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V. Vittal, A. Rose, M.J. Kelley, T.S. Acott; Trabecular Meshwork Gene Expression in Response to Mechanical Stretch . Invest. Ophthalmol. Vis. Sci. 2003;44(13):3162.
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Purpose: Trabecular meshwork (TM) detects changes in intraocular pressure (IOP) as mechanical stretching. The TM maintains IOP homeostasis by adjusting the production of extracellular matrix (ECM) macromolecules such as matrix metalloproteinase-2 (MMP-2), membrane type matrix metalloproteinase-1 (MT1-MMP) and tissue inhibitor of metalloproteinase-2 (TIMP-2), which are involved in ECM remodeling. Studies were conducted to evaluate additional changes in TM gene expression in response to mechanical stretching. Methods: Porcine TM cells were subjected to mechanical stretch for 12, 24 and 48 hours and total RNA was extracted from the cells. Changes in gene expression were evaluated using microarrays containing 5700 or 8400 cDNAs. Some of the genes that showed significant changes in expression were analyzed by quantitative RT-PCR for verification. In addition, western blot analysis of TM cell extracts after 24, 48 and 72 hours of mechanical stretch was performed to measure the levels of protein expression of selected genes. Results: On the 5700 gene microarray at 12, 24 and 48 hrs respectively, 15, 37 and 10 genes were down-regulated and 37, 63 and 39 were up-regulated using statistical significance of P≤0.05 and greater than a 50% increase or decrease in expression with stretch. Dramatic increases in gene expression were observed for vimentin, various metallothioneins and metastasis associated protein-1 (MTA1). In addition, several genes associated with the cellular cytoskeleton and the ECM were up regulated. Genes associated with the cytoskeleton include actin, tubulin, vimentin, gelsolin, transgelin, CKAP4 and those involved with ECM include fibronectin, thrombospondin, fibromodulin, tenascin C, CD44, biglycan, cadherin and collagen types III, IX and XV. Moderate increases in expression were also seen in small leucine-rich proteoglycan genes such as lumican, mimecan and keratocan. Among all the genes that were up-regulated, further analysis of fibronectin, various metallothioneins, lumican, mimecan and keratocan was performed by quantitative RT-PCR and similar increases in gene expression were observed. Western blot analysis of fibronectin also showed corresponding increase in protein after mechanical stretch. Conclusions: TM cells, upon sensing mechanical stretch, increase the production of various ECM and cytoskeletal macromolecules. This is in agreement with our hypothesis that the changes in IOP cause mechanical stretching, which triggers the TM to turnover the ECM in order to maintain IOP homeostasis.
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