Phenotypic and Functional Alterations in Tunneling Nanotubes Formed by Glaucomatous Trabecular Meshwork Cells.

Purpose: Trabecular meshwork (TM) cells detect and coordinate responses to intraocular pressure (IOP) in the eye. TM cells become dysfunctional in glaucoma where IOP is often elevated. Recently, we showed that normal TM (NTM) cells communicate by forming tubular connections called tunneling nanotubes (TNTs).

Tunneling Nanotubes are Novel Cellular Structures That Communicate Signals Between Trabecular Meshwork Cells.

Purpose: The actin cytoskeleton of trabecular meshwork (TM) cells plays a role in regulating aqueous humor outflow. Many studies have investigated stress fibers, but F-actin also assembles into other supramolecular structures including filopodia. Recently, specialized filopodia called tunneling nanotubes (TNTs) have been described, which communicate molecular signals and organelles directly between cells.

Mapping molecular differences and extracellular matrix gene expression in segmental outflow pathways of the human ocular trabecular meshwork.

Elevated intraocular pressure (IOP) is the primary risk factor for glaucoma, and lowering IOP remains the only effective treatment for glaucoma. The trabecular meshwork (TM) in the anterior chamber of the eye regulates IOP by generating resistance to aqueous humor outflow. Aqueous humor outflow is segmental, but molecular differences between high and low outflow regions of the TM are poorly understood.

Differential effects of caveolin-1 and -2 knockdown on aqueous outflow and altered extracellular matrix turnover in caveolin-silenced trabecular meshwork cells.

Purpose: A single nucleotide polymorphism (SNP) identified between caveolin-1 (CAV1) and caveolin-2 (CAV2) on chromosome 7 is associated with glaucoma. One function of CAVs is endocytosis and recycling of extracellular matrix (ECM) components. Here, we generated CAV-silencing lentivirus to evaluate the effects on ECM turnover by trabecular meshwork (TM) cells and to measure the effect on outflow facility in anterior segment perfusion culture.

Intraocular pressure homeostasis: maintaining balance in a high-pressure environment.

Although glaucoma is a relatively common blinding disease, most people do not develop glaucoma. A robust intraocular pressure (IOP) homeostatic mechanism keeps ocular pressures within relatively narrow acceptable bounds throughout most peoples' lives. The trabecular meshwork and/or Schlemm's canal inner wall cells respond to sustained IOP elevation and adjust the aqueous humor outflow resistance to restore IOP to acceptable levels.

Perturbation of hyaluronan synthesis in the trabecular meshwork and the effects on outflow facility.

Purpose: Hyaluronan (HA) is a major component of the aqueous outflow pathway. However, the contribution of HA to human outflow resistance remains unclear. Three HA synthase genes (HAS1-3) have been identified.

Molecular chaperone function for myocilin.

Purpose: Myocilin is thought to be a stress response protein, but its exact molecular functions have not been established. Studies were conducted to see whether myocilin can act as a general molecular chaperone.

Methods: Myocilin was isolated and purified from porcine trabecular meshwork (TM) cell culture media.

Segmental versican expression in the trabecular meshwork and involvement in outflow facility.

Purpose: Versican is a large proteoglycan with numerous chondroitin sulfate (CS) glycosaminoglycan (GAG) side chains attached. To assess versican's potential contributions to aqueous humor outflow resistance, its segmental distribution in the trabecular meshwork (TM) and the effect on outflow facility of silencing the versican gene were evaluated.

Methods: Fluorescent quantum dots (Qdots) were perfused to label outflow pathways of anterior segments.

Differential effects of ADAMTS-1, -4, and -5 in the trabecular meshwork.

Purpose: Matrix metalloproteinases (MMPs) degrade extracellular matrix (ECM) and increase outflow facility in anterior segment perfusion culture. One group is the ADAMTSs (a disintegrin and metalloproteinase with thrombospondin type 1 motifs). In this study, the authors examined the effects of ADAMTS-1, -4, and -5 on outflow facility and investigated their mRNA levels and protein expression in the trabecular meshwork (TM).

Extracellular matrix turnover and outflow resistance.

Normal homeostatic adjustment of elevated intraocular pressure (IOP) involves remodeling the extracellular matrix (ECM) of the trabecular meshwork (TM). This entails sensing elevated IOP, releasing numerous activated proteinases to degrade existing ECM and concurrent biosynthesis of replacement ECM components. To increase or decrease IOP, the quantity, physical properties and/or organization of new components should be somewhat different from those replaced in order to modify outflow resistance.

Specialized podosome- or invadopodia-like structures (PILS) for focal trabecular meshwork extracellular matrix turnover.

Purpose: There are distinctive areas of colocalization of matrix metalloproteinase (MMP)-2 and -14 on trabecular meshwork (TM) cells that resemble podosomes or invadopodia. Studies were conducted to determine whether TM cells exhibit podosome- or invadopodia-like structures (PILS) and whether they produce focal extracellular matrix (ECM) turnover.

Methods: Porcine and human TM cells and perfused anterior segment organ cultures were studied.

Effects of modifiers of glycosaminoglycan biosynthesis on outflow facility in perfusion culture.

Purpose: Glycosaminoglycans (GAGs) have been implicated in the regulation of outflow resistance of aqueous humor flow through the trabecular meshwork (TM). Their role was further investigated by assessment of the effects of chlorate, an inhibitor of sulfation, and beta-xyloside, which provides a competitive nucleation point for addition of disaccharide units, in anterior segment perfusion culture.

Methods: Outflow facility was measured in perfused porcine and human anterior organ cultures treated with 20 or 50 mM sodium chlorate, or 1 mM beta-xyloside.

Synergism of TNF and IL-1 in the induction of matrix metalloproteinase-3 in trabecular meshwork.

Purpose: TNF and IL-1 increase matrix metalloproteinase-3 (MMP-3) expression in the trabecular meshwork (TM). TNF-alpha, in combination with IL-1alpha or IL-1beta, produces highly synergistic MMP-3 increases. Possible mechanisms for this synergism in TM cells were investigated.

Signaling pathways used in trabecular matrix metalloproteinase response to mechanical stretch.

Purpose: Trabecular meshwork (TM) matrix metalloproteinase (MMP), and tissue inhibitor (TIMP) changes in response to mechanical stretching appear to be central to intraocular pressure (IOP) homeostasis. Studies were conducted to define the signal transduction pathway responsible for the increases in MMP-2 and -14 that occur in response to mechanical stretching of TM cells.

Methods: Porcine TM cells were subjected to mechanical stretching, and changes in MMP-2 and -14 levels were determined by gelatin zymography and Western immunoblot analysis.