Pathogenesis Of Primary Open-angle Glaucoma Research Articles

Primary Open Angle Glaucoma and Vascular Risk Factors: a review of population based studies from 1990 to 2017

AbstractPurposeGlaucoma is one of the leading causes of blindness worldwide, and as the proportion of those over age 40 increases, so will the prevalence of glaucoma. The pathogenesis of primary open angle glaucoma (POAG) is unclear and multiple ocular risk factors have been proposed, including intraocular pressure, ocular perfusion pressure, ocular blood flow, myopia, central corneal thickness, and disc hemorrhages. The purpose of this review was to analyse the correlation between systemic vascular risk factors and POAG pathogenesis based on major epidemiological studies.MethodologyWe used the PubMed database to identify studies including the term glaucoma in association with the following keywords: vascular risk factors, systemic risk factors, blood pressure, systemic hypertension, systemic hypotension, diabetes, dipping, vasospasm, age and migraine.ResultsThe search identified 270 unique articles. After including only relevant English‐language articles only 66 studies were reported in the analysis. Reports presenting the association between POAG and systemic vascular risk factors included a total of over 50,000 patients. Several epidemiological studies confirmed the importance of vascular risk factors in the pathogenesis and progression of glaucomatous optic neuropathy, including hypertension, blood pressure dipping. The pooled analysis confirmed hypertension as the most significant risk factor for primary open angle glaucoma.ConclusionsVascular risk factors play a significant role in the pathogenesis and progression of glaucoma. The association between POAG and diabetes mellitus type 2 or migraine still needs to be evaluated in future studies.

Microbial dysbiosis and microbiota-gut-retina axis: The lesson from brain neurodegenerative diseases to primary open-angle glaucoma pathogenesis of autoimmunity.

In recent years, microbiota-associated neurodegenerative diseases have been exploited and provided new insight into disease pathogenesis. However, primary open-angle glaucoma (POAG), known as a complex neurodegenerative disease resulting from retinal ganglion cell death and optic nerve damage, can cause irreversible blindness and visual field loss. POAG, which shares several similarities with Parkinson's disease (PD) and Alzheimer's disease (AD), has limited studies and slow progression in the understanding of pathogenesis when compared to PD and AD. In this review, we summarized the current knowledge of POAG and commensal microbiota, combined with several lines of evidence in PD and AD to propose a possible hypothesis for POAG pathogenesis: microorganisms cause glaucoma via gut-retina axis, resulting in autoantibodies and autoreactive T cells that lead to autoimmunity. Furthermore, dual-hit hypothesis, an example of a commensal pathogen that causes PD, was partially exported in POAG. Finally, future perspectives are suggested to expand understanding of POAG.

Thrombospondin-2 is up-regulated by TGFβ2 and increases fibronectin expression in human trabecular meshwork cells

Elevated intraocular pressure (IOP) is a major risk factor for the development of primary open-angle glaucoma (POAG). This is from an increased aqueous humour (AH) outflow resistance through the trabecular meshwork (TM). The pathogenic mechanisms leading to the increase in TM outflow resistance are poorly understood but are thought to be from a dysregulation of the TM extracellular matrix (ECM) environment. ECM modification and turnover are crucial in regulating the resistance to aqueous outflow. ECM turnover is influenced by a complex interplay of growth factors such as transforming growth factors (TGFβ) family and matrix metalloproteinases (MMPs). Elevated TGFβ2 levels result in an increase in ECM deposition such as fibronectin leading to increased resistance. Fibronectin is a major component of TM ECM and plays a key role in its maintenance. Thrombospondins (TSP)-1 and −2 are important regulators of the ECM environment. TSP-1 has been implicated in the pathogenesis of POAG through activation of TGFβ2 within the TM. TSP-2 does not contain the catalytic domain to activate latent TGFβ, but is able to mediate the activities of MMP 2 and 9, thereby influencing ECM turnover. TSP-2 knock out mice show lower IOP levels compared to their wild type counterparts, suggesting the involvement of TSP-2 in the pathogenesis of POAG but its role in the pathogenesis of POAG remains unclear. The purpose of this study was to investigate the role of TSP-2 in trabecular meshwork ECM regulation and hence the pathogenesis of POAG.TSP-1 and TSP-2 expressions in immortalised glaucomatous TM cells (GTM3) and primary human non-glaucomatous (NTM) and glaucomatous cells (GTM) were determined by immunocytochemistry, immuno-blot analysis and qPCR following treatment with TGFβ2 and Dexamethasone. The level of ECM protein fibronectin was determined in TM cells using immuno-blot analysis following treatment with TSP-1 or -2.TM cells secrete TSP-1 and -2 under basal conditions at the protein level and TSP-2 mRNA and protein levels were increased in response to TGFβ2 three days post treatment. Exogenous treatment with TSP-2 up-regulated the expression of fibronectin protein in GTM3 cells, primary NTM and GTM cells. TSP-1 did not affect fibronectin protein levels in GTM3 cells. This suggests that the role of TSP-2 might be distinct from that of TSP-1 in the regulation of the TM cell ECM environment. TSP-2 may be involved in the pathogenesis of POAG and contribute to increased IOP levels by increasing the deposition of fibronectin within the ECM in response to TGFβ2.

Activation of αvβ3 Integrin Alters Fibronectin Fibril Formation in Human Trabecular Meshwork Cells in a ROCK-Independent Manner.

PurposeFibronectin fibrillogenesis is an integrin-mediated process that may contribute to the pathogenesis of primary open-angle glaucoma (POAG). Here, we examined the effects of αvβ3 integrins on fibrillogenesis in immortalized TM-1 cells and human trabecular meshwork (HTM) cells.MethodsTM-1 cells overexpressing wild-type β3 (WTβ3) or constitutively active β3 (CAβ3) integrin subunits were generated. Control cells were transduced with an empty vector (EV). Deoxycholic acid (DOC) extraction of monolayers, immunofluorescence microscopy, and On-cell western analyses were used to determine levels of fibronectin fibrillogenesis and fibronectin fibril composition (EDA+ and EDB+ fibronectins) and conformation. αvβ3 and α5β1 Integrin levels were determined using fluorescence-activated cell sorting (FACS). Cilengitide and an adenovirus vector expressing WTβ3 or CAβ3 integrin subunits were used to examine the role of αvβ3 integrin in HTM cells. The role of the canonical α5β1 integrin–mediated pathway in fibrillogenesis was determined using the fibronectin-binding peptide FUD, the β1 integrin function-blocking antibody 13, and the Rho kinase (ROCK) inhibitor Y27632.ResultsActivation of αvβ3 integrin enhanced the assembly of fibronectin into DOC-insoluble fibrils in both TM-1 and HTM cells. The formation of fibronectin fibrils was dependent on α5β1 integrin and could be inhibited by FUD. However, fibrillogenesis was unaffected by Y27632. Fibrils assembled by CAβ3 cells also contained high levels of EDA+ and EDB+ fibronectin and fibronectin that was stretched.Conclusionsαvβ3 Integrin signaling altered the deposition and structure of fibronectin fibrils using a β1 integrin/ROCK-independent mechanism. Thus, αvβ3 integrins could play a significant role in altering the function of fibronectin matrices in POAG.

Особенности содержания трансформирующих факторов роста – бета 1,2,3 (TGF-β1, TGF-β2, TGF-β3) во внутриглазной жидкости при первичной открытоугольной глаукоме

Purpose. To study transforming growth factors – beta 1,2,3 (TGF-β1, TGF-β2, TGF-β3) in intraocular aqueous humor (AH) of patients with the advanced stage of primary open angle glaucoma (POAG). Material and methods. The study included an examination of 56 patients with advanced stage of POAG which comprised the main group. The diagnosis of POAG was confirmed based on ophthalmological examination. The control group consisted of 30 patients with uncomplicated cataract. TGF-β isoforms concentrations were detected by flow fluorimetry using Bio-Plex Pro™ TGF-β 3-plex Assay and double-beam laser analyzer Bio-Plex 200, Bio-Rad, USA. Three isoforms of TGF-β (TGF-β1, TGF-β2, TGF-β3) were simultaneously detected in AH. Results. The study revealed a significant increase of TGF-β isoforms concentrations in AH of the advanced POAG compared to AH of the group with uncomplicated cataract: TGF-β1 (3.1 times higher), TGF-β2 (1.4 times) and TGF-β3 (6.9 times). The following direct significant correlations were found: between TGF-β1 and TGF-β2 (r=0.41, p<0.03), between TGF-β1 and TGF-β3 (r=0.83, p<0.004), and between TGF-β2 and TGF-β3 (r=0.35, p<0.04). Conclusion. We determined a simultaneous increase of concentrations of the studied representatives of super-family transforming growth factors, which have an anti-inflammatory activity, potential to stimulate proliferation, cellular growth, synthesis of proteins of extracellular matrix, etc. These results prove their importance in the POAG pathogenesis.

Primary Open Angle Glaucoma: The Pathophysiolgy, Mechanisms, Future Diagnostic and Therapeutic Directions

Primary open angle glaucoma (POAG) is a multifactorial chronic optic neuropathy, characterized by progressive loss of retinal ganglion cells (RGC), leading to structural damage to the optic nerve head (ONH), retinal nerve fiber layer (RNFL), with visual field defects. It is occasioned by major risk factors of high intraocular pressure (IOP) and age. The pathogenesis of POAG is the imbalance between the production and drainage of the aqueous humour (AH). The resultant fluid back-up increases the IOP with consequent optic nerve damage, causing POAG. Modern diagnosis, using scanning laser polarimetry (SLP), confocal scanning laser ophthalmoscopy (CSLO), optical coherence tomography (OCT) etc, plays a vital role in the assessment of the ONH, RNFL and the macular, in POAG. OCT operates on the principles of interferometery, utilizing light beams and their pattern of back-scattering, to build high resolution cross-sectional images of ocular tissues. It gives an objective evaluation of structural alterations in the ONH or macular area of the retina in vivo. POAG is managed medically using classes of drugs like α-adrenergic agonists, beta-adrenergic receptor antagonist, prostaglandin agonists, carbonic anhydrase inhibitors, ccholinergic agonists etc. Surgical intervention is indicated with laser or incisional surgeries, when medical option fails. Future pharmacotherapeutic directions in POAG management consider transgenic model, genetic model, neuroregeneration (stem cell technique), neurodegeneration (Seeing glaucoma as a neurologic disorder much like in Parkinson's and Alzheimer's diseases and the mechanisms that cause the degeneration of RGCs), autoimmune response and T-cell autoimmune response attack.

Comprehensive bioinformatics analysis of trabecular meshwork gene expression data to unravel the molecular pathogenesis of primary open-angle glaucoma.

PurposePerforming bioinformatics analyses using trabecular meshwork (TM) gene expression data in order to further elucidate the molecular pathogenesis of primary open‐angle glaucoma (POAG), and to identify candidate target genes.MethodsA systematic search in Gene Expression Omnibus and ArrayExpress was conducted, and quality control and preprocessing of the data was performed with ArrayAnalysis.org. Molecular pathway overrepresentation analysis was performed with PathVisio using pathway content from three pathway databases: WikiPathways, KEGG and Reactome. In addition, Gene Ontology (GO) analysis was performed on the gene expression data. The significantly changed pathways were clustered into functional categories which were combined into a network of connected genes.ResultsNinety‐two significantly changed pathways were clustered into five functional categories: extracellular matrix (ECM), inflammation, complement activation, senescence and Rho GTPase signalling. ECM included pathways involved in collagen, actin and cell–matrix interactions. Inflammation included pathways entailing NF‐κB and arachidonic acid. The network analysis showed that several genes overlap between the inflammation cluster on the one hand, and the ECM, complement activation and senescence clusters on the other hand. GO analysis, identified additional clusters, related to development and corticosteroids.ConclusionThis study provides an overview of the processes involved in the molecular pathogenesis of POAG in the TM. The results show good face validity and confirm findings from histological, biochemical, genome‐wide association and transcriptomics studies. The identification of known points of action for drugs, such as Rho GTPase, arachidonic acid, NF‐κB, prostaglandins and corticosteroid clusters, supports the value of this approach to identify potential drug targets.

Сontent of cytokines and growth factors in the intraocular fluid of patients with primary open-angle glaucoma

Purpose. To study the content of cytokines and growth factors in the intraocular fluid of patients with developed stage of primary open-angle glaucoma (POAG).Materials and methods. 56 patients with a verified diagnosis developed stage of primary open-angle glaucomawere examined. The control group consisted of 30 patients with a diagnosis of uncomplicated cataract. A concentration of 17 cytokinesand 3 isoforms of the transforming growth factor (TGF) β was determined using a Bio-Plex Pro™ Human Cytokine 17-plex Assay and Bio-Plex Pro™ and TGFβ 3-plex Assay kit by flow-through fluorometry on a Bio-Plex 200, Bio-Rad double beam laser analyzer, USA.Results. Astatistically significant increase was shown in the concentrations of cytokines and growth factors (interleukins (IL) 4, 6, 7, 8, 12, 17, TGFβ 1, 2, 3, macrophage inflammatory protein 1 β) in the intraocular fluid of patients with developed stage of the primary open-angle glaucoma in respect to data obtained from the study of the intraocular fluid of the persons with uncomplicated cataract, as well as a statistically significant decrease in the concentrations IL-2, IL-10, granulocyte-macrophage colony-stimulating factor.Conclusion. In the pathogenesis of primary open-angle glaucoma, the activity of the local chronic inflammatory process is determined. This is confirmed by statistically significant changes in the studied cytokines and growth factors. Increase in the concentrations of the studied representatives of the superfamily of transforming growth factors-beta, which have anti-inflammatory activity, the ability to stimulate proliferation, cell growth, synthesis of extracellular matrix proteins, etc., attests to their importance in the mechanisms of primary open-angle glaucoma development. Increase concentrations of IL-7 in the intraocular fluid of patients with primary open-angle glaucoma allows us to assume participation in the pathogenesis of the primary open-angle glaucoma of this autocrine mediator of activation of the growth of lymphatic structures.

Caveolin-1 regulates human trabecular meshwork cell adhesion, endocytosis, and autophagy.

Impaired trabecular meshwork (TM) outflow is implicated in the pathogenesis of primary open-angle glaucoma (POAG). We previously identified the association of a caveolin-1 (CAV1) variant with POAG by genome-wide association study. Here we report a study of CAV1 knockout (KO) effect on human TM cell properties. We generated human CAV1-KO TM cells by CRISPR/Cas9 technology, and we found that the CAV1-KO TM cells less adhered to the surface coating than the wildtype TM cells by 69.34% ( P < 0.05), but showed no difference in apoptosis. Higher endocytosis ability of dextran and transferrin was also observed in the CAV1-KO TM cells (4.37 and 1.89-fold respectively, P < 0.001), compared to the wildtype TM cells. Moreover, the CAV1-KO TM cells had higher expression of extracellular matrix-degrading enzyme genes ( ADMTS13 and MMP14) as well as autophagy-related genes ( ATG7 and BECN1) and protein (LC3B-II) than the wildtype TM cells. In summary, results from this study showed that the CAV1-KO TM cells have reduced adhesion with higher extracellular matrix-degrading enzyme expression, but increased endocytosis and autophagy activities, indicating that CAV1 could be involved in the regulation of adhesion, endocytosis, and autophagy in human TM cells.

Association between rs4938723 polymorphism and the risk of primary open-angle glaucoma (POAG) in a Chinese population.

In this study, we aimed to investigate the association between rs4938723 single-nucleotide polymorphism (SNP) and the risk of primary open-angle glaucoma (POAG) in a Chinese population to clarify the molecular mechanism underlying the pathogenesis of POAG. Taqman assays and statistical analyses were utilized to analysis the associations between rs4938723 SNP/rs1042522 SNP and the risk of POAG. Luciferase assays, in-silicon analyses, real-time PCR, Western Blot, 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide (MTT) assay, and flow cytometry were conducted to establish a molecular pathway underlying the protective effect of rs4938723 T > C SNP against the risk of POAG. The maximum intraocular pressure (IOP) was evidently higher in the case group compared with that in the control group. The risk of POAG was evidently decreased among carriers of the CT/CC genotypes under a dominant model. In addition, the risk of POAG was also significantly decreased in the carriers of the CC genotype. In contrary, no significant association was found between the risk of POAG and the rs1042522 SNP in TP53 gene. In addition, rs4938723 (especially rs4938723 T > C) SNP could elevate the transcription efficiency of miR-34b promoter and the suppression of TP53, a virtual target of miR-34b. Therefore, the presence of anti-miR-34b suppressed cell proliferation and promoted cell apoptosis, thus establishing a molecular mechanism underlying the protective effect of rs4938723 polymorphism T > C against the risk of POAG. Rs4938723 polymorphism T > C could reduce the incidence of POAG by downregulating TP53 expression, thus leading to suppressed cell apoptosis and enhanced cell proliferation.

Comprehensive measurements of hydroxylinoleate and hydroxyarachidonate isomers in blood samples from primary open-angle glaucoma patients and controls

We previously reported that lower systemic antioxidant capacity is involved in primary open-angle glaucoma (POAG) and exfoliation syndrome pathogeneses as measured by ferric-reducing activity. In the present study, we measured hydroxylinoleate (HODE) and hydroxyarachidonate (HETE) isomer serum levels after sample reduction and saponification to investigate POAG pathogenesis. POAG patients (n = 198) were recruited and divided into normal- and high-tension glaucoma groups (n = 84 and 114, respectively) depending on intraocular pressure. Total HODE (/linoleic acid) and HETE (/arachidonic acid) serum levels were significantly higher in the POAG group (211.9 ± 143.0 and 181.0 ± 164.1 µmol/mol, respectively) than in controls (167.1 ± 105.2 and 132.5 ± 139.7 µmol/mol, p = 0.0025 and 0.0101, respectively). The associations between HODEs/HETEs and glaucoma were further confirmed by multivariate analyses after adjusting for differences in demographic parameters. Among the HODE isomers, the levels of 9- and 13-(Z,E)-HODEs (p = 0.0014) and singlet oxygen-specific products (i.e., 10- and 12-(Z,E)-HODEs, p = 0.0345) were higher in the POAG group than in controls, while free radical-mediated oxidation-specific products (i.e., 9- and 13-(E,E)-HODEs, p = 0.0557) demonstrated a marginal difference. Enzymatic and singlet oxygen-mediated fatty acid oxidation may be major pathways of oxidation process in glaucoma subjects.

Levels of serum trace elements in patients with primary open-angle glaucoma

PurposeGlaucoma disease is known as multifactorial. Trace elements seemed to be linked via oxidative stress mediated changes to the complex glaucoma pathophysiology. Thus, it was the aim of this study to investigate serum levels of trace elements in primary open-angle glaucoma patients (POAG). Patients and methodsPeripheral venous blood samples were taken from a total of 40 subjects: 22 primary open-angle glaucoma patients (mean age 58.1 ± 13.9, female 8, male 14) and 18 controls (mean age 38.9 ± 11.6, 6 female 6, male 12). Serum samples of cadmium, cobalt, copper, iron, lead, manganese and zinc were analyzed by Inductively-Coupled-Plasma-Optical Emission Spectrometry (Cu, Fe, Zn) and Inductively-Coupled-Plasma-sectorfield-Mass-Spectrometry (Cd, Co, Mn, Pb, Se). Data were analyzed using ANCOVA and presented as log transformed LS-mean. ResultsPatients with POAG had significantly increased serum levels of iron (2.98 ± 0.03 μg/L vs 2.98 ± 0.03 μg/L) when compared to controls, and of cadmium (1.57 ± 0.05 ng/L vs. 1.40 ± 0.06 ng/L) considering the interaction between age and the class variable (control versus POAG). A gender effect was seen for cadmium, cobalt, copper, and iron in controls and POAG patients. Iron concentration was reduced in dependency of age for both genders in normals, however lesser in POAG patients. No difference was seen in serum levels of lead, manganese, and zinc between patients with POAG and controls. ConclusionA significant elevation of serum cadmium and iron levels in POAG patients as well as an additional gender effect of cadmium, cobalt, copper, and iron in normals and POAG patients, may argue for a potential role of these trace elements in the pathogenesis of primary open-angle glaucoma.

The association of mitochondrial DNA haplogroups with POAG in African Americans

Mitochondrial dysfunction has been implicated in the pathogenesis of primary open-angle glaucoma (POAG). However, the potential significance of mitochondrial DNA (mtDNA) haplogroups to POAG has not been evaluated in the overaffected African American population. To investigate the association of mtDNA haplogroups with POAG and its phenotypic characteristics, genotyping data from 4081 African American subjects (1919 cases and 2162 controls) was analyzed using 1293 positions on mtDNA. The overall frequency of mtDNA haplogroups in the Primary Open-Angle African American Glaucoma Genetics (POAAGG) study cohort was 37% L3, 29% L2, 21% L1, 4% L0, and 10% non-African haplogroups (non-L). When all haplogroups (L0, L1, L2, and non-L) were compared against theL3 reference group, after adjusting by age and principal component of ancestry, the non-L3 haplogroups showed higher risk of POAG (OR-1.19, p = 0.02), with a particularly strong association among males (OR = 1.41, p = 0.003). More specifically the non-L group was associated with higher POAG risk than the L3 haplogroup (OR = 1.77, p = 0.007, Bonferroni adjusted p = 0.027) and to the L3e (n = 256, OR = 1.92, p = 0.007, Bonferroni adjusted p = 0.029). No significant association was found when genders were analyzed together or in female only analysis. There were no significant differences in various POAG endophenotypes across mtDNA haplogroups. This study expands our knowledge of mitochondrial genetics and mtDNA haplogroup associations in African American POAG. Further work is needed to better understand the functional role of mtDNA polymorphisms and their interactions with nuclear genes that affect POAG.

Roles of Toll-Like Receptor 4 for Cellular Pathogenesis in Primary Open-Angle Glaucoma: A potential therapeutic strategy

In recent years, glaucoma has been proposed as an autoimmune disease and an understanding immune-regulation concept has been applied for novel glaucoma therapy. Current evidence suggests an innate immunity is a keystone step for primary open angle glaucoma (POAG) pathogenesis resulting from trabecular meshwork (TM) cell fibrosis and retinal ganglion cell (RGC) death. Toll-like receptor 4 (TLR4) is a common player in the innate immunity, which appears on the TM and RGC of POAG. The activation of TLR4 regulates several molecules involving both fibrosis and cell death. Inhibition of TLR4 decreases TGF-β2-induced fibrosis in TM cells and enhances cell survival of RGC in both optic nerve crush and ischemia models. In this review, we will summarize the molecular mechanisms of TLR4 related to POAG pathogenesis. An understanding of this mechanism may provide novel development of therapeutic strategies for POAG.

A Metabolomics Profiling of Glaucoma Points to Mitochondrial Dysfunction, Senescence, and Polyamines Deficiency.

To determine the plasma metabolomic signature of primary open-angle glaucoma (POAG). We compared the metabolomic profiles of plasma from individuals with POAG (n = 36) with age- and sex-matched controls with cataract (n = 27). A targeted metabolomics study was performed using the standardized p180 Biocrates Absolute IDQ p180 kit with a QTRAP 5500 mass spectrometer. Multivariate analyses were performed using principal component analysis (PCA) and the least absolute shrinkage and selection operator (LASSO) method. Among the 151 metabolites accurately measured, combined univariate and multivariate analyses revealed 18 discriminant metabolites belonging to the carbohydrate, acyl-carnitine, phosphatidylcholine, amino acids, and polyamine families. The metabolomic signature of POAG points to three closely interdependent pathophysiologic conditions; that is, defective mitochondrial oxidation of energetic substrates, altered metabolism resembling that observed in senescence, and a deficiency in spermidine and spermine, both polyamines being involved in the protection of retinal ganglion cells. Our results highlight a systemic and age-related mitochondrial defect in the pathogenesis of POAG.

Analysis of CYP1B1 sequence alterations in patients with primary open-angle glaucoma of Saudi origin.

Cytochrome P450 Family 1 Subfamily B Member 1 (CYP1B1; OMIM# 601771) gene encodes one of the cytochrome P450 family of enzymes. CYP1B1 mutations have been associated primarily with primary congenital glaucoma (PCG). Similar studies were reported in juvenile open-angle glaucoma, Rieger’s and Peters anomalies. Reports of likely pathogenic sequence alterations in families affected with adult-onset primary open-angle glaucoma (POAG) triggered this investigation. We screened unrelated POAG cases and healthy controls for mutations in CYP1B1 using automated Sanger sequencing to identify five known polymorphisms and one CYP1B1 mutation (p.G61E) in a heterozygous status. The p.G61E mutation is known to cause PCG in a homozygous or compound heterozygous form, and thus, its presence here in a heterozygous form indicates carrier status. These findings suggest that CYP1B1 may have no major role in the pathogenesis of POAG, at least, in the Saudi population. However, further investigations are needed to validate these findings in a larger cohort.

The screening of pathogenic locus in a primary open angle glaucoma pedigree

Objective To screen the pathogenic locus and gene in a primary open angle glaucoma(POAG), and to provide a basis for molecular genetic study of POAG. Methods A POAG pedigree with 35 members was diagnosed in Sichuan peoples' Hospital from January to August 2005.The disease history and clinical data were collected.Genome-wide scan was performed for the families.Specific software was used to calculate the LOD value, which based on the allele (haploid) typing result with two-point method to definite the positive loci by the largest LOD value. Results The POAG family had 35 members of 4 generations.18 patients were diagnosed as juvenile open angle glaucoma from visual disc shape abnormality and loss of typical visual field.All of the patients in this family suffered various degrees of binocular vision loss and vision loss in childhood, with poorly visual function.The LOD values of 3 short tandom repeat (STR) markers on chromosome 2 were greater than 3.0, they were D2S2369 (LOD value 4.003 3), D2S2332 (LOD value 3.840 2) and D2S337 (LOD value 4.752 0). There was a genetic linkage near the three genetic markers in the family.The primary glaucoma positive locus was a in chromosome p15 to chromosome p16.2, and the genetic distance was about 9 Mb, locating in between the markers D2S2369 and D2S2397. Conclusions GLCIH is a pathogenic locus for this POAG pedigree, which supplies an evidence for elucidating the pathogenesis of POAG. Key words: Primary open angle glaucoma; Locus; Genome-wide scan

Interleukin-6–mediated trans-signaling inhibits transforming growth factor-β signaling in trabecular meshwork cells

Glaucoma is one of the major causes of blindness, and transforming growth factor-β2 (TGF-β2) has been found to be elevated in the aqueous humor of eyes with primary open-angle glaucoma (POAG). TGF-β2 in aqueous humor causes the glaucoma-related fibrosis of human trabecular meshwork (HTM), suggesting an important role of TGF-β in POAG pathogenesis. Here, we sought to elucidate the effects of IL-6 trans-signaling on TGF-β signaling in HTM cells. Using a multiplex immunoassay, POAG patients decreased IL-6 levels and increased soluble IL-6 receptor (sIL-6R) levels compared with the controls. In in vitro experiments, we observed that the IL-6 level was increased in the conditioned medium of HTM cells after TGF-β2 stimulation. To elucidate the relationship between TGF-β2 and IL-6 in HTM cells, we conducted Western blotting and immunohistochemical analyses, and we noted that the combination of IL-6 and sIL-6R (IL6/sIL-6R) suppressed TGF-β-induced up-regulation of α-smooth muscle actin in HTM cells, whereas IL-6 alone did not. This suggests that trans-signaling, not classic signaling, of IL-6 suppresses TGF-β-induced fibrosis of HTM. IL6/sIL-6R also suppressed TGF-β-mediated activation of myosin light chain 2 (MLC2), Smad2, and p38. Of note, these inhibitory effects of IL6/sIL-6R on TGF-β were partly reduced by siRNA-mediated knockdown of STAT3. Moreover, IL-6/sIL-6R partly inhibited TGF-β-induced activation of the Smad-sensitive promoter detected with luciferase reporter gene assays and up-regulation of TGFRI and TGFRII, evaluated by quantitative real-time RT-PCR. Strikingly, overexpression of TGFRI and TGFRII diminished these inhibitory effects of IL-6/sIL-6R. We conclude that of IL-6-mediated trans-signaling potently represses TGF-β signaling in HTM cells.

Proteomic Analysis of Aqueous Humor from Primary Open Angle Glaucoma Patients on Drug Treatment Revealed Altered Complement Activation Cascade.

Primary open angle glaucoma (POAG) is a complex disease and a leading cause of irreversible blindness, and its underlying pathophysiology remains poorly understood. Proteomic characterization of the protein composition of aqueous humor (AH) may identify prognostic candidate proteins involved in pathogenesis and progression of the disease. To delineate the possible mechanisms that lead to POAG, this study adopted state-of-art mass spectrometric technique and analyzed AH of POAG and their respective controls. In total, more than 1000 proteins were identified with false discovery rate of less than 1%. Numerous proteins of complement cascade, immunoglobulin, neuronal and amyloidogenic proteins, which were part of processes like acute-phase and inflammatory response, humoral immune and acute inflammatory response, regulation of complement activation and protein processing were identified. Proteins of complement system underwent significant changes, which correlate to pathogenic events characterizing POAG, including altered complement cascade, astrocyte activation, neural degeneration, and apoptosis. Further, protein modification such as deamidation of complement subcomponent was noted, particularly in POAG. Proteomic analysis of AH allows a better understanding of the mechanism involved in the pathogenesis of POAG.

MicroRNA Profiling in Glaucoma Eyes With Varying Degrees of Optic Neuropathy by Using Next-Generation Sequencing.

To explore the microRNA (miRNA) profile and its putative role in glaucomatous optic neuropathy by using next-generation sequencing. Aqueous humor (AH) samples were collected from 19 primary open-angle glaucoma (POAG) eyes and 17 cataract eyes before surgery. Next-generation sequencing was performed for RNA samples extracted from 18 AH samples, and the bioinformatics approach was applied for samples with adequate clean data output. The other 18 samples were used for quantitative PCR validation of sequencing results. In total, 12 (six POAG and six cataract controls) samples with sufficient clean data output after sequencing were used for further data analysis. Four hundred sixty-six and 480 mature miRNAs were detected in the POAG and cataract control groups, respectively. Among them, 164 miRNAs were detected in all POAG samples, and 96 miRNAs were detected in all cataract control samples. Furthermore, 88 miRNAs were identified as differently expressed between POAG and cataract control eyes. In addition, 16 miRNAs were differently expressed between POAG eyes with severe visual field damage and eyes with moderate visual field damage. This differential expression was predicted to regulate thiamine metabolism, purine metabolism, and transcriptional misregulation. Relative expression patterns of hsa-miR-184, hsa-miR-486-5p, and hsa-miR-93-5p were confirmed by quantitative PCR. This study comprehensively demonstrated the miRNA expression profile in the AH of POAG eyes, especially the differential expression of miRNA in eyes with varying degrees of visual field damage, which, together with the underlying miRNA-related pathways, indicate new targets for the pathogenesis and progression of POAG.