Reduced Scleral TIMP-2 Expression Is Associated With Myopia Development: TIMP-2 Supplementation Stabilizes Scleral Biomarkers of Myopia and Limits Myopia Development.

Purpose: The purpose of this study was to determine the endogenous regulation pattern of tissue inhibitor of metalloproteinase-2 (TIMP-2) in the tree shrew sclera during myopia development and investigate the capacity of exogenous TIMP-2 to inhibit matrix metalloproteinase-2 (MMP-2) in vitro and both scleral collagen degradation and myopia development in vivo. Methods: TIMP-2 expression in the sclera during myopia development was assessed using polymerase chain reaction. In vitro TIMP-2 inhibition of MMP-2 was investigated using a gelatinase activity plate assay and zymography. Tree shrews were injected with a collagen precursor before undergoing monocular form deprivation and concurrent daily subconjunctival injections of either TIMP-2 or vehicle to the form-deprived eye. In vivo ocular biometry changes were monitored, and scleral tissue was collected after 12 days and assayed for collagen degradation. Results: The development of myopia was associated with a mean reduction in TIMP-2 mRNA expression after 5 days of form deprivation (P < 0.01). Both activation and activity of MMP-2 were inhibited by TIMP-2 with an IC50 of 10 to 20 and 2 nM, respectively. In vivo exogenous addition of TIMP-2 significantly reduced myopia development (P < 0.01), due to reduced vitreous chamber elongation (P < 0.01). In vivo TIMP-2 treatment also significantly inhibited posterior scleral collagen degradation relative to vehicle-treated eyes (P < 0.01), with levels similar to those in control eyes. Conclusions: Myopia development in mammals is associated with reduced expression of TIMP-2, which contributes to increased degradative activity in the sclera. It follows that replenishment of this TIMP-2 significantly reduced the rate of both scleral collagen degradation and myopia development.

How does atropine exert its anti-myopia effects?

In the following point-counterpoint article, internationally-acclaimed myopia researchers were challenged to defend the two opposing sides of the topic defined by the title; their contributions, which appear in the order, Point followed by Counterpoint, were peer-reviewed by both the editorial team and an external reviewer. Independently of the invited authors, the named member of the editorial team provided an Introduction and Summary, both of which were reviewed by the other members of the editorial team. By their nature, views expressed in each section of the Point-Counterpoint article are those of the author concerned and may not reflect the views of all of the authors.

Eyes in various species can shorten to compensate for myopic defocus.

PURPOSE: We demonstrated that eyes of young animals of various species (chick, tree shrew, marmoset, and rhesus macaque) can shorten in the axial dimension in response to myopic defocus. METHODS: Chicks wore positive or negative lenses over one eye for 3 days. Tree shrews were measured during recovery from induced myopia after 5 days of monocular deprivation for 1 to 9 days. Marmosets were measured during recovery from induced myopia after monocular deprivation, or wearing negative lenses over one or both eyes, or from wearing positive lenses over one or both eyes. Rhesus macaques were measured after recovery from induced myopia after monocular deprivation, or wearing negative lenses over one or both eyes. Axial length was measured with ultrasound biometry in all species. RESULTS: Tree shrew eyes showed a strong trend to shorten axially to compensate for myopic defocus. Of 34 eyes that recovered from deprivation-induced myopia for various durations, 30 eyes (88%) shortened, whereas only 7 fellow eyes shortened. In chicks, eyes wearing positive lenses reduced their rate of ocular elongation by two-thirds, including 38.5% of eyes in which the axial length became shorter than before. Evidence of axial shortening in rhesus macaque (40%) and marmoset (6%) eyes also occurred when exposed to myopic defocus, although much less frequently than that in eyes of tree shrews. The axial shortening was caused mostly by the reduction in vitreous chamber depth. CONCLUSIONS: Eyes of chick, tree shrew, marmoset, and rhesus macaque can shorten axially when presented with myopic defocus, whether the myopic defocus is created by wearing positive lenses, or is the result of axial elongation of the eye produced by prior negative lens wear or deprivation. This eye shortening facilitates compensation for the imposed myopia. Implications for human myopia control are significant.

Regulation of scleral metabolism in myopia and the role of transforming growth factor-beta.

Myopia is one of the most prevalent ocular conditions and is the result of a mismatch between the power of the eye and axial length of the eye. In the vast majority of cases the structural cause of myopia is an excessive axial length of the eye, or more specifically the vitreous chamber depth. In about 3% of the general population in Europe, USA and Australia, the degree of myopia is above 6 dioptres and is termed high myopia. In South East Asia the figure is closer to 20% of the general population with high myopia. The prevalence of sight threatening ocular pathology is markedly increased in eyes with high degrees of myopia (>-6 D). This results from the excessive axial elongation of the eye which, by necessity, must involve the outer coat of the eye, the sclera. Current theories of refractive development acknowledge the pivotal role of the sclera in the control of eye size and the development of myopia. This review details the major structural, biochemical and biomechanical changes that underlie abnormal development of the mammalian sclera in myopia. In describing the changes in regulation of sclera metabolism in myopia, the pivotal role of transforming growth factor-ß signalling is highlighted as the responsible factor for certain critical events in myopia development that ultimately result in the scleral pathology observed in high myopia.

Muscarinic antagonist control of myopia: evidence for M4 and M1 receptor-based pathways in the inhibition of experimentally-induced axial myopia in the tree shrew.

PURPOSE: The broadband muscarinic antagonist atropine is effective in stopping the progression of myopia in animals and humans. The partially selective M(1)/M(4) antagonist pirenzepine also slows progression of myopia, although not as effectively as atropine. Due to the supra maximal doses utilized in these studies, it is unclear if this antimyopia effect occurs through a receptoral-based mechanism, and if so, which receptors are involved. Studies in chicks indicate the involvement of the M(4) muscarinic receptor. The current study investigated the effect of the highly selective muscarinic antagonists Muscarinic Toxin 3 (MT3) (M(4) selective) and Muscarinic Toxin 7 (MT7) (M(1) selective) on experimental myopia in a mammalian model. METHODS: Tree shrews (n = 23) underwent daily intravitreal injections of MT3, MT7, or vehicle (phosphate buffered saline) for five days in the treated eye, combined with deprivation of vision with a translucent occluder (MD). The contralateral eye was unocccluded and underwent intravitreal injections of vehicle for the same period. Two additional groups (n = 10) underwent daily intravitreal injections of MT7 or vehicle for 10 days in the treated eye combined with negative lens (-9.5 diopter [D]) defocus (LIM). The control eye was injected with saline and wore a plano lens. RESULTS: Both MT3 and MT7 treatment reduced the development of deprivation-induced myopia (treated-control eye [T-C]; vehicle-MD; -4.3 ± 0.6 D versus MT3-MD; -0.7 ± 0.2 D and MT7-MD; -0.7 ± 0.4 D; P < 0.001). MT7 treatment was effective at inhibiting lens-induced myopia (T-C; vehicle-LIM; -4.6 ± 0.5 D versus MT7-LIM; 0.2 ± 0.2 D; P < 0.05). CONCLUSIONS: The findings demonstrate that inhibition of form-deprivation myopia by muscarinic antagonists involves both M(4) and M(1) muscarinic receptor signaling pathways in mammals.

The effect of daily transient +4 D positive lens wear on the inhibition of myopia in the tree shrew.

PURPOSE: Negative-lens-induced defocus causes accelerated ocular elongation and myopia, whereas positive-lens-induced defocus produces reduced ocular elongation and hyperopia. Short durations of positive lens wear result in markedly stronger temporal effects than do short periods of negative lens wear in the chick model of refractive development. In mammalian and nonhuman primate models, there have been equivocal results in inhibiting myopia by short periods of positive lens wear when compared with data from the chick model. The purpose of the present study was an evaluation of full-time -9.5 D negative lens wear interrupted by short periods of daily +4 D positive lens wear in preventing experimental myopia in the tree shrew. METHODS: One treatment group wore negative lenses (-9.5 D) binocularly for 23 hours a day (10 hours of which were spent in total darkness), interrupted by 1 hour of wearing positive lenses (+4 D) binocularly for 12 days. Another group of animals wore negative lenses (-9.5 D) binocularly for 23 hours a day, interrupted by two 30-minute periods of positive lens (+4 D) wear daily, again for 12 days. The animals were raised on a 14-hour/10-hour light-dark cycle. Animals wearing -9.5 D lenses binocularly, interrupted by 0-powered lenses for either 1 hour or two 30-minute periods daily for 12 days, acted as controls. RESULTS: Continuous wear of -9.5 D lenses binocularly induced a -10.8 D myopic shift in refraction. Full-time wear of -9.5 D lenses binocularly, interrupted by 1 hour of 0-power lens wear binocularly, caused a myopic shift of 3.6 D over 12 days, whereas wearing -9.5 D lenses, interrupted by 1 hour every day of +4.0 D lens wear binocularly, whether it was continuous or divided into two 30-minute periods, caused a myopic shift of only 0.7 D over 12 days. CONCLUSIONS: Daily intermittent +4 D positive lens wear effectively inhibits experimentally induced myopia and may prove a viable approach for preventing myopia progression in children.

The M4 muscarinic antagonist MT-3 inhibits myopia in chick: evidence for site of action.

PURPOSE: It is well established that the broad-band muscarinic antagonist, atropine is effective at inhibiting the progression of myopia and does so by preventing the elongation of the vitreous chamber of the eye. However, uncertainty remains as to whether this effect occurs through a receptoral mechanism and, if so, which muscarinic receptor subtype mediates this effect. Previous work, in avian and mammalian models of myopia, implicates the M1 and M4 receptors as potential targets. The current study used physiologically relevant concentrations of highly selective muscarinic antagonists (MT-3 and MT-7) to further characterise the role of the M4 receptor in the control of myopia in the chick model of refractive development. METHODS: Nine groups of week-old chicks underwent 5 days of monocular deprivation, with a translucent occluder, to induce myopia. These animals had either no injection, scleral puncture with a needle, or daily intravitreal injections of MT-3 (M4-selective), MT-7 (M1-selective) or vehicle. Three concentrations of each antagonist were delivered (250 nm, 2.5 µm and 10 µm). After the treatment period, keratometry, retinoscopy and A-Scan ultrasound were used to assess ocular biometry. RESULTS: MT-3 treatment produced a significant dose-dependent reduction in relative myopia (treated-control eye) compared to vehicle treatment (vehicle -10.1 ± 1.1 D vs 10 µm MT-3 -4.0 ± 1.5 D, p < 0.01). The majority of this effect was due to reduced relative vitreous chamber elongation in drug treated eyes (vehicle +0.26 ± 0.04 mm, 10 µm MT-3 +0.08 ± 0.07 mm, p < 0.05). In contrast, MT-7 had no significant effect on the development of myopia (MT-7 10 µm: myopia, -12.1 ± 0.8 D and vitreous chamber depth, +0.23 ± 0.07 mm). Calculations indicate that the experimentally achieved concentrations of MT-3 at intraocular receptors necessary to inhibit 50% of myopia development (between 5 and 50 nm) were consistent with published in vitro affinity constants for the M4 receptor and below those for the M1 receptor. Histology demonstrated that MT-3 at the doses used had no gross effects on the retina, indicating a non-toxic mode of action. CONCLUSIONS: In the chick, which lacks a homologue of the mammalian M1 receptor, the above findings represent compelling evidence that muscarinic antagonists prevent myopia progression through an M4-receptor mediated mechanism, most likely located in the retina.

Retinal thinning in tree shrews with induced high myopia: optical coherence tomography and histological assessment.

This study determined retinal thinning in a mammalian model of high myopia using optical coherence tomography (OCT) and histological sections from the same retinal tissue. High myopia was induced in three tree shrews (Tupaia belangeri) by deprivation of form vision via lid suture of one eye, with the other eye a control. Ocular biometry data was obtained by Ascan ultrasonography, keratometry and retinoscopy. The Zeiss StratusOCT was used to obtain Bscans in vivo across the retina. Subsequently, eyes were enucleated and retinas fixed, dehydrated, embedded and sectioned. Treated eyes developed a high degree of axial myopia (-15.9 ± 2.3D; n = 3). The OCT analysis showed that in myopic eyes the nasal retina thinned more than the temporal retina relative to the disc (p=0.005). Histology showed that the retinas in the myopic eyes comprise all layers but were thinner than the retinas in normal and control eyes. Detailed thickness measurements in corresponding locations of myopic and control eyes in superior nasal retina using longitudinal reflectivity profiles from OCT and semithin vertical histological sections showed the percentage of retinal thinning in the myopic eyes was similar between methods (OCT 15.34 ± 5.69%; histology 17.61 ± 3.02%; p = 0.10). Analysis of retinal layers revealed that the inner plexiform, inner nuclear and outer plexiform layers thin the most. Cell density measurements showed all neuronal cell types are involved in retinal thinning. The results indicate that in vivo OCT measurements can accurately detect retinal thinning in high myopia.

The effect of pirenzepine on positive- and negative-lens-induced refractive error and ocular growth in chicks.

PURPOSE: The selective muscarinic antagonist pirenzepine inhibits experimentally induced myopia in avian and mammalian species, including nonhuman primates and adolescent humans. Transient positive lens defocus has a potent inhibitory effect on negative-lens-induced myopia in avian and mammalian models. The purpose of the present study was to determine the influence of daily treatment with pirenzepine on ocular growth and refractive error in chicks wearing positive lenses. METHODS: The chicks were allocated to one of eight groups (n = 6 each group) on the basis of whether they wore +10 or -10 D lenses monocularly and whether they received daily intravitreal injections of pirenzepine (700 µg) or vehicle (phosphate-buffered saline) in the lens-defocused eye. In vivo refractive and biometric data were collected, and glycosaminoglycan synthesis in the sclera was assessed. RESULTS: Pirenzepine did not alter the level of positive-lens-induced hyperopia in chicks wearing +10 D lenses compared with that in the vehicle control group (+8.1 ± 0.6 D vs. +8.9 ± 2.4 D, mean ± SEM; P = 0.76). In contrast, pirenzepine caused significant inhibition of negative-lens-induced myopia compared with that in the vehicle group (-1.1 ± 1.5 D vs. -8.8 ± 1.1 D; P = 0.001). Glycosaminoglycan synthesis in the posterior sclera was significantly increased in the negative-lens-treated groups and showed small decreases in the positive-lens-treated groups. CONCLUSIONS: The influence of pirenzepine on ocular growth in chicks differed by sign of lens defocus, with pirenzepine blocking negative-lens effects on ocular growth, but not positive-lens effects. The most likely reason that hyperopia was not enhanced by pirenzepine treatment was that the rapid compensatory eye growth associated with positive lenses eliminated the imposed myopic defocus, and the clear retinal image prevented any additional hyperopia from developing.

Inhibition of matrix metalloproteinase activity in the chick sclera and its effect on myopia development.

PURPOSE: To investigate the contribution of matrix degradation in the two-layer avian sclera to the development of myopia. METHODS: Tissue inhibitor of metalloproteinase-2 (TIMP-2) was used to inhibit chick scleral collagen degradation with (3)H-proline, a marker for this effect. Ex vivo scleral culture experiments confirmed TIMP-2 doses for in vivo experimentation. Ocular growth and refractive response to exogenous TIMP-2 (11.25, 2.25, and 0.45 picomoles, plus vehicle only) were monitored in 7-day-old chicks during the induction of myopia over 4 days with a translucent occluder. Collagen degradation was assessed, in whole sclera and in separated scleral layers by using the same paradigm (11.25 picomoles TIMP-2; vehicle only). RESULTS: Approximately 60% of collagen degradation was inhibited with low (2 nM) doses of TIMP-2 in the ex vivo sclera. Degradative activity in the in vivo chick sclera increased significantly (46%) during myopia development, with all the altered activity confined to the fibrous layer. Addition of TIMP-2 significantly reduced (by 46%) this accelerated scleral collagen degradation, also by acting in the fibrous layer. TIMP-2 had no significant effect on (3)H-proline incorporated in the cartilaginous scleral layer and cornea. Despite inhibiting collagen degradation TIMP-2 had no significant effect on myopia development. CONCLUSIONS: Increased collagen degradation is a feature of scleral remodeling in chick myopia development, but is confined to the fibrous scleral layer. Significant inhibition of this collagenolytic activity with TIMP-2 has little effect on refractive error development, suggesting that collagen degradation in the sclera contributes little to the development of myopia in the chick.

Effects of a head-mounted display on the oculomotor system of children.

PURPOSE: This study evaluated the effects of short term and extended viewing of virtual imagery using the Binocular Viewer (new generation bi-ocular viewer) on the visual system of children, and compared these effects with that of viewing a high definition television (HDTV) display. METHODS: Sixty children aged 5 to 16 years viewed 30 min of virtual imagery using the Binocular Viewer and a HDTV display on two occasions. Sixteen subjects, aged 13 to 16 years, completed a third session of extended viewing (80 min) with the Binocular Viewer. Oculomotor function and symptoms were assessed previewing, immediately postviewing, and 10 min postviewing. RESULTS: Thirty minutes of Binocular Viewer use resulted in symptom increases (p < 0.05) immediately postviewing ("feeling tired," "feeling sleepy," "difficulty concentrating," and "sore/aching eyes") however most symptoms had dissipated by 10-min postviewing. There were no significant symptom differences between viewing with the Binocular Viewer and the HDTV display at either time point. An increase in symptoms (p < 0.05) immediately postviewing was recorded after 80 min of Binocular Viewer use ("feeling tired," "feeling bored," "feeling sleepy," and "tired eyes"), however only "feeling tired" and "feeling bored" remained significantly increased (p < 0.05) 10-min postviewing. Near unaided visual acuity demonstrated a significant and consistent reduction immediately (p < 0.01) and at 10 min (p < 0.05) following 30 min of Binocular Viewer use and immediately following 80 min of use (p < 0.01). Near unaided VA was also significantly reduced (p < 0.01) immediately after 30 min of HDTV display use. CONCLUSIONS: Virtual imagery viewing with the Binocular Viewer in children aged 5 to 16 years had few additional adverse effects when compared to viewing a more conventional HDTV display. The Binocular Viewer was comfortable to wear for up to 80 min of viewing. The consistent reduction in near vision for both viewing durations with the Binocular Viewer requires further investigation.

Developmental eye movement test: what is it really measuring?

PURPOSE: The Developmental Eye Movement (DEM) test is commonly used as a clinical visual-verbal ocular motor assessment tool. However, while the DEM test ratio has been reported to correlate with horizontal saccadic eye movements, there have been no published comparative studies of the DEM test and objective eye movement measures. The aim of this study was to compare DEM test performance with explicit quantification of saccadic eye movements, reading performance, symptomatology and visual processing speed, to assess the validity of the DEM test in clinical practice. METHODS: One hundred fifty-eight children aged 8 to 11 years completed the DEM test and a battery of eye movement tasks, recorded by a Microguide 1000 infrared eye tracker. All subjects completed a symptomatology survey. Reading performance and visual processing data was collected for 77 and 75 children, respectively. RESULTS: One hundred twenty-nine of the 158 subjects (81.65%) passed the DEM test. There was no significant correlation between any component of DEM test performance and quantitative eye movement parameters (gain, latency, asymptotic peak velocity, and number of corrective saccades) or symptomatology. There were significant correlations between DEM test outcome and reading performance, and with visual processing speed. CONCLUSIONS: DEM test performance does not correlate with saccadic eye movement skills or symptomatology. However, it is related to reading performance and visual processing speed. This study suggests that although DEM test times may not correlate directly with eye movement parameters, they do correlate with aspects of reading performance and thus may serve a diagnostic role in clinical practice.

Regulation of scleral cell contraction by transforming growth factor-beta and stress: competing roles in myopic eye growth.

Reduced extracellular matrix accumulation in the sclera of myopic eyes leads to increased ocular extensibility and is related to reduced levels of scleral transforming growth factor-beta (TGF-beta). The current study investigated the impact of this extracellular environment on scleral cell phenotype and cellular biomechanical characteristics. Scleral cell phenotype was investigated in vivo in a mammalian model of myopia using the myofibroblast marker, alpha-smooth muscle actin (alpha-SMA). In eyes developing myopia alpha-SMA levels were increased, suggesting increased numbers of contractile myofibroblasts, and decreased in eyes recovering from myopia. To understand the factors regulating this change in scleral phenotype, the competing roles of TGF-beta and mechanical stress were investigated in scleral cells cultured in three-dimensional collagen gels. All three mammalian isoforms of TGF-beta altered scleral cell phenotype to produce highly contractile, alpha-SMA-expressing myofibroblasts (TGF-beta3>TGF-beta2>TGF-beta1). Exposure of cells to the reduced levels of TGF-beta found in the sclera in myopia produced decreased cell-mediated contraction and reduced alpha-SMA expression. These findings are contrary to the in vivo gene expression data. However, when cells were exposed to both the increased stress and the reduced levels of TGF-beta found in myopia, increased alpha-SMA expression was observed, replicating in vivo findings. These results show that although reduced scleral TGF-beta is a major contributor to the extracellular matrix remodeling in the myopic eye, it is the resulting increase in scleral stress that dominates the competing TGF-beta effect, inducing increased alpha-SMA expression and, hence, producing a larger population of contractile cells in the myopic eye.

Relationship of the optical coherence tomography signal to underlying retinal histology in the tree shrew (Tupaia belangeri).

PURPOSE: To interpret the retinal origin of the optical coherence tomography (OCT) signal by objectively (i.e., minimal investigator bias) aligning in vivo OCT longitudinal reflectivity profiles (LRPs) with corresponding vertical histologic sections. METHODS: The Zeiss StratusOCT system was used to obtain retinal B-scans in vivo in eyes from adult tree shrews. Subsequently, the retinas were fixed and embedded. Semithin vertical sections through the retina were obtained from the same locations as the LRPs. A statistical correlation procedure that accounted for axial tissue shrinkage determined the best relationship between features in the LRP and sublaminae boundaries in corresponding histology sections. RESULTS: For the optimal relationship, the three regions of high reflectivity in the inner OCT signal corresponded to (1) the nerve fiber and ganglion cell layers, (2) the inner plexiform layer and amacrine cell somas, and (3) the outer plexiform layer. The two regions of low reflectivity in the inner OCT signal corresponded to (1) the somas of Müller, bipolar, and horizontal cells in the inner nuclear layer and (2) the outer nuclear layer. The outer OCT signal had a region of high reflectivity that corresponded to the photoreceptor inner and outer segments, the pigment epithelium, Bruch's membrane, and at least part of the choriocapillaris. CONCLUSIONS: These results provide a clear interpretation for the OCT signal in terms of the underlying retinal anatomy. This interpretation can be used in vivo to identify sublaminae affected by retinal disease and has implications for the origin of the inner OCT signal in human retina.

Biomechanics of the sclera in myopia: extracellular and cellular factors.

PURPOSE: Excessive axial elongation of the eye is the principal structural cause of myopia. The increase in eye size results from active remodelling of the sclera, producing a weakened scleral matrix. The present study will detail the biomechanics of the sclera and highlight the matrix and cellular factors important in the control of eye size. METHODS: Scleral elasticity (load vs. tissue extension) and creep rate (tissue extension vs. time) have been measured postmortem in human eyes. Animal models of myopia have allowed the direct relevance of scleral biomechanics to be investigated during myopia development. Recently, data on tissue matrices incorporating scleral fibroblasts have highlighted the role of cellular contraction in scleral biomechanics. RESULTS: Scleral elasticity is increased in eyes developing myopia, with a reduction in the failure load of the tissue. Scleral creep rate is increased in the sclera from eyes developing myopia, and reduced in eyes recovering from myopia. These changes in biomechanical properties of the sclera occur early in the development of myopia (within 24 h). Alterations in scleral biomechanics during myopia development have been attributed to changes in matrix constituents, principally reduced collagen content. Although the biochemical structure of the sclera plays a critical role in defining the mechanical properties, recent studies investigating the cellular mechanics of the sclera, implicate myofibroblasts in scleral biomechanics. Scleral myofibroblasts have the capacity to contract the matrix and are regulated by tissue stress and growth factors such as transforming growth factor-beta. Changes in these regulatory factors have been observed during myopia development, implicating cellular factors in the resultant weakened sclera. CONCLUSIONS: Changes in the biomechanical properties of the sclera are important in facilitating the increase in axial length that results in myopia. Understanding the matrix and cellular factors contributing to the weakened sclera may aid in the development of a clinically appropriate treatment for myopia.

The effect of positive lens defocus on ocular growth and emmetropization in the tree shrew.

Optical defocus influences postnatal ocular development in animal models. Induced negative lens defocus results in accelerated ocular elongation and myopia. Positive lens-induced defocus findings across animal models have been inconsistent. Specifically, in the tree shrew, positive lens-induced defocus has produced equivocal results. This study evaluated the response of the tree shrew to induced positive lens defocus. One treatment group wore positive lenses binocularly, which were increased in power sequentially from +2 to +4, +6, +8, and +9.5 D over 8 weeks. Other groups wore +4, +6, and +9.5 D lenses, respectively, for 8 weeks. Animals wearing zero-powered (plano) lenses served as controls. Refractive error and ocular dimensions were measured at the start of treatment and every week thereafter. Sequentially increasing positive lens power caused a relative hyperopia of +5.6 D (p < 0.01) compared to the plano lens group (+10.9 +/- 1.8 D vs +5.3 +/- 0.5 D). Constant +4 D lens wear produced +6.9 D relative hyperopia, while +6 and +9.5 D lens wear did not induce hyperopia. Lens-induced defocus changes in refractive state were significantly correlated with vitreous chamber depth changes. The threshold for consistent responses to positive lens defocus in tree shrew was between +4 and +6 D. The results will enable targeted investigation of the efficacy of positive lens defocus in inhibiting myopic ocular growth.

Expression of collagen-binding integrin receptors in the mammalian sclera and their regulation during the development of myopia.

PURPOSE: The sclera has a collagen-rich extracellular matrix that undergoes significant biochemical and biomechanical remodeling during myopic eye growth. The integrin family of cell surface receptors play critical roles in extracellular matrix and biomechanical remodeling in connective tissues. This study identified the major collagen-binding integrin receptors in the mammalian sclera and investigated their mRNA expression during the development of and recovery from experimental myopia. METHODS: The presence of the alpha1, alpha2, and beta1 integrin subunits was examined by using tree-shrew-specific primers and RT-PCR. Scleral expression of alpha1beta1 and alpha2beta1 receptor proteins was further investigated by using Western blot analysis and immunocytochemistry. Myopia was induced monocularly by occluding pattern vision and scleral tissue collected after 24 hours and 5 days. In a subset of the 5-day treatment group, vision was restored for 24 hours before tissue was isolated. Total RNA was extracted, and integrin subunit expression levels were assessed with quantitative real-time PCR. RESULTS: The presence of the major collagen-binding integrin subunits alpha1, alpha2, and beta1 was confirmed by RT-PCR in both scleral tissue and cultured scleral fibroblasts. Both the alpha1 and alpha2 integrin subunit proteins were identified in tree shrew scleral tissues, and integrin receptor expression was localized to scleral fibroblast focal adhesions. After only 24 hours of myopia induction, a time when no structural elongation has occurred, significant decreases were observed in the expression of the alpha1 (-36%) and beta1 (-44%) integrin subunits. After 5 days of myopia induction, alpha1 integrin expression had returned to baseline levels, whereas the alpha2 subunit showed a significant decrease in expression (-52%). The 5-day integrin profiles were maintained during recovery from the induced myopia, with only alpha2 integrin showing a statistically significant relative decrease in expression (-41%). CONCLUSIONS: The mammalian sclera expresses the major collagen-binding integrin subunits. The alpha1 and beta1 subunit expression was decreased early during the development of myopia, whereas the regulation of alpha2 integrin occurred at a later time point. The differential regulation of alpha1beta1 and alpha2beta1 during the development of myopia may reflect specific roles for these receptors in the scleral extracellular matrix and biomechanical remodeling that accompanies myopic eye growth.

Altered visual sensitivity in axial high myopia: a local postreceptoral phenomenon?

PURPOSE: The present study investigated retinal integrity in high myopia using spatial psychophysical tasks. METHODS: Ten axial high myopes (-8.5 to -11.5 D) and 10 age-matched control subjects (+/-1.0 D) were recruited. All participants underwent clinical examination and ocular biometry and demonstrated no visible macular disease with visual acuities better than 6/12. Foveal summation thresholds were determined for white and S-cone-isolating spots of various diameters up to 5.4 degrees and spatial contrast sensitivity to luminance sine wave gratings (0.5-9.7 cyc/deg). Data were analyzed after correction for the magnification induced by eye size and correcting lens power. RESULTS: Spatial summation for both white and S-cone-isolating spots showed a generalized loss of sensitivity at all spot sizes in myopes relative to control subjects (P = 0.01). Critical areas at maximum summation were significantly larger in myopes, for S-cone isolating spots only, after image size correction (P = 0.048). Sensitivity at maximum summation correlated negatively with vitreous chamber depth for both targets (P = 0.005). Sensitivities for S-cone and luminance spots also correlated (P < 0.001), indicating widespread dysfunction. Myopes displayed contrast sensitivity losses at high spatial frequencies (P

Characterization of the integrin receptor subunit profile in the mammalian sclera.

PURPOSE: During the increased eye growth that results in myopia, the sclera undergoes biochemical and biomechanical remodeling. The cell surface integrin receptor family has important roles during tissue remodeling, regulating the extracellular matrix environment and cellular biomechanical properties. As integrin receptors may have a role in remodeling during myopia, this study detailed subunit gene expression in the mammalian sclera. METHODS: Several tissues, including sclera, were isolated from the tree shrew, a mammalian model used in eye growth studies. Total RNA was purified, reverse transcribed and primers for the alpha- and beta-integrin subunits were designed to the published human sequence in areas of high inter-species homology. PCR was used to amplify products of predetermined size and all tree shrew integrin subunits were sequenced to confirm their identity. Multiple PCR conditions were used to identify the scleral integrin subunits, and positive control tissues were included to reduce the possibility of false negative results. RESULTS: Integrin PCR products corresponding to the beta1-, beta4-, beta5-, and beta8-integrin subunits and the alpha-integrin subunits, alpha1-6-, alpha9-11- and alphav-integrin were identified in the sclera and in scleral fibroblast cultures. The respective sequences showed a high identity (>81%) to their human counterparts. The beta2-, beta3-, beta6-, beta7-, alpha7-, and alpha8-integrin subunits were not detected in tree shrew scleral samples, despite being present in the respective positive controls. Association of the 4 beta-integrin subunits with the 10 alpha-integrin subunits suggests that the mammalian sclera is capable of expressing 13 of the 24 identified integrin receptors. CONCLUSIONS: This is the first systematic description of the integrin subunit expression profile in the sclera. Due to the multiple roles of integrin receptors during tissue remodeling, the identification of these scleral integrins is an important preliminary step in determining the role of these receptors during normal eye growth and myopia development.