Guinea Pig Eye Research Articles

Biomechanical changes occur in myopic choroidal stroma and mirror those in the adjacent sclera

Retina-derived growth signals relayed from the choroid to the sclera cause remodeling of the extracellular scleral matrix, resulting in myopic ocular elongation. However, to the best of our knowledge, no studies have assessed changes in choroidal stromal biomechanical properties during myopia progression. Here we utilized 7 µm-resolution scanning acoustic microscopy (SAM) to assess biomechanical properties (bulk modulus (K) and mass density (rho)) of choroidal stroma from guinea pig eyes with form-deprivation (FD) induced myopia. The choroidal stroma had considerable intrinsic strength arising from its biomechanical properties and these were differentially affected by myopia in central and peripheral regions. Choroidal stromal biomechanical values were also highly correlated with those in adjacent scleral regions, and the choroidal stromal-scleral association was stronger in myopic eyes. Biomechanical changes observed in the choroidal stroma of myopic eyes were mirrored to those observed in the adjacent sclera. These findings suggest that choroidal stromal remodeling may accompany myopia and open the door to the source of the signals that cause scleral remodeling in myopia.

A Decrease in Autophagy Increases the Level of Collagen Type I Expression in Scleral Fibroblasts.

Autophagy dysregulation triggers extracellular matrix remodeling via changes in cellular collagen levels and protease secretion. However, the effect of autophagy on scleral extracellular matrix remodeling in the context of myopia is not fully understood. In this study, we measured the level of autophagy in sclera of form deprivation myopic guinea pigs; we also sought a correlation between the level of autophagy in human scleral fibroblasts and the extent of COL1A1 synthesis. We measured the level of COL1A1 expression and the levels of autophagic protein markers in scleral tissues in vivo using a form deprivation myopic guinea pig model. Rapamycin and chloroquine were respectively used to activate and inhibit autophagy in cultured human scleral fibroblasts. COL1A1 gene and protein expression levels were analyzed via quantitative real-time polymerase chain reaction, Western blotting, and immunofluorescence. Levels of autophagy-related proteins were assessed via Western blotting. The sclera of form deprivation myopic guinea pig eyes exhibited decreased expression of COL1A1 and increased expression level of autophagy. After chloroquine exposure, human scleral fibroblasts exhibited decreased autophagy and increased COL1A1 expression. Inhibition of scleral fibroblast autophagy increased COL1A1 expression at the gene and protein levels, thus explaining the effect of autophagy on collagen synthesis by scleral fibroblasts.

Temporal bright light at low frequency retards lens-induced myopia in guinea pigs.

Bright light conditions are supposed to curb eye growth in animals with experimental myopia. Here we investigated the effects of temporal bright light at very low frequencies exposures on lens-induced myopia (LIM) progression. Myopia was induced by application of -6.00 D lenses over the right eye of guinea pigs. They were randomly divided into four groups based on exposure to different lighting conditions: constant low illumination (CLI; 300 lux), constant high illumination (CHI; 8,000 lux), very low frequency light (vLFL; 300/8,000 lux, 10 min/c), and low frequency light (LFL; 300/8,000 lux, 20 s/c). Refraction and ocular dimensions were measured per week. Changes in ocular dimensions and refractions were analyzed by paired t-tests, and differences among the groups were analyzed by one-way ANOVA. Significant myopic shifts in refractive error were induced in lens-treated eyes compared with contralateral eyes in all groups after 3 weeks (all P < 0.05). Both CHI and LFL conditions exhibited a significantly less refractive shift of LIM eyes than CLI and vLFL conditions (P < 0.05). However, only LFL conditions showed significantly less overall myopic shift and axial elongation than CLI and vLFL conditions (both P < 0.05). The decrease in refractive error of both eyes correlated significantly with axial elongation in all groups (P < 0.001), except contralateral eyes in the CHI group (P = 0.231). LFL condition significantly slacked lens thickening in the contralateral eyes. Temporal bright light at low temporal frequency (0.05 Hz) appears to effectively inhibit LIM progression. Further research is needed to determine the safety and the potential mechanism of temporal bright light in myopic progression.

Biomechanical properties of the myopic choroid

Retina-derived growth signals relayed from the choroid to the sclera cause remodeling of the extracellular matrix, resulting in myopic ocular elongation. However, no studies have assessed changes in choroidal biomechanical properties during myopia progression. The present study utilized 7-μm-resolution scanning acoustic microscopy (SAM) to assess biomechanical properties of choroids in guinea pig eyes with form-deprivation (FD) induced myopia. Specifically, neonatal guinea pigs underwent unilateral FD for 1 week (resulting in moderate to high myopia). 12-μm-thick serial cryosections of eyes were scanned with SAM and two-dimensional maps of bulk modulus (K), mass density (rho) were calculated. We found that the choroid had considerable intrinsic strength arising from its biomechanical properties and these were differentially affected by myopia in central and peripheral regions. Choroidal biomechanical values were also highly correlated with those in adjacent scleral regions, and the choroidal-scleral association was stronger in myopic eyes. In conclusion, biomechanical changes observed in the choroid of myopic eyes were mirrored to those observed in the adjacent sclera. These new findings suggest that the choroid remodeling may accompany myopia and opens the door to the source of the signals that cause scleral remodeling in myopia.

Biomechanical and microstructural changes in posterior myopic Guinea pig

The biomechanical and microstructural properties of the posterior sclera are drivers of myopia progression to high/pathologic myopia, a leading cause of blindness, affecting over 2.3 billion people worldwide. We investigated the effect of myopia on mechanical properties of the posterior sclera in guinea-pig (GP) eyes using scanning-acoustic-microscopy (SAM) with 3-μm spatial resolution. Form-deprivation (FD) was used to induce myopia in the right eyes of six GPs between 4 and 12 days of age, and 6-μm-thick scleral cryosections were scanned using a custom-made SAM. Bulk modulus (K) parameter-maps were estimated from SAM radiofrequency echo signals. We studied the effects of myopia on SAM maps in the nasal versus temporal regions of the posterior sclera. In the induced myopia eyes, K-values in the nasal region were 0.126 GPa lower than that in the temporal region (p = 0.009), whereas in the control eye, K did not significantly differ between regions (p &amp;gt; 0.07). Scleral biomechanical properties changed depending on eye region and induced myopia level. These results demonstrate that SAM-maps are important to elucidate myopia pathogenesis, providing insight to the location and timing of scleral stiffness reduction, as reflected in lower K-values in myopic eyes. [Work supported by the NIH grant R01GM143388 (J.M.), UON grant G2100649 (S.M.), and NMRC CIRG19nov-0030 (Q.V.H.).]

Quantitative collagen fiber assessment in myopic guinea pig eye sclera using a cylindrical Gaussian form factor at 80 MHz

Myopia (also called near-sightedness) is an irreversible ocular abnormality that occurs when the eye is excessively elongated, resulting in defocused images and vision. When myopia progresses to pathological myopia, the possibility of permanent vision loss is extremely high. Therefore, it is important to identify early myopic eyes likely to progress to pathologic myopia and vision loss to start treatment and intervene as soon as possible. As myopia progresses, posterior eye elongation is caused by changes in the microstructural properties of the sclera including decreased collagen density and fibril diameters. High-frequency quantitative ultrasound (QUS) allows measuring biomechanical parameters associated with changes in tissue microstructure. This study investigated microstructural changes occurring in the posterior sclera in myopic guinea pig eyes using two independent QUS approaches at 80-MHz: envelope statistics quantification using the Homodyned-K distribution and backscatter coefficient quantification using a cylindrical Gaussian form factor. QUS parameters suggest changes in collagen fiber size occur in myopic sclera and is associated with microstructural changes in the scleral collagen fiber homogeneity and density. In conclusion, high-frequency QUS has the potential to quantitatively characterize microstructural changes that occur in the posterior sclera during myopia development and/or progression.

Modifying Escherichia coli to mimic Shigella for medical microbiology laboratory teaching: a new strategy to improve biosafety in class.

Laboratory teaching of medical microbiology involves highly pathogenic microorganisms, thus posing potential biosafety risks to the students and the teacher. To address these risks, non/low-pathogenic microorganisms were modified to mimic highly pathogenic ones or highly pathogenic microorganisms were attenuated directly using the CRISPR/Cas9 technology. This study describes the modification of Escherichia coli DH5α to mimic Shigella and its evaluation as a safe alternative for medical laboratory teaching. To generate E. coli DH5α△FliC△tnaA2a, the tnaA and FliC genes in E. coli DH5α were knocked out using CRISPR/Cas9 technology; a plasmid bearing the O-antigen determinant of S. flexneri 2a was then constructed and transformed. Acid tolerance assays and guinea pig eye tests were used to assess the viability and pathogenicity, respectively. Questionnaires were used to analyze teaching effectiveness and the opinions of teachers and students. The survey revealed that most teachers and students were inclined towards real-time laboratory classes than virtual classes or observation of plastic specimens. However, many students did not abide by the safety regulations, and most encountered potential biosafety hazards in the laboratory. E. coli DH5α△FliC△tnaA2a was biochemically and antigenically analogous to S. flexneri 2a and had lower resistance to acid than E. coli. There was no toxicity observed in guinea pigs. Most of teachers and students were unable to distinguish E. coli DH5α△FliC△tnaA2a from pure S. flexneri 2a in class. Students who used E. coli DH5α△FliC△tnaA2a in their practice had similar performance in simulated examinations compared to students who used real S. flexneri 2a, but significantly higher than the virtual experimental group. This approach can be applied to other high-risk pathogenic microorganisms to reduce the potential biosafety risks in medical laboratory-based teaching and provide a new strategy for the development of experimental materials.

Effects of riboflavin/ultraviolet-A(UVA) scleral crosslinking on the mechanical behavior of the scleral fibroblasts of lens-induced myopia Guinea pigs

Myopia is becoming increasingly severe, and studies have shown that the cellular mechanics of scleral fibroblasts are altered following myopia. Scleral UVA-Riboflavin Collagen Crosslinking（sCXL） is a promising treatment for myopia prevention and control of axial growth. Understanding the mechanical properties of scleral fibroblasts is crucial, as it influences the cellular response and limits the extent of molecular deformation triggered. Thus, our study aimed to investigate the effect of mechanical properties of scleral fibroblasts in a lens-induced myopic guinea pig model following sCXL. For this purpose, we performed the 0.1% riboflavin/UVA scleral crosslinking (365 nm,3 mW/cm2,30 min) in the right eyes of guinea pigs in Group CXL. In Group LIM, the right eyes were only administrated negative lens for 6 weeks. No treatment was performed in both eyes of the guinea pigs in group Control. The scleral fibroblasts were isolated and cultured from the scleral tissue at the cross-linking area in Group CXL and the corresponding area in Group LIM and control. The curve of the length of microtubules inhaled by cells under negative pressure was measured by a microaspiration-based isolation technique, and the equilibrium Young's modulus and apparent viscosity of scleral fibroblasts were calculated by formula fitting. The equilibrium Young's modulus of scleral fibroblasts in group CXL was significantly lower than that in the LIM group (P < 0.01, two-sample t-test between pairs）, and there was no significant difference between groups CXL and control. The results show that sCXL can effectively moderate the phenomenon that scleral fibroblasts are not easy to deform after myopia. The apparent viscosity modulus in the CXL group was higher than the groups' control and LIM. Taken together, our data demonstrate the biomechanics of the scleral fibroblasts altered after Riboflavin/UVA scleral collagen cross-linking in a lens-induced myopia model.

Intraocular complement activation is related to retinal vascular and neuronal degeneration in myopic retinopathy.

To investigate the relationship between the intraocular levels of complement proteins and myopia-related retinal neuronal and vascular degeneration. Aqueous humour from 147 myopic patients, including 60 low-myopia and 87 high-myopia were collected during Implantable Collamer Lens implantation surgery. All participants received comprehensive ophthalmic examinations, including logMAR best corrected visual acuity, axial length measurement, fundus photography and ocular B-scan ultrasonography. The myopic eyes were further classified into simple myopia (SM, n = 78), myopic posterior staphyloma (PS, n = 39) and PS with myopic chorioretinal atrophy (PS + CA, n = 30). Retinal thickness and vascular density in the macula (6 mm × 6 mm) and optic nerve head (4.5 mm × 4.5 mm) were measured using Optical Coherence Tomography (OCT) and OCT angiography (OCTA). The levels of complement proteins including C1q, C3, C3b/iC3b, C4, CFB, CFH, C2, C4b, C5, C5a, CFD, MBL and CFI in the aqueous humour were measured using the Luminex Multiplexing system. The real-time RT-PCR was conducted to examine the expression of complement genes (C1q, C2, C3, C4, CFI and CFD) in the guinea pig model of long-term form deprivation-induced myopic retinal degeneration. OCTA showed that retinal neuronal thickness and vascular density in superficial and deep layers of the macular zone as well as vascular density in the optic nerve head were progressively decreased from SM to PS and PS + CA (p < 0.05). The aqueous humour levels of C1q, C3, C3b/iC3b, C4, CFB, CFH, C2, C4b, C5 and CFI were significantly higher in high-myopic eyes compared to those in low-myopic eyes. Further subgroup analysis revealed the highest levels of complement components/fragments in the PS + CA group. The intraocular levels of complement factors particularly C3b/iC3b and C4 were negatively correlated with macular zone deep layer retinal thickness and vascular density and optic nerve head vascular density. The expression of C2, C3 and C4 genes was significantly higher in guinea pig eyes with myopic retinal degeneration compared to control eyes. The intraocular classical pathway and alternative pathway of the complement system are partially activated in pathological myopia. Their activation is related to the degeneration of retinal neurons and the vasculature in the macula and the vasculature in the optic nerve head.

Isolation of Retinal Pigment Epithelial Cells from Guinea Pig Eyes.

This protocol describes the isolation of cells of the retinal pigment epithelium (RPE) from the eyes of young pigmented guinea pigs for potential application in molecular biology studies, including gene expression analyses. In the context of eye growth regulation and myopia, the RPE likely plays a role as a cellular relay for growth modulatory signals, as it is located between the retina and the two walls of the eye, such as the choroid and sclera. While protocols for isolating the RPE have been developed for both chicks and mice, these protocols have proven not to be directly translatable to the guinea pig, which has become an important and widely used mammalian myopia model. In this study, molecular biology tools were used to examine the expression of specific genes to confirm that the samples were free of contamination from the adjacent tissues. The value of this protocol has already been demonstrated in an RNA-Seq study of RPE from young pigmented guinea pigs exposed to myopia-inducing optical defocus. Beyond eye growth regulation, this protocol has other potential applications in studies of retinal diseases, including myopic maculopathy, one of the leading causes of blindness in myopes, in which the RPE has been implicated. The main advantage of this technique is that it is relatively simple andonce perfected, yields high-quality RPE samples suitable for molecular biology studies, including RNA analysis.

Atropine-eluting silicone contact lenses for myopia control.

Myopia, also known as nearsightedness, is one of the prime reasons for vision impairment worldwide. Atropine in topical ophthalmic solutions (e.g., 0.01% atropine sulfate eye drops) is the primary medical treatment for controlling myopia, especially for pseudomyopia or true myopia in rapid progress. However, aqueous atropine solution is unstable and easily breaks down to tropic acid, which will result in vision blur. Drug-eluting contact lenses (CLs) have been explored as a potentially superior alternative to effectively control the drug release and improve the drug efficacy. In this work, an atropine-eluting contact lens was developed by encapsulating an atropine implant in a silicon-based contact lens, towards functioning in vision correction and controlling myopia. The safety and effectiveness of this atropine-eluting contact lens were verified with rabbit and guinea pig models. The results showed that the lenses reduced the side effects like mydriasis and no other adverse events were observed in rabbit eyes. More importantly, atropine-loaded lenses could effectively delay the progress of form-deprivation myopia with guinea pig eyes as the model. Thus, we concluded that atropine-eluting CLs prepared by implantation technology may be an option for the treatment of myopia.

EFEMP1 is a potential biomarker of choroid thickness change in myopia.

To explore the possible molecular mechanism by which epidermal growth factor-containing fibulin-like extracellular matrix protein 1 (EFEMP1) regulates choroid thickness (CT) in the development of myopia. In total, 131 subjects were divided into the emmetropia (EM) group, non-high myopia (non-HM) group and high myopia (HM) group. Their age, refraction, intraocular pressure, and other ocular biometric parameters were collected. A 6 × 6 mm area centered on the optic disc was scanned by coherent optical tomography angiography (OCTA) to measure CT, and the tear concentrations of EFEMP1 were quantified using enzyme-linked immunosorbent assay (ELISA) analysis. Twenty-two guinea pigs were divided into the control group and the form-deprivation myopia (FDM) group. The right eye of the guinea pig in the FDM group was covered for 4 weeks, and the diopter and axial length of the right eye of the guinea pig were measured before and after the treatment. After the measurement, the guinea pig was euthanized, and the eyeball was removed. Quantitative reverse transcription polymerase chain reaction, western blotting assays and immunohistochemistry were used to assess the expression of EFEMP1 in the choroid. There were significant differences in CT among the three groups (p < 0.001). CT was positively correlated with age in HM (r = -0.3613, p = 0.0021), but no significant correlation with SE (p > 0.05) was observed. Furthermore, there were increased levels of EFEMP1 in the tears of myopic patients. After 4 weeks of covering the right eye of the FDM guinea pigs, there was a significant increase in axial length and a decrease in diopter (p < 0.05). The mRNA and protein expression of EFEMP1 was significantly increased in the choroid. Choroidal thickness was significantly thinner in myopic patients, and the expression level of EFEMP1 in the choroid increased during the development of FDM. Therefore, EFEMP1 may be involved in the regulation of choroidal thickness in myopia patients.

Effects of riboflavin/ultraviolet-A scleral collagen cross-linking on regional scleral thickness and expression of MMP-2 and MT1-MMP in myopic guinea pigs.

To investigate the effects of scleral collagen cross-linking (SXL) using riboflavin and ultraviolet A (UVA) light on the scleral thickness of different regions and expression of matrix metalloproteinase 2 (MMP-2) and membrane-type MMP-1 (MT1-MMP) in guinea pigs with lens-induced myopia. Forty-eight 4-week-old guinea pigs were assigned to three groups (n = 16 per group): SXL group, lens-induced myopia (LIM) group, and control group. The sclera of the right eye of the guinea pig in the SXL group was surgically exposed, riboflavin was dropped on the treatment area for 10 minutes before the 30-minute UVA irradiation. The same surgical procedure was performed in the LIM group without UVA irradiation. The -10.00 D lenses were then placed on the right eyes of guinea pigs in the SXL and LIM groups for six weeks. The control group received no treatment. The left eyes were untreated in all groups. The ocular axial length (AXL) and refraction were measured at 4 weeks and 10 weeks of age. 10-week-old guinea pigs were sacrificed, and the right eyes were enucleated and evenly divided for preparation of hematoxylin and eosin (HE) stained sections, quantitative real-time polymerase chain reaction (qPCR) and western blotting. The scleral thickness of different regions was measured on HE stained sections. The temporal half of the sclera was harvested to measure the expression of MMP-2 and MT1-MMP by qPCR and western blotting. The AXL was significantly shorter, and the degree of myopic refraction was significantly lower in the SXL group than those in the LIM group at 10 weeks of age. The scleral thickness of the cross-linked area was significantly greater in the SXL group than that of the corresponding area in the LIM group, while the scleral thickness of the untreated nasal side was not significantly different between the SXL group and the LIM group. The expression of MMP-2 and MT1-MMP of the cross-linked sclera was significantly downregulated compared with that of the corresponding area in the LIM group. Riboflavin/UVA SXL could slow myopia progression and thicken the cross-linked sclera in guinea pigs, which might be related to the downregulation of MMP-2 and MT1-MMP expression during the scleral remodeling process.

Unilateral corneal insult also alters sensory nerve activity in the contralateral eye

AbstractWhen the ocular surface of one eye is inflamed or injured, its corneal sensory nerve activity is modified, which in turn evokes ocular discomfort, irritation, and pain sensations referred to the affected eye. Some patients with unilateral ocular inflammation, infection, or surgery also report discomfort and pain in the contralateral eye. Are these sensations experienced at the fellow eye due to alterations of its sensory nerve activity?To test this hypothesis, we recorded in vitro the activity of corneal mechanonociceptor, polymodal nociceptor and cold thermoreceptor nerves of both eyes of guinea pigs previously subjected unilaterally to three different experimental inflammatory and lesion conditions: UV‐induced photokeratitis, microkeratome corneal surgery, and chronic tear deficiency caused by removal of the main lacrimal gland. Recordings from the eyes of naïve animals were used as control.Compared with the naïve guinea pigs, animals subjected to unilateral UV‐induced mild corneal inflammation, showed inhibition of the spontaneous and stimulus‐evoked activity of cold thermoreceptors, and increased activity (sensitization) of nociceptors affecting both the ipsilateral and the contralateral eye. Unilateral microkeratome surgery affected the activity of nociceptors mostly, inducing sensitization in both eyes. Chronic eye dryness by removal of the main lacrimal gland increased cold thermoreceptor activity in both eyes. These results show that after unilateral eye damage, corneal sensory nerve activity is altered in both eyes, although to a lesser degree in the contralateral eye.This is the first direct demonstration that unilateral corneal nerve insult, and especially ocular surface inflammation, functionally affects the activity of the different types of corneal sensory nerves in both the ipsilateral and contralateral eyes. The mechanisms underlying the changes of contralateral sensory nerve activity remain to be determined, although available data support the involvement of the immune system. The present results have two main implications: (a) contralateral eyes cannot be considered as a control in experimental designs of both preclinical and clinical studies; and, (b) clinicians must consider the convenience of treating both eyes of patients with unilateral ocular conditions to avoid pain and secondary undesirable effects in the fellow eye.

Biometric measurement with a commercially available swept-source optical coherence tomography in myopia model species

BackgroundBiometric parameters play an important role in studies on myopia. This study aimed to explore the application of a commercially available two-dimensional visualized swept-source optical coherence tomography (OCT) system, for in vivo biometric measurement in multiple myopia model species. MethodsIn this study, chickens, guinea pigs, and C57BL/6 mice underwent eye imaging with the commercially available OCT (CASIA2), and the original images were used to calculate the central corneal thickness, anterior chamber depth (ACD), lens thickness (LT), vitreous chamber depth (VCD), and axial length (AL). The retinal thickness and choroidal thickness were also calculated in chicken eyes. The repeatability of the biometric measurement outcomes was analyzed. ResultsExcellent repeatable AL measurements were obtained for all three species, with an intraclass correlation coefficient (ICC) of ≥0.941 and a within-subject standard deviation of ≤0.055. Excellent repeatability was found in chicken eyes for ACD, LT, and VCD, with an ICC of ≥0.932; in guinea pig eyes for ACD and VCD, with an ICC of ≥0.934; and in mouse eyes for LT, with an ICC of ≥0.941. ConclusionsIt is effective to use commercially available OCT to measure biometric parameters in chickens, guinea pigs, and C57BL/6 mice. This methodology could potentially increase the accuracy and efficiency of future myopia animal experiments.

Biomechanical changes in myopic sclera correlate with underlying changes in microstructure

Myopia alters the microstructural and biomechanical properties of the posterior sclera, which is characterized as a layered structure with potentially different inter-layer collagen fibril characteristics. Scanning acoustic microscopy (SAM) has been used to investigate how the micron-scale bulk mechanical properties of the posterior sclera are affected by myopia. Other investigators have employed second harmonic generation (SHG) imaging to characterize the collagen microstructure of tissues. In the present study, SAM and SHG imaging were used to investigate the existence of biomechanically-distinct scleral layers and identify relationships between mechanical properties and tissue microstructure in myopic guinea pig (GP) eyes. Diffusers were worn over the right eyes of six, 1-week-old GPs for one week to induce unilateral form-deprivation myopia. GPs were euthanized, enucleated, and eyes were cryosectioned. Twelve-micron-thick adjacent vertical cryosections were scanned with SAM or SHG. SAM maps of bulk modulus, mass density, and acoustic attenuation were estimated. A fiber-extraction algorithm applied to SHG images estimated collagen fiber length, width, straightness, alignment, and number density. Results revealed that the posterior sclera may exhibit biomechanically distinct layers that are affected differently in myopia. Specifically, a layered structure was observed in the mechanical-parameter maps of control eyes that was less apparent in myopic eyes. Collagen fibers in myopic eyes had smaller diameters and were more aligned. Myopia-associated biomechanical changes were most significant in the outermost and innermost scleral layers. SAM-measured mechanical parameters were correlated with collagen fiber microstructure, particularly fiber length, alignment, and number density, which may imply the biomechanical parameters estimated from SAM measurements are related to tissue microstructure. Interestingly, some changes were greatest in more-peripheral regions, suggesting interventions to strengthen the sclera may be effective away from the optic nerve and efficacy may be achieved best when intervention is applied to the outermost layer.

Scleral Cross-Linking in Form-Deprivation Myopic Guinea Pig Eyes Leads to Glaucomatous Changes.

PurposeTo investigate the potential glaucomatous changes caused by scleral cross-linking (CXL) in a guinea pig form-deprivation (FD) myopia model.MethodsEighty 4-week-old tricolor guinea pigs were divided into four groups: FD only, genipin CXL only, FD plus CXL, and control. Refractive error, axial length (AL), intraocular pressure (IOP), and structural and vasculature optic disc changes in optical coherence tomography (OCT) and OCT angiography (OCTA) were measured at baseline and day 21. CXL efficacy was evaluated by scleral rigidity Young's modulus values. Histological and molecular changes in the anterior chamber angle, retina, and sclera were assessed.ResultsBaseline parameters were similar among groups (P > 0.05). The FD plus CXL group at day 21 had the least increase of AL (0.14 ± 0.08 mm) and highest IOP elevation (31.5 ± 3.6 mmHg) compared with the FD-only group (AL: 0.68 ± 0.17 mm; IOP: 22.2 ± 2.6 mmHg) and the control group (AL: 0.24 ± 0.09 mm; IOP: 17.4 ± 1.8 mmHg) (all P < 0.001). OCT and OCTA parameters of the optic disc in the FD plus CXL group at day 21 showed glaucomatous changes and decreased blood flow signals. Sclera rigidity increased in the CXL and FD plus CXL groups. Advanced glycation end products deposited extensively in the retina, choroid, and sclera of FD plus CXL eyes.ConclusionsCXL causes increased IOP and subsequent optic disc, anterior segment, and scleral changes while inhibiting myopic progression and axial elongation in FD guinea pig eyes. Therefore, applying CXL to control myopia raises safety concerns.

Decreased Choroidal Blood Perfusion Induces Myopia in Guinea Pigs.

PurposeThe development of myopia in guinea pigs can be inhibited by attenuating scleral hypoxia by increasing choroidal blood perfusion (ChBP). In this study, we reduced ChBP through surgical and pharmacological methods to determine the effect on myopia development. We also determined whether ChBP was reduced by quinpirole, a drug that enhances form-deprivation myopia (FDM).MethodsChBP was reduced in the right eyes of guinea pigs via transection of the temporal ciliary arteries or daily injections of phenylephrine into the inferior peribulbar space for one week during normal ocular growth. Other guinea pigs were subjected to two weeks of monocular FDM—with facemasks, along with daily injections of quinpirole, a dopamine D2 receptor agonist, to enhance the FDM. Changes in refraction, axial length, ChBP, and choroidal thickness (ChT) were measured in both treated and fellow eyes of the treatment and control groups. Scleral hypoxia labeling with pimonidazole adducts and α-smooth muscle actin (α-SMA) protein were also measured.ResultsSurgical and pharmacological reduction of ChBP induced myopia development in the treated eyes. These treatments rendered the scleral hypoxia and increased scleral α-SMA expression. Furthermore, quinpirole injections, which increased the magnitude of myopia, augmented the FDM-associated reductions in ChBP and ChT and increased the levels of scleral hypoxia and α-SMA protein.ConclusionsDecreased ChBP in guinea pigs leads to scleral hypoxia and scleral myofibroblast transdifferentiation with increased α-SMA expression, ultimately resulting in myopia development. In future clinical trials, ChBP reduction can serve as a potential biomarker for early detection of myopia development.

Unilateral Corneal Insult Also Alters Sensory Nerve Activity in the Contralateral Eye.

After the unilateral inflammation or nerve lesion of the ocular surface, the ipsilateral corneal sensory nerve activity is activated and sensitized, evoking ocular discomfort, irritation, and pain referred to the affected eye. Nonetheless, some patients with unilateral ocular inflammation, infection, or surgery also reported discomfort and pain in the contralateral eye. We explored the possibility that such altered sensations in the non-affected eye are due to the changes in their corneal sensory nerve activity in the contralateral, not directly affected eye. To test that hypothesis, we recorded the impulse activity of the corneal mechano- and polymodal nociceptor and cold thermoreceptor nerve terminals in both eyes of guinea pigs, subjected unilaterally to three different experimental conditions (UV-induced photokeratitis, microkeratome corneal surgery, and chronic tear deficiency caused by removal of the main lacrimal gland), and in eyes of naïve animals ex vivo. Overall, after unilateral eye damage, the corneal sensory nerve activity appeared to be also altered in the contralateral eye. Compared with the naïve guinea pigs, animals with unilateral UV-induced mild corneal inflammation, showed on both eyes an inhibition of the spontaneous and stimulus-evoked activity of cold thermoreceptors, and increased activity in nociceptors affecting both the ipsilateral and the contralateral eye. Unilateral microkeratome surgery affected the activity of nociceptors mostly, inducing sensitization in both eyes. The removal of the main lacrimal gland reduced tear volume and increased the cold thermoreceptor activity in both eyes. This is the first direct demonstration that unilateral corneal nerve lesion, especially ocular surface inflammation, functionally affects the activity of the different types of corneal sensory nerves in both the ipsilateral and contralateral eyes. The mechanisms underlying the contralateral affectation of sensory nerves remain to be determined, although available data support the involvement of neuroimmune interactions. The parallel alteration of nerve activity in contralateral eyes has two main implications: a) in the experimental design of both preclinical and clinical studies, where the contralateral eyes cannot be considered as a control; and, b) in the clinical practice, where clinicians must consider the convenience of treating both eyes of patients with unilateral ocular conditions to avoid pain and secondary undesirable effects in the fellow eye.

Dynamic Changes of AREG in the Sclera during the Development of Form-Deprivation Myopia in Guinea Pigs

ABSTRACTPurposeTo investigate amphiregulin (AREG) expression in the sclera during the development of form-deprivation myopia (FDM) and after the recovery of FDM in guinea pigs.MethodsSixty-four 2-week-old guinea pigs were randomly divided into the control and FDM groups. The right eyes of animals in FDM group were covered for 2 weeks (2 W) and 4 weeks (4 W), or were covered for 4 weeks and then uncovered for the subsequent 2 weeks (6 W). The diopters and axial lengths (AL) in the right eyes of guinea pigs were measured. Reverse transcription polymerase chain reaction (RT-PCR), immunofluorescence, and western blotting assays were used to detect the relative mRNA and protein expressions of AREG in the sclera of guinea pigs.ResultsBefore masking (0 W), the refraction and AL in the right eyes of guinea pigs in the control and FDM groups did not differ significantly (both p > .05). Myopic shift was induced in guinea pigs with the diopters gradually decreased and AL increased in the FDM group. While no significant difference was found in control group at different time points, the relative AREG mRNA and protein expression levels in the FDM group were significantly increased in 2 W and 4 W and then decreased after 4 weeks of covering followed by uncovering for 2 weeks (all p < .05).ConclusionsAREG was expressed in the sclera of guinea pigs. Moreover, the expression levels of AREG increased during the development of FDM and downregulated after recovery of FDM. Therefore, AREG may be involved in the regulation of scleral remodeling in myopia.