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J. A. Dyck, Y. Guo, T. Doser, W. O. Cepurna, E. C. Johnson, J. C. Morrison; Retinal Ganglion Cell Layer (RGC) Gene Expression Changes in a Rat Glaucoma Model Include the Dramatic Upregulation of Activating Transcription Factor 3 (ATF3). Invest. Ophthalmol. Vis. Sci. 2009;50(13):2753.
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Glaucoma is characterized by elevated intraocular pressure (IOP) exposure, optic nerve axon loss and RGC death, but relationships between these components remains unclear. Previously we used laser capture microdissection (LCM) and microarray analysis to characterize gene expression changes in the retinal ganglion cell layer (RGCL). Here we use quantitative real-time PCR (qPCR) to confirm changes for some the most significantly upregulated genes: ATF3, lipocalin 2 (Lcn2), heme oxygenase 1(Hmox1) and signal transducer and activator of transcription 3 (STAT3), and the downregulation of neurofilament L (Nefl), a RGC axonal protein. Additionally, because transcription factors (TF) are the most upregulated gene class, we determined the expression levels Jun and Junb, two TF not on the arrays.
Unilateral IOP elevation was produced in Brown Norway rats by episcleral vein injection of hypertonic saline. RGC layers were collected by LCM, RNA extracted, amplified and reverse transcribed for qPCR. RGC gene expression was compared between three retinal groups: controls (n=13), 20 early injury (<30% optic nerve axon loss) and 17 extensive injury (>30% axon loss) by ANOVA. Regression analysis was used to correlate expression levels with axon loss.
As shown (figure), expression changes discovered by array analysis (N=24) are confirmed by this larger qPCR analysis. In addition, significant positive linear correlations were found between optic nerve axon loss and ATF3 (r2=0.24), Lcn2 (r2=0.43), STAT3 (r2=0.21), Jun (r2=0.20), and Junb (r2=0.19), all p<0.01.
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