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Translational Vision Science &... Aug 2020To use second harmonic generation imaging and fluorescence recovery after photobleaching to demonstrate alterations in scleral collagen structure and permeability after...
PURPOSE
To use second harmonic generation imaging and fluorescence recovery after photobleaching to demonstrate alterations in scleral collagen structure and permeability after crosslinking in rat and human eyes.
METHODS
Excised rat and human scleras were imaged ex vivo with an inverted two-photon excitation fluorescence microscope before and after photochemical crosslinking using riboflavin and 405-nm laser light. Fluorescence recovery after photobleaching was applied to measure the diffusion of fluorescein isothiocyanate-dextran across the sclera.
RESULTS
Crosslinking caused scleral collagen fibers to become wavier and more densely packed, with surface collagen being more affected than deeper collagen fibers. Crosslinked sclera showed significantly decreased permeability in the irradiation zone and also extended as far as 250 µm outside the irradiation zone.
CONCLUSIONS
Photochemical crosslinking induced changes in scleral structure and permeability that extended to tissue even outside the irradiation zone.
TRANSLATIONAL RELEVANCE
Ultrastructural changes associated with the emerging clinical technique of photochemical scleral crosslinking have not been well characterized. We demonstrate not only changes in scleral collagen by second harmonic generation imaging but also the associated functional changes in tissue permeability by fluorescence recovery after photobleaching. We report the novel finding of reduced permeability extending well beyond the direct irradiation zone. This has implications for control in the clinical setting.
Topics: Animals; Collagen; Humans; Light; Permeability; Rats; Riboflavin; Sclera
PubMed: 32934895
DOI: 10.1167/tvst.9.9.45 -
Current Eye Research Jun 2020: Corneal collagen cross-linking by ultraviolet light activation of riboflavin has been used clinically to enhance corneal stiffness. We sought to determine if...
: Corneal collagen cross-linking by ultraviolet light activation of riboflavin has been used clinically to enhance corneal stiffness. We sought to determine if cross-linking differentially affects scleral regions.: Adjacent, parallel strips of sclera were cut from superolateral, superomedial, inferolateral, and inferomedial quadrants of posterior and equatorial sclera of 12 human cadaver eyes. One of each pair served as control while the other was cross-linked by immersion in 0.1% riboflavin and 365 nm exposure at 6 mW/cm irradiance for 30 min. Behavior of strips was characterized using a microtensile load cell. Preloaded strips were imaged using orthogonally mounted cameras and optical coherence tomography to determine specimen dimensions including cross-sectional area. Tension was measured during 0.1 mm/s constant rate elongation.: Young's modulus (YM), the slope of the relationship relating tensile stress to strain, was calculated at 8% strain, and increased significantly after cross-linking ( < .001). In posterior sclera, mean (± standard error of mean, SEM) YM is increased in the superolateral, superomedial, inferolateral, and inferomedial quadrants by 46 ± 15%, 32 ± 11%, 67 ± 20%, and 53 ± 11%, respectively. In equatorial sclera, YM is increased by 139 ± 43%, 68 ± 27%, 143 ± 92%, and 68 ± 14%, respectively. The YM of pooled equatorial quadrants increased significantly more than that of the pooled posterior quadrants.: Scleral collagen cross-linking by ultraviolet activation of riboflavin differentially increases scleral YM more in the equatorial than posterior sclera, and most in the lateral, equatorial sclera. Cross-linking might be used to arrest progressive myopia or to prevent staphyloma formation.
Topics: Biomechanical Phenomena; Collagen; Cross-Linking Reagents; Elastic Modulus; Humans; Photochemotherapy; Photosensitizing Agents; Riboflavin; Sclera; Tensile Strength; Tomography, Optical Coherence; Ultraviolet Rays
PubMed: 31735063
DOI: 10.1080/02713683.2019.1694157 -
Experimental Eye Research May 2023Collagen is the main load-bearing component of cornea and sclera. When stretched, both of these tissues exhibit a behavior known as collagen fiber recruitment. In...
Collagen is the main load-bearing component of cornea and sclera. When stretched, both of these tissues exhibit a behavior known as collagen fiber recruitment. In recruitment, as the tissues stretch the constitutive collagen fibers lose their natural waviness, progressively straightening. Recruited, straight, fibers bear substantially more mechanical load than non-recruited, wavy, fibers. As such, the process of recruitment underlies the well-established nonlinear macroscopic behavior of the corneoscleral shell. Recruitment has an interesting implication: when recruitment is incomplete, only a fraction of the collagen fibers is actually contributing to bear the loads, with the rest remaining "in reserve". In other words, at a given intraocular pressure (IOP), it is possible that not all the collagen fibers of the cornea and sclera are actually contributing to bear the loads. To the best of our knowledge, the fraction of corneoscleral shell fibers recruited and contributing to bear the load of IOP has not been reported. Our goal was to obtain regionally-resolved estimates of the fraction of corneoscleral collagen fibers recruited and in reserve. We developed a fiber-based microstructural constitutive model that could account for collagen fiber undulations or crimp via their tortuosity. We used experimentally-measured collagen fiber crimp tortuosity distributions in human eyes to derive region-specific nonlinear hyperelastic mechanical properties. We then built a three-dimensional axisymmetric model of the globe, assigning region-specific mechanical properties and regional anisotropy. The model was used to simulate the IOP-induced shell deformation. The model-predicted tissue stretch was then used to quantify collagen recruitment within each shell region. The calculations showed that, at low IOPs, collagen fibers in the posterior equator were recruited the fastest, such that at a physiologic IOP of 15 mmHg, over 90% of fibers were recruited, compared with only a third in the cornea and the peripapillary sclera. The differences in recruitment between regions, in turn, mean that at a physiologic IOP the posterior equator had a fiber reserve of only 10%, whereas the cornea and peripapillary sclera had two thirds. At an elevated IOP of 50 mmHg, collagen fibers in the limbus and the anterior/posterior equator were almost fully recruited, compared with 90% in the cornea and the posterior sclera, and 70% in the peripapillary sclera and the equator. That even at such an elevated IOP not all the fibers were recruited suggests that there are likely other conditions that challenge the corneoscleral tissues even more than IOP. The fraction of fibers recruited may have other potential implications. For example, fibers that are not bearing loads may be more susceptible to enzymatic digestion or remodeling. Similarly, it may be possible to control tissue stiffness through the fraction of recruited fibers without the need to add or remove collagen.
Topics: Humans; Intraocular Pressure; Glaucoma; Extracellular Matrix; Collagen; Tonometry, Ocular; Sclera; Biomechanical Phenomena
PubMed: 36935071
DOI: 10.1016/j.exer.2023.109446 -
PloS One 2022To evaluate corneo-scleral junction (CSJ) using anterior segment optical coherence tomography (AS-OCT) and describe the pattern of cornea and sclera interfusion based on... (Observational Study)
Observational Study
PURPOSE
To evaluate corneo-scleral junction (CSJ) using anterior segment optical coherence tomography (AS-OCT) and describe the pattern of cornea and sclera interfusion based on tissue reflectivity.
METHODS
This prospective observational study enrolled candidates for vision correction. Eyes with previous ocular surgery or irregular corneas were excluded. Temporal and nasal CSJ width and reflectivity patterns were assessed with AS-OCT horizontal scans. Correlations between manual and automated variables and multivariate linear regression analyses with age and spherical equivalent were performed.
RESULTS
101 right eyes were analysed. Temporal CSJ was wider (median 1.62; 1.13 to 2.22 mm) compared to the nasal side (median 1.18; 0.73 to 1.80 mm) (p<.0001). The temporal CSJ width showed negative correlation with ipsilateral anterior chamber angle measurements and positive correlation with horizontal visible iris diameter (HVID). These relationships were not statistically significant for the nasal CSJ width. No significant correlations with age or refractive error were observed at both sides. The pattern of temporal CSJ reflectivity was mostly V- or U-shaped. The eyes with V-shaped temporal CSJ had significantly larger HVID than the eyes with irregular temporal CSJ. The nasal CSJ presented irregular reflectivity in 47% of cases.
CONCLUSIONS
The temporal CSJ was wider and had regular (V or U-shaped) reflectivity patterns, while nasal CSJ was narrower and more irregular. The CSJ width was independent of age and refractive error and could not be predicted from other parameters. The HVID measurement accuracy may benefit from CSJ analysis based on AS-OCT.
Topics: Humans; Tomography, Optical Coherence; Sclera; Cornea; Anterior Chamber; Refraction, Ocular; Refractive Errors
PubMed: 36490278
DOI: 10.1371/journal.pone.0278884 -
PloS One 2018There is extensive knowledge on the relationship of posterior scleral biomechanics and intraocular pressure (IOP) load on glaucomatous optic neuropathy; however, the...
There is extensive knowledge on the relationship of posterior scleral biomechanics and intraocular pressure (IOP) load on glaucomatous optic neuropathy; however, the role for biomechanical influence of the perilimbal scleral tissue on the aqueous humor drainage pathway, including the distal venous outflow system, and IOP regulation is not fully understood. The purpose of this work is to study the outflow characteristics of perfused porcine eyes relative to the biomechanical properties of the perilimbal sclera, the posterior sclera and the cornea. Enucleated porcine eyes from eleven different animals were perfused with surrogate aqueous at two fixed flow rates while monitoring their IOP. After perfusion, mechanical stress-strain and relaxation tests were conducted on specimens of perilimbal sclera, posterior sclera, and cornea from the same perfused eyes. Statistical analysis of the data demonstrated a strong correlation between increased tangent modulus of the perilimbal sclera tissues and increased perfusion IOP (R2 = 0.74, p = 0.0006 at lower flow rate and R2 = 0.71, p = 0.0011 at higher flow rate). In contrast, there were no significant correlations between IOP and the tangent modulus of the other tissues (Posterior sclera: R2 = 0.17 at lower flow rate and R2 = 0.30 at higher flow rate; cornea: R2 = 0.02 at lower flow rate and R2<0.01 at higher flow rate) nor the viscoelastic properties of any tissue (R2 ≤ 0.08 in all cases). Additionally, the correlation occurred for IOP and not net outflow facility (R2 ≤ 0.12 in all cases). These results provide new evidence that IOP in perfused porcine eyes is strongly influenced by the tangent modulus, sometimes called the tissue stiffness, of the most anterior portion of the sclera, i.e. the limbus.
Topics: Animals; Biomechanical Phenomena; Elasticity; Intraocular Pressure; Materials Testing; Mechanical Phenomena; Sclera; Swine; Viscosity
PubMed: 29718942
DOI: 10.1371/journal.pone.0195882 -
American Journal of Ophthalmology Oct 2021To optimize the flanged belt-loop technique of scleral fixation through biomechanical testing and report clinical outcomes of resultant modifications.
OBJECTIVE
To optimize the flanged belt-loop technique of scleral fixation through biomechanical testing and report clinical outcomes of resultant modifications.
DESIGN
Experimental study.
METHODS
The force to disinsert flanged polypropylene suture from human cadaveric sclera was assessed using a tensile testing machine and compared to the breaking strengths of 9-0 and 10-0 polypropylene. The effects of modifying suture gauge (5-0, 6-0, 7-0, or 8-0), amount of suture cauterized (0.5 or 1.0 mm), and sclerotomy size (27, 30, 32, 33 gauge) were investigated. Belt-loop intrascleral fixation using 6-0 and 7-0 polypropylene with 30 and 32 gauge needles, respectively, was performed in 4 patients. Main outcome measures were flanged suture disinsertion forces in cadaveric sclera.
RESULTS
The average force to disinsert a flange created by melting 1.0 mm of 5-0, 6-0, 7-0, and 8-0 polypropylene suture from human cadaveric sclera via 27, 30, 32, and 33 gauge needle sclerotomies was 3.0 ± 0.5 N, 2.1 ± 0.3 N, 0.9 ± 0.2 N, and 0.4 ± 0.1 N, respectively. The disinsertion forces for flanges formed by melting 0.5 mm of the same gauges were 72%-79% lower (P < .001). In comparison, the breaking strengths of 9-0 and 10-0 polypropylene were 0.91 ± 0.4 N and 0.52 ± 0.03 N. Belt-loop fixation using 6-0 and 7-0 polypropylene with 30 and 32 gauge sclerotomies demonstrated good outcomes at 6 months.
CONCLUSIONS
The flanged belt-loop technique is a biomechanically sound method of scleral fixation using 1.0 mm flanges of 5-0 to 7-0 polypropylene paired with 27, 30, and 32 gauge sclerotomies. In contrast, 8-0 polypropylene and 0.5 mm flanges of any suture gauge will likely be unstable with this technique.
Topics: Humans; Lens Implantation, Intraocular; Lenses, Intraocular; Polypropylenes; Sclera; Suture Techniques; Sutures
PubMed: 33945819
DOI: 10.1016/j.ajo.2021.04.017 -
ELife Mar 2022Hallmark social activities of humans, such as cooperation and cultural learning, involve eye-gaze signaling through joint attentional interaction and ostensive...
Hallmark social activities of humans, such as cooperation and cultural learning, involve eye-gaze signaling through joint attentional interaction and ostensive communication. The gaze-signaling and related cooperative-eye hypotheses posit that humans evolved unique external eye morphologies, including uniformly white sclera (the whites of the eye), to enhance the visibility of eye-gaze for conspecifics. However, experimental evidence is still lacking. This study tested the ability of human and chimpanzee participants to discriminate the eye-gaze directions of human and chimpanzee images in computerized tasks. We varied the level of brightness and size in the stimulus images to examine the robustness of the eye-gaze directional signal against simulated shading and distancing. We found that both humans and chimpanzees discriminated eye-gaze directions of humans better than those of chimpanzees, particularly in visually challenging conditions. Also, participants of both species discriminated the eye-gaze directions of chimpanzees better when the contrast polarity of the chimpanzee eye was reversed compared to when it was normal; namely, when the chimpanzee eye has human-like white sclera and a darker iris. Uniform whiteness in the sclera thus facilitates the visibility of eye-gaze direction even across species. Our findings thus support but also critically update the central premises of the gaze-signaling hypothesis.
Topics: Animals; Eye Movements; Fixation, Ocular; Hominidae; Humans; Pan troglodytes; Sclera
PubMed: 35256053
DOI: 10.7554/eLife.74086 -
Journal of the Royal Society, Interface Apr 2015Residual deformations strongly influence the local biomechanical environment in a number of connective tissues. The sclera is known to be biomechanically important in...
Residual deformations strongly influence the local biomechanical environment in a number of connective tissues. The sclera is known to be biomechanically important in healthy and diseased eyes, such as in glaucoma. Here, we study the residual deformations of the sclera, as well as the adjacent choroid and retina. Using freshly harvested porcine eyes, we developed two approaches of quantifying residual deformations in the spherically shaped tissues of interest. The first consisted of punching discs from the posterior wall of the eye and quantifying the changes in the area and eccentricity of these samples. The second consisted of cutting a ring from the equatorial sclera and making stress-relieving cuts in it. Measurements of curvature were made before and after the stress-relieving cuts. Using the first approach, we observed a 42% areal contraction of the choroid, but only modest contractions of the sclera and retina. The observed contractions were asymmetric. In the second approach, we observed an opening of the scleral rings (approx. 10% decrease in curvature). We conclude that residual bending deformations are present in the sclera, which we speculate may be due to radially heterogeneous growth and remodelling of the tissue during normal development. Further, residual areal deformations present in the choroid may be due to the network of elastic fibres in this tissue and residual deformations in the constituent vascular bed. Future studies of ocular biomechanics should attempt to include effects of these residual deformations into mechanical models in order to gain a better understanding of the biomechanics of the ocular wall.
Topics: Analysis of Variance; Animals; Biomechanical Phenomena; Choroid; Eye Diseases; Female; Ocular Physiological Phenomena; Retina; Sclera; Swine
PubMed: 25740853
DOI: 10.1098/rsif.2014.1101 -
Experimental Eye Research May 2020Scleral fibroblast activation occurs in glaucomatous and myopic eyes. Here we perform an unbiased screen to identify kinase inhibitors that reduce fibroblast activation...
Scleral fibroblast activation occurs in glaucomatous and myopic eyes. Here we perform an unbiased screen to identify kinase inhibitors that reduce fibroblast activation to diverse stimuli in vitro and to in vivo intraocular pressure (IOP) elevation. Primary cultures of peripapillary scleral (PPS) fibroblasts from two human donors were screened using a library of 80 kinase inhibitors to identify compounds that inhibit TGFβ-induced extracellular matrix (ECM) synthesis. Inhibition of myofibroblast differentiation was verified by alpha smooth muscle actin (αSMA) immunoblot and collagen contraction assay. Inhibition of IOP-induced scleral fibroblast proliferation was assessed by ELISA assay for proliferating cell nuclear antigen (PCNA). The initial screen identified 7 inhibitors as showing>80% reduction in ECM binding. Three kinase inhibitors were verified to reduce TGFβ-induced αSMA expression and cellular contractility (rottlerin, PP2, tyrphostin 9). The effect of three Src inhibitors, bosutinib, dasatinib, and SU-6656, on myofibroblast differentiation was evaluated, with only dasatinib significantly inhibiting TGFβ-induced ECM synthesis, αSMA expression, and cellular contractility at nanomolar dosages. Subconjunctival injection of dasatinib reduced IOP-induced scleral fibroblast proliferation compared to control (4.9 ± 11.1 ng/sclera with 0.1 μM versus 88.7 ± 38.6 ng/sclera in control, P < 0.0001). Dasatinib inhibits scleral myofibroblast differentiation and there is pharmacologic evidence that this inhibition is not solely due to Src-kinase inhibition.
Topics: Aged; Aged, 80 and over; Animals; Cell Differentiation; Cells, Cultured; Dasatinib; Disease Models, Animal; Female; Glaucoma; Humans; Male; Mice; Myofibroblasts; Protein Kinase Inhibitors; Sclera; Signal Transduction
PubMed: 32179077
DOI: 10.1016/j.exer.2020.107999 -
Biomedicine & Pharmacotherapy =... Dec 2018The aim of this study was to investigate the effects of pilocarpine-induced ciliary body constant contraction for a long time period on the elastic modulus and collagen...
The aim of this study was to investigate the effects of pilocarpine-induced ciliary body constant contraction for a long time period on the elastic modulus and collagen in corneal and scleral tissues in the early developmental stage. Twelve one-month-old New Zealand white rabbits were randomly monocularly treated with pilocarpine to cause ciliary body constant contraction. After 1- and 2-months, the sclera tissues and the eyeballs were obtained to assess the cornea and three regions of the sclera-anterior, equatorial and posterior. The corneal tissues and the three regions of the scleral tissues were cut into strips for elastic modulus measurement using an Instron 5544, the size distribution of collagen fibrils was examined using electron microscopy, the samples were homogenized, and the concentration of hydroxyproline was measured to determine the collagen content. For corneal tissues, long-term pilocarpine-induced ciliary body constant contraction did not affect the elastic modulus and collagen. For scleral tissues, long-term pilocarpine-induced ciliary body constant contraction affected the elastic modulus, diameter of collagen fibrils and collagen content in the equatorial and posterior sclera, however, the anterior sclera were unaffected. These results suggested that pilocarpine-induced ciliary body contraction could affect the scleral structure and lead to deformation of the sclera and eyeball, thereby affecting visual functions function in the process of ocular emmetropization.
Topics: Animals; Ciliary Body; Collagen; Cornea; Elastic Modulus; Hydroxyproline; Pilocarpine; Rabbits; Sclera
PubMed: 30372887
DOI: 10.1016/j.biopha.2018.10.032