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M Salvador-Silva, P Yang, S Aoi, MR Hernandez; Oligonucleotide Microarray Analysis of Gene Expression in Human Optic Nerve Head Astrocytes Exposed to Hydrostatic Pressure . Invest. Ophthalmol. Vis. Sci. 2002;43(13):2887.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: Microarray analysis was performed to identify genes involved in signal transduction and/or cytoskeletal interactions in human optic nerve head (ONH) astrocytes exposed to elevated hydrostatic pressure (HP). Methods: Cultured ONH astrocytes were grown under to 60 mm Hg HP for 6, 24 or 48 h. Control cultures were subjected to atmospheric pressure. Total RNA was extracted. Labeled, purified and fragmented cRNA was hybridized to human Genome U95Av2 chip (∼12,000 genes) using GeneChip Instrument System (Affymetrix). Data were analyzed by GeneSpring Analysis Software. Results: Our results identified differential expression of genes involved in cell adhesion and astrocyte migration. Seven genes involved in signal transduction and cytoskeleton organization showed significant upregulation after exposure to HP compared to controls. Upregulation of potassium-chloride co-transporter 1 (KCC1), ephrin B receptor (Eph-B) and actin binding LIM protein (AbLIM) was observed after 24 h HP. Expression of activating transcription factor (ATF) showed a linear increase throughout 48 h HP. Upregulation of G-protein-coupled receptors (GPCR-45 and peptide YY) and p-21 activated kinase-3 (PAK-3) was observed after 48 h HP. Specific gene products were confirmed by Western blot analysis on cell lysates. Conclusion: Upregulation of genes involved in signal transduction and cytoskeletal organization in human ONH astrocytes are consistent with changes in cell shape, cell adhesion and migration observed in human ONH astrocytes exposed to HP. Increased expression of KCC1, an ion co-transporter, and/or Eph-B, a receptor tyrosine kinase, which are located in the cell membrane, may occur in response to HP-related stress. Increased expression of G-protein coupled receptors and PAK-3, a stress activated kinase, may modulate cell adhesion and migration through interactions with the actin cytoskeleton. Increased expression of ATF, a nuclear transcription factor, suggests a role in transcriptional regulation of gene expression during exposure to HP.
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