The Structural Layers of the Porcine Iris Exhibit Inherently Different Biomechanical Properties.

Purpose: The purpose of this study was to isolate the structural components of the ex vivo porcine iris tissue and to determine their biomechanical properties. Methods: The porcine stroma and dilator tissues were separated, and their dimensions were assessed using optical coherence tomography (OCT). The stroma underwent flow test (n = 32) to evaluate for permeability using Darcy's Law (ΔP = 2000 Pa, A = 0.0391 mm2), and both tissues underwent stress relaxation experiments (ε = 0.5 with initial ramp of δε = 0.1) to evaluate for their viscoelastic behaviours (n = 28). Viscoelasticity was characterized by the parameters ß (half width of the Gaussian distribution), τm (mean relaxation time constant), E0 (instantaneous modulus), and E∞ (equilibrium modulus). Results: For the stroma, the hydraulic permeability was 9.49 ± 3.05 × 10-6 mm2/Pa · s, and the viscoelastic parameters were ß = 2.50 ± 1.40, and τm = 7.43 ± 4.96 s, with the 2 moduli calculated to be E0 = 14.14 ± 6.44 kPa and E∞ = 6.08 ± 2.74 kPa. For the dilator tissue, the viscoelastic parameters were ß = 2.06 ± 1.33 and τm = 1.28 ± 1.27 seconds, with the 2 moduli calculated to be E0 = 9.16 ± 3.03 kPa and E∞ = 5.54 ± 1.98 kPa. Conclusions: We have established a new protocol to evaluate the biomechanical properties of the structural layers of the iris. Overall, the stroma was permeable and exhibited smaller moduli than those of the dilator muscle. An improved characterization of iris biomechanics may form the basis to further our understanding of angle closure glaucoma.

Intravitreal Neuroglobin Mitigates Primate Experimental Glaucomatous Structural Damage in Association with Reduced Optic Nerve Microglial and Complement 3-Astrocyte Activation.

Current management of glaucomatous optic neuropathy is limited to intraocular pressure control. Neuroglobin (Ngb) is an endogenous neuroprotectant expressed in neurons and astrocytes. We recently showed that exogenous intravitreal Ngb reduced inflammatory cytokines and microglial activation in a rodent model of hypoxia. We thus hypothesised that IVT-Ngb may also be neuroprotective in experimental glaucoma (EG) by mitigating optic nerve (ON) astrogliosis and microgliosis as well as structural damage. In this study using a microbead-induced model of EG in six Cynomolgus primates, optical coherence imaging showed that Ngb-treated EG eyes had significantly less thinning of the peripapillary minimum rim width, retinal nerve fibre layer thickness, and ON head cupping than untreated EG eyes. Immunohistochemistry confirmed that ON astrocytes overexpressed Ngb following Ngb treatment. A reduction in complement 3 and cleaved-caspase 3 activated microglia and astrocytes was also noted. Our findings in higher-order primates recapitulate the effects of neuroprotection by Ngb treatment in rodent EG studies and suggest that Ngb may be a potential candidate for glaucoma neuroprotection in humans.

Incidence of COVID-19 Vaccination-Related Uveitis and Effects of Booster Dose in a Tertiary Uveitis Referral Center.

Background: We report vaccine and booster-related uveitis in Singapore, a country with high vaccination and booster rates to highlight the differences and potential role of prophylactic treatment for sight-threatening infectious uveitis. Methods: Clinical data extracted from the de-identified uveitis database in Singapore National Eye Center. Six patients (eight eyes) developed uveitis within 14 days after undergoing COVID-19 vaccination (primary and/or booster). Results: All patients received two doses of COVID-19 vaccination, and 1.39% (6/431) developed COVID-19 vaccine-related uveitis. Fifty-percent% (3/6) with non-infectious anterior uveitis (NIAU) presented with a non-granulomatous anterior uveitis (AU). The remaining (3/6) presenting with a granulomatous AU were diagnosed with reactivation of cytomegalovirus, varicella-zoster virus and toxoplasma chorioretinitis, respectively. All the patients responded to definitive treatment specific to their diagnosis. The mean visual acuity at presentation was 0.36 ± 0.20 logMAR and improved to 0.75 ± 0.09 (p = 0.009). Mean time from vaccination to uveitis was 9.7 (range: 3-14) days. All patients developed uveitis after second vaccination dose. 16.67% (1/6) patients had a recurrence after the third booster dose. None of the three patients with infectious uveitis developed recurrence but had received maintenance therapy up to or during the booster. Conclusion: Uveitis after COVID-19 vaccination is uncommon. In our series, a higher rate of reactivations of latent infections was seen. With definitive treatment, all cases were self-limited without systemic sequelae. Prophylactic treatment during booster vaccine may prevent reactivation of sight-threatening infections and reduce morbidity although risk-benefits should be considered for individual patients given the low rate of occurrence.

The Management of Ocular Surface Squamous Neoplasia (OSSN).

The rise of primary topical monotherapy with chemotherapeutic drugs and immunomodulatory agents represents an increasing recognition of the medical management of ocular surface squamous neoplasia (OSSN), which may replace surgery as the standard of care in the future. Currently, there is no consensus regarding the best way to manage OSSN with no existing guidelines to date. This paper seeks to evaluate evidence surrounding available treatment modalities and proposes an approach to management. The approach will guide ophthalmologists in selecting the most appropriate treatment regime based on patient and disease factors to minimize treatment related morbidity and improve OSSN control. Further work can be done to validate this algorithm and to develop formal guidelines to direct the management of OSSN.

Common variants in SOX-2 and congenital cataract genes contribute to age-related nuclear cataract.

Nuclear cataract is the most common type of age-related cataract and a leading cause of blindness worldwide. Age-related nuclear cataract is heritable (h2 = 0.48), but little is known about specific genetic factors underlying this condition. Here we report findings from the largest to date multi-ethnic meta-analysis of genome-wide association studies (discovery cohort N = 14,151 and replication N = 5299) of the International Cataract Genetics Consortium. We confirmed the known genetic association of CRYAA (rs7278468, P = 2.8 × 10-16) with nuclear cataract and identified five new loci associated with this disease: SOX2-OT (rs9842371, P = 1.7 × 10-19), TMPRSS5 (rs4936279, P = 2.5 × 10-10), LINC01412 (rs16823886, P = 1.3 × 10-9), GLTSCR1 (rs1005911, P = 9.8 × 10-9), and COMMD1 (rs62149908, P = 1.2 × 10-8). The results suggest a strong link of age-related nuclear cataract with congenital cataract and eye development genes, and the importance of common genetic variants in maintaining crystalline lens integrity in the aging eye.

Longitudinal assessment of optic nerve head changes using optical coherence tomography in a primate microbead model of ocular hypertension.

In humans, the longitudinal characterisation of early optic nerve head (ONH) damage in ocular hypertension (OHT) is difficult as patients with glaucoma usually have structural ONH damage at the time of diagnosis. Previous studies assessed glaucomatous ONH cupping by measuring the anterior lamina cribrosa depth (LCD) and minimal rim width (MRW) using optical coherence tomography (OCT). In this study, we induced OHT by repeated intracameral microbead injections in 16 cynomolgus primates (10 unilateral; 6 bilateral) and assessed the structural changes of the ONH longitudinally to observe early changes. Elevated intraocular pressure (IOP) in OHT eyes was maintained for 7 months and serial OCT measurements were performed during this period. The mean IOP was significantly elevated in OHT eyes when compared to baseline and compared to the control eyes. Thinner MRW and deeper LCD values from baseline were observed in OHT eyes with the greatest changes seen between month 1 and month 2 of OHT. Both the mean and maximum IOP values were significant predictors of MRW and LCD changes, although the maximum IOP was a slightly better predictor. We believe that this model could be useful to study IOP-induced early ONH structural damage which is important for understanding glaucoma pathogenesis.

Effects of Exogenous Neuroglobin (Ngb) on retinal inflammatory chemokines and microglia in a rat model of transient hypoxia.

Neuroglobin is an endogenous neuroprotective protein. We determined the safety of direct delivery of Neuroglobin in the rat retina and its effects on retinal inflammatory chemokines and microglial during transient hypoxia. Exogenous Neuroglobin protein was delivered to one eye and a sham injection to the contralateral eye of six rats intravitreally. Fundus photography, Optical Coherence Topography, electroretinogram, histology and Neuroglobin, chemokines level were determined on days 7 and 30. Another 12 rats were subjected to transient hypoxia to assess the effect of Neuroglobin in hypoxia exposed retina by immunohistochemistry, retinal Neuroglobin concentration and inflammatory chemokines. Intravitreal injection of Neuroglobin did not incite morphology or functional changes in the retina. Retinal Neuroglobin protein was reduced by 30% at day 7 post hypoxia. It was restored to normoxic control levels with intravitreal exogenous Neuroglobin injections and sustained up to 30 days. IL-6, TNFα, IL-1B, RANTES, MCP-1 and VEGF were significantly decreased in Neuroglobin treated hypoxic retinae compared to non-treated hypoxic controls. This was associated with decreased microglial activation in the retina. Our findings provide proof of concept suggesting intravitreal Neuroglobin injection is non-toxic to the retina and can achieve the functional level to abrogate microglial and inflammatory chemokines responses during transient hypoxia.

Transscleral cyclophotocoagulation and its histological effects on the conjunctiva.

Micropulse transscleral cyclophotocoagulation (MP-TCP) is increasingly being used as an initial procedure prior to conjunctival filtration surgeries. However, it is uncertain whether MP-TCP may cause inflammation and scarring of the bulbar conjunctiva. Thus, we aimed to study the histological effects of MP-TCP (compared to controls and continuous wave [CW]-TCP) on the conjunctiva. Our study included 10 Dutch Belted Rabbits that underwent TCP in their right eyes (n = 5, MP-TCP; n = 5, CW-TCP), while their left eyes served as controls. The rabbits were euthanised at 4 weeks, and their dissected globes underwent histopathological and immunohistochemical examination. We observed greater conjunctival inflammation in MP-TCP or CW-TCP-treated eyes compared to controls, but not between each other. The majority of the lymphocytic infiltrates were CD4 T-cells. Increased conjunctival fibrosis was evident in MP-TCP or CW-TCP-treated eyes, to similar extents, compared to controls. However, the increased staining for myofibroblasts was not statistically significant in TCP-treated eyes. We concluded that MP-TCP causes significantly greater overall conjunctival inflammation and scarring compared to controls, similar to CW-TCP. As these are risk-factors for fibrosis and failure of the conjunctival bleb, further studies are required to explore the effect, if any, of post-TCP conjunctival changes on future bleb morphology and survival.

Corneal Inlay Damage After Cosmetic Laser Treatment of the Eyelid With Long-Pulsed Nd:YAG Laser.

Importance: Protective eyewear or corneal shields are recommended during cosmetic facial laser treatment. Objective: To describe the occurrence of corneal inlay damage following treatment to the eyelids and face with an Nd:YAG laser. Design, Setting, and Participants: This observational case report includes a single incident case cared for at a tertiary care center. A 58-year-old man who had undergone bilateral uncomplicated myopic laser in situ keratomileusis surgery in 2003 and corneal inlay implant in the nondominant left eye in 2013 experienced decreased visual acuity (VA) and pain in the left eye after the application of 2 passes of the Nd:YAG laser to his face and both eyelids for facial tightening. At presentation, the uncorrected VA was counting fingers OS and 20/20 OD. Slitlamp biomicroscopy showed a corneal epithelial defect overlying a deformed corneal inlay, peripheral scattered pigmentary deposits, corneal haze, and brown discoloration of the lamellar pocket of the inlay. He underwent explant of the inlay and debridement 48 hours later because of deteriorating VA and increasing corneal haze. Exposure: Application of long-pulsed Nd:YAG laser at 1064 nm to the middle one-third of both upper eyelids and the periorbital region in a man with a corneal inlay implant. Corneal shields were not worn during the procedure. The peak penetration depth of this laser system is approximately 4 mm. The mean (SD) thickness of the upper eyelid in Asian eyes is 1.127 (0.238) mm. Main Outcomes and Measures: Improvement of corneal inlay damage. Results: In this 58-year-old man, 3 months after the inlay explant, the intrastromal discoloration had resolved. There was still residual corneal haze, but the patient was able to achieve a best-corrected VA of 20/25 OS for distance and J3 (Snellen equivalent, 20/30) OS for near. Conclusions and Relevance: Although the exact cause and effect cannot be determined from a single case, our findings suggest that a history of corneal inlay implant should be asked about prior to any long-pulsed Nd:YAG laser treatment to the periorbital skin and eyelids. Furthermore, these findings suggest that laser treatment to the eyelids should be avoided and that protective eyewear or corneal shields are recommended during cosmetic facial laser treatment in all patients.

Permeability of the porcine iris stroma.

Current literature has not considered or provided any data on the permeability of the iris stroma. In this study, we aimed to determine the hydraulic permeability of porcine irides from the isolated stroma. Fifteen enucleated porcine eyes were acquired from the local abattoir. The iris pigment epithelium was scraped off using a pair of forceps and the dilator muscles were pinched off using a pair of colibri toothed forceps. We designed an experimental setup, based on Darcy's law, and consisting of a custom 3D-printed pressure column using acrylonitrile butadiene styrene (ABS) plastic. PBS solution was passed through the iris stroma in a 180° arc shape, with a column height of approximately 204 mm (2000 Pa). Measurements of iris stromal thickness were conducted using optical coherence tomography (OCT). To measure flow rate, we measured the mass (volume) of PBS solution using a mass balance in approximately 1 min. Histology was performed using hematoxylin and eosin (H&E) and anti-smooth muscle antibody (anti-α-SMA) for validation. The permeability experiments demonstrated that the iris stroma is a biphasic tissue that allows fluid flow. Our image processing results determined the area of flow to be 7.55 mm2 and the tissue thickness to be between 180 and 430 µm. The hydraulic permeability of the porcine stroma, calculated using Darcy's law, was 5.13 ±â€¯2.39 × 10-5 mm2/Pa•s. Histological and immunochemical studies confirmed that the tissues used for this permeability study were solely iris stroma. Additionally, anti-α-SMA staining revealed staining specific for stromal blood vessels, with the notable absence of dilator and sphincter muscle staining. Our study combined experimental microscopic data with the theory of biphasic materials to investigate the hydraulic permeability of the iris stroma. This work will serve as a basis on which to validate future biomechanical studies of human irides with which may ultimately aid disease diagnosis and inform the design of novel treatments.

Biomechanical Properties of Bruch's Membrane-Choroid Complex and Their Influence on Optic Nerve Head Biomechanics.

Purpose: The purpose of this study was to measure the rupture pressure and the biomechanical properties of porcine Bruch's membrane (BM)-choroid complex (BMCC) and the influences of BM on optic nerve head (ONH) tissues. Methods: The biomechanical properties of BMCC were extracted through uniaxial tensile tests of 10 BMCC specimens from 10 porcine eyes; the rupture pressures of BMCC were measured through burst tests of 20 porcine eyes; and the influence of BM on IOP-induced ONH deformations were investigated using finite element (FE) analysis. Results: Uniaxial experimental results showed that the average elastic (tangent) moduli of BMCC samples at 0% and 5% strain were 1.60 ± 0.81 and 2.44 ± 1.02 MPa, respectively. Burst tests showed that, on average, BMCC could sustain an IOP of 82 mm Hg before rupture. FE simulation results predicted that, under elevated IOP, prelamina tissue strains increased with increasing BM stiffness. On the contrary, lamina cribrosa strains showed an opposite trend but the effects were small. Conclusions: BMCC stiffness is comparable or higher than those of other ocular tissues and can sustain a relatively high pressure before rupture. Additionally, BM may have a nonnegligible influence on IOP-induced ONH deformations.

Immunopathology of Virus-Induced Anterior Uveitis.

Herpes simplex virus, varicella zoster virus, human cytomegalovirus, and rubella virus are the most common causes of virus-induced anterior uveitis. They can present in a variety of entities not only with typical but also overlapping clinical characteristics. These viral infections are commonly associated with ocular infiltration of T cells and B/plasma cells, and expression of cytokines and chemokines typical of a proinflammatory immune response. The infections differ in that the herpes viruses cause an acute lytic infection and inflammation, whereas rubella virus is a chronic low-grade infection with slowly progressing immunopathological responses. The outcome of an intraocular viral infection may largely be guided by the characteristics of the virus, which subsequently dictates the severity and type of the immune response, and the host immune status.

Genetic association study of exfoliation syndrome identifies a protective rare variant at LOXL1 and five new susceptibility loci.

Exfoliation syndrome (XFS) is the most common known risk factor for secondary glaucoma and a major cause of blindness worldwide. Variants in two genes, LOXL1 and CACNA1A, have previously been associated with XFS. To further elucidate the genetic basis of XFS, we collected a global sample of XFS cases to refine the association at LOXL1, which previously showed inconsistent results across populations, and to identify new variants associated with XFS. We identified a rare protective allele at LOXL1 (p.Phe407, odds ratio (OR) = 25, P = 2.9 × 10-14) through deep resequencing of XFS cases and controls from nine countries. A genome-wide association study (GWAS) of XFS cases and controls from 24 countries followed by replication in 18 countries identified seven genome-wide significant loci (P < 5 × 10-8). We identified association signals at 13q12 (POMP), 11q23.3 (TMEM136), 6p21 (AGPAT1), 3p24 (RBMS3) and 5q23 (near SEMA6A). These findings provide biological insights into the pathology of XFS and highlight a potential role for naturally occurring rare LOXL1 variants in disease biology.

Serum IgG2 and tissue IgG2 plasma cell elevation in orbital IgG4-related disease (IgG4-RD): Potential use in IgG4-RD assessment.

AIMS: To determine the role of serum and tissue IgG2 in orbital biopsies with the histological features of IgG4-related disease (IgG4-RD) in comparison with non-IgG4-related orbital inflammatory disorders (OID), including autoimmune disorders. METHODS: This is an international (Sheffield, UK, and Singapore) collaborative, retrospective case review of 69 patients (38 from Singapore National Eye Centre and 31 from Royal Hallamshire Hospital, Sheffield) with orbital inflammatory biopsies between 2002 and 2016. Clinical information and histology were reviewed and cases were classified into three groups: Group 1: IgG4-RD orbital inflammation (n=43); Group 2: idiopathic OID (n=12) and Group 3: autoimmune OID (n=14). Serum IgG1, IgG2, IgG3 and IgG4 levels were collated where available and immunohistochemistry (IHC) for tissue IgG2 plasma cells was performed. RESULTS: Dual IHC showed IgG2 plasma cells as a distinct population from IgG4 plasma cells. Significant (twofold) serum IgG2 elevation was noted among IgG4-RD (group 1), idiopathic (group 2) and autoimmune OID (group 3). Similarly, significant elevation of tissue IgG2 plasma cells was also seen among IgG4-RD (group 1), idiopathic and autoimmune OID (groups 2 and 3). CONCLUSIONS: Significant elevations of serum IgG2 and tissue IgG2 plasma cells are present in orbital IgG4-RD in comparison with non-IgG4 orbital inflammation (idiopathic and autoimmune OID), suggesting that IgG2 may play a role in IgG4-RD. A serum IgG2 cut-off >5.3 g/L was found to be 80% sensitive and 91.7% specific for orbital IgG4-RD, with an accuracy of 0.90. Tissue IgG2 and IgG4 subclass reporting may provide additional insight regarding the 'IgG4-RD' pathogenesis.

Clinical Features and Treatment Outcomes of Orbital Inflammatory Disease in Singapore: A 10-Year Clinicopathologic Review.

PURPOSE: 1) To assess the clinical profile and treatment outcomes of orbital inflammatory disease in the local population, and 2) classify patients using current histopathological criteria. METHODS: Ten-year retrospective clinicopathologic review of patients diagnosed with orbital inflammatory disease who underwent tissue biopsy from January 2001 to December 2011 at a tertiary referral centre in Singapore. Data collection included patient demographics, clinical presentation, investigations, systemic disease, histopathology review, clinical classification, medical and surgical management, response to treatment and recurrence rates. RESULTS: The study comprised 70 patients. Thirty-seven (52.9%) had nonspecific inflammation distributed as follows: lacrimal (n = 23), diffuse (n = 5), anterior (n = 5), myositic (n = 4). Thirty-three (47.1%) had specific inflammation of the following subtypes: idiopathic sclerosing inflammation (n = 9), granulomatous disorders (n = 8), transitional lesions (n = 5), vasculitis (n = 4), and others (n = 7). A total of 76.8% of patients received oral prednisolone, with a median duration of three months. Response to treatment was good in 71.9% of patients. Recurrence occurred in 22 (32.8%) patients at a mean interval of 20 months after completion of treatment, and was higher in myositic and vasculitic subtypes. There was no significant correlation between duration of treatment and recurrence. CONCLUSIONS: This study has re-emphasized the importance and utility of orbital biopsy and histopathologic typing for optimal management of orbital inflammatory disease. It has also improved the knowledge of the rate and response to treatment of its various subtypes.

Histological features of Cytomegalovirus-related corneal graft infections, its associated features and clinical significance.

AIM: To describe the histological features of ITALIC! Cytomegalovirus (CMV)-related corneal graft infections, its associated features and clinical significance. METHODS: This was a retrospective histological study of 48 consecutive cases of failed repeat penetrating keratoplasty cases with a clinical diagnosis of allograft rejection from 2011 to 2013. CMV infection was confirmed with CMV antibody immunohistochemistry (IHC) and electron microscopy. Additional CD163 and CD68 IHCs for macrophages were also performed. Clinical data and previous graft histology were then reviewed. RESULTS: Mean incidence of CMV infection in corneal graft rejection buttons was 6.3% per year. 3/48 graft buttons were CMV antibody positive. Histological features of CMV graft infection include: (1) stromal keratocytes with cytopathic changes; (2) lack of inflammation, only occasional macrophages present and (3) absence of vascularisation. None of the patients had a history of active CMV infection. CONCLUSION: CMV infection is not limited as endotheliitis, but extends into the corneal stroma, and is a potential reservoir for graft infection, especially in partial thickness endothelial surgery. Clinical features are often non-specific, although glaucoma was present in our patients. CMV-infected grafts showed CD163-positive M2 macrophages in close association with the infected keratocytes, suggesting that the macrophage may be important in CMV graft infection. Histological examination with CMV IHC is a useful method to detect CMV infection postoperatively. Post penetrating keratoplasty, CMV systemic treatment with valganciclovir can prevent graft infection and failure. Boston keratoprosthesis may be a potential alternative surgery in active CMV infections that obviates the need for systemic therapy.

A common variant near TGFBR3 is associated with primary open angle glaucoma.

Primary open angle glaucoma (POAG), a major cause of blindness worldwide, is a complex disease with a significant genetic contribution. We performed Exome Array (Illumina) analysis on 3504 POAG cases and 9746 controls with replication of the most significant findings in 9173 POAG cases and 26 780 controls across 18 collections of Asian, African and European descent. Apart from confirming strong evidence of association at CDKN2B-AS1 (rs2157719 [G], odds ratio [OR] = 0.71, P = 2.81 × 10(-33)), we observed one SNP showing significant association to POAG (CDC7-TGFBR3 rs1192415, ORG-allele = 1.13, Pmeta = 1.60 × 10(-8)). This particular SNP has previously been shown to be strongly associated with optic disc area and vertical cup-to-disc ratio, which are regarded as glaucoma-related quantitative traits. Our study now extends this by directly implicating it in POAG disease pathogenesis.

A common variant mapping to CACNA1A is associated with susceptibility to exfoliation syndrome.

Exfoliation syndrome (XFS) is the most common recognizable cause of open-angle glaucoma worldwide. To better understand the etiology of XFS, we conducted a genome-wide association study (GWAS) of 1,484 cases and 1,188 controls from Japan and followed up the most significant findings in a further 6,901 cases and 20,727 controls from 17 countries across 6 continents. We discovered a genome-wide significant association between a new locus (CACNA1A rs4926244) and increased susceptibility to XFS (odds ratio (OR) = 1.16, P = 3.36 × 10(-11)). Although we also confirmed overwhelming association at the LOXL1 locus, the key SNP marker (LOXL1 rs4886776) demonstrated allelic reversal depending on the ancestry group (Japanese: OR(A allele) = 9.87, P = 2.13 × 10(-217); non-Japanese: OR(A allele) = 0.49, P = 2.35 × 10(-31)). Our findings represent the first genetic locus outside of LOXL1 surpassing genome-wide significance for XFS and provide insight into the biology and pathogenesis of the disease.