• animal models
• anti-inflammatory steroids
• corticosteroids
• eye
• glaucoma
• glucocorticoid receptor
• ocular hypertension
• primary open-angle glaucoma
• steroid glaucoma

• Humans
• Mice
• Animals
• Glucocorticoids/pharmacology
• Receptors, Glucocorticoid/metabolism
• Glaucoma, Open-Angle/chemically induced
• Glaucoma, Open-Angle/pathology
• Hydrocortisone
• Glaucoma/metabolism
• Ocular Hypertension/metabolism
• Iatrogenic Disease

• R01 EY016242 NEI NIH HHS
• R01 EY030967 NEI NIH HHS
• NIH EY016242 and EY030967 NEI NIH HHS

• Cross-links
• Extracellular Matrix
• Glaucoma
• Proteomics
• Segmental outflow
• Trabecular meshwork

• Humans
• Trabecular Meshwork
• Extracellular Matrix
• Glaucoma
• Aqueous Humor
• Ocular Hypertension

• R01 EY030238 NEI NIH HHS
• R01 EY019643 NEI NIH HHS
• R01 EY025721 NEI NIH HHS
• R01 DC002368 NIDCD NIH HHS
• R01 EY008247 NEI NIH HHS
• P30 CA069533 NCI NIH HHS
• P30 EY010572 NEI NIH HHS
• R01 EY026048 NEI NIH HHS
• R01 EY032590 NEI NIH HHS
• R01 EY016134 NEI NIH HHS

We investigated whether an inverse relationship exists between intraocular pressure (IOP) and effective filtration area (EFA) in the trabecular meshwork (TM) in a steroid-induced ocular hypertensive (SIOH) mouse model and the morphological changes associated with the reduction of EFA. C57BL/6 mice (n = 15 per group) received either 0.1% dexamethasone (DEX) or saline eye drops twice daily for five weeks. IOP was measured weekly. Fluorescent tracers were injected into the anterior chamber to label EFA at the endpoint. Injected eyes were fixed and processed for confocal microscopy. EFA in the TM was analyzed. Light and electron microscopy were performed in high- and low-tracer regions of six eyes per group. The mean IOP was ~4 mm Hg higher in DEX-treated than saline-treated control eyes (p < 0.001) at the endpoint. EFA was reduced in DEX-treated eyes compared to controls (p < 0.01) and negatively correlated with IOP (R² = 0.38, p = 0.002). Reduced thickness of juxtacanalicular tissue (JCT) and increased abnormal extracellular matrix in the JCT were found to be associated with reduced EFA. Our data confirm the inverse relationship between EFA and IOP, suggesting that morphological changes in the JCT contribute to the reduction of EFA, thus elevating IOP in SIOH mouse eyes.

• confocal microscopy
• effective filtration area
• intraocular pressure
• morphology
• steroid-induced ocular hypertensive mouse model
• trabecular meshwork
• transmission electron microscopy

• Glaucoma
• PLGA microspheres
• animal model
• dexamethasone
• neurodegeneration

• Animal model
• Glaucoma
• In vivo
• Intraocular pressure
• Pathogenesis
• Rodent

• Animals
• Disease Models, Animal
• Glaucoma/diagnosis
• Glaucoma/physiopathology
• Intraocular Pressure/physiology
• Optic Nerve/pathology
• Retinal Ganglion Cells/pathology

• R01 EY016242 NEI NIH HHS
• R01 EY030366 NEI NIH HHS
• R01 EY030967 NEI NIH HHS
• R21 EY024635 NEI NIH HHS

Adeno-associated viruses (AAVs) are the vector of choice for gene therapy in the eye, and self-complementary AAVs (scAAVs), which do not require second-strand DNA synthesis, can be transduced into cells of the trabecular meshwork (TM). The scAAV transduction patterns in the anterior segment of normotensive eyes have been investigated previously, but those in ocular hypertensive (OHT) eyes have not. We assessed the transduction efficiencies of AAV serotypes 2, 5, and 8 in the anterior-segment structures of the eyes of Sprague-Dawley rats with OHT by circumlimbal suturing, followed 3 days later by intracameral injection of scAAV serotype 2 (scAAV2), scAAV5, or scAAV8 packaged with EGFP. The transduction of scAAV2 and scAAV5 in the TM of OHT rats was markedly enhanced after 1 month, and transduction of scAAV5 was more efficient than that of scAAV2; transduction of scAAV8 into the TM did not occur. The transduction of scAAV2, scAAV5, and scAAV8 was enhanced in the ciliary body, iris, and corneal endothelium of the OHT eyes for 3 months. The expression levels of receptors for scAAV2 and scAAV5 were significantly increased in the OHT compared with control eyes. The results demonstrated that scAAV2 and scAAV5 target the ciliary body and TM in OHT eyes, and that the OHT-related changes in anterior-segment structures enhance scAAV transduction.

• adeno-associated virus
• gene therapy
• glaucoma

• chronic ocular hypertension
• glaucoma
• rodent models

• Glaucoma
• Intraocular pressure
• Model
• Steroids

• Animals
• Cell Culture Techniques
• Dexamethasone/adverse effects
• Disease Models, Animal
• Glaucoma/chemically induced
• Glaucoma/physiopathology
• Glucocorticoids/adverse effects
• Humans
• Intraocular Pressure/drug effects
• Ocular Hypertension/chemically induced
• Ocular Hypertension/physiopathology
• Organ Culture Techniques
• Trabecular Meshwork/drug effects
• Trabecular Meshwork/pathology
• Triamcinolone Acetonide/adverse effects

• R01 EY025543 NEI NIH HHS

• Bevacizumab
• Clearance
• Intraocular pressure
• Schlemm's canal
• Trabecular meshwork

• Angiogenesis Inhibitors/analysis
• Angiogenesis Inhibitors/pharmacokinetics
• Animals
• Anterior Chamber/metabolism
• Bevacizumab/analysis
• Bevacizumab/pharmacokinetics
• Choroidal Neovascularization/drug therapy
• Choroidal Neovascularization/metabolism
• Cornea/metabolism
• Disease Models, Animal
• Intravitreal Injections
• Iris/metabolism
• Male
• Rats
• Rats, Inbred BN
• Time Factors
• Trabecular Meshwork/chemistry
• Trabecular Meshwork/metabolism
• Vascular Endothelial Growth Factor A/metabolism

• Glaucoma
• Glucocorticoid
• Intraocular pressure
• Ocular hypertension
• Steroid glaucoma

• Animal models
• Glaucoma
• Viral vectors

Proteins have important functional roles in the body, which can be altered in disease states. The eye is a complex organ rich in proteins; in particular, the anterior eye is very sophisticated in function and is most commonly involved in ophthalmic diseases. Proteomics, the large scale study of proteins, has greatly impacted our knowledge and understanding of gene function in the post-genomic period. The most significant breakthrough in proteomics has been mass spectrometric identification of proteins, which extends analysis far beyond the mere display of proteins that classical techniques provide. Mass spectrometry functions as a “mass analyzer” which simplifies the identification and quantification of proteins extracted from biological tissue. Mass spectrometric analysis of the anterior eye proteome provides a differential display for protein comparison of normal and diseased tissue. In this article we present the key proteomic findings in the recent literature related to the cornea, aqueous humor, trabecular meshwork, iris, ciliary body and lens. Through this we identified unique proteins specific to diseases related to the anterior eye.

• Mass spectrometry
• Proteomics
• Ocular
• Glaucoma

• R01 EY016112 NEI NIH HHS

• Animals
• Biomedical Research
• Humans
• Manuscripts as Topic
• Ophthalmology
• Review Literature as Topic
• Serial Publications/trends

• dexamethasone
• fibronectin
• human trabecular meshwork cells
• latrunculin-A

The intention of this review is to provide an overview of current methodologies employed in the rapidly developing field of ocular proteomics with emphasis on sample preparation, two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) and mass spectrometry (MS). Appropriate sample preparation for the diverse range of cells and tissues of the eye is essential to ensure reliable results. Current methods of protein staining for 2D-PAGE, protein labelling for two-dimensional difference gel electrophoresis, gel-based expression analysis and protein identification by MS are summarised. The uses of gel-free MS-based strategies (MuDPIT, iTRAQ, ICAT and SILAC) are also discussed. Proteomic technologies promise to shed new light onto ocular disease processes that could lead to the discovery of strong novel biomarkers and therapeutic targets useful in many ophthalmic conditions.

In addition to their well-documented ocular therapeutic effects, glucocorticoids (GCs) can cause sight-threatening side-effects including ocular hypertension presumably via morphological and biochemical changes in trabecular meshwork (TM) cells. In the present study, we directly compared the glucocorticoid receptor (GR) potency for dexamethasone (DEX), fluocinolone acetonide (FA) and triamcinolone acetonide (TA), examined the expression of known GRα and GRβ isoforms, and used gene expression microarrays to compare the effects of DEX, FA, and TA on the complete transcriptome in two primary human TM cell lines.

GR binding affinity for DEX, FA, and TA was measured by a cell-free competitive radio-labeled GR binding assay. GR-mediated transcriptional activity was assessed using the GeneBLAzer beta-lactamase reporter gene assay. Levels of GRα and GRβ isoforms were assessed by Western blot. Total RNA was extracted from TM 86 and TM 93 cells treated with 1 μM DEX, FA, or TA for 24 hr and used for microarray gene expression analysis. The microarray experiments were repeated three times. Differentially expressed genes were identified by Rosetta Resolver Gene Expression Analysis System.

The GR binding affinity (IC₅₀) for DEX, FA, and TA was 5.4, 2.0, and 1.5 nM, respectively. These values are similar to the GR transactivation EC₅₀ of 3.0, 0.7, and 1.5 nM for DEX, FA, and TA, respectively. All four GRα translational isoforms (A-D) were expressed in TM 86 and TM 93 total cell lysates, however, the C and D isoforms were more highly expressed relative to A and B. All four GRβ isoforms (A-D) were also detected in TM cells, although GRβ-D isoform expression was lower compared to that of the A, B, or C isoforms. Microarray analysis revealed 1,968 and 1,150 genes commonly regulated by DEX, FA, and TA in TM 86 and TM 93, respectively. These genes included RGC32, OCA2, ANGPTL7, MYOC, FKBP5, SAA1 and ZBTB16. In addition, each GC specifically regulated a unique set of genes in both TM cell lines. Using Ingenuity Pathway Analysis (IPA) software, analysis of the data from TM 86 cells showed that DEX significantly regulated transcripts associated with RNA post-transcriptional modifications, whereas FA and TA modulated genes involved in lipid metabolism and cell morphology, respectively. In TM 93 cells, DEX significantly regulated genes implicated in histone methylation, whereas FA and TA altered genes associated with cell cycle and cell adhesion, respectively.

Human trabecular meshwork cells in culture express all known GRα and GRβ translational isoforms, and GCs with similar potency but subtly different chemical structure are capable of regulating common and unique gene subsets and presumably biologic responses in these cells. These GC structure-dependent effects appear to be TM cell-lineage dependent.