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PubMed2002

Translimbal laser photocoagulation to the trabecular meshwork as a model of glaucoma in rats.

PURPOSE: To develop and characterize a model of pressure-induced optic neuropathy in rats. METHODS: Experimental glaucoma was induced unilaterally in 174 Wistar rats, using a diode laser with wavelength of 532 nm aimed at the trabecular meshwork and episcleral veins (combination treatment group) or only at the trabecular meshwork (trabecular group) through the external limbus. Intraocular pressure (IOP) was measured by a tonometer in rats under ketamine-xylazine anesthesia. Possible retinal vascular compromise was evaluated by repeated fundus examinations and by histology. The degree of retinal ganglion cell (RGC) loss was assessed by a masked, semiautomated counting of optic nerve axons. Effects of laser treatment on anterior ocular structures and retina were judged by light microscopy. RESULTS: After the laser treatment, IOP was increased in all eyes to higher than the normal mean IOP of 19.4 +/- 2.1 mm Hg (270 eyes). Peak IOP was 49.0 +/- 6.1 mm Hg (n = 108) in the combination group that was treated by a laser setting of 0.7 seconds and 0.4 W and 34.0 +/- 5.7 mm Hg (n = 46) in the trabecular group. Mean IOP after 6 weeks was 25.5 +/- 2.9 mm Hg in glaucomatous eyes in the combination group compared with 22.0 +/- 1.8 mm Hg in the trabecular group. IOP in the glaucomatous eyes was typically higher than in the control eyes for at least 3 weeks. In the combination group, RGC loss was 16.1% +/- 14.4% at 1 week (n = 8, P = 0.01), 59.7% +/- 25.7% at 6 weeks (n = 88, P < 0.001), and 70.9% +/- 23.6% at 9 weeks (n = 12, P < 0.001). The trabecular group had mean axonal loss of 19.1% +/- 14.0% at 3 weeks (n = 9, P = 0.004) and 24.3% +/- 20.2% at 6 weeks (n = 25, P < 0.001), increasing to 48.4% +/- 32.8% at 9 weeks (n = 12, P < 0.001). Laser treatment led to closure of intertrabecular spaces and the major outflow channel. The retina and choroid were normal by ophthalmoscopy at all times after treatment. Light microscopic examination showed only loss of RGCs and their nerve fibers. CONCLUSIONS: Increased IOP caused by a laser injury to the trabecular meshwork represents a useful and efficient model of experimental glaucoma in rats.

Animals↗
PubMed1999

Stimulation of maxi-K channels in trabecular meshwork by tyrosine kinase inhibitors.

PURPOSE: Muscarinic agonists contract and tyrosine kinase inhibitors relax precontracted trabecular meshwork, a smooth muscle-like tissue involved in the regulation of aqueous humor outflow. The effect of tyrosine kinase inhibitors on membrane currents of cells stimulated by acetylcholine was examined. METHODS: Cells from bovine trabecular meshwork were studied using both the perforated patch-clamp technique with nystatin and the single-channel technique. RESULTS: Application of the tyrosine kinase inhibitor genistein (5 x 10(-5) M) on trabecular meshwork cells stimulated with acetylcholine resulted in a reversible increase in outward current to 578%+/-154% (n = 16) of the initial current level. The effect of genistein was dose dependent. Reversal potential was hyperpolarized by 15+/-3 mV (n = 9). Tyrphostin 51, a synthetic inhibitor of tyrosine kinases, had the same effect (433%+/-46%; n = 7). Daidzein, a nonactive structural analogue of genistein, had no effect (n = 4). The stimulation of outward current by tyrosine kinase inhibitors was blocked by substitution of tetraethylammonium (TEA+) for potassium, whereas the potassium channel blockers glibenclamide (K-ATP) and apamin (low-conductance calcium-activated potassium channel) had no effect. Blockage of the high-conductance calcium-activated potassium channel (maxi-K) by charybdotoxin or iberiotoxin (10(7) M) suppressed 86%+/-18% (n = 4) of the response. Depleting the cells of calcium did not have an effect on the current stimulated by genistein. In the excised inside-out configuration, open probability increased to 417%+/-39% (n = 3) after exposure to genistein. CONCLUSIONS: In trabecular meshwork, tyrosine kinase inhibitors activate maxi-K (K(Ca)) channels. Hyperpolarization caused by efflux of potassium could lead to the relaxation of trabecular meshwork by tyrosine kinase inhibitors.

Acetylcholine↗
PubMed1981

Age-related changes in trabecular meshwork cellularity.

To investigate whether alterations in the cellular component of the trabecular meshwork occur with age, we evaluated trabecular meshwork cellularity and absolute cell number, using specimens obtained from patients ranging in age from newborn to 81 years. We obtained reproducible, quantitative measurements of these parameters, using montages of 1 micrometer meridional sections. A progressive decrease in cellularity (58%) and absolute cell number (47%) in the trabecular meshwork occurred over the 81-year period studied. Regression analysis suggested that the decrease in cellularity and cell number is linear function of age, resulting from a loss of approximately 0.58% of all cells per year. This change in cellularity parallels that seen in the corneal endothelium. implying that trabecular cells, like corneal endothelial cells, may have a limited reparative capacity.

Adolescent↗
PubMed1995

Glycoconjugates of the human trabecular meshwork: a lectin histochemical study.

Twelve specimens of resin-embedded human trabecular meshwork were probed with a panel of 21 biotinylated lectins, using an avidin-biotin peroxidase revealing system, in order to determine the normal pattern of saccharide expression in this tissue. High-mannose, intermediate and hybrid N-linked glycans, and complex N-linked bisected and non-bisected bi/tri-antennate glycans, as shown by the binding of Canavalia ensiformis (ConA), Pisum sativum (PSA), Lens culinaris (LCA) agglutinins and Phaseolus vulgaris erythroagglutinin (ePHA), were strongly expressed by the canal of Schlemm endothelium and juxtacanalicular tissue, but less so by the corneoscleal meshwork. Highly branced complex glycans were not found, as there was no binding by Phaseolus vulgaris leukoagglutinin (IPHA). Sialyl residues, especially those alpha 2,6-linked as demonstrated by strong Sambucus nigra (SNA) lectin staining, were also abundant in this area. N-acetyllactosamine sequences and some O-linked glycans were present in the trabecular meshwork, as shown by Solanum tuberosum (STA), Datura stramonium (DSA), and Jacalin (Jac) lectin binding, while fucose residues were not detected by Tetragonolobus purpureas (LTA) or Ulex europaeus-1 (UEA-1) agglutinins. These results indicate similarities with renal glomerular and vascular endothelium, although the lack of binding with UEA-1 agglutinin suggests differences which may relate to the specialized function of the trabecular meshwork. This study provides a baseline for comparative analysis of the glycans of human trabecular meshwork in pathological conditions such as primary open-angle glaucoma.

Adult↗
PubMed2004

Effect of CD44 suppression by antisense oligonucleotide on attachment of human trabecular meshwork cells to HA.

The effects of suppression of CD44 by CD44-specific antisense oligonucleotide on attachment of human trabecular meshwork cells to hyaluronic acid (HA) were observed and the possible relationship between CD44 and primary open-angle glaucoma (POAG) investigated. CD44-specific antisense oligonucleotide was delivered with cationic lipid to cultured human trabecular meshwork cells. The expression of CD44 suppressed by CD44-specific antisense oligonucleotide was detected by RT-PCR and Western blotting. The effect of CD44 suppression by specific antisense oligonucleotide on attachment of trabecular meshwork cells to HA was measured by MTT assay. Results showed that expression of CD44 was suppressed by CD44-specific antisense oligonucleotide. Antisense oligonucleotide also suppressed the adhesion of human trabecular meshwork cells to HA in a concentration dependent manner. It was concluded that attachment of human trabecular meshwork cells to HA was decreased when CD44 was suppressed by specific antisense oligonucleotide. CD44 might play a role in pathogenesis of POAG by affecting the adhesion of trabecular meshwork cells to HA.

Cell Adhesion↗
PubMed1989

The effect of organ culture on human trabecular meshwork.

The effect of organ culture on the trabecular meshwork was studied in a series of human eyes using a perfusion culture system. One eye of a pair was cultured while the fellow eye was immediately fixed in glutaraldehyde. Culture periods ranged from 2 to 28 days, during which a mean intraocular pressure of 27 mmHg +/- 15 was maintained. The perfused culture medium appeared to leave the eye via Schlemm's canal and collector channels, mimicking the in vivo situation. The trabecular meshwork was well maintained with this system in 21 of 25 eyes. Cells remained in position on trabecular lamellae, cellular organelles usually remained normal, and the lamellae remained intact. Culture-induced changes were noted, with some cells developing intracytoplasmic lipid vacuoles and other cells developing swollen mitochondria. In addition, scattered focal cell necrosis was observed, most often in the uveal meshwork. Meshwork cellularity (nuclei:solid tissue) was determined with an image analysis system; there was an average cell loss of 20-40% in the cultured eyes as compared to their respective fellow control eyes. Overall, organ culture appears to maintain the human trabecular meshwork for at least 28 days, allowing controlled experimental studies to be performed.

Aged↗
PubMed2002

Effect of transforming growth factor-beta(2) on extracellular matrix synthesis in bovine trabecular meshwork cells.

OBJECTIVE: To investigate the effect of transforming growth factor-beta(2) (TGF-beta(2)) in human aqueous humor on extracellular matrix synthesis in cultured bovine trabecular meshwork cells. METHODS: Cultured 3-5 passage bovine trabecular meshwork cells were divided into control group and experimental group. After treated with 0 ng/L (control), 0.32 x 10(3) ng/L, 1.00 x 10(3) ng/L, 3.20 x 10(3) ng/L TGF-beta(2) for 48 h, collagen, fibronectin and hyaluronic acid synthesis in bovine trabecular meshwork cells were examined respectively by (3)H-proline incorportion and liquid scintillation technique, enzyme linked immunosorbent assay (ELISA) and radioimmunoassay (RIA). RESULTS: In comparison with the control group, TGF-beta(2) significantly promoted the synthesis of collagen at 0.32 x 10(3) ng/L (P < 0.05), 1.00 x 10(3) ng/L (P < 0.01), 3.20 x 10(3) ng/L (P < 0.01), (3)H-proline incorporation increased in a concentration-dependent manner; and 1.00 x 10(3) ng/L (P < 0.05), 3.20 x 10(3) ng/L (P < 0.01) TGF-beta(2) significantly promoted the synthesis of fibronectin in cultured bovine trabecular meshwork cells. 1.00 x 10(3) ng/L (P < 0.01) and 3.20 x 10(3) ng/L (P < 0.01) TGF-beta(2) significantly inhibited the synthesis of hyaluronic acid in the cultured cells. CONCLUSION: The data suggest that TGF-beta(2) play important roles in extracellular matrix age-related changes of normal trabecular meshwork and abnormal deposition of the extracellular matrix in the trabecular meshwork, especially the juxtacanalicular tissue of primary open-angle glaucoma patients.

Animals↗
PubMed1999

Expression of integrin receptors in the human trabecular meshwork.

PURPOSE: To examine the expression of integrin receptors in the human trabecular meshwork. METHODS: The expression of integrins in human tissues was visualized by immunohistochemical staining using integrin-specific antibodies. Immunoprecipitation was performed after biotin labeling of cell surface proteins. Reverse transcriptase-polymerase chain reaction was used to detect the presence of mRNA species for integrin subunits. RESULTS: Human trabecular meshwork tissues obtained from donors 2 to 65 years old stained positively for integrins alpha1, alpha3, alpha4, alpha5, alpha6, alphav, beta1, beta3, beta4 and beta5. The staining was observed mostly around the edges of trabecular beams in association with trabecular meshwork cells. The staining intensity did not appear to vary with donor age. In addition, immunoprecipitation of tissue extracts revealed the presence of integrin alpha2 and confirmed the absence of beta2. Results from polymerase chain reactions were consistent with these findings. CONCLUSIONS: A spectrum of integrin receptors that may have important roles in the cell-matrix interactions are demonstrated in the human trabecular meshwork. The repertoire identified in tissues is similar to that found in cultured cells except that the beta4 expression in tissues is lost in cultures.

Adult↗
PubMed1983

[Immunofluorescence studies of the human trabecular meshwork].

Accumulation of extracellular material in the trabecular meshwork is responsible for increased resistance to outflow in the human eye. By means of the indirect immunofluorescence technique the authors detected extracellular collagen type IV, fibronectin and laminin in the trabecular meshwork and the wall of Schlemm's canal of glaucoma patients and controls. Collagen type IV is the so-called "basement membrane collagen" present in the lens capsule, Bowman's and Descemet's membranes. Fibronectin is a glycoprotein with many biologic activities; it can promote cell attachment and forms complexes with collagen. Laminin was isolated in 1979 and is present in the lamina lucida of basement membranes. The increased production and pathological metabolism of these three substances might be of some importance in the pathogenesis of primary open-angle glaucoma.

Collagen↗
PubMed2002

[Expression of tissue inhibitors of metalloproteinase in monkey trabecular meshwork cells and ciliary muscle cells].

OBJECTIVE: To identify tissue inhibitors of metalloproteinase (TIMP) released by normal monkey trabecular meshwork cells and ciliary muscle cells in vitro and to detect the roles of matrix metalloproteinase (MMP) and TIMP playing in trabecular outflow and uveoscleral outflow in normal eyes. METHODS: The expressions of TIMP and MMP in monkey trabecular meshwork and ciliary muscle cultures were detected by the technology of reverse zymography. RESULTS: (1) TIMP-1, TIMP-2 and TIMP-3 were found in both trabecular meshwork and ciliary muscle cultured cells. (2) The ratio of MMP/TIMP in trabecular meshwork cells was much higher than that in ciliary muscle cells. CONCLUSIONS: The different ratio of MMP/TIMP in these two types of cells suggests that the capability of degrading extracellular matrix in trabecular meshwork cells be much higher than that in ciliary muscle cells, that may be one of the reasons why conventional trabecular outflow is stronger than uveoscleral outflow.

Animals↗
PubMed2003

Bradykinin enhancement of PGE2 signalling in bovine trabecular meshwork cells.

Kinins and prostaglandins activate signalling pathways in cells of the trabecular meshwork and have opposing effects on outflow resistance to aqueous humor. Consequently, interactions between these pathways may be important in the regulation of intraocular pressure. In the present study, the influence of bradykinin on PGE(2) signalling was examined in primary cultures of bovine trabecular meshwork cells. Incubation of cells with bradykinin produced a concentration-dependent (EC(50)=3.6+/-0.7 nM) elevation of intracellular free Ca(2+). At a maximal concentration of 100 nM, the increase in Ca(2+) was rapid, peaking in 30 sec, and then slowly returned to baseline. This effect was completely inhibited in cells pretreated with the selective B(2) kinin receptor antagonist, Hoe-140. Treatment of trabecular meshwork cells with PGE(2), in comparison, had no effect on cellular Ca(2+) but produced a concentration-dependent increase in adenosine 3', 5'-cyclic monophosphate (cAMP) formation. Bradykinin had no effect on basal cAMP. However, incubation of cells with PGE(2) in combination with bradykinin resulted in a 3- to 5-fold enhancement of PGE(2)-stimulated cAMP production. Bradykinin enhancement of cAMP stimulation was concentration-dependent with an EC(50) of 3.6+/-1.8 nM. Treatment of cells with bradykinin increased the response maximum for PGE(2) signalling, while the EC(50) for PGE(2) was not changed. This action of bradykinin was again blocked in cells pretreated with Hoe-140. Bradykinin also produced a 2- to 3-fold increase in isoproterenol and cholera toxin-stimulated cAMP accumulation. However, when adenylyl cyclase was stimulated directly with forskolin, bradykinin failed to alter cAMP production. These results indicate that bradykinin activates B(2) kinin receptors in trabecular meshwork cells to amplify PGE(2)-stimulated cAMP formation by facilitating the interaction between activated G(s) and the catalytic unit of adenylyl cyclase.

Animals↗
PubMed1976

Immunofluorescent studies on the trabecular meshwork in open-angle glaucoma.

The trabecular meshwork of eyes with open-angle glaucoma has been demonstrated to have an increase in gamma globulin and plasma cells, raising the question of an immunogenic mechanism in this disorder. In the present study, however, immunofluorescence assays on the trabecular meshwork of eyes with open-angle glaucoma were negative for specific immunoglobulins and for complement components that would result specifically from an antigen-antibody reaction. The study fails to provide any evidence in support of an immunogenic mechanism in open-angle glaucoma.

Adult↗
PubMed1985

The effect of dexamethasone on the synthesis of collagen in normal human trabecular meshwork explants.

This study demonstrates that the trabecular meshwork cells of the human eye incorporate 3H-Proline into collagen during in vitro incubation. Addition of dexamethasone to the incubation mixture produced a marked decrease in this incorporation. Dexamethasone was active at 10(-8) M and higher concentrations. The specificity of the hormone effect was demonstrated by its inability to alter 3H-leucine incorporation in these cells. These results indicate that dexamethasone decreases the synthesis of collagen, a major component of the extracellular matrix, in the trabecular meshwork.

Collagen↗
PubMed2001

Tear in the trabecular meshwork caused by an airsoft gun.

PURPOSE: To report a case of a tear in the trabecular meshwork caused by an airsoft gun, a toy that propels a plastic bullet. METHODS: Case report. RESULTS: A 7-year-old Japanese boy sustained an ocular injury to the right eye from an airsoft gun. Ophthalmic examination 1 hour after the injury showed a best-corrected visual acuity in the injured eye of hand motion, corneal abrasion and edema, hyphema, and commotio retinae. Gonioscopy 6 days after the injury revealed a tear in the trabecular meshwork as well as an angle recession. Ultrasound biomicroscopy (UBM) strongly suggested that the tear extended into Schlemm's canal. Corneal abrasion and edema, hyphema, and commotio retinae resolved over 10 days, and best-corrected visual acuity improved to 20/15. Two months after the injury, the trabecular meshwork had not healed. CONCLUSION: Airsoft guns can cause a full-thickness tear in the trabecular meshwork, which may contribute to development of late-onset glaucoma. UBM is useful to evaluate the tomographic features of the disrupted trabecular meshwork. The potential force of airsoft guns to cause substantial ocular injuries should be recognized. Wearing ocular protection should be mandatory while playing with airsoft guns.

Child↗
PubMed1990

Glycosaminoglycans of human trabecular meshwork in perfusion organ culture.

The synthetic profile of glycosaminoglycans (GAGs) of human trabecular meshwork in perfusion organ culture was studied in a series of 34 human eyes. The anterior segments of these eyes were cultured for periods of two to 28 days and received medium containing 3H-glucosamine and 35S-sulfate during the final 48 hours of culture. The meshwork was then dissected and the GAGs isolated and subjected to sequential enzyme digestion. Active labelling of hyaluronic acid, chondroitin sulfate, dermatan sulfate, keratan sulfate, and heparan sulfate was found in all time periods. Eyes cultured seven and 14 days had similar incorporation profiles to "fresh" eyes (cultured 48 hours to allow for labelling). Eyes cultured 21 days showed an increase in dermatan sulfate labelling and a slight decrease in keratan sulfate labelling when compared with "fresh" eyes. Light microscopic autoradiography confirmed the trabecular meshwork incorporation of the radiolabelled precursors at all time periods. Thus, the trabecular meshwork remains metabolically active and GAG synthetic profiles remain reasonably similar to fresh eyes for up to three weeks in a perfusion organ culture system. This system may serve as a model for future studies of human trabecular meshwork GAGs.

Adult↗
PubMed1997

Differences in the trabecular meshwork between Belgian and Congolese patients with open-angle glaucoma.

PURPOSE: To study the alterations of the trabecular meshwork of Belgian and Congolese patients with open angle glaucoma (OAG). METHODS: A trabeculectomy was performed in 27 OAG patients from Belgium and 24 from Congo; the trabecular specimens were fixed, embedded, stained, and studied by light microscopy. These specimens were compared with the trabecular meshworks from 5 Belgian non-glaucomatous eyes. RESULTS: The mean number (+/- standard deviation) of trabecular cells per field was 69 (+/- 10) in Belgian normal eyes, 34 (+/- 17) in Belgian OAG patients, and 10 (+/- 6) in Congolese OAG patients. The trabecular meshwork was collapsed and Schlemm's canal was closed in the cases with a diminished number of cells. Pigmentation was present in 79% of the Congolese and in 35% of the Belgian specimens. CONCLUSION: The trabecular meshwork from OAG patients has a lower cellularity than normal, and the effect is much more pronounced in the trabecular meshwork from Congolese patients. This may be a possible explanation for the racial differences in OAG and the more severe forms of glaucoma in black people.

Aged↗
PubMed1998

Effects of ascorbic acid on levels of fibronectin, laminin and collagen type 1 in bovine trabecular meshwork in organ culture.

PURPOSE: Fibronectin, laminin and collagen type I are important extracellular matrix products of trabecular meshwork cells. This study was performed to examine the effects of ascorbic acid, a significant component in the aqueous humor, on the levels of these proteins in trabecular meshwork cells maintained in organ culture. METHODS: The anterior segment of freshly enucleated bovine eyes was perfused in a modified organ culture system. Three cultures were set up simultaneously. One received serum-free medium containing 100 micrograms/ml of ascorbic acid, one received 250 micrograms/ml of ascorbic acid and one served as a control. After 72 h, the tissues were processed for paraffin sections and immunostaining was conducted using an avidin-biotin-peroxidase complex method. Western blot and dot blot assays were performed on tissue extracts. RESULTS: Compared with the controls, the staining for fibronectin and laminin was markedly enhanced in trabecular meshwork tissues treated with both concentrations of ascorbic acid. Increased collagen type I production by trabecular meshwork cells was also demonstrated in the presence of ascorbic acid. Western blot and dot blot results confirmed the immunostaining findings. CONCLUSIONS: Ascorbic acid promotes production of fibronectin, laminin and collagen type I by trabecular meshwork cells. The organ culture results are consistent with those obtained previously from tissue culture studies.

Animals↗
PubMed1997

Hyaluronan in the human trabecular meshwork.

PURPOSE: Hyaluronan (HA) is a high-molecular weight glycosaminoglycan that can affect water and solute fluxes in the extracellular matrix. The distribution of HA in the human trabecular meshwork of nonglaucomatous eyes was examined to help understand the potential role of HA in the regulation of aqueous outflow resistance. METHODS: Histolocalization of HA was established in situ in the trabecular meshwork of human eyes with no known diseases of the anterior segment. A specific biotinylated HA-binding peptide was used as a probe for this study, with enhanced sensitivity of HA detection achieved by modifications of the fixation and staining procedures. RESULTS: Evaluation of HA staining in the aqueous outflow pathway in comparison to that in other ocular structures (e.g., the vitreous) showed pronounced staining in the trabecular meshwork. The staining intensity was similar between various layers of the meshwork. Both the filtering and the anterior nonfiltering portions of the trabecular meshwork showed pronounced HA staining. The staining was localized primarily to the trabecular meshwork endothelial cells. CONCLUSIONS: Pronounced HA staining observed in the various layers of the trabecular meshwork suggests that substantial amount of HA is present in the nonglaucomatous outflow pathway. The staining pattern suggests that HA is associated with the endothelial cells lining the trabecular beams. This finding supports potential roles for this glycosaminoglycan in the regulation of the physiological aqueous outflow resistance or in the maintenance of the outflow channels or both. Histochemical localization of HA in the various layers of the non-glaucomatous meshwork provides a useful basis for future comparative studies of HA distribution and relative amounts in the trabecular meshworks of eyes affected by various types of glaucoma.

Aged↗