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H. Miao, P. Du, S. Lin, A. A. Wise, M. Hernandez; Gene Expression Profile of Optic Nerve Head (ONH) Astrocytes From Normal and Glaucomatous Caucasian (CA) and African American (AA) Donors. Invest. Ophthalmol. Vis. Sci. 2007;48(13):3659.
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© ARVO (1962-2015); The Authors (2016-present)
To discover susceptibility genes to human glaucoma by comparing gene expression profiles in ONH astrocytes from normal and glaucomatous AA and CA donor eyes.
We used Hu133A & Hu133Av2 Affymetrix chips to compare gene expression levels in primary cultures of ONH astrocytes from 12 CA (age 60±10) and 12 AA (age 58±12) normal eyes; 7 CA (age 73±10 ) and 6 AA (age 59±6) glaucomatous eyes. Eighty chips were normalized with RMA in R using Bioconductor. Significant differential gene expression levels were detected using mixed effects modeling and Significance Analysis of Microarray (SAM). Clustering and pathway analysis was applied to selected genes to discover co-regulated gene sets in both ethnic groups and in disease. Differential expression was confirmed by quantitative RT-PCR, western blot and ELISA. Cell adhesion and migration assays tested physiological responses. Glutathione (GSH) assay detected levels of intracellular GSH.
Multiple analyses selected 105 genes differentially expressed between normal AA and CA (p< 0.01). Up-regulated genes were involved in signaling (8 genes), response to stress (7 genes) and transcription regulation (7genes). Down-regulated genes in AA were signaling (4), CNS related genes (6), ECM genes (5) and cell adhesion (5). A total of 25 genes were differentially expressed in AA and CA glaucomatous astrocytes including genes involved in signaling, cell shape, adhesion and motility. Enigma, a PDZ and LIM domain containing protein that regulates actin organization (p< 0.00005) and Plectin 1 (PLEC1) an intermediate filament involved in cytoskeleton-membrane attachment (p< 7.18E-09) were over-expressed in glaucomatous astrocytes. Unique genes that were differentially expressed in AA glaucomatous astrocytes included: RAB5B, a Ras GTP binding protein involved in vesicular trafficking (p< 8.31E-07); TFDP1, a transducer of TGF-ß signaling; CEP350, a PPAR -interacting protein (p< 2.36E-06) and HABP4, a novel intracellular hyaluronan binding protein (p< 0.0003). GSH levels in normal AA were lower compared to normal CA astrocytes and similar to glaucomatous astrocytes consistent with the increased levels of GSH metabolizing enzymes in normal AA and in glaucomatous astrocytes.
Our data show that astrocytes from AA and CA normal and glaucomatous donors display distinct expression profiles suggesting subtle changes in gene expression in ONH astrocytes unique to the development and/or progression of POAG in AA.
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