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Immunogenicity against intravitreally administered brolucizumab has been previously described and associated with cases of severe intraocular inflammation, including retinal vasculitis/retinal vascular occlusion (RV/RO). The presence of antidrug antibodies (ADAs) in these patients led to the initial hypothesis that immune complexes could be key mediators. Although the formation of ADAs and immune complexes may be a prerequisite, other factors likely contribute to some patients having RV/RO, whereas the vast majority do not. To identify and characterize the mechanistic drivers underlying the immunogenicity of brolucizumab and the consequence of subsequent ADA-induced immune complex formation, a translational approach was performed to bridge physicochemical characterization, structural modeling, sequence analysis, immunological assays, and a quantitative systems pharmacology model that mimics physiological conditions within the eye. This approach revealed that multiple factors contributed to the increased immunogenic potential of brolucizumab, including a linear epitope shared with bacteria, non-natural surfaces due to the single-chain variable fragment format, and non-native drug species that may form over prolonged time in the eye. Consideration of intraocular drug pharmacology and disease state in a quantitative systems pharmacology model suggested that immune complexes could form at immunologically relevant concentrations modulated by dose intensity. Assays using circulating immune cells from treated patients or treatment-naïve healthy volunteers revealed the capacity of immune complexes to trigger cellular responses such as enhanced antigen presentation, platelet aggregation, endothelial cell activation, and cytokine release. Together, these studies informed a mechanistic understanding of the clinically observed immunogenicity of brolucizumab and associated cases of RV/RO.

In October 2019, Novartis launched brolucizumab, a single-chain variable fragment molecule targeting vascular endothelial growth factor A, for the treatment of neovascular age-related macular degeneration. In 2020, rare cases of retinal vasculitis and/or retinal vascular occlusion (RV/RO) were reported, often during the first few months after treatment initiation, consistent with a possible immunologic pathobiology. This finding was inconsistent with preclinical studies in cynomolgus monkeys that demonstrated no drug-related intraocular inflammation, or RV/RO, despite the presence of preexisting and treatment-emergent antidrug antibodies (ADAs) in some animals. In this study, the immune response against brolucizumab in humans was assessed using samples from clinical trials and clinical practice. In the brolucizumab-naïve population, anti-brolucizumab ADA responses were detected before any treatment, which was supported by the finding that healthy donors can harbor brolucizumab-specific B cells. This suggested prior exposure of the immune system to proteins with structural similarity. Experiments on samples showed that naïve and brolucizumab-treated ADA-positive patients developed a class-switched, high-affinity immune response, with several linear epitopes being recognized by ADAs. Only patients with RV/RO showed a meaningful T cell response upon recall with brolucizumab. Further studies in cynomolgus monkeys preimmunized against brolucizumab with adjuvant followed by intravitreal brolucizumab challenge demonstrated that high ADA titers were required to generate ocular inflammation and vasculitis/vascular thrombosis, comparable to RV/RO in humans. Immunogenicity therefore seems to be a prerequisite to develop RV/RO. However, because only 2.1% of patients with ADA develop RV/RO, additional factors must play a role in the development of RV/RO.

Purpose: To evaluate whether topical acrizanib (LHA510), a small-molecule vascular endothelial growth factor receptor inhibitor, could suppress the need for anti-vascular endothelial growth factor therapy over a 12-week period in patients with neovascular age-related macular degeneration., Design: A phase 2 multicenter randomized double-masked, vehicle-controlled proof-of-concept study., Methods: Trial includes n = 90 patients with active choroidal neovascularization due to neovascular age-related macular degeneration and under anti-vascular endothelial growth factor treatment. All patients received an intravitreal injection of ranibizumab at baseline and were retreated when there was evidence of disease recurrence (rescue). Patients were randomized 1:1 to receive topical LHA510 or vehicle for 12 weeks. Drops were administered twice a day for 8 weeks and then 3 times a day for the last 4 weeks., Main Outcome Measure: The primary outcome was the number of patients requiring rescue over 84 days of topical dosing. Key secondary outcome measures were time to first rescue, total number of ranibizumab injections, changes in central subfield thickness, and changes of visual acuity from baseline to day 84., Results: The extended per protocol set included 70 patients of whom 25 of 33 patients in the LHA510 group (75.8%) and 25 of 37 patients in the placebo group (67.6%) required rescue by day 84 (P = .8466). Secondary and subgroup analysis did not support evidence of efficacy. Twenty-one of 46 patients administered LHA510 developed a reversible corneal haze that resolved with cessation of treatment and did not recur in patients restarted at once daily frequency., Conclusion: In spite of extensive optimization for topical efficacy, LHA510 failed to demonstrate clinical efficacy., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

Age-related macular degeneration (AMD) is one of the most common causes of visual impairment in the elderly, with a complex and still poorly understood etiology. Whole-genome association studies have discovered 34 genomic regions associated with AMD. However, the genes and cognate proteins that mediate the risk, are largely unknown. In the current study, we integrate levels of 4782 human serum proteins with all genetic risk loci for AMD in a large population-based study of the elderly, revealing many proteins and pathways linked to the disease. Serum proteins are also found to reflect AMD severity independent of genetics and predict progression from early to advanced AMD after five years in this population. A two-sample Mendelian randomization study identifies several proteins that are causally related to the disease and are directionally consistent with the observational estimates. In this work, we present a robust and unique framework for elucidating the pathobiology of AMD., (© 2022. The Author(s).)

We sought to refine understanding about associations identified in prior studies between angiotensin-II receptor blockers, metformin, selective serotonin reuptake inhibitors, fibric-acid derivatives, or calcium channel blockers and progression to glaucoma filtration surgery for open-angle glaucoma (OAG). We used new-initiator, active-comparator cohort designs to investigate these drugs in two data sources. We adjusted for confounders using stabilized inverse-probability-of-treatment weights and evaluated results using "intention-to-treat" and "as-treated" follow-up approaches. In both data sources, Kaplan-Meier curves showed trends for more rapid progression to glaucoma filtration surgery in patients taking calcium channel blockers compared with thiazides with as-treated (MarketScan P = 0.15; Medicare P = 0.03) and intention-to-treat follow-up (MarketScan P < 0.01; Medicare P = 0.10). There was suggestion of delayed progression for selective serotonin reuptake inhibitor compared with tricyclic antidepressants in Medicare, which was not observed in MarketScan. Our study provided support for a relationship between calcium channel blockers and OAG progression but not for other investigated drugs., (© 2019 The Authors Clinical Pharmacology & Therapeutics © 2019 American Society for Clinical Pharmacology and Therapeutics.)

The future of next generation therapeutics for glaucoma is strong. The recent approval of two novel intraocular pressure (IOP)-lowering drugs with distinct mechanisms of action is the first in over 20 years. However, these are still being administered as topical drops. Efforts are underway to increase patient compliance and greater therapeutic benefits with the development of sustained delivery technologies. Furthermore, innovations from biologics- and gene therapy-based therapeutics are being developed in the context of disease modification, which are expected to lead to more permanent therapies for patients. Neuroprotection, including the preservation of retinal ganglion cells (RGCs) and optic nerve is another area that is actively being explored for therapeutic options. With improvements in imaging technologies and determination of new surrogate clinical endpoints, the therapeutic potential for translation of neuroprotectants is coming close to clinical realization. This review summarizes the aforementioned topics and other related aspects.

Purpose: To evaluate whether inhibition of the proinflammatory cytokines IL-1β or IL-17A by canakinumab or secukinumab, respectively, influence the signs and symptoms of dry eye., Methods: In a randomized, double-masked, placebo-controlled, outpatient clinical trial, 72 patients with moderate to severe dry eye were randomly assigned in a 1:1:1 ratio to treatment with a single intravenous dose of canakinumab, of secukinumab, or of placebo. Signs and symptoms of dry eye were evaluated on the treatment day and 1 week, 4 weeks, and 8 weeks after treatment. The prespecified primary efficacy endpoint was corneal staining in the study eye 4 weeks after treatment. Secondary endpoints included tear production (Schirmer test), tear film breakup time, conjunctival redness, the ocular surface disease index (OSDI), the frequency of a desire for a topical ocular lubricant, and visual acuity., Results: Of the 71 patients included in the analysis of safety, the rate of adverse events was similar between treatment groups. The course of corneal staining scores from baseline to 4 weeks, respectively, were for canakinumab 1.46 to 1.33 (P = 0.62 compared with placebo), for secukinumab 1.46 to 1.23 (P = 0.22), and for placebo 1.68 to 1.42. There were no changes in the other measures of efficacy beyond what was within the range expected for stochastic day-to-day variation., Conclusions: The results suggest that the inhibition of IL-1β or IL-17A obtained by systemic administration of neutralizing drugs does not influence the severity of dry eye.

Purpose: Secukinumab, a fully human anti-interleukin-17A monoclonal antibody, exhibited promising activity in a proof-of-concept study when administered in intravenous (IV) doses to patients with active, chronic, noninfectious uveitis. This study compared the efficacy and safety of different IV and subcutaneous (SC) doses of secukinumab in patients with noninfectious uveitis., Design: Multicenter, randomized, double-masked, dose-ranging, phase 2 clinical trial., Participants: Thirty-seven patients with active noninfectious intermediate uveitis, posterior uveitis, or panuveitis who required corticosteroid-sparing immunosuppressive therapy., Methods: Patients were randomized to secukinumab 300 mg SC every 2 weeks for 4 doses, secukinumab 10 mg/kg IV every 2 weeks for 4 doses, or secukinumab 30 mg/kg IV every 4 weeks for 2 doses. Intravenous or SC saline was administered to maintain masking. Efficacy was assessed on day 57 (2-4 weeks after last dose)., Main Outcome Measures: Percentage of patients with treatment response, defined as (1) at least a 2-grade reduction in vitreous haze score or trace or absent vitreous haze in the study eye without an increase in corticosteroid dose and without uveitis worsening or (2) reduction in corticosteroid dosages to prespecified levels without uveitis worsening. Percentage of patients with remission, defined as anterior chamber cell and vitreous haze scores of 0 or 0.5+ in both eyes without corticosteroid therapy or uveitis worsening., Results: Secukinumab 30 mg/kg IV and 10 mg/kg IV, compared with the 300 mg SC dose, produced higher responder rates (72.7% and 61.5% vs. 33.3%, respectively) and remission rates (27.3% and 38.5% vs. 16.7%, respectively). Statistical and clinical superiority for the 30 mg/kg IV dose compared with the 300 mg SC dose was established in a Bayesian probability model. Other measures, including time to response onset, change in visual acuity, and change in vitreous haze score, showed numeric trends favoring IV dosing. Secukinumab, administered in IV or SC formulations, appeared safe and was well tolerated., Conclusions: Intravenous secukinumab was effective and well tolerated in noninfectious uveitis requiring systemic corticosteroid-sparing immunosuppressive therapy. Greater activity with IV dosing suggests that patients may not receive sufficient drug with SC administration. High-dose IV secukinumab may be necessary to deliver secukinumab in therapeutic concentrations., (Copyright © 2015 American Academy of Ophthalmology. Published by Elsevier Inc. All rights reserved.)

Purpose: Exfoliation syndrome (ES) is commonly associated with glaucoma, premature cataracts, and other ocular and systemic pathologies. LOXL1 gene variants are significantly associated with ES; however, the role of the protein in ES development remains unclear. The purpose of this study was to characterize the ocular phenotype in Loxl1(-/-) (null) mice., Methods: Loxl1 null mice and strain-matched controls (C57BL) were evaluated by clinical and histologic analyses., Results: Anterior segment histology showed a pronounced vesiculation of the anterior lens in the null mice. The lesions were subcapsular and in direct apposition with the posterior iris surface. Fluorescein angiography showed increased diffusion of fluorescein into the anterior chamber of the null mice compared with age-matched controls (P = 0.003, two-tailed, unequal variance t-test), suggesting compromise of the blood-aqueous barrier. Intraocular pressure measurements were within the normal range (16.5 ± 2.0 mm Hg) in null mice up to 1 year of age. Immunohistochemistry showed decreased elastin in the iris and ciliary body in the null mouse compared with controls., Conclusions: Elimination of LOXL1 in mice impairs the blood-aqueous humor barrier in the ocular anterior segment and causes lens abnormalities consistent with cataract formation, but does not result in deposition of macromolecular material or glaucoma. These results show that mice lacking LOXL1 have some ES features but that complete disease manifestation requires other factors that could be genetic and/or environmental.

Purpose: To assess the association between single nucleotide polymorphisms (SNPs) of the gene region containing cyclin-dependent kinase inhibitor 2B antisense noncoding RNA (CDKN2B-AS1) and glaucoma features among primary open-angle glaucoma (POAG) patients., Design: Retrospective observational case series., Methods: We studied associations between 10 CDKN2B-AS1 SNPs and glaucoma features among 976 POAG cases from the Glaucoma Genes and Environment (GLAUGEN) study and 1971 cases from the National Eye Institute Glaucoma Human Genetics Collaboration (NEIGHBOR) consortium. For each patient, we chose the feature from the eye with the higher value. We created cohort-specific multivariable models for glaucoma features and then meta-analyzed the results., Results: For 9 of the 10 protective CDKN2B-AS1 SNPs with minor alleles associated with reduced disease risk (eg, the G allele at rs2157719), POAG patients carrying these minor alleles had smaller cup-to-disc ratio (0.05 units smaller per G allele at diagnosis; 95% CI: -0.08, -0.03; P = 6.23E-05) despite having higher intraocular pressure (IOP) (0.70 mm Hg higher per G allele at DNA collection; 95% CI: 0.40, 1.00; P = 5.45E-06). For the 1 adverse rs3217992 SNP with minor allele A associated with increased disease risk, POAG patients with A alleles had larger cup-to-disc ratio (0.05 units larger per A allele at diagnosis; 95% CI: 0.02, 0.07; P = 4.74E-04) despite having lower IOP (-0.57 mm Hg per A allele at DNA collection; 95% CI: -0.84, -0.29; P = 6.55E-05)., Conclusion: Alleles of CDKN2B-AS1 SNPs, which influence risk of developing POAG, also modulate optic nerve degeneration among POAG patients, underscoring the role of CDKN2B-AS1 in POAG., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Purpose: To examine the effects of caffeinated coffee consumption on intraocular pressure (IOP), ocular perfusion pressure (OPP), and ocular pulse amplitude (OPA) in those with or at risk for primary open-angle glaucoma (POAG)., Methods: We conducted a prospective, double-masked, crossover, randomized controlled trial with 106 subjects: 22 with high tension POAG, 18 with normal tension POAG, 20 with ocular hypertension, 21 POAG suspects, and 25 healthy participants. Subjects ingested either 237 ml of caffeinated (182 mg caffeine) or decaffeinated (4 mg caffeine) coffee for the first visit and the alternate beverage for the second visit. Blood pressure (BP) and pascal dynamic contour tonometer measurements of IOP, OPA, and heart rate were measured before and at 60 and 90 min after coffee ingestion per visit. OPP was calculated from BP and IOP measurements. Results were analysed using paired t-tests. Multivariable models assessed determinants of IOP, OPP, and OPA changes., Results: There were no significant differences in baseline IOP, OPP, and OPA between the caffeinated and decaffeinated visits. After caffeinated as compared with decaffeinated coffee ingestion, mean mm Hg changes (± SD) in IOP, OPP, and OPA were as follows: 0.99 (± 1.52, P<0.0001), 1.57 (± 6.40, P=0.0129), and 0.23 (± 0.52, P<0.0001) at 60 min, respectively; and 1.06 (± 1.67, P<0.0001), 1.26 (± 6.23, P=0.0398), and 0.18 (± 0.52, P=0.0006) at 90 min, respectively. Regression analyses revealed sporadic and inconsistent associations with IOP, OPP, and OPA changes., Conclusion: Consuming one cup of caffeinated coffee (182 mg caffeine) statistically increases, but likely does not clinically impact, IOP and OPP in those with or at risk for POAG.

Calcineurin (CaN) activation is critically involved in the regulation of spine morphology in response to oligomeric amyloid-β (Aβ) as well as in synaptic plasticity in normal memory, but no existing techniques can monitor the spatiotemporal pattern of CaN activity. Here, we use a spectral fluorescence resonance energy transfer approach to monitor CaN activation dynamics in real time with subcellular resolution. When oligomeric Aβ derived from Tg2576 murine transgenic neurons or human AD brains were applied to wild-type murine primary cortical neurons, we observe a dynamic progression of CaN activation within minutes, first in dendritic spines, and then in the cytoplasm and, in hours, in the nucleus. CaN activation in spines leads to rapid but reversible morphological changes in spines and in postsynaptic proteins; longer exposure leads to NFAT (nuclear factor of activated T-cells) translocation to the nucleus and frank spine loss. These results provide a framework for understanding the role of calcineurin in synaptic alterations associated with AD pathogenesis.

Purpose: We previously reported that calcineurin, a Ca(2+)/calmodulin-dependent serine/threonine phosphatase, is activated and proposed that it participates in retinal ganglion cell (RGC) apoptosis in two rodent ocular hypertension models. In this study, we tested whether calcineurin activation by itself, even in the absence of ocular hypertension, is sufficient to cause RGC degeneration., Methods: We compared RGC and optic nerve morphology after adeno-associated virus serotype 2 (AAV2)-mediated transduction of RGCs with constitutively active calcineurin (CaNCA) or unactivated, wild-type calcineurin (CaNwt). Retinas and optic nerves were harvested 7-16 weeks after injection of the AAV into mouse vitreous. In flatmounted retinas, the transduced RGCs were identified with immunohistochemistry. The morphology of the RGCs was revealed by immunostaining for neurofilament SMI32 or by using GFP-M transgenic mice. A modified Sholl analysis was applied to analyze the RGC dendritic morphology. Optic nerve damage was assessed with optic nerve grading according to the Morrison standard., Results: CaNwt and CaNCA were highly expressed in the injected eyes. Compared to the CaNwt-expressing RGCs, the CaNCA-expressing RGCs had smaller somas, smaller dendritic field areas, shorter total dendrite lengths, and simpler dendritic branching patterns. At 16 weeks, the CaNCA-expressing eyes had greater optic nerve damage than the CaNwt-expressing eyes., Conclusions: Calcineurin activation is sufficient to cause RGC dendritic degeneration and optic nerve damage. These data support the hypothesis that calcineurin activation is an important mediator of RGC degeneration, and are consistent with the hypothesis that calcineurin activation may contribute to RGC neurodegeneration in glaucoma.

Purpose: The purpose of this study was to evaluate the incidence of glaucoma in eyes with severe chemical burn, before and after keratoprosthesis., Methods: A retrospective chart review of 28 eyes of 23 patients with severe ocular chemical burns who had undergone Boston Keratoprosthesis (BKPro) surgery at the Massachusetts Eye and Ear Infirmary, between 1990 and 2010. The incidence and severity of the outcome of glaucoma, preoperatively and postoperatively, were reviewed. Related issues, such as type of chemical burn; visual acuity (VA); device retention rate; number and nature of previous, concomitant, and subsequent procedures; and incidence of other postoperative complications, were reviewed for a median follow-up time of 57 months., Results: The number of eyes with a preoperative history or signs of glaucoma was 21, 9 of which had glaucoma progression after BKPro implantation. In addition, 2 more eyes developed glaucoma postoperatively. Preoperative vision was counting fingers or worse in all eyes. Best-corrected postoperative VA ranged from no light perception to 20/20. Seventeen eyes (61%) achieved 20/60 or better VA at some point during their follow-up, but only 9 (32%) maintained 20/60 at the last follow-up. Of the 28 eyes, 6 had the BKPro replaced once and 1 had it replaced twice. Superimposed, 8 of the most severely burned patients developed retinal detachment postoperatively., Conclusions: Glaucoma is very common in eyes with severe chemical burns. A keratoprosthesis can rehabilitate vision, but postoperative glaucoma can be difficult to manage.

The Publisher regrets that this article is an accidental duplication of an article that has already been published, doi:10.1016/j.exer.2010.04.002. The duplicate article has therefore been withdrawn., (Copyright © 2011. Published by Elsevier Ltd.. All rights reserved.)

Low levels of hypoxia have been suggested to be a mechanism of retinal damage in glaucoma. To test the hypothesis that the activation of the hypoxia-responsive transcription factor hypoxia inducible factor-1alpha (HIF-1alpha) is involved in the pathophysiology of glaucoma, we used a rat model of glaucoma to study (1) HIF-1alpha retinal protein levels by immunoblot analysis, (2) cellular localization of HIF-1alpha in the retina by immunohistochemistry, and (3) expression of retinal HIF-1 gene targets by quantitative real-time polymerase chain reaction. Glaucoma was unilaterally induced in rats by injecting hypertonic saline in episcleral veins. We find that HIF-1alpha protein was increased in the retina following elevation of intraocular pressure, specifically in Müller glia and astrocytes but not in activated microglia. Eight established HIF-1 target genes were measured in experimental glaucoma. Retinal Epo, Flt-1, Hsp-27, Pai-1, and Vegfa mRNA levels were increased and Et-1, Igf2, and Tgfbeta3 levels were decreased in the glaucomatous retinas. Thus, the increase in HIF-1alpha levels in Müller glia and astrocytes is accompanied by a marked up regulation of some, but not all, HIF-1 transcriptional targets. These data support the hypothesis that HIF-1alpha becomes transcriptionally active in astrocytes and Müller cells but not microglia or neurons in glaucoma, arguing against a global hypoxia stimulus to the retina.

Purpose: To evaluate the elastin gene (ELN) as a secondary risk factor for pseudoexfoliation syndrome (PXFS) and the associated glaucoma pseudoexfoliation glaucoma (PXFG)., Methods: One hundred seventy-eight unrelated patients with PXFS, including 132 patients with PXFG, and 113 unrelated controls were recruited from the Massachusetts Eye and Ear Infirmary. All the patients and controls were white of European ancestry. Three tag SNPs (rs2071307, rs3823879, and rs3757587) that capture the majority of alleles in ELN were genotyped. Single-SNP association was analyzed using Fisher exact test. Haplotype analysis and the set-based test were used to assess the association for the whole gene. Interaction analysis was done between the ELN SNP rs2071307 and LOXL1 SNP rs2165241 using logistic regression. Multiple comparisons were corrected using the Bonferroni method., Results: All 3 ELN tag SNPs were not significantly associated with PXFS and PXFG (P>0.20). The minor allele frequencies in PXFS, PXFG, and controls were 40.7%, 39.8%, and 45.6%, respectively for rs2071307, 6.7%, 6.3%, and 5.4% for rs3823879, and 14.8%, 16.2%, and 13.6% for rs3757587. Haplotype analysis and the set-based test did not find significant association of ELN with PXFS (P=0.94 and 0.99, respectively). No significant interaction effects on PXFS were identified between the ELN and LOXL1 SNPs (P=0.55)., Conclusions: Our results suggest that common polymorphisms of ELN are not associated with PXFS and PXFG in white populations. Further studies are required to identify secondary genetic factors that contribute to PXFS.

Purpose: A significant number of survivors of hematopoietic stem-cell transplantation (HSCT) report enduring adverse effects of treatment, including illness-related post-traumatic stress disorder (PTSD) symptoms and general distress. We report results of a randomized clinical trial that tested the effects of a 10-session, telephone-administered cognitive-behavioral therapy (CBT) intervention on PTSD, depression, and distress symptoms., Methods: Survivors who had undergone HSCT 1 to 3 years earlier (N = 408) were assessed for study eligibility. Those who met study eligibility criteria (n = 89) completed a baseline assessment that included a clinical interview and self-report measures of PTSD symptoms (the primary outcome) and depression and general distress (the secondary outcomes). Next, they were randomly assigned to CBT or an assessment-only condition. Survivors in the CBT group completed 10 individual telephone-based CBT sessions (T-CBT) that included strategies to reduce PTSD symptoms, depression, and general distress. Follow-up assessments occurred at 6, 9, and 12 months after the baseline assessment., Results: Linear mixed-model analyses revealed that, compared with HSCT survivors in the assessment-only condition, survivors who completed T-CBT reported fewer illness-related PTSD symptoms, including less avoidance (P < .001) and fewer intrusive thoughts (P < .05) as well as less general distress and fewer depressive symptoms (P < .05) even after controlling for potential demographic and medical covariates. These results were consistent across the three follow-up assessments., Conclusion: A brief, telephone-administered CBT intervention developed for HSCT survivors is an efficacious treatment for reducing illness-related PTSD symptoms and general distress.

Purpose: Intraocular pressure (IOP) is an important risk factor in glaucoma. Gene expression changes were studied in glaucomatous rat retinal ganglion cells (RGCs) to elucidate altered transcriptional pathways., Methods: RGCs were back-labeled with Fluorogold. Unilateral IOP elevation was produced by injection of hypertonic saline into the episcleral veins. Laser capture microdissection (LCM) was used to capture an equal number of RGCs from normal and glaucomatous retinal sections. RNA was extracted and amplified, labeled, and hybridized to rat genome microarrays, and data analysis was performed. After selected microarray data were confirmed by RT-qPCR and immunohistochemistry, biological pathway analyses were performed., Results: Significant changes were found in the expression of 905 genes, with 330 genes increasing and 575 genes decreasing in glaucomatous RGCs. Multiple cellular pathways were involved. Ingenuity pathway analysis demonstrated significant changes in cardiac beta-adrenergic signaling, interferon signaling, glutamate receptor signaling, cAMP-mediated signaling, chemokine signaling, 14-3-3-mediated signaling, and G-protein-coupled receptor signaling. Gene set enrichment analysis showed that the genes involved in apoptotic pathways were enriched in glaucomatous RGCs. The prosurvival gene Stat3 was upregulated in response to elevated IOP, and immunohistochemistry confirmed that Stat3 and phosphorylated-Stat3 levels were increased in RGCs in experimental glaucoma. In addition, the expression of several prosurvival genes normally expressed in RGCs was decreased., Conclusions: There are extensive changes in gene expression in glaucomatous RGCs involving multiple molecular pathways, including prosurvival and prodeath genes. The alteration in the balance between prosurvival and prodeath may contribute to RGC death in glaucoma.

Glaucoma is a disease in which retinal ganglion cells (RGCs) die leading ultimately to blindness. Over the past decade and a half, information has begun to emerge regarding specific molecular responses of the retina to conditions of elevated intraocular pressure (IOP). It is now clear that the state of the RGC in glaucoma depends on a balance of pro-survival and pro-death pathways in the retina and details of these responses are still being worked out. In this review, we will discuss the evidence supporting the involvement of specific apoptotic cascades as well as the insults that trigger RGC apoptosis. In addition, we will present evidence supporting the existence of endogenous protective mechanisms as well as exogenous neuroprotective strategies., (Copyright 2010 Elsevier Ltd. All rights reserved.)

Purpose: Glaucoma is a neurodegenerative disease in which elevated intraocular pressure (IOP) leads to progressive loss of retinal ganglion cells (RGCs) and blindness. Calcium dyshomeostasis has been suggested to play a role in the pathologic events that lead to RGC loss, though the details of these events are not well understood. Calcium-induced activation of calpain has been shown to contribute to neuronal death in a wide variety of neurodegenerative diseases. The authors hypothesize that similar events occur in glaucoma., Methods: The authors used a well-established rat model of experimental glaucoma. Retinal tissues were harvested after 5 or 10 days of elevated IOP and were subjected to immunoblot analysis, immunoprecipitation, and MALDI-ProTOF/MS peptide fingerprint mapping. Immunohistochemistry was used to localize calpain activation., Results: The authors present four independent lines of evidence that calpain is activated in experimental glaucoma. First, they showed that a 55-kDa autocatalytic active form of calpain is detected on immunoblot analysis. Second, they demonstrated the cleavage of two well-established calpain substrates, spectrin and calcineurin, only in eyes with elevated IOP. Third, they used MALDI-ProTOF to analyze cleaved calcineurin and immunoblot analysis of spectrin cleavage products and showed that both substrates were cleaved by calpain in experimental glaucoma. Fourth, they used immunohistochemistry to show that calpain-mediated spectrin cleavage occurs in RGCs under conditions of elevated IOP., Conclusions: These data support the hypothesis that calpain is activated under conditions of elevated intraocular pressure and provide further details of the pathologic events leading to RGC loss in glaucoma.

Amyloid beta (Abeta)-containing plaques are surrounded by dystrophic neurites in the Alzheimer's disease (AD) brain, but whether and how plaques induce these neuritic abnormalities remain unknown. We tested the hypothesis that soluble oligomeric assemblies of Abeta, which surround plaques, induce calcium-mediated secondary cascades that lead to dystrophic changes in local neurites. We show that soluble Abeta oligomers lead to activation of the calcium-dependent phosphatase calcineurin (CaN) (PP2B), which in turn activates the transcriptional factor nuclear factor of activated T cells (NFAT). Activation of these signaling pathways, even in the absence of Abeta, is sufficient to produce a virtual phenocopy of Abeta-induced dystrophic neurites, dendritic simplification, and dendritic spine loss in both neurons in culture and in the adult mouse brain. Importantly, the morphological deficits in the vicinity of Abeta deposits in a mouse model of AD are ameliorated by CaN inhibition, supporting the hypothesis that CaN-NFAT are aberrantly activated by Abeta and that CaN-NFAT activation is responsible for disruption of neuronal structure near plaques. In accord with this, we also detect increased levels of an active form of CaN and NFATc4 in the nuclear fraction from the cortex of patients with AD. Thus, Abeta appears to mediate the neurodegeneration of AD, at least in part, by activation of CaN and subsequent NFAT-mediated downstream cascades.

Purpose: Glaucoma is a frequent and often severe problem in patients needing keratoprosthesis (KPro) surgery. Standard glaucoma shunts in these cases often develop a very dense capsule that obstructs flow and causes intraocular pressure elevation. Therefore, we developed shunts that deliver aqueous to distant epithelialized cavities where an obstructing capsule is less likely to form. In this retroactive study of such shunts, the risk of postoperative infection (especially endophthalmitis) has been assessed., Methods: Ahmed shunts were connected with a tube to the lacrimal sac or ethmoid sinuses. In subsequent designs the polypropylene plate was eliminated, the valve was enclosed and a distal tube added (New World Medical Inc, Rancho Cucamonga, CA). The latter shunts were connected to the maxillary sinus or lower lid fornix. Between 2001 and 2005 the devices were implanted in 34 patients with very severe ocular disease. (31 had KPro implanted). Thus, the distal tube was led to the lacrimal sac (2 cases), to the ethmoid sinuses (6), to the maxillary sinus (16 cases), and to the lower lid fornix (10 cases). The patients were followed for cumulatively 145 shunt years, with a mean follow-up of 4 years and 3 months. They were instructed to administer low-dose topical antibiotics indefinitely., Results: Only 1 case of acute bacterial endophthalmitis occurred, thus an incidence of 0.7% per shunt year. A slow-growing Mycobacterium was cultured in a disintegrating eye and 1 bacterial maxillary sinusitis occurred, both in autoimmune diseases. Three valves became exposed and had to be removed (1 replaced). Four eyes developed hypotony, probably from valve failure., Conclusions: The shunt arrangement, which connects the anterior chamber to potentially microbe-populated cavities or to the lower lid fornix and its flora, might be suspected to allow rapid retrograde invasion of infective agents, resulting in endophthalmitis. In this series of KPro patients, however, the incidence of severe infection was very low, in fact comparable to that after standard trabeculectomy.

Purpose: To evaluate the efficacy and safety of diode laser transscleral cyclophotocoagulation (DLTSC) to control intraocular pressure (IOP) in keratoprosthesis patients with uncontrolled glaucoma., Patients and Methods: Between 1993 and 2007, 18 eyes of 18 patients underwent DLTSC, either before (n=3), during (n=1), or after (n=14) keratoprosthesis surgery. Keratoprosthesis type I was used in 72%. All but one of these patients received an Ahmed Glaucoma Valve, either with or after the keratoprosthesis placement. Best-corrected visual acuity, IOP (assessed by digital palpation), number of medications, and complications were recorded preoperatively, at day 7, at 1, 3, and 6 months then every 6 months postoperatively., Results: Mean follow-up was 26.6+/-19.6 months (mean+/-SD) and mean age was 50.1+/-15.6 years. Glaucoma was identified in 11 eyes before keratoprosthesis surgery and in 7 eyes after. Mean postoperative IOP was significantly reduced at 6, 12, 24, 36, and 48 months after DLTSC. DLTSC was repeated in 6 eyes. At final visit, mean best-corrected visual acuity was not decreased and there were no statistically significant differences in the number of glaucoma medications. Two patients had complications after DLTSC: a conjunctival dehiscence and a fungal endophthalmitis., Conclusions: DLTSC has beneficial long-term effects in the control of IOP and can be considered in the management of keratoprosthesis patients with refractory glaucoma.

Purpose: Although ischemia has previously been suggested to contribute to the pathogenesis of glaucoma, neovascularization is not implicated in glaucoma. Because vascular endothelial growth factor-A (VEGF-A) is a key mediator in neovascularization response, we investigated the levels of the major pro-angiogenic (VEGF-A164) and anti-angiogenic VEGF-A subtypes (VEGF-A165b) in the retina during experimental glaucoma., Methods: Glaucoma was induced unilaterally in rats by injecting 1.9 M hypertonic saline solution in the episcleral veins. The contralateral eye served as the control. The intraocular pressure (IOP) of each eye was measured via Tonopen in conscious rats. Eyes were enucleated either on the 5th or the 10th day of elevated IOP. Whole retinal lysates were separated by SDS-PAGE and transferred to PVDF membranes. Levels of VEGF-A164 and VEGF-A165b were analyzed by western blotting using specific antibodies. In a different group of rats, retinal ganglion cells were retrogradely labeled by injecting Fluorogold in the superior colliculus a week before the induction of glaucoma. After the eyes were enucleated on the fifth day of elevated IOP, posterior eye cups were sectioned using a cryostat. Levels and localization of VEGF-A164 and VEGF-A165b were examined in retinal sections by immunohistochemistry., Results: VEGF-A164 levels remained unchanged between the control and glaucomatous retinas after five days (p=0.341) and 10 days of elevated IOP (p=0.117). The presence of the anti-angiogenic VEGF-A isoform has not been previously reported in the rat. An antibody specific to VEGF-A165b detected the anti-angiogenic protein in the rat retina. VEGF-A165b levels were significantly increased (2.33+/-0.44 fold, p=0.014) in the glaucomatous retinas compared to those in controls after five days of elevated IOP. VEGF-A165b levels were not different (p=0.864) between the control and glaucomatous retinas following 10 days of elevated IOP. Expression of both VEGF-A164 and VEGF-A165b were observed in the retinal ganglion cells (RGC) and inner nuclear layer (INL)., Conclusions: Five day elevation of IOP leads to an increase in the anti-angiogenic VEGF-A165b levels but not in the pro-angiogenic VEGF-A164 levels in the glaucomatous retina. VEGF-A165b levels return to baseline after 10 days of elevated IOP, and VEGF-A164 levels remain unchanged. We speculate that the short-term elevation of VEGF-A165b levels and/or the unchanged levels of VEGF-A164 contribute to the lack of neovascularization in the glaucomatous retina.

Purpose: To describe periorbital changes induced by chronic topical therapy with daily bimatoprost 0.03% (Lumigan, Allergan Inc., Irvine, CA, U.S.A.)., Methods: A clinical investigation of 5 nonconsecutive patients with unilateral glaucoma treated daily with topical bimatoprost 0.03% for up to 4 years prior to presentation., Results: In eyes treated with bimatoprost 0.03% the authors noted periorbital fat atrophy, deepening of the upper eyelid sulcus, relative enophthalmos, loss of the lower eyelid fullness, and involution of dermatochalasis compared with the fellow untreated eye. By inspecting old photographs the authors confirmed that these unilateral changes were not present prior to starting bimatoprost. In addition, these changes were partially reversible after discontinuation of the medication, whenever that was possible. In 2 cases imaging studies confirmed the clinical impression that these findings were not related to primary orbital pathology., Conclusions: Physicians and patients should be aware of the potential of bimatoprost 0.03% to produce periorbital changes.

Retinal ganglion cells (RGCs) are the only output neurons of the retina, and their degeneration after damage to the optic nerve or in glaucoma is a well established system for studying apoptosis in the central nervous system. Frequently used procedures for assessing RGC number in retinal flat mounts suffer from two problems: RGC densities are not uniform across retinal flat mounts, and density measures may therefore not reflect total number, and flat mounts do not allow efficient use of tissue. To overcome these problems we developed a stereological method for efficiently assessing RGC number in cryostat sections of the retina. We empirically demonstrate that only approximately 1:20 sections need be assessed to accurately estimate the total number of RGCs in the rat retina, providing ample tissue for additional studies in the same retina and saving considerably on more exhaustive sampling strategies. Using this method, we estimate that there are 86,282+/-4759 RGCs in the normal Brown Norway rat retina. These counts match well with estimates of axon counts in optic nerve. In a pilot study of experimental glaucoma, we determined a reduction of RGCs to 53,862+/-4272 (p<0.05). The current technique should prove advantageous to assess neuroprotective strategies in these experimental models.

Background: Pseudoexfoliation syndrome is a major risk factor for glaucoma in many populations throughout the world. Using a U.S. clinic-based case control sample with broad ethnic diversity, we show that three common SNPs in LOXL1 previously associated with pseudoexfoliation in Nordic populations are significantly associated with pseudoexfoliation syndrome and pseudoexfoliation glaucoma., Methods: Three LOXL1 SNPs were genotyped in a patient sample (206 pseudoexfoliation, 331 primary open angle glaucoma, and 88 controls) from the Glaucoma Consultation Service at the Massachusetts Eye and Ear Infirmary. The SNPs were evaluation for association with pseudeoexfoliation syndrome, pseudoexfoliation glaucoma, and primary open angle glaucoma., Results: The strongest association was found for the G allele of marker rs3825942 (G153D) with a frequency of 99% in pseudoexfoliation patients (with and without glaucoma) compared with 79% in controls (p = 1.6 x 10-15; OR = 20.93, 95%CI: 8.06, 54.39). The homozygous GG genotype is also associated with pseudoexfoliation when compared to controls (p = 1.2 x 10-12; OR = 23.57, 95%CI: 7.95, 69.85). None of the SNPs were significantly associated with primary open angle glaucoma., Conclusion: The pseudoexfoliation syndrome is a common cause of glaucoma. These results indicate that the G153D LOXL1 variant is significantly associated with an increased risk of pseudoexfoliation and pseudoexfoliation glaucoma in an ethnically diverse patient population from the Northeastern United States. Given the high prevalence of pseudooexfoliation in this geographic region, these results also indicate that the G153D LOXL1 variant is a significant risk factor for adult-onset glaucoma in this clinic based population.

Purpose: The role of heat shock proteins (Hsp) in injury response has been well established, but it is now becoming apparent that the phosphorylation state of Hsp27 may be a critical determinant of its ability to act in a protective capacity. In this study, the expression of Hsp27 and its phosphorylation were evaluated in an experimental glaucoma model in Brown Norway rats and in a spontaneous model of glaucoma in the DBA/2J mouse., Methods: The intraocular pressure (IOP) of one eye was elevated by injecting 1.9 M saline into the episcleral vein, as previously described in Brown Norway rats. IOP was measured in awake Brown Norway rats before surgery and every other day after saline injection. After 10 days of elevated IOP, the retinas were either removed for Western blot analysis or fixed for immunohistochemistry (IHC). IOP measurement in 7-month-old female DBA/2J mice was performed by direct cannulation. Retinas of eyes with and without elevated IOP were collected for Western blot analysis., Results: Western blot results showed a significant increase in total Hsp27 (3.9-fold; P < 0.05; n = 8) and phosphorylated Hsp27 (pHsp27) (2.1-fold; P < 0.05; n = 6) in high IOP eyes in the experimental rat glaucoma model, and similar increases were observed in DBA/2J mice with elevated IOP (3.1-fold and 2.2-fold for Hsp27 and pHsp27, respectively; P < 0.05; n = 5). In rats, increased Hsp27 and pHsp27 immunoreactivity were observed in the nerve fiber layer of elevated IOP eyes and colocalized with glial fibrillary acidic protein (GFAP) and with vimentin staining, suggesting that glial cells contribute to the increase in Hsp27 and pHsp27 seen in experimental glaucoma. No change in Hsp70 or Hsp90 was observed., Conclusions: These results confirm previous reports of elevated Hsp27 in experimental glaucoma and extend them to the DBA/2J mouse. In addition, a significant increase occurred in Hsp27 phosphorylation with elevated IOP in both models of glaucoma. IHC studies show that the increases in Hsp27 and pHsp27 occur primarily in glial cells.

Purpose: Photoreceptor apoptosis is associated with retinal detachment (RD) induced photoreceptor degeneration. Previously, we demonstrated the importance of caspase activation for RD-induced photoreceptor death in a rat model of RD. However, extracellular signals that precede the activation of caspases and photoreceptor degeneration remain unclear. The aim of this study is to characterize the molecular and cellular responses that occur after RD. The expression of cytokines, chemokines, and growth factors were examined in a rat model of RD., Methods: RD was induced in adult rats by subretinal injection of sodium hyaluronate. Retinal tissues were collected at various times (1, 3, 6, 24, and 72 h) after the induction of detachment. To screen for expressional changes in response to RD, major candidates for cytokines, chemokines, and growth factors were broadly examined by quantitative real time polymerase chain reaction (QPCR). To identify the cellular sources of the expressed genes, cells from various layers of the retina were obtained using laser capture microdissection (LCM), and their mRNAs were isolated. Protein expression was quantified by immunohistochemistry and Enzyme Linked-Immuno-Sorbent Assay (ELISA). To assess the potential of early response genes after RD to induce photoreceptor degeneration, exogenous recombinant proteins were subretinally injected and the photoreceptor cell death was assessed using a TdT-dUTP terminal nick-end labeling (TUNEL) assay at 24 h after RD., Results: At 72 h after RD a significant increase in mRNA levels for tumor necrosis factor alpha (TNF-alpha), interleukin-1beta (IL-1beta), monocyte chemotactic protein-1 (MCP-1), and basic fibroblast growth factor (bFGF) were detected in the neural retina. LCM revealed increased expression of mRNA for bFGF and MCP-1 in all retinal layers, though bFGF was especially evident in the outer nuclear layer (ONL) and MCP-1 in the inner nuclear layer (INL). TNF-alpha was increased in the ONL and the INL, and IL-1beta was increased in the ganglion cell layer. Time course experiments showed that TNF-alpha, IL-1beta and MCP-1 increased within 1 h after RD, while bFGF was increased by 24 h. Increased protein expression for TNF-alpha, IL-1beta, and MCP-1 was demonstrated by ELISA at 6 h after RD. Immunohistochemistry showed TNF-alpha and bFGF expression in the whole retina, with IL-1beta specifically expressed in astrocytes and MCP-1 in Müller cells. Subretinal administration of MCP-1 significantly increased TUNEL-positive cells in the ONL 24 h after RD, while injection of vehicle control, TNF-alpha, or IL-1beta showed no effect., Conclusions: Retinal glial cells, including astrocytes and Müller cells, are a major source of cytokine induction after RD. The increased expression and release of MCP-1 may be an important cause of photoreceptor degeneration associated with RD. This study helps to understand the mechanisms of RD-induced photoreceptor degeneration. Our results may provide new therapeutic targets to prevent photoreceptor degeneration following RD.

Objective: To better understand the course of glaucoma during pregnancy in women with preexisting disease., Methods: Retrospective case series of 28 eyes of 15 women with glaucoma followed up during pregnancy. Data were analyzed for age, race/ethnicity, medications, glaucoma type, intraocular pressure (IOP), and visual fields before, during, and after pregnancy., Results: In 16 (57.1%) of 28 eyes, IOP was stable during pregnancy, with no progression of visual field loss. In 5 eyes (17.9%), visual field loss progressed during pregnancy, while IOP remained stable or increased. In 5 eyes (17.9%), IOP increased during pregnancy, but there was no progression of visual field loss. In 2 eyes (7.1%), data were inconclusive because of medication noncompliance and preexisting severe visual field loss. Glaucoma medications were used by 13 of 15 patients to control glaucoma during pregnancy. The classes of medications used most frequently were beta-blockers, alpha2-adrenergic agents, cholinergic agents, and topical carbonic anhydrase inhibitors., Conclusions: The course of glaucoma during pregnancy is variable, and women must be monitored closely during pregnancy. Medications may be necessary to control IOP and to prevent vision loss during pregnancy.

Our purpose was to evaluate the accuracy, reproducibility and predictive ability of two non-invasive tonometers developed for intraocular pressure (IOP) measurements in the mouse. The prototype impact-rebound tonometer (I-R) and a prototype optical interferometry tonometer (OIT) utilizing a fiberoptic pressure sensor, were compared. Enucleated eyes from C57/BL6 mice were used for the calibration. The anterior chamber was cannulated and the IOP was adjusted in increments of 5 cm of H2O (open stopcock method). A calibration curve was generated for each individual eye along with a master calibration curve for all eyes. Two operators measured the IOP. The instruments were then used in alternating order to measure the IOP in C57/BL6 and in DBA2/J animals. The same eyes were subsequently cannulated and the error of the non-invasive tonometers was determined. Both tonometers yielded almost equivalent ex vivo calibration curves with individual R2 of 0.9878 and 0.9902 respectively. Both instruments were highly reproducible. In vivo the I-R tonometer underestimated while the OIT overestimated the IOP. This error was systematic and therefore predictable. The confidence intervals of this error were determined by comparing the IOP estimates provided by each tonometer with the measurements obtained invasively by cannulation in vivo. The 95% CI of the error were 2.36 mmHg for the I-R and 2.62 mmHg for the OIT respectively. Non-invasive tonometry in the mouse is feasible. Both non-invasive instruments provide accurate and reproducible measurements with the OIT requiring calibration curves for each individual investigator.

Purpose: Thy1 is a surface glycoprotein uniquely expressed in retinal ganglion cells (RGCs) in retina. The aim of this study was to investigate the expression change of Thy1 in a model of experimental glaucoma., Methods: The change of protein and mRNA levels of Thy1 in the retina were studied using stereological counts of back-labeled RGCs, Western blot analysis, immunohistochemistry, and laser capture microdissection (LCM) of RGCs with quantitative PCR analysis of mRNA in a model of experimental glaucoma. LCM after optic nerve crush was also performed to evaluate Thy1 mRNA levels after a different injury., Results: After 10 days of elevated IOP, there was a 34% loss of RGC number, Thy1 protein decreased 60% in eyes with elevated intraocular pressure (IOP), and Thy1 mRNA levels decreased 51% in RGCs. Both protein and mRNA level change of Thy1 is to a much greater extent than RGC number loss., Conclusions: The current results confirm that Thy 1 mRNA levels do not reflect the number of RGCs present and extend this to include a parallel decrease in Thy1 protein levels. These results suggest that Thy1 serves as an early marker of RGC stress, but not a marker of RGC loss, in models of retinal damage.

In glaucoma, retinal ganglion cells (RGCs) die by apoptosis, generally attributed to an elevated intraocular pressure (IOP). We now describe the impact of elevated IOP in the rat on expression of caspase 8 and caspase 9, initiators of the extrinsic and intrinsic caspase cascades, respectively. Activation of both caspases was demonstrated by the presence of cleaved forms of the caspases and the detection of cleaved Bid and PARP, downstream consequences of caspase activation. Surprisingly, the absolute level of procaspase 9 was also elevated after 10 days of increased IOP. To examine the cause of increased levels of the procaspase, we used laser capture microdissection to capture Fluorogold back-labeled RGCs and real-time polymerase chain reaction to measure mRNA changes of initiating caspases. The mRNA levels of both caspase 8 and caspase 9 were increased specifically in RGCs. These data suggest that elevated IOP activates a transcriptional up-regulation and activation of initiating caspases in RGCs and triggers apoptosis through both extrinsic and intrinsic caspase cascades.

Increased intraocular pressure (IOP) leads, by an unknown mechanism, to apoptotic retinal ganglion cell (RGC) death in glaucoma. We now report cleavage of the autoinhibitory domain of the protein phosphatase calcineurin (CaN) in two rodent models of increased IOP. Cleaved CaN was not detected in rat or mouse eyes with normal IOP. In in vitro systems, this constitutively active cleaved form of CaN has been reported to lead to apoptosis via dephosphorylation of the proapoptotic Bcl-2 family member, Bad. In a rat model of glaucoma, we similarly detect increased Bad dephosphorylation, increased cytoplasmic cytochrome c (cyt c), and RGC death. Oral treatment of rats with increased IOP with the CaN inhibitor FK506 led to a reduction in Bad dephosphorylation and cyt c release. In accord with these biochemical results, we observed a marked increase in both RGC survival and optic nerve preservation. These data are consistent with a CaN-mediated mechanism of increased IOP toxicity. CaN cleavage was not observed at any time after optic nerve crush, suggesting that axon damage alone is insufficient to trigger cleavage. These findings implicate this mechanism of CaN activation in a chronic neurodegenerative disease. These data demonstrate that increased IOP leads to the initiation of a CaN-mediated mitochondrial apoptotic pathway in glaucoma and support neuroprotective strategies for this blinding disease.

Retinal ganglion cells die by apoptosis after optic nerve crush. FK506 has been shown to be neuroprotective in this model but the mechanism(s) by which it exerts these actions remains unknown. We and others have shown that caspase 9 is cleaved in the retina in other injury models and we hypothesized that the neuroprotection observed with FK506 was mediated by interference with caspase 9 activation. The present study examined the cellular localization of caspase 9 cleavage after intraorbital optic nerve crush in rats, the time course of caspase 9 cleavage after optic nerve crush and the ability of orally administered FK506 to block caspase 9 cleavage after optic nerve crush. We show by immunohistochemistry that cleaved caspase 9 is present in retinal ganglion cells (identified by prior backlabelling) after optic nerve crush. Immunoblot analysis showed that caspase 9 cleavage is significantly elevated 5 and 8 days after optic nerve crush. We show that orally administered FK506 reaches the retina and is pharmacologically active in retinal tissue. Furthermore, the oral administration of FK506 5 mg kg(-1) day(-1) blocks the cleavage of caspase 9 at both time points. These data suggest that caspase 9 activation may play an important role in retinal ganglion cell death following optic nerve crush and that the neuroprotection seen with FK506 may be mediated by interfering with the activation of caspase 9.

Objective: To evaluate the risk of cataract formation in young patients after initial trabeculectomy., Design: Retrospective, noncomparative case series., Participants: Thirty-four eyes from 27 patients undergoing initial trabeculectomy at the Glaucoma Consultation Service, Massachusetts Eye and Ear Infirmary (mean age, 43.7 years; range, 12-54 years)., Intervention: Follow-up averaged 42.6 months (range, 11-90 months)., Methods: Lens status was observed before surgery and at 3 months; 6 months; and 1, 2, 3, 4, 5, and 6 years after initial trabeculectomy., Main Outcome Measure: The main outcome measure was defined as cataract extraction for visually significant lenticular opacifications that developed after trabeculectomy., Results: The rate of cataract extraction after initial trabeculectomy was 24% (n = 8). The average time from trabeculectomy to cataract extraction was 26 months (range, 5-58 months). Progression of lenticular opacities occurred throughout the follow-up period. There was no increased rate of cataract formation in subjects with uveitic and steroid-induced glaucoma when compared with all other types of glaucoma. In the patients with both eyes in the study, the first eye was a predictor of cataract progression in the fellow eye., Conclusions: Cataract is a common complication after trabeculectomy in young patients. The 24% rate of cataract extraction after trabeculectomy reported in this study is a significant risk of which young patients contemplating surgery should be aware.

Purpose: We investigated retinal ganglion cell (RGC) death and activation of caspase 9 in rats with experimental glaucoma., Methods: Elevated intraocular pressure (IOP) was induced in rats using the Morrison model. Surviving backlabeled RGC were counted and TUNEL staining detected apoptosis. Procaspase 9 expression and activated caspase 9 were studied by immunoblot and immunohistochemistry., Results: IOP correlated with surviving RGC. TUNEL-positive RGC were observed in animals with elevated IOP. Procaspase 9 levels increased with IOP intensity. Cleaved caspase 9 was detected by immunoblot only in rats with peak IOP above 35 mm Hg for > or =6 days. Cleaved caspase 9 staining was seen only in the ganglion cell layer of retinas from rats with peak IOP > or =32 mm Hg., Conclusions: RGC loss is correlated with IOP in experimental glaucoma. These results support activation of caspase 9, the intrinsic caspase cascade, in RGC death in experimental glaucoma.

The optic neuropathy of glaucoma appears to be multifactorial in etiology. Both mechanical and vasogenic theories remain viable explanations for the observed nerve damage and the destructive effect of trophic withdrawal could be espoused by either theory. Each theory feeds into the final common pathway of cell death; even the immune system may kill cells in glaucoma by apoptosis. This knowledge has led to the hope that glaucomatous nerve damage can be curtailed or even prevented with the use of neuroprotective agents. Indeed, there are many intriguing new therapeutic possibilities on the horizon. Until evidence from clinical trials demonstrates the effectiveness of these agents however, control of IOP will remain the mainstay of treatment for glaucoma.

Objective: To compare the efficacy of brimonidine 0.2% with apraclonidine 0.5% in preventing intraocular pressure (IOP) elevations after anterior segment laser surgery., Design: Double-masked, randomized clinical trial., Participants: Sixty-six patients underwent either laser peripheral iridotomy, argon laser trabeculoplasty, or neodymium:yttrium-aluminum-garnet laser capsulotomy., Intervention: Eyes received either one drop of brimonidine 0.2% or apraclonidine 0.5% before laser surgery., Main Outcome Measures: Intraocular pressure, heart rate, and blood pressure were measured before laser surgery and at 1 hour, 3 hours, 24 hours, and 1 week after laser surgery., Results: Before the laser treatment, 33 patients (50.0%) received brimonidine 0.2% and 33 patients (50.0%) received apraclonidine 0.5%. Eight of 33 patients (24.2%) in the brimonidine-treated group and 9 of 33 patients (27.3%) in the apraclonidine group had postoperative IOP increases of 5 mmHg or more. This was not statistically different (P = 0.80). By the time of last follow-up examination, 3 of 33 patients (9.1%) in the brimonidine-treated group and 3 of 33 patients (9.1%) in the apraclonidine group had IOP increases of 10 mmHg or more. This was also not statistically different (P > or = 0.95). The mean IOP reduction from baseline in the brimonidine group (-2.8 +/- 2.8 mmHg) was not statistically different (P = 0.55) compared with the mean IOP reduction in the apraclonidine group (-3.6 +/- 3.3 mmHg). There were no statistically significant changes in mean heart rate or blood pressure in either group except for a slight reduction in diastolic blood pressure at 1 hour (P = 0.005) in the brimonidine group (-5.2 +/- 7.4 mmHg) compared with the apraclonidine group (-0.2 +/- 6.4 mmHg). There were no clinically significant side effects noted in either group., Conclusions: A single preoperative drop of brimonidine 0.2% is as effective as apraclonidine 0.5% in preventing IOP elevation immediately after anterior segment laser surgery.

Although uncommon, SO is a fearful postoperative complication because of its potential to blind both eyes. It can result not only from penetrating ocular surgery but also from nonpenetrating ocular procedures. Thus, it is important to consider in any patient who has undergone ocular surgery and develops bilateral uveitis, particularly because prompt, sufficient treatment is required to maximize visual outcome. It is also important to note that the disease may present with a spectrum of clinical findings, none of which is pathognomonic. Thus, suspicion is important for making the diagnosis. Treatment should address the T-cell-mediated nature of the disease. With appropriate treatment, visual acuity of no less than 20/60 is likely. However, before the start of treatment, which consists of immunosuppressants, infection must be ruled out and potential side effects of treatments must be considered. Furthermore, any patient with a history of SO needs ample immunosuppressant coverage for ocular procedures. Better understanding of the pathogenesis of the disease may lead to safer treatments that result in improved visual outcome and a cure. Meanwhile, because of its relapsing nature, SO requires continual, close surveillance, even after many years of quiescence.