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Alterations in gene expression induced by cyclic mechanical stress in trabecular meshwork cells.

ABSTRACT: To investigate the changes in gene expression induced by cyclic mechanical stress (CMS) in trabecular meshwork (TM) cells.Human TM cultures from three donors were plated on type I collagen-coated flexible silicone bottom plates and subjected to 15% stretching, one cycle per second for 6 h. Non-stressed parallel control cultures were incubated under the same conditions in the absence of CMS. Total RNA from each culture was amplified (1 round of amplification) and hybridized to Operon Human Oligo Arrays version 3.0 (35 K probes). Differences in gene expression induced by CMS were analyzed using Genespring 7.2. quantitative polymerase chain reaction (Q-PCR) was used to confirm changes in the expressions of 12 selected genes. The effects of chemical inhibitors for p38, ERK (extracellular signal-regulated kinase), JNK (Jun N-terminal kinase), PKA (protein kinase A), PI3K (phosphoinositide 3-kinase), and P2 (purinergic 2) receptors on the induction of MMP3 (matrix metalloproteinase 3), HSP70 (heat shock protein 70), ECSM1 (endothelial cell specific molecule 1), BMP2 (bone morphogenetic protein 2), VEGFC (vascular endothelial growth factor C), and IL-8 (interleukin 8) were evaluated in porcine TM cells subjected to the same regime of CMS as that used in human cells.CMS induced extensive gene expression changes (664 genes, p < or = 0.05) twofold or higher in cultured TM cells. Many of these changes were related to extracellular matrix (ECM) synthesis and remodeling including the upregulation of two metalloproteinases (MMP3 and MMP10). Cytoskeleton and cell adhesion genes were also affected by CMS as well as genes known to be involved in cellular protection against stress including several members of the HSP70 family. Inhibition of PI3K/AKT and P2 receptors pathways significantly reduced the induction of MMP3 and IL-8 whereas the inhibition of the PKA/cAMP pathway decreased ECSM1 and BMP2.CMS activated many genes that could influence the aqueous humor outflow facility, specifically genes involved in ECM synthesis and remodeling (e.g. MMPs), cytoskeletal organization, and cell adhesion. Induction of MMP3 has the potential to increase the aqueous humor outflow facility and could be part of a homeostatic mechanism involved in the maintenance of normal intraocular pressure (IOP) levels. Other observed changes are more likely to be related to general cellular responses to stress (e.g., HSP70, ECSM1, and BMP2). Although these latter changes may initially help to repair mechanical damage, some of them such as the induction of BMP2 could eventually increase tissue rigidity and compromise the ability of the TM to maintain normal levels of outflow resistance.

SUBMITTER: Luna C

PROVIDER: S-EPMC2654047 | biostudies-literature | 2009

REPOSITORIES: biostudies-literature

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Alterations in gene expression induced by cyclic mechanical stress in trabecular meshwork cells.

Luna Coralia C Li Guorong G Liton Paloma B PB Epstein David L DL Gonzalez Pedro P

Molecular vision 20090311

<h4>Purpose</h4>To investigate the changes in gene expression induced by cyclic mechanical stress (CMS) in trabecular meshwork (TM) cells.<h4>Methods</h4>Human TM cultures from three donors were plated on type I collagen-coated flexible silicone bottom plates and subjected to 15% stretching, one cycle per second for 6 h. Non-stressed parallel control cultures were incubated under the same conditions in the absence of CMS. Total RNA from each culture was amplified (1 round of amplification) and h ...[more]

PMID: 19279691

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Project description:BackgroundTrabecular meshwork and Schlemm's canal are the tissues appointed to modulate the aqueous humour outflow from the anterior chamber. The impairment of their functions drives to an intraocular pressure increase. The selective laser trabeculoplasty is a laser therapy of the trabecular meshwork able to decrease intraocular pressure. The exact response mechanism to this treatment has not been clearly delineated yet. The herein presented study is aimed at studying the gene expression changes induced in trabecular meshwork cells by selective laser trabeculoplasty (SLT) in order to better understand the mechanisms subtending its efficacy.Methodology/principal findingsPrimary human trabecular meshwork cells cultured in fibroblast medium underwent selective laser trabeculoplasty treatment. RNA was extracted from a pool of cells 30 minutes after treatment while the remaining cells were further cultured and RNA was extracted respectively 2 and 6 hours after treatment. Control cells stored in incubator in absence of SLT treatment were used as reference samples. Gene expression was evaluated by hybridization on miRNA-microarray and laser scanner analysis. Scanning electron microscopic examination was performed on 2 Trabecular meshwork samples after SLT at 4(th) and 6(th) hour from treatment. On the whole, selective laser trabeculoplasty modulates in trabecular meshwork the expression of genes involved in cell motility, intercellular connections, extracellular matrix production, protein repair, DNA repair, membrane repair, reactive oxygen species production, glutamate toxicity, antioxidant activities, and inflammation.Conclusions/significanceSLT did not induce any phenotypic alteration in TM samples. TM is a complex tissue possessing a great variety of function pivotal for the active regulation of aqueous humour outflow from the anterior chamber. SLT is able to modulate these functions at the postgenomic molecular level without inducing damage either at molecular or phenotypic levels.

Dataset Information

Alterations in gene expression induced by cyclic mechanical stress in trabecular meshwork cells.

Publications

Alterations in gene expression induced by cyclic mechanical stress in trabecular meshwork cells.

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