Evaluation of trabecular meshwork-specific promoters in vitro and in vivo using scAAV2 vectors expressing C3 transferase

AIM:To evaluate the potential of two trabecular meshwork(TM)-specific promoters,Chitinase 3-like 1(Ch3L1)and matrix gla protein(MGP),for improving specificity and safety in glaucoma gene therapy based on self-complementary AAV2(scAAV2)vector technologies.METHODS:An scAAV2 vector with C3 transferase(C3)as the reporter gene(scAAV2-C3)was selected.The scAAV2-C3 vectors were driven by Ch3L1(scAAV2-Ch3L1-C3),MGP(scAAV2-MGP-C3),enhanced MGP(scAAV2-eMGP-C3)and cytomegalovirus(scAAV2-CMV-C3),respectively.The cultured primary human TM cells were treated with each vector at different multiplicities of infections.Changes in cell morphology were observed by phase contrast microscopy.Actin stress fibers and Rho GTPases/Rho-associated protein kinase pathway-related molecules were assessed by immunofluorescence staining,real-time quantitative polymerase chain reaction and Western blot.Each vector was injected intracamerally into the one eye of each rat at low and high doses respectively.In vivo green fluorescence was visualized by a Micron III Retinal Imaging Microscope.Intraocular pressure(IOP)was monitored using a rebound tonometer.Ocular responses were evaluated by slit-lamp microscopy.Ocular histopathology analysis was examined by hematoxylin and eosin staining.RESULTS:In TM cell culture studies,the vectormediated C3 expression induced morphologic changes,disruption of actin cytoskeleton and reduction of fibronectin expression in TM cells by inhibiting the Rho GTPases/Rhoassociated protein kinase signaling pathway.At the same dose,these changes were significant in TM cells treated with scAAV2-CMV-C3 or scAAV2-Ch3L1-C3,but not in cells treated with scAAV2-eMGP-C3 or scAAV2-MGP-C3.At lowinjected dose,the IOP was significantly decreased in the scAAV2-Ch3L1-C3-injected eyes but not in scAAV2-MGPC3-injected and scAAV2-eMGP-C3-injected eyes.At highinjected dose,significant IOP reduction was observed in the scAAV2-eMGP-C3-injected eyes but not in scAAV2-MGP-C3-injected eyes.Similar to scAAV2-CMV-C3,scAAV2-Ch3L1-C3 vector showed efficient transduction both in the TM and corneal endothelium.In anterior segment tissues of scAAV2-eMGP-C3-injected eyes,no obvious morphological changes were found except for the TM.Inflammation was absent.CONCLUSION:In scAAV2-transduced TM cells,the promoter-driven efficiency of Ch3L1 is close to that of cytomegalovirus,but obviously higher than that of MGP.In the anterior chamber of rat eye,the transgene expression pattern of scAAV2 vector is presumably affected by MGP promoter,but not by Ch3L1 promoter.These findings would provide a useful reference for improvement of specificity and safety in glaucoma gene therapy using scAAV2 vector.

基于MeshWorks和Optimus的白车身断面尺寸分析优化

为提高商用车车身前期设计阶段的开发效率,保证项目开发节点按时通过,在车身设计阶段引入参数化建模和联合仿真优化技术。基于MeshWorks软件对梁结构的整体宽度、翻边宽度、整体高度和板件厚度进行参数化建模,再通过Optimus软件搭建基于模态分析与刚度分析的优化流程,进行白车身轻量化设计。优化方案可减重14.6 kg,减重幅度5.1%,结构第1阶扭转频率提升4.4%,弯曲刚度提升2.8%,前端扭转刚度提升0.9%,后端扭转刚度提升1.5%。在性能小幅提升的情况下,车身轻量化效果明显。

Apoptosis in the iris and trabecular meshwork of medically treated and untreated primary open angle glaucoma patients

AIM:To compare the trabecular meshwork(TM)and iris apoptosis of treated and untreated primary open angle glaucoma(POAG)patients.METHODS:Eight treatment-naive,newly diagnosed(group 1)and 11 medlcaiy treated(group 2)patients with POAG were included in the study.Each patient underwent a limbus-based trabeculectomy.The TM and peripheral iris specimens were dissected out and were snap-frozen in liquid nitrogen and stored at-80t until they were assayed.Apoptosis in each group was assesed by TUNEL method.RESULTS:The mean patient age was 60.6±5.8 years(53-68 years)vs 58.9±8.9 years(47-70 years)in group 1and group 2(P=0.859).The mean treatment time in group 2 was 22.2±7.3 months(12-34 months).Apoptotic indexes in TM and iris were significantly higher in POAG patients using medication(group 2)compared to treatment-naive POAG patients(group 1)(P=0.004,0.015;respectively).CONCLUSION:Long term administration of topical antiglaucoma medications causes additional toxic effects on TM.

Fluid-Structure Interaction Simulation of Aqueous Outflow System in Response to Juxtacanalicular Meshwork Permeability Changes with a Two-Way Coupled Method

Elevated intraocular pressure appears to have a broader impact on increased resistance to aqueous humor outflow through the conventional aqueous outflow system(AOS).However,there is still no consensus about exact location of the increased outflow resistance of aqueous humor,and the mechanism is not perfect.In addition,it is difficult to accurately obtain hydrodynamic parameters of aqueous humor within the trabecular meshwork outflow pathways based on the current technology.In this paper,a two-way fluid-structure interaction simulation was performed to study the pressure difference and velocity in the superficial trabecular meshwork,juxtacanalicular meshwork(JCM)and Schlemm’s canal in response to JCM permeability changes.We obtained the JCM permeability of normal intraocular pressure varied between 1×10?15 m2 and 10×10?15 m2 while permeability of the JCM ranged from 2×10?16 m2 and 3×10?16 m2 under conditions of high intraocular pressure.The study indicated that the fluid dynamics parameters in trabecular meshwork and Schlemm’s canal are most significantly affected by the changes of JCM permeability.Moreover,the study demonstrates that the finite element modeling of AOS provides a practical means for studying the outflow dynamics and the biomechanical environment of the AOS.

Inhibition of latrunculin-A on dexamethasone-induced fibronectin production in cultured human trabecular meshwork cells

AIM: To determine the effects of a low dose latrunculin (LAT)-A on dexamethasone (Dex)-induced upregulation of extracellular matrix proteins fibronectin (FN) in cultured human trabecular meshwork (HTM) cells.METHODS: HTM cells were cultured to confluent and incubated with 0.4μmol/L Dex and/or 0.05μmol/L LAT-A.FN expression in HTM cells was evaluated by Western blot and immunofluorescence microscopy.RESULTS: Dex up-regulated FN production in HTM cells,failed to do so when co-incubated with LAT-A.LAT-A decreased production of FN in cultured HTM cells.CONCLUSION: This study indicated that LAT-A may modulate the expression of fibronectin in trabecular meshwork to achieve treatment for steroids and other types of glaucoma.It has an important prospect as an intraocular pressurelowering drug.

Insulin-like Growth Factor-Ⅰ Gene Cloning and Protein Expression in Bovine Trabecular Meshwork Tissue and Cells

Whether cultured bovine trabecular meshwork cells and trabecular tissue ex vivo express insulin like growth factor I (IGF Ⅰ) messenger RNA (mRNA) and protein was investigated. Total RNA of cultured bovine trabecular meshwork cells as well as trabecular meshwork tissue freshly excised from bovine eyes was extracted, and reverse transcriptase polymerase chain reaction (RT PCR) was used to detect IGF Ⅰ mRNA. RT PCR product was verified by sequencing. Immunohistochemical stain was used to detect IGF Ⅰ protein. The results showed that a single PCR amplified product was obtained, and the sequence was homologous to the known sequence.. IGF Ⅰ immunostain was positive in the cytoplasm of trabecular meshwork cells. It was concluded that trabecular meshwork cells produce IGF Ⅰ and contribute to the presence of IGF Ⅰ in trabecular meshwork microenvironment as well as aqueous humor. Trabecular meshwork cells were affected by IGF Ⅰ not only through paracrine, but also autocrine action. Whether abnormal down regulations in IGF Ⅰ production may contribute to the pathogenesis of primary open angle glaucoma and the possibility of promoting the autocrine action of IGF Ⅰ by trabecular meshwork cells to treat the diesease is worth further investigation.

Gene transfer to human trabecular meshwork cells in vitro and ex vivo using HIV-based lentivirus

AIM: To investigate whether the enhanced green fluorescent protein(EGFP) reporter gene could be transferred into human trabecular meshwork(HTM) cells by a HIV-based lentivirus both in vitro and ex vivo.METHODS: The HIV-based lentivirus that contains an EF1-α promoter driving EGFP expression cassette was constructed following the standard molecular cloning methods. The cultured HTM cells were transduced at a range of multiplicity of infection(MOI) with HIV-based lentivirus. EGFP positive cell populations were detected by flow cytometry. Human anterior eye segments were cultured with perfusion culture system and transfected by HIV-based lentivirus with a 1 ×108transducing unit(TU) virus in perfusion liquid. The intraocular pressure was recorded every 8h for 21 d. The expression of EGFP in the anterior segment of the human eye was detected by fluorescence microscopy. Furthermore, the distribution of EGFP expression was confirmed by anti-EGFP immunohistochemical staining.RESULTS: The HIV-based lentivirus which contains an EF1-α promoter driving EGFP expression cassette was constructed successfully. After HTM cells were transduced with HIV-based lentivirus containing EGFP in vitro, the ratio of EGFP positive cells to the total cell number reached 92.3%, with the MOI of 15. After the lentivirus containing EGFP were used to transduce human anterior eye segments, the EGFP could be directly detected by fluorescence microscopy in vivo.Immunohistochemistry staining revealed that 88.19%EGFP-positive trabecular meshwork(TM) cells were observed in the human anterior segment. Nevertheless,the intraocular pressure in the lentivirus-transduced group kept constant when compared with control group(P >0.05).CONCLUSION: EGFP gene could be efficiently transferred into HTM cells both in vitro and ex vivo by using HIV-based lentivirus.

Changes in Schlemm’s canal,trabecular meshwork,and relevant parameters in the early stage after SMILE of myopia patients

AIM:To observe the changes in Schlemm’s canal(SC),trabecular meshwork(TM),and anterior chamber relevant parameters after small incision lenticule extraction(SMILE) of myopia patients.METHODS:A total of 58 eyes from 30 patients who underwent SMILE were divided into a low and moderate myopia group(group A,32 eyes) and a high myopia group(group B,26 eyes).The diameter and area of the SC,the width and thickness of TM obtained by CIRRUS HD-OCT5000,and the related anterior chamber parameters obtained by Pentacam anterior segment analysis system,accommodation amplitude(AMP) were observed pre-and postoperatively.The preoperative intraocular pressure(IOP) and postoperative correction of intraocular pressure(IOPcc) were measured.RESULTS:The diameter and area of the SC in the two groups were significantly increased postoperatively(all P<0.01).The TM width of the patients in the two groups were increased at 1mo after surgery(both P<0.01),but the TM thickness did not change(P>0.05).The corneal curvature,central anterior chamber depth,and anterior chamber volume decreased after SMILE surgery(all P<0.01).There was a weak negative correlation between the SC area change and AMP change in group A(r=-0.362,P<0.01).The postoperative IOP decreased after correction by Shah formula(P<0.05).CONCLUSION:SC and TM in myopia patients change in the early postoperative stage of SMILE and the IOP is decline.

Effects of Nitric Oxide on Proliferation and Apoptosis of Cultured Bovine Trabecular Meshwork Cell

The effects of different doses of nitric oxide (NO) on the proliferation and apoptosis of the cultured bovine trabecular meshwork (TM) cells were studied. L arginine and N G nitro L arginine methyl (L NAME) were incubated with TM cells for 48 h. In the control group, no medicine was given. In the experimental groups, concentrations of L arginine and L NAME were 1×10 -7 mol/L, 1×10 -6 mol/L, 1×10 -5 mol/L, 1×10 -4 mol/L, 1×10 -3 mol/L and 1×10 -2 mol/L, respectively. NO 2 - in supernate, the proliferation and apoptosis of TM cells and mRNA expression of bcl 2 and bax were measured by Griess reagent, terminal deoxynucleotidyl transferase mediated dUTP nick end labeling (TUNEL), MTT assay and in situ hybridization,respectively. The results showed that L arginine with concentration ≥1×10 -4 mol/L could induce apoptosis of the TM cells and inhibit the proliferation of TM cells through increasing the NO levels, down regulating bcl 2 mRNA expression and up regulating bax mRNA expression; L NAME with concentration ≥1×10 -5 mol/L could induce the proliferation of the TM cells through suppressing the production of NO. It was concluded that NO in high level could induce apoptosis of the TM cells and suppress the proliferation of the TM cells.

Effects of Hydrostatic Pressure on Cultured Bovine Trabecular Meshwork Cells

In order to explore the effects of pressure on trabecular meshwork cells, bovine trabecular meshwork cells were cultured in vitro and subjected to different levels of hydrostatic pressure. The cellular morphology, ultrastructure and phagocytosis were studied with inverted phase-contrast microscopy, light microscopy and transmission electron microscopy, etc. It was found that the cells subjected to 2. 0 kPa or 2. 67 kPa for 48 h had no remarkable difference as compared with the controls in terms of parameters observed. Those under 4. 0 kPa for 24 h showed slight changes in structure and a mild decrease in phagocytic function. The damage appeared more severe if the pressure was higher or lasted longer. From the above we conclude that trabecular meshwork cells can only bear pressure below a certain level. They may be destroyed structurally or impaired functionally by pressure over this level.

Effects of dexamethasone and HA1077 on actin cytoskeleton and β-catenin in cultured human trabecular meshwork cells

AIM:To investigate the effects of dexamethasone(DEX) and 1-(5-isoquinolinesulfonyl)-homopiperazine(HA1077) on actin cytoskeleton and β-catenin in cultured human trabecular meshwork(HTM) cells.METHODS: The HTM cells were separated from human eyeball and cultured in vitro.They were divided into control group,DEX(1×10^(-6)mol/L) group,HA1077(3×10^(-5)mol/L)group,and DEX(1×10^(-6)mol/L) and HA1077(3×10^(-5)mol/L)group.Actin cytoskeleton and β-catenin in HTM cells of the four groups were examined by immunofluorescence and Western blot analyses.RESULTS: In DEX group,there were reorganization of actin cytoskeleton and formation of cross linked actin networks(CLANs),which were partially reversed in DEX and HA1077 group.DEX treatment also induced an increased expression of β-catenin,which was obviously reduced in DEX and HA1077 group.Meanwhile,the cultured HTM cells in HA1077 group had lower expression of β-catenin than that in the control group. CONCLUSION: Our results show that HA1077 can reverse the changes of actin organization and expression of β-catenin induced by DEX in cultured HTM cells,suggesting that HA1077 may play an important role in increasing outflow and reducing intraocular pressure.

A Study of Toxicity of 5-Fluorouracil on Bovine Trabecular Meshwork Cells in Vitro

In order to explore whether the conventional use of 5 fluorouracil (5 Fu) had any toxic effects on trabecular meshwork cells, bovine trabecular meshwork cells were cultured in vitro and exposed to 5 Fu at different concentrations. The cellular morphology, ultrastructure, mortality and phagocytosis were studied under light microscopy, transmission electron microscopy and methods of Wright’s stain. It was found that the toxic effects of 5 Fu on the cells were in a dose dependent mode. 1×10 -1 mg/ml of 5 Fu caused a large part of cells rounded up, while 1×10 -3 mg/ml of the drug only a rough appearance of the cell surface. Exposure to 1×10 -2 mg/ml of 5 Fu made mitochrone swollen and rough endoplasmic reticulum enlarged, with the cell mortality being 50.5 %. The latex microspheres engulfed in cytoplasm in cells receiving 1×10 -1 and 1×10 -2 mg/ml of 5 Fu were significantly decreased as compared with those in the control group ( P <0.01). It was concluded that the safe concentration of 5 Fu on bovine trabecular meshwork cells was 1×10 -3 mg/ml and the conventional dosage of 5 Fu in clinical practice would not cause injury to trabecular meshwork cells.

Effect of Transforming Growth Factor-β_2 on Phagocytosis in Cultured Bovine Trabecular Meshwork Cells

The effect of transforming growth factor β 2 (TGF β 2) on phagocytosis in bovine trabecular meshwork cells in vitro was investigated. After the cultured bovine trabecular meshwork cells were treated with 0 ng/ml, 0.32 ng/ml, 1 ng/ml, 3.2 ng/ml TGF β 2 for 24 h, latex beads were added into the incubation medium, and the numbers of the latex beads in 20 adjacent cells were counted under a microscope 24 h later, after treatment with Wright’s stain. Our results showed that the average numbers of the latex beads in the trabecular meshwork cells treated with TGF β 2 of different concentrations were 53.1±1.7 beads/cell, 56.4±2.9 beads/cell and 77.9±6.5 beads/cell respectinvely, in comparison with 45.5±3.3 beads/cell of the control group. TGF β 2 significantly increased the number of the latex beads phagocytosed by cultured bovine trabecular meshwork cells in a dose dependent manner. TGF β 2 could promote the phagocytosis of bovine trabecular meshwork cells in vitro . It may be involved in the cellularity decrease of the trabecular meshwork in the patients of primary open angle glaucoma through promoting the phagocytosis of trabecular meshwork cells.

Expression of Mesenger RNA for Transforming Growth Factor-β_1 in Bovine Trabecular Meshwork

Purpose: To investigate the relationship between transforming growth factor-β1(TGF-β1) and primary open-angle glaucoma, we have determined whether trabec-ular tissues have the expression of messenger RNA for TGF-β1.Methods: Total RNA of 24 newborn bovine trabecular tissue were extracted byGuanidine isothiocyanate method. The TGF-β33 plasmid was brought into E. col-ibacillius HB101 and amplificated. After Bam HI endolase degradation and labelwith a-32p-dATP the RNA was hybridized with the cDNA (complementary DNA)probe and examined by autoradiography.Results: The presence of mRNA for TGF-β1 in bovine trabecular meshwork wasconfirmed.Conclusions: The TGF-β1 present in normal aqueous humor must be at least partlyderived from the trabecular meshwork. It offered a basis for understanding therelationship between abnormal synthesis, activation and clearance of TGF-β1 andthe pathogenesis of primary open-angle glaucoma (POAG) in molecular biology.Eye Science 1996; 12:1-4.

Extracellular matrix gene expression in human trabecular meshwork cells following mechanical fluid flow stimulation

AIM: To investigate changes in extracellular matrix(ECM) gene expression in human trabecular meshwork(HTM) cells in response to mechanical fluid flow stimulation.METHODS: HTM cells were grown on a glass plate coated with 0.02% type Ⅰ collagen(COL) and exposed to shear stress(0, 0.2, 1.0 dyne/cm;) for 12 h.Changes in genes related to the ECM were evaluated by real-time reverse transcriptase-polymerase chain reaction.Phosphorylation of Smad2 protein was investigated by Western blotting.RESULTS: After mechanical stimulation, COL type 4 alpha 2, COL type 6 alpha 1, and fibronectin-1 mRNA were significantly higher than the static control(P<0.05, <0.05, and <0.01, respectively).The metalloproteinase-2 and plasminogen activator inhibitor-1 mRNA were significantly higher than the static control(P<0.05 and <0.01, respectively), while the differences in the tissue inhibitors of metalloproteinases-2 mRNA were not significant.The phosphorylation of Smad2 levels was significantly higher compared to the static control cells.CONCLUSION: Changes in the expressions of genes associated ECM metabolism result in HTM cells after mechanical stimulation.The mechanical stimulation of the aqueous humor to the trabecular meshwork may promote ECM turnover and contribute to intraocular pressure homeostasis.

Glaucoma secondary to trabecular meshwork precipitates:a case report of Grant’s syndrome

Dear Editor,We write to report a case of glaucoma secondary to trabecular meshwork(TM)precipitates.Secondary glaucoma is a common complication of uveitis,which usually presented as elevated intraocular pressure(IOP)with inflammatory signs,such as ciliary congestion,anterior chamber glare and cells and so on.

Expression profile analysis to identify potential gene changes induced by dexamethasone in the trabecular meshwork

AIM:To investigate potential gene changes in trabecular meshwork(TM)induced by dexamethasone(DEX)in steroidinduced glaucoma(SIG).METHODS:The expression data of 24 cases from a public functional genomics data were sorted to identify the mechanisms of action of DEX on the TM.The relationships of the differentially expressed genes(DEGs)were enriched using Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)analysis.In addition,the hub genes were screened by the Search Tool for the Retrieval of Interacting Genes Database(STRING)and Cytoscape tools.Finally,human TM cells(HTMCs)were treated with DEX to preliminarily explore the function of hub genes.RESULTS:Totally 47 DEGs,including 21 downregulated and 26 upregulated genes were identified.The primary enriched results of the DEGs consisted of inflammatory response,extracellular matrix(ECM),negative regulation of cell proliferation,TNF signalling pathway and the regulation of tr yptophan channels by inflammator y mediators.Subsequently,pro-melanin-enriched hormone(PMCH)and Bradykinin B1 receptor(BDKRB1)were screened as hub genes.It is verified in GSE37474 data set.Western blot and quantitative real-time polymerase chain reaction(q PCR)results showed that protein and RNA expression levels of BDKRB1 were significantly decreased after DEX treatment,while PMCH was not significantly changed.CONCLUSION:BDKRB1 may be a key gene involved in SIG onset,providing a suitable therapeutic target for improving the prognosis of SIG patients.

Effect of Dexamethasone and Aquaporin-1 Antisense Oligonucleotides on the Aquaporin-1 Expression in Cultured Human Trabecular Meshwork Cells

在 aquaporin-1 (AQP1 ) 的表示的变化在房间与 dexamethasone 和 transfected withanti 感觉 oligonucleotides (AS-ODN ) 对待的有教养的人的 trabecular raeshwork (HTM ) 的 mRNA 和蛋白质被学习，并且 AQP1 规定 incortico-steroid-glaucoma 的含意和禁止 AQP1 表示的 AS-ODN 的可能性被评估。在 vitro 的有教养的 HTM 房间分别地与 oligonucleotides 与不同集中 ofdexamethasone 和 transfected 被对待 5 天。然后， HTM 房间的全部的 RNA 和蛋白质被提取。AQP1 mRNA 和蛋白质的变化由 RT-PCR 和西方的污点是表明的品质上和份量上。乐队紧张被成像分析检测。在西方的污点的 RT-PCR 和那些的结果之间有一种平行关系。在对待 dexamethasone 的组的 AQP1 mRNA 和蛋白质的表示层次我们 reincreased 开始并且当 dexamethasone 集中被促进，以后减少了。在 0。04 μ g /mL 并且 0。4 个μ g /mL 组， AQP1 的层次比在控制高组(0 μ g /mL ) 。Inthe 4 μ g / mL 和 40 个μ g /mL 组， AQP1 表达式层次比在控制 group.AS-ODN 低能下面调整以一种剂量依赖者方式的 AQP1 mRNA 和蛋白质的表达式。在 5 μ g /mL，下面规定效率到达了最大值。在在所有感觉 oligonucleotides 组和控制组之间的 AQP1 mRNA 和蛋白质的表示没有统计上重要的差别。dexamethasone 可以导致涉及 corticosteroid 绿内障的出现的 AQP1 表示 inHTM 房间的变化，这被建议。在到某程度的 HTM 房间的 AS-ODN 罐头下面调整 theAQP1 表示。

Effect of persistent high intraocular pressure on microstructure and hydraulic permeability of trabecular meshwork

As the aqueous humor leaves the eye, it first passes through the trabecular meshwork(TM). Increased flow resistance in this region causes elevation of intraocular pressure(IOP), which leads to the occurrence of glaucoma. To quantitatively evaluate the effect of high IOP on the configuration and hydraulic permeability of the TM, second harmonic generation(SHG) microscopy was used to image the microstructures of the TM and adjacent tissues in control(normal) and high IOP conditions. Enucleated rabbit eyes were perfused at a pressure of 60 mm Hg to achieve the high IOP. Through the anterior chamber of the eye, in situ images were obtained from different depths beneath the surface of the TM. Porosity and specific surface area of the TM in control and high IOP conditions were then calculated to estimate the effect of the high pressure on the permeability of tissue in different depths. We further photographed the histological sections of the TM and compared the in situ images. The following results were obtained in the control condition, where the region of depth was less than55 μm with crossed branching beams and large pores in the superficial TM. The deeper meshwork is a silk-like tissue with abundant fluorescence separating the small size of pores. The total thickness of pathway tissues composed of TM and juxtacanalicular(JCT) is more than 100 μm. After putting a high pressure on the inner wall of the eye, the TM region progressively collapses and decreases to be less than 40 μm. Fibers of the TM became dense, and the porosity at 34 μm in the high IOP condition is comparable to that at 105 μm in the control condition. As a consequent result, the permeability of the superficial TM decreases rapidly from 120 μm2to 49.6 μm2and that of deeper TM decreases from 1.66 μm2to0.57 μm2. Heterogeneity reflected by descent in permeability reduces from 12.4 μm of the control condition to 3.74 μm of the high IOP condition. The persistently high IOP makes the TM region collapse from its normal state, in which the collagen fibers of the TM are arranged in regular to maintain the physiological permeability of the outflow pathway. In the scope of pathologically high IOP, the microstructure of the TM is sensitive to pressure and hydraulic permeability can be significantly affected by IOP.

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.