Alterations in anterior lens capsule structure and LTBP-2 expression in primary angle-closure glaucoma.

OBJECTIVE: This study investigated the role of latent-transforming growth factor ß-binding protein 2 (LTBP-2) in primary angle-closure glaucoma (PACG) by analysing its expression and the ultrastructure of the anterior lens capsule in PACG patients with age-related cataract (ARC). METHODS: Tissue samples of the anterior lens capsule were collected from patients undergoing cataract phacoemulsification surgery. Patients in the experimental group were diagnosed with primary angle-closure (PAC) combined with ARC (PAC+ARC) and PACG combined with ARC (PACG+ARC). The control group consisted of patients with only ARC. The techniques used included scanning electron microscopy, real-time fluorescence quantitative polymerase chain reaction (RT-qPCR), western blotting and immunofluorescence. RESULTS: Ultrastructural analysis revealed disordered connections in PAC+ARC, loose connections in PACG+ARC and well-ordered connections in ARC. RT-qPCR and western blotting showed significantly lower LTBP-2 mRNA and protein expression in PAC+ARC and PACG+ARC than in ARC, with PAC+ARC having the lowest levels. Immunofluorescence confirmed these findings, showing varying LTBP-2 fluorescence intensities across groups. CONCLUSION: The study identified ultrastructural changes in the anterior lens capsules in PACG accompanied by reduced LTBP-2 expression, especially in PAC+ARC patients. This suggests a potential role for LTBP-2 in PACG development, warranting further investigation.

Untargeted metabolomics in the aqueous humor reveals the involvement of TAAR pathway in glaucoma.

Understanding the metabolic dysfunctions and underlying complex pathological mechanisms of neurodegeneration in glaucoma could help discover disease pathways, identify novel biomarkers, and rationalize newer therapeutics. Therefore, we aimed to investigate the local metabolomic alterations in the aqueous humor and plasma of primary glaucomatous patients. This study cohort comprised primary open-angle glaucoma (POAG), primary angle-closure glaucoma (PACG), and cataract control groups. Aqueous humor and plasma samples were collected from patients undergoing trabeculectomy or cataract surgery and subjected to high-resolution mass spectrometry (HRMS) analysis. Spectral information was processed, and the acquired data were subjected to uni-variate as well as multi-variate statistical analyses using MetaboAnalyst ver5.0. To further understand the localized metabolic abnormalities in glaucoma, metabolites affected in aqueous humor were distinguished from metabolites altered in plasma in this study. Nine and twelve metabolites were found to be significantly altered (p < 0.05, variable importance of projection >1 and log2 fold change ≥0.58/≤ -0.58) in the aqueous humor of PACG and POAG patients, respectively. The galactose and amino acid metabolic pathways were locally affected in the PACG and POAG groups, respectively. Based on the observation of the previous findings, gene expression profiles of trace amine-associated receptor-1 (TAAR-1) were studied in rat ocular tissues. The pharmacodynamics of TAAR-1 were explored in rabbits using topical administration of its agonist, ß-phenyl-ethylamine (ß-PEA). TAAR-1 was expressed in the rat's iris-ciliary body, optic nerve, lens, and cornea. ß-PEA elicited a mydriatic response in rabbit eyes, without altering intraocular pressure. Targeted analysis of ß-PEA levels in the aqueous humor of POAG patients showed an insignificant elevation. This study provides new insights regarding alterations in both localized and systemic metabolites in primary glaucomatous patients. This study also demonstrated the propensity of ß-PEA to cause an adrenergic response through the TAAR-1 pathway.

Aqueous Humor Cytokine Response in the Contralateral Eye after First-Eye Cataract Surgery in Patients with Primary Angle-Closure Glaucoma, High Myopia or Type 2 Diabetes Mellitus.

BACKGROUND: Bilateral sequential cataract surgery within a short period is becoming more prevalent because of the efficiency and safety of modern cataract surgery. It has been reported that the first surgical eye might affect the contralateral eye. This study investigated the cytokines involved in the immunopathogenesis of pre-existing ocular or systemic conditions, as well as the inflammatory biomarkers in response to topical stimuli, by analyzing the cytokine profile of aqueous humor (AH) from cataract patients without these morbidities as control and with type 2 diabetes mellitus (DM), primary angle-closure glaucoma (PACG) or high myopia (HM) in each eye at the beginning of first (defined as baseline) and second eye cataract surgery. METHODS: Forty patients were recruited in this cohort study (10/group). Bilateral sequential cataract surgeries were conducted at intervals of 12.08 ± 1.2 days. Aqueous humor samples (100-200 µL/eye) were separately collected from 40 first-eyes and 40 second-eyes at the beginning of the cataract surgeries. Twenty-seven selected cytokines were detected with Luminex-multiplex immunoassay. The concentrations of cytokines in the aqueous humor and their association with pre-existing ocular or systemic conditions were analyzed and compared between and within the groups. RESULTS: Before first-eye surgery (baseline), the levels of interleukin (IL)-1ra, IL-13 and tumor necrosis factor (TNF)-alpha were significantly increased in PACG compared with controls. The levels of IL-13 were increased while that of IL-15 were decreased in HM. Compared with controls, 11 cytokines were significantly increased in DM. In the AH of the contralateral eye after first-eye cataract surgery, basic fibroblast growth factor (bFGF) was significantly more abundant in PACG and HM, while the levels of monocyte chemoattractant protein-1 (MCP-1) and interferon gamma-induced protein 10 (IP-10) were decreased in PACG. We also identified 6 significantly upregulated cytokines in DM compared with controls. Compared with baseline, there was an overlap of 5 altered cytokines in the AH of contralateral eyes after first-eye surgery between the four groups. Some were exclusively altered in each subgroup, with 1 in the control group, 4 cytokines in the PACG and HM groups, and none in the DM group. CONCLUSIONS: From the initial profile, it is observed that patients with pre-existing ocular or systemic conditions have some degree of inflammation in their eyes before surgery and in the contralateral eye after the first eye cataract surgery, which could be peculiar of the morbid conditions of the patients. Inflammation was more detectable in patients with type 2 DM before surgery. PACG and HM patients showed stronger intraocular inflammatory reactions to topical stimuli compared with controls and DM patients. Our data suggest that ophthalmologists should pay closer attention to inflammatory responses, especially in cataract patients with pre-existing conditions, although the clinical significance of these changes following surgery remains to be further investigated.

Primary angle closure glaucoma is characterized by altered extracellular matrix homeostasis in the iris.

PURPOSE: To characterize the proteome of the iris in primary angle closure glaucoma (PACG). EXPERIMENTAL DESIGN: In this cross-sectional study, iris samples were obtained from surgical iridectomy of 48 adults with PACG and five normal controls. Peptides from iris were analysed using liquid chromatography-tandem mass spectrometry on an Orbitrap Q Exactive Plus mass spectrometer. Verification of proteins of interest was conducted using selected reaction monitoring on a triple quadrupole mass spectrometer. The main outcome was proteins with a log2 two-fold difference in expression in iris between PACG and controls. RESULTS: There were 3,446 non-redundant proteins identified in human iris, of which 416 proteins were upregulated and 251 proteins were downregulated in PACG compared with controls. Thirty-two upregulated proteins were either components of the extracellular matrix (ECM) (fibrillar collagens, EMILIN-2, fibrinogen, fibronectin, matrilin-2), matricellular proteins (thrombospondin-1), proteins involved in cell-matrix interactions (integrins, laminin, histidine-rich glycoprotein, paxillin), or protease inhibitors known to modulate ECM turnover (α-2 macroglobulin, tissue factor pathway inhibitor 2, papilin). Two giant proteins, titin and obscurin, were up- and down-regulated, respectively, in the iris in PACG compared with controls. CONCLUSIONS AND CLINICAL RELEVANCE: This proteomic study shows that ECM composition and homeostasis are altered in the iris in PACG.

Inflammatory cytokines levels in aqueous humour and surgical outcomes of trabeculectomy in patients with prior acute primary angle closure.

PURPOSE: To quantify the levels of three inflammatory cytokines in the aqueous humour of patients with prior acute primary angle closure (APAC) and investigate their correlation with surgical outcomes of trabeculectomy. METHODS: In this prospective cohort study, aqueous humour samples were collected from 44 prior APAC eyes. Analyte concentrations of monocyte chemoattractant protein-1 (MCP-1), vascular endothelial growth factor (VEGF) and interleukin-6 (IL-6) were measured using multiplexed immunoassay kits. Intraocular pressure was measured using Goldmann application tonometry. RESULTS: Forty-four prior APAC eyes were followed up for 24 months after trabeculectomy and divided into success and failure groups according to surgical outcomes. Monocyte chemoattractant protein-1 (MCP-1) levels in the aqueous humour were significantly higher in the failure group (p = 0.0118). Univariate and multivariate analyses showed that MCP-1 level was a significant risk factor for trabeculectomy outcomes (univariate analysis: p = 0.016, odds ratio = 14.538; multivariate analysis: p = 0.023, odds ratio = 13.718). When prior APAC eyes were divided according to MCP-1 levels, the overall success rate was significantly higher in eyes with low MCP-1 levels than eyes with high MCP-1 levels (p = 0.0249). CONCLUSION: In prior APAC patients, the MCP-1 level in the aqueous humour predicts trabeculectomy results. Therefore, modulation of MCP-1 expression may have potential clinical applications after filtration surgery.

Higher TGF-ß1, TGF-ß2, MMP-2, and TIMP-1 Levels in the Aqueous Humor of Patients with Acute Primary Angle Closure.

PURPOSE: To assess the quantitative differences in the levels of members of the transforming growth factor (TGF-ß), matrix metalloproteinase (MMP), and tissue inhibitor of MMP (TIMP) families in the aqueous humor (AH) between patients with acute primary angle closure (APAC) and those with cataract only. METHODS: AH samples were collected from 26 patients with APAC and cataract as well as 26 patients with age-related cataract only. Multiplex assays were used to measure the concentrations of TGF-ß1, TGF-ß2, and TGF-ß3; MMP-1, MMP-2, MMP-7, MMP-9, and MMP-10; and TIMP-1 and TIMP-2. RESULTS: The concentrations of TGF-ß1, TGF-ß2, MMP-2, (p < 0.001), and TIMP-1 were significantly higher (all p < 0.001) in AH samples from patients with APAC versus cataract only. Conversely, the AH concentrations of MMP-7 (p = 0.524), MMP-9 (p = 0.103), MMP-10 (p = 0.111), and TIMP-2 (p = 0.059) did not significantly differ between the groups. The concentrations of TGF-ß3 and MMP-1 were below the respective detection limits in most AH samples. CONCLUSION: The AH levels of TGF-ß1, TGF-ß2, MMP-2, and TIMP-1 were elevated in APAC eyes. Such altered protein levels could induce abnormal deposition of extracellular matrix in the trabecular meshwork, resulting in an increase in aqueous outflow resistance and, thereby, providing a possible explanation of the mechanism of residual glaucoma after cataract surgery.

Aqueous Inflammation and Ischemia-Related Biomarkers in Neovascular Glaucoma with Stable Iris Neovascularization.

Purpose: To characterize the aqueous levels of inflammation and ischemia-related biomarkers in a spectrum of retinal ischemic conditions, including neovascular glaucoma (NVG) with stable iris neovascularization after pan retinal photocoagulation (PRP) and anti-VEGF treatment. Methods: Aqueous samples were collected from 139 eyes including NVG (n = 12), stable NVG (n = 26), CRVO (n = 11), NPDR (n = 18), PACG (n = 18), PDR (n = 25), BRVO (n = 7) and cataract (n = 22). The levels of VEGF-A, IL-8 and EPO were measured with ELISA. Results: Aqueous VEGF-A significantly decreased after anti-VEGF and PRP, from 983.79 ± 821.16 pg/ml in the NVG group (n = 11) to 256.50 ± 51.14 pg/ml in the stable NVG group (n = 24) (P = .015). Aqueous VEGF-A in stable NVG group (256.50 ± 51.14 pg/ml, n = 24) was significantly higher (ANOVA, P < .001) than in CRVO (212.10 ± 19.84 pg/ml, n = 7, P = .017), NPDR (221.18 ± 38.21 pg/ml, n = 14, P = .015), BRVO (213.14 ± 48.50 pg/ml, n = 6, P = .028) and cataract group (185.30 ± 34.35 pg/ml, n = 22, P < .001). Aqueous IL-8 in stable NVG group (74.82 ± 10.78 pg/ml, n = 24) was significantly higher (ANOVA, P < .001) than in CRVO (65.19 ± 15.34 pg/ml, n = 11, P = .032) and cataract group (54.11 ± 12.28 pg/ml, n = 22, P < .001). Aqueous EPO in stable NVG group (17.48 ± 3.02 pg/ml, n = 24) was significantly higher (ANOVA, P < .001) than in BRVO (14.98 ± 2.57 pg/ml, n = 7, P = .034) and cataract group (13.50 ± 2.65 pg/ml, n = 22, P < .001). Aqueous concentrations of VEGF-A and IL-8 correlated positively with IOP (r = 0.413, P < .001, r = 0.349, P < .001, respectively, r = correlation coefficient). VEGF-A correlated positively with IL-8 and EPO (P < .001, P = .002, respectively). IL-8 correlated positively with EPO (P < .001). Conclusions: The aqueous levels of VEGF-A, IL-8 and EPO in NVG patients with stable iris neovascularization, who had received PRP and anti-VEGF, were still significantly higher than in control groups with some retinal ischemic conditions.

Lowered Decorin With Aberrant Extracellular Matrix Remodeling in Aqueous Humor and Tenon's Tissue From Primary Glaucoma Patients.

Purpose: To evaluate the inflammatory cytokine, growth factors, extracellular matrix (ECM) remodeling genes, profibrotic and antifibrotic molecules in patients undergoing glaucoma filtration surgery (GFS). Additionally, the effect of preoperative antiglaucoma medications (AGMs) and postoperative bleb status were related to these parameters. Methods: Tenon's tissue and aqueous humour (AH) were collected from 207 patients undergoing GFS with primary open-angle glaucoma (POAG) (n = 77), primary angle-closure glaucoma (PACG) (n = 62), and cataract controls (n = 68). Monocyte chemoattractant protein-1 (MCP-1), connective tissue growth factor (CTGF), transforming growth factor ß1/2 (TGF-ß1/2), lysyl oxidase (LOX), lysyl oxidase L2 (LOXL2), elastin (ELN), collagen type 1 α 1 (COL1A1), secreted protein acidic and rich in cysteine (SPARC), α-smooth muscle actin (α-SMA), and decorin (DCN) were determined in tenon's tissue by real-time PCR and in AH using ELISA. Results: A significant increase was observed in the transcripts of MCP-1, TGF-ß2, and SPARC in POAG and PACG (P < 0.05); CTGF, TGF-ß1, LOX, LOXL2, ELN, COL1A1, and α-SMA in PACG (P < 0.05) compared with control. DCN transcript was significantly decreased in POAG and PACG (P < 0.05) compared with control. The protein levels of CTGF, TGF-ß1/ß2, ELN, SPARC, and LOXL2 was significantly elevated in POAG and PACG (P < 0.05); DCN was decreased (P < 0.05) compared with control. These parameters showed significant association with duration of preoperative AGMs and postoperative bleb status. Conclusions: This study demonstrates increased expression of growth factors and ECM molecules, both at protein and transcript levels in GFS patients. A decreased DCN in AH seems striking, and if restored might have a therapeutic role in minimizing postoperative scarring to improve GFS outcome.

Aqueous humor levels of TGFß2 and SFRP1 in different types of glaucoma.

BACKGROUND: To assess bioactive transforming growth factor-ß2 (TGFß2) and secreted frizzled-related protein-1 (SFRP1) levels in aqueous humor (AH) of different types of glaucoma. METHODS: AH samples were obtained immediately before ophthalmic surgery with a 27-gauge needle attached to a microsyringe from 126 eyes (105 patients) divided into five groups: cataract (control), primary open-angle glaucoma (POAG), chronic angle-closure glaucoma (CACG), primary angle-closure suspects (PACS), and acute angle-closure glaucoma (AACG). Bioactive TGFß2 and SFRP1 levels were assayed by ELISA. RESULTS: The concentration of TGFß2 in AH of POAG patients, but not CACG, PACS, or AACG patients, was significantly higher than control eyes. However, within the AACG group, although the TGFß2 levels in AH did not differ significantly from the control level when all AACG patients were grouped together, there were differences when the AACG patients were divided into high and normal intraocular pressure (IOP); TGFß2 of AACG patients with high IOP (> 21 mmHg) was significantly higher than those with normal IOP. AH levels of SFRP1 were not significantly different among the groups. However, a statistical significant, negative correlation between SFRP1 and IOP existed in the POAG group. POAG patients with high IOP had lower levels of SFRP1 than those with normal IOP. In contrast, a significant, positive correlation between SFRP1 level and IOP was detected in the AACG group. AACG patients with high IOP had a higher level of SFRP1 than those with normal IOP. Concentrations of TGFß2 and SFRP1 did not correlate significantly with each other, or with age. CONCLUSIONS: These results indicate that AH levels of TGFß2 and SFRP1 showed different profiles in different types of glaucomas.

Detection of Proteins Associated with Extracellular Matrix Regulation in the Aqueous Humour of Patients with Primary Glaucoma.

Purpose: The protein composition of aqueous humour (AH) has held significant relevance and remains to be the prime sample in the discovery of biomarkers in glaucoma. The purpose of this study is to analyze the AH protein concentrations in primary open angle glaucoma (POAG) and primary angle closure glaucoma (PACG) and further examine the proteome changes compared to cataract control. Methods: AH was collected from 90 POAG, 72 PACG, 78 cataracts (controls) in this study. The total protein was quantified using Bradford's assay. Samples were subjected to trypsin digestion followed by liquid chromatography-mass spectrometry (LC-MS) for proteomic studies (n = 3 per group). The extracellular matrix has a major influence on the AH outflow, and the regulator proteins osteopontin (OPN), cathepsin D, and cystatin C detected by mass spectrometry are validated in AH samples by Western blot and turbidimetric immunoassay. Results: We observed a significant increase in protein levels of POAG (p = .0009); interestingly, a similar increase in PACG compared to cataract (p < .0001) and POAG (p = .02). Proteomics analysis identified 184, 190, and 299 proteins in control, POAG and PACG. OPN was increased in POAG (p = .0319) and PACG (p = .0103) compared to control. The precursor form of cathepsin D was increased in POAG and decreased in PACG, though not significant compared to control. Cystatin C was also increased in both POAG (p = .0310) and PACG (p = .0125) compared to control. Conclusion: In this study, we report for the first time that PACG cohort had higher total protein compared to controls. A qualitative comparison of proteomes revealed increased numbers of proteins identified in PACG. We assume that elevated levels of OPN and cystatin C in POAG and PACG along with altered cathepsin levels may contribute to ECM aberration in glaucoma.

Comparative evaluation of the aqueous humor proteome of primary angle closure and primary open angle glaucomas and age-related cataract eyes.

PURPOSE: To analyze and compare the total proteome of aqueous humor (AH) from patients having primary angle closure glaucoma (PACG), primary open angle glaucoma (POAG) and age-related cataract. MATERIALS AND METHODOLOGY: Aqueous humor was collected from age-matched PACG, POAG and cataract patients who underwent surgery, and it was immediately stored at - 80 °C until analysis. From each sample, 25 µg of total protein was subjected to trypsin digestion and subsequently LC-MS/MS analysis was performed for the deep proteome analysis. The data acquired after the LC-MS/MS analysis were analyzed using Proteome Discoverer 1.4. The identified peptide matches were validated using percolator, at less than 1% false discovery rates. RESULTS: A total of 625, 594 and 636 proteins were identified in PACG, POAG and cataract groups, respectively (n = 9 in each group). The inter-group comparison among all these groups showed that 246 proteins were identified in all the three groups. An average of 236 ± 42, 218 ± 40 and 214 ± 62 proteins from each AH sample of PACG, POAG and cataract, respectively, was identified. There were 53 proteins commonly found in all 9 PACG AH, 59 proteins in POAG AH and 42 proteins in 9 cataracts AH samples. In the individual analysis, there were 28 proteins found in all the samples analyzed representing the "constitutive AH proteome." Spectral counting analysis of 246 proteins identified in all three group types showed significant differences in protein abundance. In proteins unique to PACG AH, 7 proteins viz. ARHGEF12, APC2, WAS, PIK3CG, ITGB1, MSN and PFN1 out of 226 were found in "Regulation of Actin Cytoskeleton" pathway, whereas in POAG 5 out of 206 proteins viz. ADCY2, ITPR1, MAPK3, MAP3K2 and TUBB1 were found in "Gap Junction" pathway. CONCLUSIONS: A qualitative as well as a quantitative comparison of proteomes of AH from PACG, POAG and age-related cataract eyes showed significant differences, thus providing clues to the disease pathophysiology.

Matricellular Protein Levels in Aqueous Humor and Surgical Outcomes of Trabeculectomy.

Purpose: The purpose of this study was to quantify levels of three matricellular proteins in the aqueous humor in patients with previous acute primary angle closure (APAC) and investigate their correlation with bleb morphology and surgical outcomes of trabeculectomy. Methods: In this prospective study, aqueous humor samples were collected from 40 previous APAC eyes. Concentrations of three matricellular proteins-secreted protein acidic and rich in cysteine (SPARC), thrombospondin-2, and osteopontin-were measured using multiplexed immunoassays kits. Intraocular pressure was measured using Goldmann application tonometry. Bleb morphology was assessed using anterior segment optical coherence tomography, and bleb score was calculated according to bleb size and reflectivity. Results: When previous APAC eyes were divided according to surgical outcome 18 months after trabeculectomy, SPARC protein was significantly higher in aqueous humor in the failure group (P = 0.009). When previous APAC eyes were divided according to SPARC level, eyes with low SPARC levels had significantly higher overall success rate compared with eyes with high SPARC levels (P = 0.005 for complete success and P = 0.018 for qualified success). Multiple logistic regression analyses showed that eyes with higher levels of SPARC were more likely to have a failed filtration surgery than were eyes with lower levels of SPARC. For complete success, P = 0.006 and odds ratio (OR) = 6.458; for qualified success, P = 0.033 and OR = 2.608. The level of SPARC was found to have a positive correlation with bleb score (P < 0.001, R2 = 0.471). Conclusions: In previous APAC patients, the SPARC level in aqueous humor is a prognostic factor for surgical results of trabeculectomy. Modulation of SPARC expression may have potential clinical applications after filtration surgery.

Matricellular Proteins Play a Potential Role in Acute Primary Angle Closure.

PURPOSE: To quantify levels of matricellular proteins in aqueous humor samples from acute primary angle closure (APAC) and non-glaucomatous cataract eyes and investigate their correlation with intraocular pressure (IOP) fluctuation. MATERIALS AND METHODS: Aqueous humor samples were collected from 63 eyes including 29 current APAC eyes, 12 previous APAC eyes, and 22 cataract eyes. Concentrations of four main matricellular proteins (SPARC, tenascin-C, thrombospondin-2, and osteopontin) were measured using multiplexed immunoassay kits. Correlations between matricellular proteins and age, sex, and IOP were then detected using Spearman's rank correlation coefficient. RESULTS: The levels of SPARC, thrombospondin-2, and osteopontin were significantly elevated in the APAC group as compared to the cataract group (p < 0.001, p < 0.001, and p = 0.009, respectively). Further separation of the APAC group into current and previous APAC groups showed that only the differences of SPARC and thrombospondin-2 between the current APAC and cataract groups were significant (both p < 0.001). All four matricellular proteins were found to have a positive correlation with IOP in the current APAC group but no correlation was found in the previous APAC or cataract groups. CONCLUSIONS: The levels of matricellular proteins were significantly elevated in the current APAC eyes and positively correlated to IOP. Further studies are necessary to investigate the molecular mechanisms and histological evidence of pathogenesis in matricellular proteins in APAC.

Primary angle closure glaucoma (PACG) susceptibility gene PLEKHA7 encodes a novel Rac1/Cdc42 GAP that modulates cell migration and blood-aqueous barrier function.

PLEKHA7, a gene recently associated with primary angle closure glaucoma (PACG), encodes an apical junctional protein expressed in components of the blood aqueous barrier (BAB). We found that PLEKHA7 is down-regulated in lens epithelial cells and in iris tissue of PACG patients. PLEKHA7 expression also correlated with the C risk allele of the sentinel SNP rs11024102 with the risk allele carrier groups having significantly reduced PLEKHA7 levels compared to non-risk allele carriers. Silencing of PLEKHA7 in human immortalized non-pigmented ciliary epithelium (h-iNPCE) and primary trabecular meshwork cells, which are intimately linked to BAB and aqueous humor outflow respectively, affected actin cytoskeleton organization. PLEKHA7 specifically interacts with GTP-bound Rac1 and Cdc42, but not RhoA, and the activation status of the two small GTPases is linked to PLEKHA7 expression levels. PLEKHA7 stimulates Rac1 and Cdc42 GTP hydrolysis, without affecting nucleotide exchange, identifying PLEKHA7 as a novel Rac1/Cdc42 GAP. Consistent with the regulatory role of Rac1 and Cdc42 in maintaining the tight junction permeability, silencing of PLEKHA7 compromises the paracellular barrier between h-iNPCE cells. Thus, downregulation of PLEKHA7 in PACG may affect BAB integrity and aqueous humor outflow via its Rac1/Cdc42 GAP activity, thereby contributing to disease etiology.

Multiplex Cytokine Analysis of Aqueous Humor from the Patients with Chronic Primary Angle Closure Glaucoma.

PURPOSE: To compare the levels of cytokines and growth factor in aqueous humor of the patients with chronic primary angle closure glaucoma (PACG) and cataract. METHODS: Aqueous humor samples were collected from 19 chronic PACG patients and compared with 14 nonglaucomatous controls presenting for cataract surgery. The levels of 27 cytokines and growth factors were measured in the aqueous samples using multiplex bead immunoassay and compared across groups. RESULTS: Significantly higher levels of interleukin (IL)-8 (p < 0.001), eotaxin (p < 0.001), interferon gamma-induced protein (IP)-10 (p < 0.001) and macrophage inflammatory protein-1-beta (MIP-1ß; p < 0.001) were observed in aqueous of chronic PACG patients compared to controls. In comparison to controls, significantly lower levels of IL-9 (p = 0.001), IL-17 (p < 0.001), tumor necrosis factor-alpha (TNF-α; p < 0.001), granulocyte-macrophage colony-stimulating factor (GM-CSF; p < 0.001), and IL-5 (p = 0.001) were observed in chronic PACG eyes. All other assayed cytokines-IL-1ß, interleukin-1 receptor antagonist (IL-1rα), IL-6, IL-7, IL-10, IL-12, IL-13, IL-15, fibroblast growth factor-basic (FGF-basic), granulocyte colony-stimulating factor (G-CSF), monocyte chemotactic protein-1 (MCP-1), macrophage inflammatory protein-1-alpha (MIP-1α), and vascular endothelial growth factor (VEGF) -showed no significant difference between the groups. CONCLUSIONS: These results suggest that the aqueous cytokine levels of chronic PACG eyes differ significantly from nonglaucomatous eyes. This is the first study reporting significantly increased levels of eotaxin, MIP-1ß, and IP-10 and lower levels of TNF-α, IL-5, IL-9, IL-17, and GM-CSF in chronic PACG patients, suggesting a plausible role of these inflammatory cytokines in its pathogenesis.

Vascular Endothelial Growth Factor is Increased in Aqueous Humor of Acute Primary Angle-Closure Eyes.

PURPOSE: To measure and compare the levels of vascular endothelial growth factor (VEGF) in the aqueous humor of patients with acute primary angle closure (APAC), primary angle-closure glaucoma (PACG), and normal cataract (controls). METHODS: Aqueous humor samples were prospectively collected from 38 APAC eyes, 36 PACG eyes, and 25 nonglaucomatous cataract control eyes. The levels of aqueous humor VEGF were measured using enzyme-linked immunoassays. The clinical characteristics of participants were also collected for correlation analysis. RESULTS: VEGF was detected in aqueous humor samples of 35 of 38 APAC patients (mean±SE of the mean, 935±258 pg/mL), 30 of 36 PACG patients (165±37.5 pg/mL), and 16 of 25 cataract controls (69.5±13.5 pg/mL). The mean concentration of VEGF in APAC eyes was 13.5 and 5.7 times higher than that in controls and PACG eyes, respectively, and these differences were statistically significant (both P<0.0167). In the correlation analysis that included all participants, the aqueous humor VEGF level was found to correlate negatively with axial length (ρ=-0.342, P=0.001), and positively with intraocular pressure (ρ=0.434, P<0.001). CONCLUSIONS: VEGF was significantly increased in aqueous humor of APAC eyes. An increase in aqueous humor VEGF may be the result of the characteristic ocular ischemia and hypoxia observed in APAC eyes as a consequence of sudden excessive increases in intraocular pressure during the acute episode.

Chemokine (C-C motif) ligand 2 and chemokine (C-C motif) ligand 7 in angle-closure glaucoma.

PURPOSE: To compare the concentrations of chemokine (C-C motif) ligand 2 (CCL2) and chemokine (C-C motif) ligand 7 (CCL7) in aqueous humour of eyes with acute primary angle-closure (APAC), primary angle-closure glaucoma (PACG) or with age-related cataract. METHODS: The prospective investigation included a study group of 56 patients (24 patients with APAC; 32 patients with PACG) undergoing glaucoma surgery and a control group of 21 patients undergoing routine cataract surgery. The interval between glaucoma attack and trabeculectomy was 19.4 ± 9.8 days. RESULTS: The concentrations of CCL2 (3037 ± 501 versus 1127 ± 141 pg/ml; p = 0.003) and CCL7 (10.5 ± 2.4 versus 2.8 ± 0.4 pg/ml; p < 0.001) were significantly higher in the APAC group than in the PACG group, while the concentrations of both cytokines did not differ significantly between the PACG group and the control group (CCL2: 1190 ± 98 pg/ml; p = 0.10; CCL7: 2.0 ± 0.6; p = 0.18). The concentrations of both cytokines were significantly and inversely correlated with the interval between the acute angle-closure attack and surgery (CCL2: p = 0.02; ρ = -0.48; CCL7: p = 0.04; ρ = -0.42). Higher CCL2 concentrations were associated with higher prevalence of surgical failure (28.6% versus 7.1%; p = 0.036). CONCLUSIONS: Concentrations of CCL2 and CCL7 as major inflammatory cytokines with potential influence on the risk of filtering surgery failure were significantly elevated after an acute glaucoma attack and decreased with longer interval to the attack. Trabeculectomy may have a lower risk of failure if performed later than earlier after an acute glaucoma attack, with lower CCL2 concentrations associated with better surgical outcome.

Aqueous humor concentration of VEGF and retinal oxygen saturation after unilateral acute primary angle closure.

PURPOSE: To measure the levels of vascular endothelial growth factor (VEGF) in aqueous humor in patients with unilateral acute primary angle closure (APAC) and retinal oxygen saturation (SO2 ) after trabeculectomy. METHODS: Twelve patients, with unilateral trabeculectomy-required APAC, were recruited as the study group. Aqueous humor samples were collected prospectively for every subject. VEGF concentrations were analysed via enzyme-linked immunosorbent assay (ELISA), and retinal SO2 was measured using oximeter (Reykjavik, Iceland) at 1 month after trabeculectomy. RESULTS: In the APAC eyes, the mean aqueous humor concentration of VEGF was 388.4 ± 260.1 pg/ml and positively correlated with preoperative intraocular pressure (ρ = 0.658, p = 0.020). In the normal controls, the mean SO2 levels in the larger arterioles and venules were 93.0 ± 5.9% and 59.5 ± 5.4% and the arteriovenous [A-V] difference was 33.5 ± 6.3%. In the study group, the arteriolar SO2 at 1 month after surgery (97.6 ± 19.4%, p = 0.147) did not differ significantly from the values in the control group. However, the mean venular SO2 was significantly lower than the control eyes (50.0 ± 9.1% versus 59.5 ± 5.4%, p = 0.004), and a remarkable increased A-V difference (47.6 ± 22.7% versus 33.5 ± 6.3%) was found in the APAC eyes (p = 0.037). CONCLUSIONS: Lower venular SO2 and increased A-V difference existed in the APAC eyes after surgery, and it is possible that the saturation was even lower during the attack. Together with the elevated VEGF concentrations in aqueous humor, these provided indirect evidence that ocular hypoxia was constantly present during APAC attack.

Current concepts in the treatment of vitreous block, also known as aqueous misdirection.

Vitreous block (VB), a rare secondary angle-closure caused by anterior rotation of the ciliary body, occurs in a variety of settings, but most frequently after glaucoma filtering surgery. The etiology remains unclear, but choroidal expansion and anterior vitreous abnormalities have been proposed. In the past, treatment of VB has yielded high rates of failure and recurrence. Advancements in surgical techniques, however, have led to improved visual outcomes. We review the history of this condition and present a stepwise approach to its diagnosis and treatment using modern imaging modalities and surgical techniques.

Soluble CD44 and vascular endothelial growth factor levels in patients with acute primary angle closure.

PURPOSE: Acute elevation of intraocular pressure (IOP) in acute primary angle closure (APAC) can cause huge damage to the variable cells in the eye; however, the mechanisms that connect the two processes still remain unclear. In this study, we aim to evaluate the levels of soluble CD44 (sCD44) and vascular endothelial growth factor (VEGF) in the aqueous humour of acute primary angle closure patients. METHODS: This study included 24 eyes of 24 APAC patients (11 eyes with current APAC and 13 eyes with previous APAC) and 15 eyes of 15 cataract subjects. Clinical data were acquired, and aqueous humour was collected. The levels of sCD44 and VEGF in the aqueous humour were determined by ELISA and magnetic bead immunoassay technique. RESULTS: The concentrations of the sCD44 and VEGF in the current APAC were 9.9 ± 8.8 ng/ml and 2440.2 ± 2107.1 ng/ml, respectively, which were significantly higher when compared to the previous APAC group (p = 0.001) and cataract (p < 0.001); however, there were no significant differences between the group with previous APAC and with cataract. Higher IOP was associated with higher concentration of sCD44 (Rho = 0.617, p = 0.001). The concentration of the VEGF in aqueous humour of APAC patients was closely related to the sCD44 levels (Rho = 0.752, p < 0.001). CONCLUSION: After acute increase of IOP in the APAC, the level of sCD44 and VEGF increased significantly in the aqueous humour. The damage due to high IOP may therefore be mediated through the release of sCD44.