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S.A. Harvey, S.C. Anderson, N. SundarRaj; Microarray Analysis of the TGF-ß Induced Phenotypic Transition in Cultured Human Corneal Stromal Cells . Invest. Ophthalmol. Vis. Sci. 2003;44(13):834.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: TGF-ß is a major factor in corneal wound healing: blockade of TGF-ß inhibits corneal opacification, edema and angiogenesis. We characterized the effects of TGF-ß1 on gene expression in cultured human corneal stromal cells (HCSCs). Methods: HCSCs (derived from three different donor eyes) were treated for 48 hours with either FGF-1 and heparin to maintain a fibroblast phenotype, or with TGF-ß1 to convert to myofibroblast phenotype. Total RNA was extracted, processed and analyzed using either Affymetrix HU95Av2 or HU133A chips. Results: relative to FGF treated cells, over 220 unique gene transcripts increased in TGF-ß1 treated cells, while over 180 decreased. TGF-ß1 treatment is known to increase elaboration of extracellular matrix (ECM), cause cytoskeletal changes (including increased expression of α-smooth-muscle actin), elicit immunosuppression, and alter rates of growth and proliferation. In six pairwise comparisons, TGF-ß1 caused the following [mean fold] consistent changes: where previously reported in the literature these are in boldface. Collagen ECM: collagens Iα1  and Iα2 [3.4], IIIα1 [7.1], IVα1  and IVα2 , Vα1  and Vα2 [3.7]; also the collagen synthetic enzymes lysyl oxidase [5.2] and proline hydroxylase [3.8]. Non-collagen ECM: biglycan [9.1], lumican [12.1], versican [4.0], perlecan [2.9], leprecan [3.0] and thrombospondin [4.3]. Cytoskeleton: actin α2 [5.7], actinin α1 [3.0], filamin A [2.5], tropomyosin 1(α) [5.7], transgelin [5.5], calponin [2.2] and caldesmon [4.7]. Immunosuppression: decreased expression of IL-1ß [-32], IL-8 [-39], and the chemokines CXCL1 [-14], CXCL2 [-36], CXCL3 [-62] CXCL5 [-44] and CXCL6 [-9.6]; also downregulation of the synthetic pathway for PGE2 (PLA2 [-4.1] / COX2 [-4.6] / PGE synthase [-8.5]). Cell signaling: down-regulation of the TGF-ß signaling protein Smad3 [-7.0] and up-regulation of the signaling GTPase RhoB [2.8] both will act to decrease TGF-ß signaling. RhoE [-3.2] which acts to disperse actin stress fibers is down-regulated presumably to preserve the myofibroblast phenotype. Finally, the largest increase (mean, 122 fold) in all six comparisons was for an angiopoietin-like protein (CDT6) originally cloned from human cornea. CDT6 is potently profibrotic and anti-angiogenic. Conclusions: Comparative microarray analyses accurately reflect known TGF-ß-elicited phenotypic changes in HCSC. We have identified numerous additional changes which will enable us to more fully characterize this phenotypic transition, providing novel therapeutic opportunities in corneal wound healing.
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