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Nitasha R Phatak, Dorota L Stankowska, Raghu R Krishnamoorthy; Overexpression of the POU Domain Transcription Factor Brn-3b leads to Upregulation of the Pro-survival Factor Bcl-2 in Rat Retinal Ganglion Cells. Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):4410.
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Brn-3b a POU domain transcription factor plays a key role in regulating retinal ganglion cell axon outgrowth during development. The anti-apoptotic factor Bcl-2 has been shown to be responsible for the intrinsic regenerative capacity of axons in neurons of the central nervous system. The purpose of this study was to determine if overexpression of Brn-3b could lead to changes in Bcl-2 expression in rat retinal ganglion cells.
Adeno-associated viral vectors (serotype 2) encoding either Brn-3b (rAAV-syn-Brn3b) or the control vector (rAAV-syn-GFP) (4x109 viral particles) were injected intravitreally in one eye of Brown Norway rats. Following two weeks (to allow for robust expression from the AAV vectors), rats were maintained for an additional week and humanely sacrificed. Rat eyes were enucleated fixed with 4% paraformaldehyde (PFA) and embedded in OCT. Frozen sections of retinas were obtained and subjected to immunostaining for Brn-3b and Bcl-2 proteins. The sections were co-stained with an antibody to beta-III tubulin, which served as a marker of retinal ganglion cells. Changes in the expression of Brn-3b and Bcl-2 were studied by using confocal microscopy.
A marked increase in immunostaining for Brn-3b was found in retinas of rats injected with rAAV-syn-Brn3b, compared to those injected with rAAV-syn-GFP. An increased immunostaining for Bcl-2 was observed in retinal ganglion cells in rats intravitreally injected with rAAV-syn-Brn3b, compared to those injected with the rAAV-syn-GFP virus.
Administration of rAAV-syn-Brn3b in rat eyes could upregulate Brn3b expression in retinal ganglion cells. Transcription factor Brn-3b could elevate the expression of Bcl-2 suggesting its potential for neuroprotection in rat retinal ganglion cells.
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