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J.B. Fileta, W. Huang, Y. Guo, C.L. Grosskreutz; Thy1 Downregulation in Retinal Ganglion Cells (RGC) in Experimental Glaucoma . Invest. Ophthalmol. Vis. Sci. 2006;47(13):1261.
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Thy1 is a surface glycoprotein uniquely expressed in RGC of the retina. This study sought to investigate protein and mRNA expression changes of Thy1 in a model of experimental glaucoma.
IOP elevation was induced in rats by hypertonic saline injections into episcleral veins using the Morrison model. IOP was measured every other day by Tonopen in conscious rats. Following 10 days of elevated IOP, retinal protein was collected or whole eyes processed for histological analysis. Protein and mRNA level changes of Thy1 in the retina were studied using Western blot analysis, immunohistochemistry (IHC), laser capture microdissection (LCM), and quantitative PCR analysis (qPCR). RGC loss was assessed using stereological counts of backlabeled RGC.
Following 10 days of elevated IOP (mean IOP = 42.21 ± 1.8 mmHg, control IOP = 18.16 ± 0.84 mmHg, n=6, p < 0.01) there was a significant change in Thy1 expression in retinas with experimental glaucoma. Elevated IOP resulted in a 34 ± 2.4 % (n=6, p < 0.01) loss of RGC and a qualitative decrease in Thy1 levels in RGC by IHC. There was a corresponding 60 ± 33 % (n=6, p < 0.01) decrease in Thy1 protein levels in whole retinal lysates. LCM was used to capture an equal number of RGC from control and glaucomatous eyes. qPCR analysis of mRNA from captured RGC revealed a 51 ± 24 % (n=6, p < 0.05) reduction of Thy1 message levels in RGC from glaucoma eyes when compared with an equal number of RGC from control eyes. While RGC cell number loss was substantial, both protein and mRNA level changes of Thy1 were affected to a much greater extent.
These results confirm earlier reports that Thy1 mRNA and protein levels are decreased in experimental glaucoma and are not a reflection of the absolute number of RGC present. We extend them to indicate that Thy1 message levels reflect a decrease in the amount of Thy1 mRNA per cell. Expression changes at the protein and message level far exceeded RGC loss. These results suggest that Thy1 serves as an early marker of RGC stress, but not a marker of RGC loss, in experimental glaucoma.
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