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William J. Curry, Sorcha Finnegan, Paul Hocking, Manir Ali, Chris Inglehearn, Alan Stitt, John McGarvey, Rene Beattie; Raman Spectroscopic Analysis Of The Wild Type And Retinal Dysplasia And Degeneration (rdd) Chick Retina. Invest. Ophthalmol. Vis. Sci. 2011;52(14):1827.
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This study used Raman spectroscopy to review the biochemistry of retinogenesis (neurogenesis and synaptogenesis) in the wild type (wt) chick retina with the onset and progression of degeneration at equivalent ages in the retinal dysplasia and degeneration (rdd) chick retina.
Rdd and wt chick embryonic (E12, 13, 17, 19) and post hatch (P1) eyes (n=6) were enucleated, immersion fixed in 4%paraformaldehyde), cryoprotected in 5% and 30% (w/v) sucrose/PBS/sodium azide (4°C). 10 µm central retinal cross-sections were analysed using a Jobin-Yvon confocal Raman microscope (633 nm) and spectra mapped (x3, 1 µm interval) across the retinal layers prior to histological staining. Background subtraction was performed in Matlab and principal component analysis (PCA) was performed in Unscrambler.
PCA identified 30 biochemical constituents in the chick retina. A fatty-acid lipid was observed in the developing inner plexiform layer (IPL) of both wt and rdd embryonic retina but was more intense in the outer nuclear layer (ONL) of the wt retina and less intense in ONL of the rdd retina. A more saturated fatty acid was observed in the OPL, INL and proximal IPL of the embryonic wt chick but was absent in the analogous rdd retina. At P1 this fatty acid was diminished and more diffuse in both the rdd and wt retina. Cytochrome was observed in the GCL and NFL of the embryonic rdd retina and several other protein signals were expressed in the ONL, IPL and INL with strong differences observed between different stages of development and between the wt and rdd chicks.
This study is the first to report Raman microscopic mapping of developing and degenerating chicken retina. Raman microscopy provided a holistic overview of distribution of dominant biochemical molecules in the wt and rdd chick retina, including different protein populations including cytochrome, lipids, and DNA. It has promise as a significant tool for understanding the onset and progression of retinal degeneration.
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