PURPOSE: To investigate the effect of beta-adrenergics on the production of nitric oxide (NO) in cultured trabecular meshwork cells. METHODS: Primarily cultured porcine trabecular meshwork cells were exposed to timolol and to propranolol at various concentrations. Cellular survival was assessed by MTT assay and the production of nitrite was assessed by Griess reaction after 24 hours and 3 days respectively. Also investigated was the effect of co-administraton of timolol and isoproterenol. RESULTS: After exposure for 24 hours, neither drug affected the cellular survival. Timolol and propranolol inhibited the production of NO significantly (p<0.05). Isoproterenol abolished timolol-induced inhibition of NO production after 24 hours. These results were similar after exposure for 3 days. CONCLUSIONS: Both timolol and propranolol inhibit the production of NO in trabecular meshwork cells and isoproterenol abolished this effect. These results suggest that beta-adrenerics involves in the production of NO in trabecular meshwork cells.
Isoproterenol ; Nitric Oxide* ; Propranolol ; Timolol ; Trabecular Meshwork*

PURPOSE: To investigate the cytotoxicities of wild type myocilin and trunctated (Q368X) myocilin in cultured human trabecular meshwork (TM) cells. METHODS: GFP tagged truncated myocilin and DsRED1 tagged wild type myocilin were expressed in TM cells using adenoviral vectors and observed colocalization by confocal microscope. Cytopathic effects in the cells were examined by light microscope and WST-1 cell proliferation assay. RESULTS: Colocalization of wild type and truncated myocilin was observed in cells co-expressing the proteins. Truncated myocilin was found to be toxic to cells, leading to deformed cellular morphology and diminished cell proliferation. CONCLUSIONS: The intracellular accumulation of truncated myocilin exhibited cytotoxicity in trabecular meshwork cells, and eventually resulted in diminished number and dysfunction of trabecular meshwork cells, which might be involved in the pathogenesis of glaucoma.
Cell Proliferation ; Glaucoma ; Humans ; Trabecular Meshwork*

PURPOSE: To investigate the effects of genistein on the survival and production of nitric oxide in cultured human trabecular meshwork cells (HTMC). METHODS: Primarily cultured HTMC were exposed to 0, 1, and 10 microm genistein using serum-deprived media. Production of nitric oxide and eNOS activity were assessed with the Griess assay and RT-PCR after exposure to genistein for 10 min and one day. Cellular survival was assessed via MTT assay after exposure to genistein for one day. RESULTS: Genistein significantly increased the production of nitric oxide after exposure for 1 min at 10 microm and for 1 day at 1 microm under serum-deprived conditions. Genistein increased eNOS activity and cellular survival in HTMC. CONCLUSIONS: Genistein increases cellular survival under serum-deprived conditions, accompanied with an increase in nitric oxide production after both short-term and long-term exposures.
Genistein ; Humans ; Nitric Oxide ; Trabecular Meshwork

We evaluated thirty normal human eyes(aged from 22 to 95) to investigate the changes with age of the cellularity and acid mucopolysaccharides in the trabecular meshwork. The cellularities were evaluated from the number of the cells per unit area of trabecular meshwork and the number of cells per unit length of trabecular meshwork. In addition, acid mucopolysaccharides(AMS) were investigated by counter staining the trabecular meshwork specimens, with Van Gieson following colloidal iron. The results were as follows: 1. With age, cellularities of the whole trabecular meshwork and the counterpart of filtration region decreased significantly(p0.05). 2. The trabecular meshwork cellularity and the absolute number of cells were decreased by 40.0% and 35.9%, respectively, for those aged from 22 to 95. 3. There was no strong relation between the change of acid mucopolysaccharides of the trabecular meshwork and aging process. The grade +1 AMS was observed in 15 eyes(50.0%) and was the most prevalent.
Aging ; Colloids ; Filtration ; Glaucoma ; Glycosaminoglycans* ; Humans ; Iron ; Trabecular Meshwork*

PURPOSE: To investigate the effects of nitroglycerin on the production of nitric oxide and its related pathway in cultured human trabecular meshwork cells (HTMC). METHODS: Primarily cultured HTMC were exposed to 10 nM nitroglycerin using 1% serum-containing media for 30 minutes. The production of nitric oxide was assessed with the Griess assay and expressions of eNOS mRNA was assessed with RT-PCR. Additionally, the cells were exposed to wortmanin and Akt1/2 kinase inhibitor to investigate the mechanism related to the production of nitric oxide. RESULTS: Nitroglycerin increased the production of nitric oxide (p < 0.05) accompanied with increased expression of eNOS mRNA. The increased production of nitric oxide and eNOS mRNA was inhibited by wortmanin and Akt1/2 kinase inhibitor. CONCLUSIONS: Low-dose nitroglycerin increased the production of nitric oxide accompanied by increased eNOS activity. Nitroglycerin drives eNOS activation via the phosphatidylinositol 3-kinase/protein kinase B pathway.
Humans ; Nitric Oxide ; Nitroglycerin ; Phosphatidylinositols ; Phosphotransferases ; RNA, Messenger ; Trabecular Meshwork

PURPOSE: To investigate the effects of tetrahydrozoline (THZ) on the survival of cultured human trabecular meshwork cells (HTMC) and the permeability of HTMC monolayer. METHODS: Primary cultured HTMC were exposed to an adrenergic agonist (0.01, 0.1, 1.0 or 10 µM THZ) for 1 day and 3 days. Carboxyfluorescein permeability through the HTMC monolayer was measured using Transwell. Cellular viability and nitric oxide (NO) production were assessed using MTT and Griess assays, respectively. RESULTS: THZ did not affect the cellular survival (p > 0.05) or NO production (p > 0.05). THZ significantly increased the carboxyfluorescein permeability through the HTMC monolayer in a dose-dependent manner compared with non-exposed control (p < 0.05) after exposure for 1 and 3 days. CONCLUSIONS: THZ does not affect the survival of HTMC but decreases the permeability of HTMC monolayer in a dose-dependent manner. Thus, THZ may possibly decrease trabecular outflow.
Adrenergic Agonists ; Humans ; Nitric Oxide ; Permeability* ; Trabecular Meshwork*

PURPOSE: To investigate the effects of benzalkonium chloride (BAC), mitomycin C (MMC) and dexamethasone (DEX) on cellular stress in cultured human trabecular meshwork cell (HTMC) monolayers. METHODS: HTMCs were cultured in the inner Transwell chamber until confluence and then were exposed to BAC, MMC or DEX for 6 hours. The carboxyfluorescein permeability through the HTMC monolayer was measured using a spectrofluorometer at 532 nm after 2 hours in the outer chamber. The 3-[4, 5 -dimethylthiazol-2-yl]-2, 5-diphenyltetrazolium bromide (MTT) assay was used to evaluate cellular viabilities. RESULTS: The carboxyfluorescein permeability through the HTMC monolayer increased and cell survival decreased with 0.002% BAC (p < 0.05). Increased permeability without decreasing cell survival occurred with 0.05 microg/mL MMC. No effect on the permeability or cell survival was observed at 0.1 or 1.0 microm DEX (p > 0.05). CONCLUSIONS: BAC and MMC induced cellular toxicity and stress at lower concentrations but did not affect survival of cultured HTMCs.
Benzalkonium Compounds* ; Cell Survival ; Dexamethasone* ; Humans ; Mitomycin* ; Permeability ; Trabecular Meshwork*

The present study investigates the role of TGF-beta1 as one of possible etiologic factors of glaucoma by studying the effect of TGF-beta1 on the secretion of matrix metalloproteinase and tissue inhibitor of matrix metalloproteinase. The cultured samples of trabecular meshwork cells were treated with TGF-beta1 in concentrations of 0, 100, 1000 and 10000 pg/ml. The samples were analysed for the secretion of MMP2 and timp2 through electrophoresis, western blot and TGF-beta1 increased, the secretions of MMP2 and TIMP2 were not significantly changed after 24 hours. But the secretion of MMP2 was decreased while the secretion of TIMP2 was increased after 72 hours. The results suggest that TGFbeta1 may be one of the etilogic factors of glaucoma.
Blotting, Western ; Electrophoresis ; Glaucoma ; Trabecular Meshwork ; Transforming Growth Factor beta1

PURPOSE: To evaluate the morphological characteristics of the transitional zone between the corneal endothelium and the trabecular meshwork by flat preparation and electron microscopy. METHODS: The materials comprised 12 eyes examined by the flat preparation and 7 eyes by the electron microscopy. The specimens were derived from the transitional tissue between the corneal endothelium and the trabecular meshwork. The specimens in the flat preparation were stained with hematoxylin-eosin and examined by light microscopy. The specimens for scanning electronic microscopy (SEM) and in transmission electronic microscopy (TEM) were examined through routine processes. RESULTS: In the specimens examined by the flat preparation, unlike peripheral corneal endothelial cells, the endothelial cell nuclei in the transitional zone were overlapped and morphologically oval. On SEM, unlike typical hexagonality and tight interdigitation of corneal endothelial cells, the endothelial cells in the transitional zone were partially successive, spaced intercellularly, and morphologically irregular. On TEM, the endothelial cells in the transitional zone were partially successive. CONCLUSIONS: The loss of cell-cell contact of endothelial cells in the transitional zone may lead to the potential proliferation capacity of endothelial cells in the transitional zone under specific conditions. Therefore, further studies on the proliferation capacity of endothelial cells in the transitional zone are needed together with more research on cell biology.
Endothelial Cells ; Endothelium, Corneal* ; Microscopy ; Microscopy, Electron ; Trabecular Meshwork

PURPOSE: To compare the effects of anti-inflammatory agents, specifically bromfenac, loteprednol, and prednisolone, on the permeability of cultured human trabecular meshwork cell (HTMC) monolayers. METHODS: HTMCs were cultured until confluency in the inner chamber of Transwell, then exposed to 1/1,000 or 1/500 diluted commercial 0.1% bromfenac, 0.5% loteprednol, and 1% prednisolone for 24 hours. The permeabilities of carboxyfluorescein through the HTMC monolayer were measured with a spectrofluorometer after 2 hours in the outer chamber. Cellular viabilities were assessed with an 3-[4,5–dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) assay. RESULTS: Bromfenac and loteprednol diluted at 1/1,000 or 1/500 did not significantly affect the cellular survival (p > 0.05). Bromfenac did not affect the permeability via the HTMC monolayer (p > 0.05) and loteprednol decreased the permeability (p < 0.05). In addition, 1/2,000 prednisolone also decreased the permeability (p < 0.05). CONCLUSIONS: Among the anti-inflammatory agents, the non-steroidal anti-inflammatory agent bromfenac did not affect the permeability, while loteprednol and prednisolone decreased the permeability through the HTMC monolayer. Thus, loteprednol and prednisolone may decrease the trabecular outflow.
Anti-Inflammatory Agents* ; Humans ; Loteprednol Etabonate ; Permeability* ; Prednisolone ; Trabecular Meshwork*