Differential global and extra-cellular matrix focused gene expression patterns between normal and glaucomatous human lamina cribrosa cells.
OBJECTIVE
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. We report the first study of global and ECM-focused gene transcription differentials between GFAP-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 four normal (n=4) and four POAG (n=4) human donors. Using Affymetrix U133A arrays the transcriptional profile between the normal and diseased groups were compared. Bioinformatic analysis was performed using robust multichip average (RMA Express) and EASE/David. Real time TaqMan PCR and immunohistochemistry analyses were performed to validate the microarray data.


RESULTS
183 genes were upregulated by greater than 1.5 fold and 220 were down regulated by greater than 1.5 fold in the POAG LC cells versus normal controls. Upregulated genes in POAG LC cells included, transforming growth factor beta 1 (TGFbeta1), secreted acid protein cysteine rich (SPARC), periostin (POSTN), thrombospondin-1 (THBS1), cartilage linking protein-1 (CRTL-1), and collagen type I (COL1A1), collagen type V (COL5A1), and collagen type XI (COL11A1). Downregulated ECM genes in POAG included fibulin 1 (FBLN1), decorin (DCN), and collagen type XVIII (COL18A1). 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 upregulated 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 in POAG and an attractive target for molecular therapeutic strategies in the future.
Kirwan
Ruaidhrí P
RP
Conway Institute of Biomolecular and Biomedical Research, University College Dublin, Ireland. ruaidhri.kirwan@ucd.ie
Wordinger
Robert J
RJ
Clark
Abbot F
AF
O'Brien
Colm J
CJ
D002460:Cell Line; D015536:Down-Regulation; D016326:Extracellular Matrix Proteins / Q000235:genetics* / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation; D005902:Glaucoma, Open-Angle / Q000235:genetics* / Q000473:pathology*; D006801:Humans; D020411:Oligonucleotide Array Sequence Analysis; D009898:Optic Disk / Q000378:metabolism* / Q000473:pathology*; D015203:Reproducibility of Results; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014158:Transcription, Genetic; D015854:Up-Regulation
D016326:Extracellular Matrix Proteins