Unknown,Transcriptomics,Genomics,Proteomics

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Transcription profiling of human GFAP-negative lamina cribrosa cells TGF-beta regulated gene transcription and protein expression

ABSTRACT: Primary open angle glaucoma (POAG) is a progressive optic neuropathy, which is a major cause of worldwide visual impairment and blindness. Pathological hallmarks of the glaucomatous optic nerve head include retinal ganglion cell axon loss and extracellular matrix (ECM) remodeling of the lamina cribrosa layer. TGF-beta is an important pro-fibrotic modulator of ECM metabolism, whose levels are elevated in human POAG lamina cribrosa tissue compared with non-glaucomatous controls. We treated human lamina cribrosa (LC) cells with TGF-beta1 (10ng/ml) for 24 hours in order to examine differential gene expression patterns in repsonse to this cytokine. In particular we focused on ECM and fibrotic genes. We found that TGF-beta1 induces expression and release of ECM components in LC cells, which may be important in regulating matrix remodeling in the lamina cribrosa. In disease states such as POAG, the LC cell may represent an important pro-fibrotic cell type and an attractive target for novel therapeutic strategies.

ORGANISM(S): Homo sapiens

SUBMITTER: Ruaidhri Patrick Kirwan

PROVIDER: E-GEOD-2705 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Transforming growth factor-beta-regulated gene transcription and protein expression in human GFAP-negative lamina cribrosa cells.

Kirwan Ruaidhrí P RP Leonard Martin O MO Murphy Madeline M Clark Abbot F AF O'Brien Colm J CJ

Glia 20051201 4

Primary open-angle glaucoma (POAG) is a progressive optic neuropathy, which is a major cause of worldwide visual impairment and blindness. Pathological hallmarks of the glaucomatous optic nerve head (ONH) include retinal ganglion cell axon loss and extracellular matrix (ECM) remodeling of the lamina cribrosa layer. Transforming growth factor-beta (TGF-beta) is an important pro-fibrotic modulator of ECM metabolism, whose levels are elevated in human POAG lamina cribrosa tissue compared with non-g ...[more]

PMID: 16078232

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Project description:Purpose: Marked extracellular matrix (ECM) remodeling occurs in the human optic nerve head in primary open angle glaucoma (POAG). The glial fibrillary acid protein (GFAP) negative lamina cribrosa cell may play an important role in this remodeling process. The authors report the first study of global and ECM-focused gene transcription differentials between GFAP-negative negative lamina cribrosa (LC) cells from normal and POAG human donors. Methods: GFAP-negative LC cell lines were generated from the optic nerve tissue of three normal (n=3) and three POAG (n=3) human donors. Using Affymetrix U133A arrays the transcriptional profile between the normal and diseased groups were compared. Bioinformatic analysis was carried out using robust multichip average (RMA Express) and EASE/David. Real time TaqMan PCR and immunohistochemistry analyses were performed to validate the microarray data. Results: 285 genes were up regulated by greater than 1.5 fold and 413 were down regulated by greater than 1.5 fold in the POAG LC cells versus normal controls. Upregulated genes in POAG LC cells included, SPARC, periostin, thrombospondin, CRTL-1, CTGF and collagen types I, III, V and VIII. Downregulated ECM genes in POAG included MMP-1, fibulin, decorin and tenacsin XB. All TaqMan PCR validation assays were significant (*p<0.05) and consistent with the array data. Immunohistochemistry of one target (periostin) confirmed its differential expression at the protein level in POAG optic nerve head tissue compared with non-glaucomatous controls. Functional annotation and over-representation analysis identified ECM genes as a statistically over-represented class of genes in POAG LC cells compared with normal LC cells. Conclusions: This study reports for the first time that POAG LC cells in-vitro demonstrate up regulated ECM and pro-fibrotic gene expression compared with normal LC cells. This may be a pathological characteristic of this cell type in POAG in-vivo. We believe that the LC cell may be a pivotal regulator of optic nerve head ECM remodeling and an attractive target for future therapeutic strategies in POAG.

Project description:Despite their emerging relevance to fully understand disease pathogenesis, we have as yet a poor understanding as to how biomechanical signals are integrated with specific biochemical pathways to determine cell behaviour. Mesothelial-to-mesenchymal transition (MMT) markers colocalized with TGF-beta1-dependent signalling and yes-associated protein (YAP) activation across biopsies from different pathologies exhibiting peritoneal fibrosis, supporting mechanotransduction as a central driving component of these class of fibrotic lesions and its crosstalk with specific signaling pathways. Transcriptome and proteome profiling of the response of mesothelial cells (MCs) to linear cyclic stretch revealed molecular changes compatible with bona fide MMT, which (i) overlapped with established YAP target gene subsets, and (ii) were largely dependent on endogenous TGF-beta1 signaling. Importantly, TGF-beta1 blockade blunts the transcriptional upregulation of the se gene signatures, but not the mechanical activation and nuclear translocation of YAP per se. We studied the role therein of caveolin-1 (Cav1), a plasma membrane mechanotransducer. Exposure of Cav1-deficient MCs to cyclic stretch led to a robust upregulation of MMT-related gene programs, which was blunted upon TGF-beta1 inhibition. Conversely, Cav1 depletion enhanced both TGF-beta1 and TGFBRI expression. Cav1 genetic deficiency exacerbated MMT and PA fibrosis in an experimental model of peritoneal ischaemic buttons. Taken together, these results support that Cav1-YAP/TAZ fine-tune the fibrotic response through the modulation of MMT, onto which TGF-beta1-dependent signaling coordinately converges. Our findings reveal a cooperation between biomechanical and biochemical signals in the triggering of MMT, representing a novel potential opportunity to intervene mechanically-induced disorders coursing with peritoneal fibrosis, such as post-surgical adhesions. This SuperSeries is composed of the SubSeries listed below.

Dataset Information

Transcription profiling of human GFAP-negative lamina cribrosa cells TGF-beta regulated gene transcription and protein expression

Publications

Transforming growth factor-beta-regulated gene transcription and protein expression in human GFAP-negative lamina cribrosa cells.

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