Evaluation of different OCT systems in quantitative imaging of human Schlemm's canal.
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ABSTRACT:
SUBMITTER: Wu X
PROVIDER: S-EPMC8792050 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:PurposeTo validate the ability of visible-light optical coherence tomography (vis-OCT) in imaging the full Schlemm's canal (SC) and its surrounding limbal vascular network in mice in vivo through a compound circumlimbal scan.MethodsWe developed an anterior segment vis-OCT system and a compound circumlimbal scanning method, which montages eight rotated raster scans. We calibrated the circumlimbal scan geometry using a three-dimensional printed phantom eyeball before imaging wild-type C57BL/6J mice. We measured SC size by segmenting SC cross sections from vis-OCT B-scan images and imaged the limbal microvascular network using vis-OCT angiography (vis-OCTA). To introduce changes in SC size, we used a manometer to adjust the intraocular pressure (IOP) to different levels. To create additional optical scattering contrast to enhance SC imaging, we surgically increased the episcleral venous pressure (EVP) and caused blood reflux into SC.ResultsUsing the compound circumlimbal scan, our anterior segment vis-OCT noninvasively imaged the full SC and limbal microvascular network in mouse for the first time. We observed an average 123% increase in SC volume when we decreased the IOP by 10 mm Hg from the baseline IOP of 7 to 10 mm Hg and an average 72% decrease in SC volume when the IOP level was elevated by 10 mm Hg from the baseline IOP. We also observed location-dependent SC size responses to IOP changes. Blood reflux caused by increased EVP enabled vis-OCTA to directly visualize SC, which matched well with the segmented SC.ConclusionsVis-OCT and vis-OCTA can accurately image the entire SC and limbal microvascular network in vivo using the compound circumlimbal scan. Vis-OCT is also able to quantitatively measure SC responses to changing IOP levels.
| S-EPMC7326574 | biostudies-literature
Project description:The distal outflow pathway of the human eye consists of the outer wall of Schlemm's canal, collector channels, and the deep-scleral, mid-scleral and episcleral vessels. It is the last region of transit for aqueous humor before returning to the venous system. While the trabecular meshwork, scleral spur, and inner wall of Schlemm's canal have been extensively analyzed to define their contributions to aqueous outflow, the role of the distal outflow pathway is not completely understood. Collector channels, emanating from Schlemm's canal were previously thought to be passive conduits for aqueous humor. However, recent studies have shown many collector channels contain flap-like appendages which move with changes in pressure. These findings, along with studies demonstrating innervation of episcleral vessels, have led to questions regarding whether other structures in the distal outflow pathway are under neural regulation and how this may influence aqueous humor outflow. This study evaluates the innervation of the outer wall of Schlemm's canal and collector channels, along with the deep-scleral, mid-scleral and episcleral vasculature with microcomputed tomography and 3-dimensional reconstruction, correlative light microscopy, immunohistochemistry, and transmission electron microscopy. Peripheral, autonomic, and sensory nerve fibers were found to be present adjacent to Schlemm's canal outer wall endothelium, collector channel endothelium, and in the different regions of the distal outflow vasculature. Nerves were more commonly identified in regions that contained collector channels when compared to regions without collector channels. These findings regarding the neural anatomy suggest an active neural regulation of aqueous humor outflow throughout the proximal and distal regions of the conventional outflow pathway.
| S-EPMC10493174 | biostudies-literature
Project description:Schlemm's canal is an important structure of the conventional aqueous humor outflow pathway and is critically involved in regulating the intraocular pressure. In this study, we report a novel finding that prospero homeobox protein 1 (Prox-1), the master control gene for lymphatic development, is expressed in Schlemm's canal. Moreover, we provide a novel in vivo method of visualizing Schlemm's canal using a transgenic mouse model of Prox-1-green fluorescent protein (GFP). The anatomical location of Prox-1⁺ Schlemm's canal was further confirmed by in vivo gonioscopic examination and ex vivo immunohistochemical analysis. Additionally, we show that the Schlemm's canal is distinguishable from typical lymphatic vessels by lack of lymphatic vessel endothelial hyaluronan receptor (LYVE-1) expression and absence of apparent sprouting reaction when inflammatory lymphangiogenesis occurred in the cornea. Taken together, our findings offer new insights into Schlemm's canal and provide a new experimental model for live imaging of this critical structure to help further our understanding of the aqueous humor outflow. This may lead to new avenues toward the development of novel therapeutic intervention for relevant diseases, most notably glaucoma.
| S-EPMC4020937 | biostudies-literature
Project description:In a healthy eye, the aqueous humour (AH) flows via the ciliary body and trabecular meshwork into the collector channels, which carry it to the episcleral veins. In glaucoma, a heterogeneous group of eye disorders affecting approximately 60 million individuals worldwide, the juxtacanalicular meshwork offers greater resistance to the outflow of the AH, leading to an increase in outflow resistance that gradually results in elevated intraocular pressure (IOP). The present review comprehensively covers the morphology of Schlemm's canal (SC) and AH pathways. The path of the AH from the anterior chamber through the trabeculum into suprascleral and conjunctival veins via collector channels is described, and the role of SC in the development of glaucoma and outflow resistance is discussed. Finally, channelography is presented as a precise method of assessing the conventional drainage pathway and facilitating localization of an uncollapsed collector and aqueous veins. Attention is also given to the relationship between aqueous and episcleral veins and heartbeat. Possible directions of future research are proposed.
| S-EPMC9293138 | biostudies-literature
Project description:Aqueous humour transport across the inner wall endothelium of Schlemm's canal likely involves flow through giant vacuoles and pores, but the mechanics of how these structures form and how they influence the regulation of intraocular pressure (IOP) are not well understood. In this study, we developed an in vitro model of giant vacuole formation in human Schlemm's canal endothelial cells (HSCECs) perfused in the basal-to-apical direction (i.e., the direction that flow crosses the inner wall in vivo) under controlled pressure drops (2 or 6 mmHg). The system was mounted on a confocal microscope for time-lapse en face imaging, and cells were stained with calcein, a fluorescent vital dye. At the onset of perfusion, elliptical void regions appeared within an otherwise uniformly stained cytoplasm, and 3-dimensional reconstructions revealed that these voids were dome-like outpouchings of the cell to form giant vacuole-like structures or GVLs that reproduced the classic "signet ring" appearance of true giant vacuoles. Increasing pressure drop from 2 to 6 mmHg increased GVL height (14 ± 4 vs. 21 ± 7 ?m, p < 0.0001) and endothelial hydraulic conductivity (1.15 ± 0.04 vs. 2.11 ± 0.49 ?l min?¹ mmHg?¹ cm?²; p < 0.001), but there was significant variability in the GVL response to pressure between cell lines isolated from different donors. During perfusion, GVLs were observed "migrating" and agglomerating about the cell layer and often collapsed despite maintaining the same pressure drop. GVL formation was also observed in human umbilical vein and porcine aortic endothelial cells, suggesting that giant vacuole formation is not a unique property of Schlemm's canal cells. However, in these other cell types, GVLs were rarely observed "migrating" or contracting during perfusion, suggesting that Schlemm's canal endothelial cells may be better adapted to withstand basal-to-apical directed pressure gradients. In conclusion, we have established an in vitro model system to study giant vacuole dynamics, and we have demonstrated that this system reproduces key aspects of giant vacuole morphology and behaviour. This model offers promising opportunities to investigate the role of endothelial cell biomechanics in the regulation of intraocular pressure in normal and glaucomatous eyes.
| S-EPMC3014386 | biostudies-literature
Project description:PurposeSchlemm's canal (SC) endothelial cells derived from donors with or without glaucoma showed different mechanical properties and gene expression. As an important contributor to the regulation of intraocular pressure (IOP) and pathogenesis of primary open-angle glaucoma (POAG), the heritable key epigenetic changes, methylation may play an important role in the physiologic function of SC cells. This study aims to identify differentially methylated CpG sites (DMSs) in primary cultures of human SC cells with or without glaucoma.MethodsWe examined the methylation pattern of seven strains of primary human cells (two glaucoma and five normal SC cell samples), which were isolated and characterized using established protocols. DNA methylation was profiled using Illumina Human Methylation 450 BeadChip. Raw data were extracted and exported using Illumina GenomeStudio software. After quantile normalization, DNA methylation data were analyzed using R package RnBeads in Bioconductor. DMSs were filtered with p ≤ 1E-5, methylation change ≥ 0.1, and false discovery rate ≤ 0.05. The closest genes and the location of each CpG site were annotated using R package FDb.InfiniumMethylation.hg19. Gene Ontology and pathway analysis was performed using WebGestalt. Selected DMSs were validated using the Zymo qMethyl kit.ResultsWe used five non-glaucoma and two glaucomatous SC cell samples to profile genome-wide DNA methylation using Illumina Infinium Methylation BeadChips. Principle component analysis showed the separation between the glaucoma and control samples. After quality control and differential analysis, we identified 298 highly significant DMSs (p ≤ 1E-5). Among them, 221 DMSs were within 1 kb of a nearby gene. Gene Ontology analysis demonstrated significant enrichment in positive regulation of cell migration, negative regulation of endothelial cell proliferation, and stress fiber and actin filament bundles. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis showed enrichment in cell adhesion and gap junctions. Several glaucoma-related genes were identified, including TGFBR3, THBS1, PITX2, DAXX, TBX3, TNXB, ANGPT1, and PLEKHA7. We also examined differentially methylated regions (DMRs) near these CpG sites and identified significant DMRs in TBX3, TNXB1, DAXX, and PITX2.ConclusionsThis study represents the first genome-wide DNA methylation profiling in cultured human primary SC cells. The DMSs were enriched in the pathways related to outflow resistance. Several DMRs were validated in glaucoma-associated genes, further suggesting the role of DNA methylation in glaucoma development. This study could provide comprehensive understanding of DNA methylation in glaucoma and its effect on aqueous humor outflow.
| S-EPMC7316632 | biostudies-literature
Project description:PurposeOcular hypertension is a major risk factor for glaucoma and the inner wall of Schlemm's canal (SC) endothelia participates in the regulation of aqueous humor outflow resistance. This study aimed to identify differentially expressed genes in primary cultures of SC cells from glaucoma patients.MethodsThis study examined SC samples from three glaucoma cases and four controls. Schlemm's canal cells were isolated from eight different postmortem human eyes. Total RNA was extracted, labeled, and hybridized to Illumina HumanWG-6 BeadChips containing probes for approximately 47,000 human transcripts. After extracting the data using Illumina GenomeStudio software, the data were normalized and analyzed using the R package limma in Bioconductor. Using Protein ANalysis THrough Evolutionary Relationships (PANTHER) software, gene ontology analysis of highly expressed genes was executed in controls and glaucoma groups separately. Pathway analysis was performed with differentially expressed genes using WebGestalt (WEB-based GEne SeT AnaLysis Toolkit). Selected genes were validated using droplet digital PCR (ddPCR).ResultsGene ontology analysis indicated similar functional categories in cases and controls. Differential analysis identified a total of 113 genes with at least 2-fold expression changes in cases. Pathway analysis indicated significant enrichment of genes in cell adhesion, heparin binding, glycosaminoglycan binding, filopodium, and extracellular matrix remodeling. Eighteen selected genes with differential expression were successfully validated using ddPCR.ConclusionsThis study represents the first genome-wide expression study of human primary SC cells from glaucoma patients and provides a potential list of targets regulating SC cell stiffness and pore formation, eventually the outflow resistance in glaucoma individuals.
| S-EPMC4614909 | biostudies-literature
Project description:Increased flow resistance is responsible for the elevated intraocular pressure characteristic of glaucoma, but the cause of this resistance increase is not known. We tested the hypothesis that altered biomechanical behavior of Schlemm's canal (SC) cells contributes to this dysfunction. We used atomic force microscopy, optical magnetic twisting cytometry, and a unique cell perfusion apparatus to examine cultured endothelial cells isolated from the inner wall of SC of healthy and glaucomatous human eyes. Here we establish the existence of a reduced tendency for pore formation in the glaucomatous SC cell--likely accounting for increased outflow resistance--that positively correlates with elevated subcortical cell stiffness, along with an enhanced sensitivity to the mechanical microenvironment including altered expression of several key genes, particularly connective tissue growth factor. Rather than being seen as a simple mechanical barrier to filtration, the endothelium of SC is seen instead as a dynamic material whose response to mechanical strain leads to pore formation and thereby modulates the resistance to aqueous humor outflow. In the glaucomatous eye, this process becomes impaired. Together, these observations support the idea of SC cell stiffness--and its biomechanical effects on pore formation--as a therapeutic target in glaucoma.
| S-EPMC4183270 | biostudies-literature
Project description:Schlemm's canal (SC) is a specialized vascular structure in the eye that functions to drain aqueous humor from the intraocular chamber into systemic circulation. Dysfunction of SC has been proposed to underlie increased aqueous humor outflow (AHO) resistance, which leads to elevated ocular pressure, a factor for glaucoma development in humans. Here, using lymphatic and blood vasculature reporter mice, we determined that SC, which originates from blood vessels during the postnatal period, acquires lymphatic identity through upregulation of prospero homeobox protein 1 (PROX1), the master regulator of lymphatic development. SC expressed lymphatic valve markers FOXC2 and integrin α9 and exhibited continuous vascular endothelial-cadherin (VE-cadherin) junctions and basement membrane, similar to collecting lymphatics. SC notably lacked luminal valves and expression of the lymphatic endothelial cell markers podoplanin and lymphatic vessel endothelial hyaluronan receptor 1 (LYVE-1). Using an ocular puncture model, we determined that reduced AHO altered the fate of SC both during development and under pathologic conditions; however, alteration of VEGF-C/VEGFR3 signaling did not modulate SC integrity and identity. Intriguingly, PROX1 expression levels linearly correlated with SC functionality. For example, PROX1 expression was reduced or undetectable under pathogenic conditions and in deteriorated SCs. Collectively, our data indicate that PROX1 is an accurate and reliable biosensor of SC integrity and identity.
| S-EPMC4153702 | biostudies-literature
Project description:Ocular hypertension in glaucoma develops due to age-related cellular dysfunction in the conventional outflow tract, resulting in increased resistance to aqueous humor outflow. Two cell types, trabecular meshwork (TM) and Schlemm's canal (SC) endothelia, interact in the juxtacanalicular tissue (JCT) region of the conventional outflow tract to regulate outflow resistance. Unlike endothelial cells lining the systemic vasculature, endothelial cells lining the inner wall of SC support a transcellular pressure gradient in the basal to apical direction, thus acting to push the cells off their basal lamina. The resulting biomechanical strain in SC cells is quite large and is likely to be an important determinant of endothelial barrier function, outflow resistance and intraocular pressure. This review summarizes recent work demonstrating how biomechanical properties of SC cells impact glaucoma. SC cells are highly contractile, and such contraction greatly increases cell stiffness. Elevated cell stiffness in glaucoma may reduce the strain experienced by SC cells, decrease the propensity of SC cells to form pores, and thus impair the egress of aqueous humor from the eye. Furthermore, SC cells are sensitive to the stiffness of their local mechanical microenvironment, altering their own cell stiffness and modulating gene expression in response. Significantly, glaucomatous SC cells appear to be hyper-responsive to substrate stiffness. Thus, evidence suggests that targeting the material properties of SC cells will have therapeutic benefits for lowering intraocular pressure in glaucoma.
| S-EPMC4268318 | biostudies-literature