GLIS1 regulates trabecular meshwork function and intraocular pressure and is associated with glaucoma in humans.
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ABSTRACT:
SUBMITTER: Nair KS
PROVIDER: S-EPMC8361148 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:PurposeIn this study, we investigate how adenosine monophosphate-activated protein kinase (AMPK) affects extracellular matrix (ECM) and cellular tone in the trabecular meshwork (TM), and examine how deletion of its catalytic α2 subunit affects IOP and aqueous humor clearance in mice.MethodsHuman TM tissue was examined for expression of AMPKα1 and AMPKα2, genomically distinct isoforms of the AMPK catalytic subunit. Primary cultured human TM cells were treated for 24 hours with the AMPK activator 5-amino-1-β-Dffff-ribofuranosyl-imidazole-4-carboxamide (AICAR), under basal or TGF-β2 stimulatory conditions. Conditioned media (CM) was probed for secreted protein acidic and rich in cysteine (SPARC), thrombospondin-1 (TSP-1), and ECM proteins, and cells were stained for F-actin. Cells underwent adenoviral infection with a dominant negative AMPKα subunit (ad.DN.AMPKα) and were similarly analyzed. Intraocular pressure, central corneal thickness (CCT), and aqueous clearance were measured in AMPKα2-null and wild-type (WT) mice.ResultsBoth AMPKα1 and AMPKα2 are expressed in TM. AICAR activated AMPKα and suppressed the expression of various ECM proteins under basal and TGF-β2 stimulatory conditions. AICAR decreased F-actin staining and increased the phospho-total RhoA ratio (Ser188). Transforming growth factor-β2 transiently dephosphorylated AMPKα. Infection with ad.DN.AMPKα upregulated various ECM proteins, decreased the phospho-total RhoA ratio, and increased F-actin staining. AMPKα2-null mice exhibited 6% higher IOP and decreased aqueous clearance compared with WT mice, without significant differences in CCT or angle morphology.ConclusionsCollectively, our data identify AMPK as a critical regulator of ECM homeostasis and cytoskeletal arrangement in the TM. Mice that are AMPKα2-null exhibit higher IOPs and decreased aqueous clearance than their WT counterparts.
| S-EPMC4023495 | biostudies-literature
Project description:Lowering intraocular pressure (IOP) delays or prevents the loss of vision in primary open-angle glaucoma (POAG) patients with high IOP and in those with normal tension glaucoma showing progression. Abundant evidence demonstrates that inhibition of contractile machinery of the trabecular meshwork cells is an effective method to lower IOP. However, the mechanisms involved in the regulation of trabecular contraction are not well understood. Although microRNAs have been shown to play important roles in the regulation of multiple cellular functions, little is known about their potential involvement in the regulation of IOP. Here, we showed that miR-200c is a direct postranscriptional inhibitor of genes relevant to the physiologic regulation of TM cell contraction including the validated targets Zinc finger E-box binding homeobox 1 and 2 (ZEB1 and ZEB2), and formin homology 2 domain containing 1 (FHOD1), as well as three novel targets: lysophosphatidic acid receptor 1 (LPAR1/EDG2), endothelin A receptor (ETAR), and RhoA kinase (RHOA). Consistently, transfection of TM cells with miR-200c resulted in strong inhibition of contraction in collagen populated gels as well as decreased cell traction forces exerted by individual TM cells. Finally, delivery of miR-200c to the anterior chamber of living rat eyes resulted in a significant decrease in IOP, while inhibition of miR-200c using an adenoviral vector expressing a molecular sponge led to a significant increase in IOP. These results demonstrate for the first time the ability of a miRNA to regulate trabecular contraction and modulate IOP in vivo, making miR-200c a worthy candidate for exploring ways to alter trabecular contractility with therapeutic purposes in glaucoma.
| S-EPMC3522713 | biostudies-literature
Project description:Alterations in stiffness of the trabecular meshwork (TM) may play an important role in primary open-angle glaucoma (POAG), the second leading cause of blindness. Specifically, certain data suggest an association between elevated intraocular pressure (IOP) and increased TM stiffness; however, the underlying link between TM stiffness and IOP remains unclear and requires further study. We here first review the literature on TM stiffness measurements, encompassing various species and based on a number of measurement techniques, including direct approaches such as atomic force microscopy (AFM) and uniaxial tension tests, and indirect methods based on a beam deflection model. We also briefly review the effects of several factors that affect TM stiffness, including lysophospholipids, rho-kinase inhibitors, cytoskeletal disrupting agents, dexamethasone (DEX), transforming growth factor-?2 (TGF-?2), nitric oxide (NO) and cellular senescence. We then describe a method we have developed for determining TM stiffness measurement in mice using a cryosection/AFM-based approach, and present preliminary data on TM stiffness in C57BL/6J and CBA/J mouse strains. Finally, we investigate the relationship between TM stiffness and outflow facility between these two strains. The method we have developed shows promise for further direct measurements of mouse TM stiffness, which may be of value in understanding mechanistic relations between outflow facility and TM biomechanical properties.
| S-EPMC6501821 | biostudies-literature
Project description:The homeostasis of extracellular matrix (ECM) and actin dynamics in the trabecular meshwork (TM) outflow pathway plays a critical role in intraocular pressure (IOP) regulation. We studied the role of cathepsin K (CTSK), a lysosomal cysteine protease and a potent collagenase, on ECM modulation and actin cytoskeleton rearrangements in the TM outflow pathway and the regulation of IOP. Initially, we found that CTSK was negatively regulated by pathological stressors known to elevate IOP. Further, inactivating CTSK using balicatib, a pharmacological cell-permeable inhibitor of CTSK, resulted in IOP elevation due to increased levels and excessive deposition of ECM-like collagen-1A in the TM outflow pathway. The loss of CTSK activity resulted in actin-bundling via fascin and vinculin reorganization and by inhibiting actin depolymerization via phospho-cofilin. Contrarily, constitutive expression of CTSK decreased ECM and increased actin depolymerization by decreasing phospho-cofilin, negatively regulated the availability of active TGFβ2, and reduced the levels of alpha-smooth muscle actin (αSMA), indicating an antifibrotic action of CTSK. In conclusion, these observations, for the first time, demonstrate the significance of CTSK in IOP regulation by maintaining the ECM homeostasis and actin cytoskeleton-mediated contractile properties of the TM outflow pathway.
| S-EPMC8616380 | biostudies-literature
Project description:Glaucoma treatment is usually initiated with topical medication that lowers the intraocular pressure (IOP) by reducing the aqueous production, enhancing the aqueous outflow, or both. However, the effect of topical IOP-lowering medications on the microstructures of the aqueous outflow pathway are relatively unknown. In this retrospective, observational study, 56 treatment-naïve patients with primary open-angle glaucoma were enrolled. Images of the nasal and temporal corneoscleral limbus were obtained using anterior segment optical coherence tomography (AS-OCT). The conjunctival vessels and iris anatomy were used as landmarks to select the same limbal area scan, and the trabecular meshwork (TM) width, TM thickness, and Schlemm's canal (SC) area were measured before and after using the IOP-lowering agents for 3 months. Among the 56 patients enrolled, 33 patients used prostaglandin (PG) analogues, and 23 patients used dorzolamide/timolol fixed combination (DTFC). After 3 months of DTFC usage, the TM width, TM thickness, and SC area did not show significant changes in either the nasal or temporal sectors. Conversely, after prostaglandin analog usage, the TM thickness significantly increased, and the SC area significantly decreased (all P < 0.01). These findings warrant a deeper investigation into their relationship to aqueous outflow through the conventional and unconventional outflow pathways after treatment with PG analogues.
| S-EPMC8437975 | biostudies-literature
Project description:Purpose:The concentration of protons in the aqueous humor (AH) of the vertebrate eye is maintained close to blood pH; however, pathologic conditions and surgery may shift it by orders of magnitude. We investigated whether and how changes in extra- and intracellular pH affect the physiology and function of trabecular meshwork (TM) cells that regulate AH outflow. Methods:Electrophysiology, in conjunction with pharmacology, gene knockdown, and optical recording, was used to track the pH dependence of transmembrane currents and mechanotransduction in primary and immortalized human TM cells. Results:Extracellular acidification depolarized the resting membrane potential by inhibiting an outward K+-mediated current, whereas alkalinization hyperpolarized the cells and augmented the outward conductance. Intracellular acidification with sodium bicarbonate hyperpolarized TM cells, whereas removal of intracellular protons with ammonium chloride depolarized the membrane potential. The effects of extra- and intracellular acid and alkaline loading were abolished by quinine, a pan-selective inhibitor of two-pore domain potassium (K2P) channels, and suppressed by shRNA-mediated downregulation of the mechanosensitive K2P channel TREK-1. Extracellular acidosis suppressed, whereas alkalosis facilitated, the amplitude of the pressure-evoked TREK-1-mediated outward current. Conclusions:These results demonstrate that TM mechanotransduction mediated by TREK-1 channels is profoundly sensitive to extra- and intracellular pH shifts. Intracellular acidification might modulate aqueous outflow and IOP by stimulating TREK-1 channels.
| S-EPMC6532698 | biostudies-literature
Project description:The receptor-associated prorenin system (RAPS) is associated with several pathologic conditions, including diabetic retinopathy, age-related macular degeneration, and uveitis. Here, we show the involvement of RAPS in the trabecular meshwork (TM) from patients with primary open-angle glaucoma (POAG) and neovascular glaucoma (NVG) due to proliferative diabetic retinopathy. Anterior chamber (AC) levels of prorenin significantly increased in both POAG and NVG, as did those of angiotensin II in NVG alone, compared to cataract. In surgically excised TM tissues, (pro)renin receptor ((P)RR) and angiotensin II type 1 receptor (AT1R) co-localized with prorenin and angiotensinogen, respectively. In screening for various genes related to glaucoma, prorenin stimulation to human TM cells exclusively upregulated cell junction constituents connexin 43 and zona occludens 1, while downregulating an extracellular matrix-degrading enzyme tissue plasminogen activator, all of which were reversed by (P)RR blockade. In contrast, angiotensin II application upregulated a pro-angiogenic factor placental growth factor alone, which was abolished by AT1R blockade. Consistently, (P)RR and AT1R co-localized with these corresponding proteins in patient TM tissues. Oxidative stress, a known etiology for glaucoma, induced the expression of prorenin and angiotensinogen in human TM cells. These data suggest the contribution of RAPS to the molecular pathogenesis of POAG and NVG through TM tissue remodeling and AC angle angiogenesis.
| S-EPMC7465309 | biostudies-literature
Project description:Glaucoma is one of the most common causes of blindness worldwide, but the risk factors of glaucoma are yet to be fully understood. We investigated the relationship between the prevalence of glaucoma and trabecular meshwork (TM) length by comparing the mean TM length of a South Korean population with that of another ethnic population. We included 250 eyes of 125 patients who underwent anterior segment optical coherence tomography at Yonsei University Gangnam Severance Hospital between January 2015 and December 2017. We measured the distance from the scleral spur to Schwalbe's line in patients with open and closed angles and calculated the TM length using the open- and closed-angle ratios in the general population. The mean TM length of the patients in our study was 752 ± 116 μm. Considering the compensated data, the estimated true mean TM length in the Korean population was 793 ± 76 μm, which was similar to the mean TM length of a previously evaluated Hispanic population, but differed significantly from those of previously evaluated Asian (Chinese), Caucasian, and African-American populations (p < 0.05). Our results support the hypothesis that the development of glaucoma would be affected by TM length.
| S-EPMC7961563 | biostudies-literature
Project description:IntroductionExtracellular matrix (ECM) materials accumulate in the trabecular meshwork (TM) tissue of patients with glaucoma, which is associated with a decrease in aqueous humor outflow and therefore an increase in intraocular pressure. To explore a potential mechanism for ECM regulation in the TM, we purified extracellular vesicles (EVs) from conditioned media of differentiated TM cells in culture isolated from non-glaucomatous and glaucomatous human donor eyes.MethodsEVs were purified using the double cushion ultracentrifugation gradient method. Fractions containing EV markers CD9 and TSG101 were analyzed using nanoparticle tracking analysis to determine their size and concentration. We then determined their proteomic cargo by mass spectrometry and compared protein profiles of EVs between normal and glaucomatous TM cells using PANTHER. Key protein components from EV preparations were validated with Western blotting.ResultsResults showed changes in the percentage of ECM proteins associated with EVs from glaucomatous TM cells compared to non-glaucomatous TM cells (5.7% vs 13.1% respectively). Correspondingly, we found that two ECM-related cargo proteins found across all samples, fibronectin and EDIL3 were significantly less abundant in glaucomatous EVs (<0.3 fold change across all groups) compared to non-glaucomatous EVs.DiscussionOverall, these data establish that ECM materials are prominent proteomic cargo in EVs from TM cells, and their binding to EVs is diminished in glaucoma.
| S-EPMC10785745 | biostudies-literature
Project description:Myocilin is highly expressed in the trabecular meshwork (TM), which plays an important role in the regulation of intraocular pressure (IOP). Myocilin abnormalities may cause dysfunction of the TM, potentially leading to increased IOP. High IOP is a well?known primary risk factor for glaucoma. Myocilin mutations are common among glaucoma patients, and they are implicated in juvenile?onset open?angle glaucoma (JOAG) and adult?onset primary open?angle glaucoma (POAG). Aggregation of aberrant mutant myocilins is closely associated with glaucoma pathogenesis. The aim of the present review was to discuss the recent findings regarding the major physiological functions of myocilin, such as intra? and extracellular proteolytic processes. We also aimed to discuss the risk factors associated with myocilin and the development of glaucoma, such as misfolded/mutant myocilin, imbalance of myocilin and extracellular proteins, and instability of mutant myocilin associated with temperature. Finally, we further outlined certain issues that are yet to be resolved, which may represent the basis for future studies on the role of myocilin in glaucoma.
| S-EPMC6317685 | biostudies-literature