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Cheri Stowell, John Klimek, Larry David, George A. Cioffi, Claude F. Burgoyne, An Zhou; Quantitative Proteomic Analyses Of The Non-human Primate Optic Nerve Head (ONH) In Early Experimental Glaucoma (EEG) And Optic Nerve Transection (ONT). Invest. Ophthalmol. Vis. Sci. 2011;52(14):4601.
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To characterize changes in protein expression within the NHP ONH at the onset of confocal scanning laser tomographic ONH surface change following the onset of laser-induced, chronic IOP-elevation (EEG) or 3 weeks after optic nerve transection (ONT).
Three weeks after unilateral surgical ONT (n=3 animals), or at the onset of reproducible ONH surface change in ipsilateral eyes exposed to mild IOP (MIOP EEG - peak IOP 40 mmHg, n=2 animals), both eyes of each NHP were enucleated. For each eye, the optic nerve head (ONH) was excised from the globe with the assistance of a 6-mm trephine and homogenized. Proteins were extracted from the homogenates and digested with trypsin. Tryptic digests of each eye were then analyzed with an LTQ ion-trap mass spectrometry (MS) system (Thermo Finnegan) with 3 technical replicates of each sample. All identified proteins were quantified using a MS spectral counting approach, and ratios of spectral counts for each protein in the ONT or EEG eye of each animal compared to its contralateral normal eye were determined. The proteins that were statistically significantly changed by GEE analysis (p<0.01) were used for further bioinformatic analyses with the assistance of the MetaCore program (GeneGo Inc).
To date, a total of 290 ONH proteins were identified and quantified in the 7 NHP subjects, among which 60 proteins were detected in all 7 pairs of eyes and did not show a change. For other proteins, differential and condition-specific changes were observed in each of the ONT, MIOP and HIOP ONHs. MetaCore gene ontology analyses of regulated proteins revealed an association of up-regulated proteins with wound healing processes in the HIOP ONH; many of these proteins were down regulated in the MIOP ONH. These proteins include neuregulin and complement component 3 protein.
ONT, MIOP and HIOP EEG eyes demonstrate ONH proteomic changes that are distinct from one another. These proteomic alterations are also different from the retinal tissues of the same eyes (Stowell et al, in revision). The observed differences may be due to differences in the mechanism and site of injury. On-going work focuses on the validation of proteomic findings and identification of cell types that account for the wound healing response in the ONH.
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