, rats were injected in the IOP-elevated eye with an adeno-associated viral vector encoding either transcription factor, Brn3b (AAV-Brn3b) or the control vector (AAV-Empty) and maintained for an additional 3 weeks of IOP elevation. Retina sections were subjected to immunohistochemical analysis of neuroregenerative markers including GAP-43, ab-LIM and acetylated tubulin. Proteins extracts were isolated from optic nerves and immunoblot analyses of GAP-43, neurofilament-M and abLIM were carried out. In a separate set of experiments, an axonal tracer (CT-B) was intravitreally injected 48 h prior to sacrifice, to assess functionality of regenerated axons due to Brn3b overexpression following IOP elevation in rats. Another batch of rats administered with AAV-Brn3b following IOP elevation was tested for visual acuity using the visual optomotor response test.
An increased immunostaining of neuroregenerative markers, GAP-43, abLIM and ac-Tuba was observed in AAV-Brn3b transduced, IOP-elevated rat eye retinas and ONHs posterior to the site of axonal injury in comparison with IOP elevated, AAV-Empty virus injected rats. Immunoblot analysis of rat optic nerves revealed a prominent increase in pro-regenerative proteins: GAP-43, abLIM and neurofilament-M in IOP elevated AAV-Brn3b injected rats, compared to empty vector injected rats. An appreciable increase in CT-B tracer along the axonal tracts, posterior to the ONH was detected in IOP elevated AAV-Brn3b injected rats indicative of an increased regenerative response and improved axonal function, mediated by Brn3b overexpression. Visual acuity of IOP elevated rats improved after intravitreal administration of AAV-Brn3b.
The data indicates that administration of AAV-Brn3b virus has the potential to reverse optic nerve damage, possibly through regeneration of the damaged axons of the optic nerve.
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