Purchase this article with an account.
R.R. Mohan, S.E. Wilson; Microarray Analysis of PDGF–Regulated Gene Expression in Human Corneal Fibroblast Cells . Invest. Ophthalmol. Vis. Sci. 2004;45(13):3776.
Download citation file:
© ARVO (1962-2015); The Authors (2016-present)
Purpose: We showed previously that platelet–derived growth factor (PDGF) regulates proliferation and chemotaxis in human corneal fibroblast (HSF) cells. Thus we sought to determine a time–dependent PDGF–regulated gene expression profile in HSF cells using gene array technology. Methods:Affymetrix HG–U133 GeneChip system was used. First passage HSF cultures were exposed to either vehicle or PDGF (20ng/ml of each PDGF isoform) for 1 hour or 8 hours. Total RNA was converted into ds–cDNA followed by the biotin–labeled cRNA using the ENZO bioarray kit. Biotin–labeled cRNA was fragmented and hybridized with U133A and U133B DNA chips. The changes in the gene expression were analyzed using MAS5.0 software after normalization of experimental data. Each assay was performed twice. Results:A quantitative analysis of 39,000 genes/EST’s showed that PDGF altered expression of 761 genes at 1 hr and 3639 genes at 8 hrs in the HSF cells. At 1 hr 589 genes showed statistically significant increase and 172 genes showed statistically significant decrease in expression. Whereas at 8 hrs 1601 genes showed statistically significant increase and 2038 genes showed statistically significant decrease in expression. Genes with a statistically significant, but marginal increase or decrease were excluded from the analysis. At 1 hr suppressor of cytokine signaling 3, RAS, dual specificity phosphatase 6, guanylate binding protein 1 interferon inducible, activating transcription factor 3 were among the top twenty genes that showed ≥2.5 fold statistically significant increase whereas cyclin–dependent kinase inhibitor 1B (p27, Kip1), calreticulin, collagen type 1 alpha 2, thrombospondin 1, profiling 2 were among the top twenty genes that showed ≤1.0 fold statistically significant decrease in expression in response to PDGF over control. However, at 8 hrs interleukin 8, dual specificity phosphatase 6, regulator of G–protein signaling 4, fibroblast growth factor 5, hepatocyte growth factor were among the top twenty genes that showed ≥3.0 fold statistically significant increase whereas cyclin G2, collagen type IV alpha 2, cartilage linking protein 1, integrin beta 3 binding protein (beta3–endonexin) genes were among the top twenty genes that showed ≥2.0 fold statistically significant decrease in expression in response to PDGF over control. Conclusions:PDGF regulates expression of multiple genes in HSF cells that have important roles in modulating the cell cycle, signal transduction, proliferation, apoptosis, adhesion and immune response. PDGF released by corneal epithelium is likely an important regulator of the stromal wound healing response. EY10056
This PDF is available to Subscribers Only