Glaucoma - Next Generation Therapeutics: Impossible to Possible.

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.

Efficacy and safety of intravenous secukinumab in noninfectious uveitis requiring steroid-sparing immunosuppressive therapy.

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.

Distinct dendritic spine and nuclear phases of calcineurin activation after exposure to amyloid-ß revealed by a novel fluorescence resonance energy transfer assay.

Calcineurin (CaN) activation is critically involved in the regulation of spine morphology in response to oligomeric amyloid-ß (Aß) 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 Aß 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.

Anti-brolucizumab immune response as one prerequisite for rare retinal vasculitis/retinal vascular occlusion adverse events.

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.

A root cause analysis to identify the mechanistic drivers of immunogenicity against the anti-VEGF biotherapeutic brolucizumab.

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.

Calcineurin activation causes retinal ganglion cell degeneration.

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.

A Randomized, Double-Masked, Multicenter Trial of Topical Acrizanib (LHA510), a Tyrosine Kinase VEGF-Receptor Inhibitor, in Treatment-Experienced Subjects With Neovascular Age-Related Macular Degeneration.

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.

A proteogenomic signature of age-related macular degeneration in blood.

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.

Amyloid beta induces the morphological neurodegenerative triad of spine loss, dendritic simplification, and neuritic dystrophies through calcineurin activation.

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.

WITHDRAWN: Reprint of: Mechanisms of retinal ganglion cell injury and defense in glaucoma.

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.

Mechanisms of retinal ganglion cell injury and defense 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.

Using Healthcare Databases to Refine Understanding of Exploratory Associations Between Drugs and Progression of Open-Angle Glaucoma.

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.

DNA sequence variants in the LOXL1 gene are associated with pseudoexfoliation glaucoma in a U.S. clinic-based population with broad ethnic diversity.

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.

Levels of vascular endothelial growth factor-A165b (VEGF-A165b) are elevated in experimental 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.

Efficient estimation of retinal ganglion cell number: a stereological approach.

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.

Hsp27 phosphorylation in experimental glaucoma.

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.

Characterization of cytokine responses to retinal detachment in rats.

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.

The course of glaucoma during pregnancy: a retrospective case series.

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.

Downregulation of Thy1 in retinal ganglion cells in experimental glaucoma.

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.

Dry Eye Signs and Symptoms Persist During Systemic Neutralization of IL-1ß by Canakinumab or IL-17A by Secukinumab.

PURPOSE: To evaluate whether inhibition of the proinflammatory cytokines IL-1ß 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-1ß or IL-17A obtained by systemic administration of neutralizing drugs does not influence the severity of dry eye.