Abstract
Abstract: :
Purpose: Proteolytic activation and inhibition of matrix metalloproteinases (MMPs) is important in the maintenance of the trabecular meshwork (TM) extracellular matrix (ECM). We determined the presence or absence of transcripts for ECM molecules, MMPs, tissue inhibitors of metalloproteinases (TIMPs), and the uPA/plasmin system in normal TM (NTM) and glaucomatous TM (GTM) cells and tissues. Methods:Total RNA was isolated from pooled NTM or GTM cell lines or tissue and used for Affymetrix Human Genome U133 GeneChip® analysis. Data were collected using Affymetrix MicroArray Suite® software and analyzed using GeneSpring® (Silicon Genetics) software. Data were normalized per chip and per gene and subsequently filtered with expression level cutoffs and 2–fold changes in gene expression. Sensitivity of detection with Affymetrix GeneChips is on the order of 1 transcript in 100,000. Results:Of the twenty–three MMP’s profiled on the U133A GeneChip, only transcripts for MMP–2, –9, and –14 were detected in both normal and glaucomatous TM cells and tissue with MMP–2 being 2–fold higher in GTM than NTM cells. Other MMPs such as MMP–1 was present in NTM cells but absent from GTM cells, MMP–3 was present in both NTM & GTM tissue, and MMP–12 was 3.5–fold higher in GTM than NTM cells. Of the four known TIMP’s (1–4), TIMP–1, –2, and –3 were present in both normal and glaucomatous TM cells and tissue and TIMP–4 was absent. TIMPs 2 and 3 were nearly 2–fold higher in GTM than NTM cells. Plasminogen activators tPA (Tissue Plasminogen Activator) and uPA (Urokinase Plasminogen Activator) were present in normal and glaucomatous TM cells and tissue whereas PAI–1 (Plasminogen Activator Inhibitor) was detectable only in TM cells. ECM components such as Collagen (COL10A1, COL15A1, & COL18A1), Decorin (variants A, C, & D), Versican, and Fibulin (FBLN–1 & –5) were higher in GTM than NTM cells. Conclusions:We detail the selective mRNA expression of ECM molecules, MMPs and potential activators and inhibitors of MMPs in the normal balance and glaucomatous imbalance of the TM.
Keywords: trabecular meshwork • extracellular matrix • gene microarray