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D.R. Hyde, S. Kassen, V. Ramanan, Y. Luo, S. Springer, T.S. Vihtelic; Changes in Gene Expression During Regeneration of the Light–Damaged Zebrafish Retina . Invest. Ophthalmol. Vis. Sci. 2005;46(13):3084.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: We wish to elucidate the molecular mechanisms that underlie the regeneration of rod and cone photoreceptors in the light–damaged zebrafish retina. Methods: Zebrafish were dark–treated for 7 days and exposed to constant intense light (30,000 lux). At various times during the light treatment (0, 16, 31, 51, 68 and 96 hrs) the retinas were analyzed by histology and immunohistochemistry to assess the level of cell damage, cell death, and the regeneration response, which is characterized by nuclear layer cell proliferation. Gene expression changes during the light treatment were identified by hybridizing the Compugen 16K Zebrafish oligo microarray with cDNA synthesized from total retinal RNA isolated at the various time points. Real–time PCR was used to confirm the changes in gene expression identified by the microarray experiments. Results: Histology and immunohistochemistry revealed signs of retinal degeneration by 16 hrs of light treatment. Pyknotic nuclei in the outer nuclear layer were observed at 31 hrs and a dramatic reduction in rhodopsin immunolabeling was observed at 68 hrs after initiating the light treatment. High levels of inner and outer nuclear layer proliferating cells were observed at 96 hrs of light treatment. Microarray analysis revealed several clusters of genes that showed a variety of expression patterns, including ones that gradually increase or decrease, plateau at intermediate time points or initially decrease and then increase in expression compared to the control levels. Real–time PCR confirmed these expression profiles for many of the genes. We identified a subset of genes with dynamic expression patterns that may be involved in stem cell self–renewal and neuronal cell differentiation. We are currently performing in situ hybridization to examine where these genes are expressed in the regenerating retina. Conclusions: We identified a number of genes that change their retinal expression profile in response to light–induced photoreceptor degeneration and during the subsequent regeneration of both rods and cones. We correlated the gene expression changes with the cellular changes in the retina. Several of these genes may be involved in pathways that are required for stem cell renewal or neuronal differentiation from these retinal stem cells. Thus, they represent good candidates for regulating the regeneration response of the light–damaged retina.
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