Immunological Aspects of Age-Related Macular Degeneration.

Increasing evidence over the past two decades points to a pivotal role for immune mechanisms in age-related macular degeneration (AMD) pathobiology. In this chapter, we will explore immunological aspects of AMD, with a specific focus on how immune mechanisms modulate clinical phenotypes of disease and severity and how components of the immune system may serve as triggers for disease progression in both dry and neovascular AMD. We will briefly review the biology of the immune system, defining the role of immune mechanisms in chronic degenerative disease and differentiating from immune responses to acute injury or infection. We will explore current understanding of the roles of innate immunity (especially macrophages), antigen-specific immunity (T cells, B cells, and autoimmunity), immune amplifications systems, especially complement activity and the NLRP3 inflammasome, in the pathogenesis of both dry and neovascular AMD, reviewing data from pathology, experimental animal models, and clinical studies of AMD patients. We will also assess how interactions between the immune system and infectious pathogens could potentially modulate AMD pathobiology via alterations in in immune effector mechanisms. We will conclude by reviewing the paradigm of "response to injury," which provides a means to integrate various immunologic mechanisms along with nonimmune mechanisms of tissue injury and repair as a model to understand the pathobiology of AMD.

Aldosterone as a mediator of severity in retinal vascular disease: Evidence and potential mechanisms.

Diabetic retinopathy (DR) and retinal vein occlusion (RVO) are the two most common retinal vascular diseases and are major causes of vision loss and blindness worldwide. Recent and ongoing development of medical therapies including anti-vascular endothelial growth factor and corticosteroid drugs for treatment of these diseases have greatly improved the care of afflicted patients. However, severe manifestations of retinal vascular disease result in persistent macular edema, progressive retinal ischemia and incomplete visual recovery. Additionally, choroidal vascular diseases including neovascular age-related macular degeneration (NVAMD) and central serous chorioretinopathy (CSCR) cause vision loss for which current treatments are incompletely effective in some cases and highly burdensome in others. In recent years, aldosterone has gained attention as a contributor to the various deleterious effects of retinal and choroidal vascular diseases via a variety of mechanisms in several retinal cell types. The following is a review of the role of aldosterone in retinal and choroidal vascular diseases as well as our current understanding of the mechanisms by which aldosterone mediates these effects.

Enhanced visualization of peripheral retinal vasculature with wavefront sensorless adaptive optics optical coherence tomography angiography in diabetic patients.

Optical coherence tomography angiography (OCTA) is a promising technique for non-invasive visualization of vessel networks in the human eye. We debut a system capable of acquiring wide field-of-view (>70°) OCT angiograms without mosaicking. Additionally, we report on enhancing the visualization of peripheral microvasculature using wavefront sensorless adaptive optics (WSAO). We employed a fast WSAO algorithm that enabled wavefront correction in <2 s by iterating the mirror shape at the speed of OCT B-scans rather than volumes. Also, we contrasted ∼7° field-of-view OCTA angiograms acquired in the periphery with and without WSAO correction. On average, WSAO improved the sharpness of microvasculature by 65% in healthy eyes and 38% in diseased eyes. Preliminary observations demonstrated that the location of 7° images could be identified directly from the wide field-of-view angiogram. A pilot study on a normal subject and patients with diabetic retinopathy showed the impact of utilizing WSAO for OCTA when visualizing peripheral vasculature pathologies.

DYNAMIC CHANGES OF THE ANTERIOR CHAMBER ANGLE PRODUCED BY INTRAVITREAL ANTI-VASCULAR GROWTH FACTOR INJECTIONS.

PURPOSE: To use anterior segment optical coherence tomography (AS-OCT) to evaluate the anterior chamber after intravitreal anti-vascular endothelial growth factor (anti-VEGF) injections. METHODS: Preinjection and immediate postinjection AS-OCT images were taken and measurements were compared including angle opening distance (AOD) and trabeculo-iris space area (TISA) at 500 µm and 750 µm from the scleral spur (AOD500, AOD750, TISA500 and TISA750, respectively), and the scleral spur angle. RESULTS: Twenty-one eyes from 21 patients were studied. There was significant narrowing of the temporal AOD500, AOD750, and temporal angle after injection (P = 0.03, 0.01, and 0.02, respectively). The percentage of narrowing of the nasal TISA500 and TISA750 was significantly greater in phakic versus pseudophakic eyes (P = 0.03 and 0.02, respectively). A higher postinjection IOP was correlated with increased narrowing of the nasal AOD500, TISA500, TISA750, and nasal angle (R = 0.22, 0.28, 0.34 and 0.20; P = 0.03, 0.01, 0.005 and 0.04, respectively) and a smaller preinjection anterior chamber depth in phakic eyes (R = 0.53, P = 0.01). CONCLUSION: After an anti-vascular endothelial growth factor injection, there is significant narrowing of the temporal anterior chamber angle in all eyes and increased narrowing of the nasal angle in phakic compared with pseudophakic eyes. Physicians performing intravitreal injections should be aware of these associations as they may contribute to our understanding of prolonged elevation of IOP after injections.

VISUAL FUNCTION MEASURES IN EARLY AND INTERMEDIATE AGE-RELATED MACULAR DEGENERATION.

PURPOSE: The objectives of this study were to evaluate 1) the feasibility of performing computerized tests of low luminance visual acuity (LLVA), cone-specific contrast (Cone Contrast Test [CCT]), contrast sensitivity, and microperimetry and 2) the test-retest repeatability of these outcomes in dry age-related macular degeneration (AMD). METHODS: This prospective study enrolled 30 subjects at a single site (8 controls, 8 early AMD, and 12 intermediate AMD). Subjects underwent LLVA, contrast sensitivity, CCT, and microperimetry with eye tracking. Low luminance deficit was defined as best-corrected visual acuity minus LLVA in EDTRS letters. Follow-up testing was administered at approximately 1 month. RESULTS: There was high test-retest repeatability at one month for all visual function metrics (intraclass correlations >0.7) except log contrast sensitivity (intraclass correlations 0.6). Compared with controls, patients with intermediate AMD showed significant deficits on best-corrected visual acuity, LLVA, low luminance deficit, percent-reduced threshold on microperimetry, and red CCT (P < 0.05), but not on contrast sensitivity, green and blue CCT. CONCLUSION: This pilot study supports the feasibility and reliability of using LLVA, microperimetry, and CCT in early dry AMD. Our data suggest these measures can be used as alternative future clinical trial endpoints. A larger, prospective natural history study of alternative visual function measures in dry AMD is warranted.

Abundance of infiltrating CD163+ cells in the retina of postmortem eyes with dry and neovascular age-related macular degeneration.

PURPOSE: Prior research in animal models has suggested that retinal macrophages play an important role in age-related macular degeneration (AMD), but studies have insufficiently characterized the distribution of retinal macrophages in various stages of human AMD. METHODS: In this case series, we analyzed H&E, periodic acid-Schiff, and CD163 and CD68 immunostained slides from 56 formaldehyde-fixed, paraffin-embedded autopsy eyes of patients over age 75: 11 age-matched, normal eyes, and 45 AMD eyes. RESULTS: Qualitative analysis of the macula and retinal periphery revealed that all eyes contained a significant number of CD163+ cells but a negligible number of CD68+ cells. In normal eyes and eyes with thin or infrequent basal laminar deposits, CD163+ cells were restricted to the inner retina. In contrast, in AMD eyes with thick basal deposits, choroidal neovascular membranes, and geographic atrophy, qualitatively there was a marked increase in the number and size of the CD163+ cells in the outer retina, sub-retinal, and sub-retinal pigment epithelium space in the macula. CONCLUSIONS: The changes in number and localization of retinal CD163+ cells in eyes with intermediate-severe AMD support a key role for macrophages in the pathogenesis and progression of the disease. A larger, quantitative study evaluating the distribution of macrophage subpopulations in postmortem AMD eyes is warranted.

Macrophage activation associated with chronic murine cytomegalovirus infection results in more severe experimental choroidal neovascularization.

The neovascular (wet) form of age-related macular degeneration (AMD) leads to vision loss due to choroidal neovascularization (CNV). Since macrophages are important in CNV development, and cytomegalovirus (CMV)-specific IgG serum titers in patients with wet AMD are elevated, we hypothesized that chronic CMV infection contributes to wet AMD, possibly by pro-angiogenic macrophage activation. This hypothesis was tested using an established mouse model of experimental CNV. At 6 days, 6 weeks, or 12 weeks after infection with murine CMV (MCMV), laser-induced CNV was performed, and CNV severity was determined 4 weeks later by analysis of choroidal flatmounts. Although all MCMV-infected mice exhibited more severe CNV when compared with control mice, the most severe CNV developed in mice with chronic infection, a time when MCMV-specific gene sequences could not be detected within choroidal tissues. Splenic macrophages collected from mice with chronic MCMV infection, however, expressed significantly greater levels of TNF-α, COX-2, MMP-9, and, most significantly, VEGF transcripts by quantitative RT-PCR assay when compared to splenic macrophages from control mice. Direct MCMV infection of monolayers of IC-21 mouse macrophages confirmed significant stimulation of VEGF mRNA and VEGF protein as determined by quantitative RT-PCR assay, ELISA, and immunostaining. Stimulation of VEGF production in vivo and in vitro was sensitive to the antiviral ganciclovir. These studies suggest that chronic CMV infection may serve as a heretofore unrecognized risk factor in the pathogenesis of wet AMD. One mechanism by which chronic CMV infection might promote increased CNV severity is via stimulation of macrophages to make pro-angiogenic factors (VEGF), an outcome that requires active virus replication.

RaScaL: A Pilot Study to Assess the Efficacy, Durability, and Safety of a Single Intervention with Ranibizumab plus Peripheral Laser for Diabetic Macular Edema Associated with Peripheral Nonperfusion on Ultrawide-Field Fluorescein Angiography.

Background and Objectives: To determine the efficacy, durability, and safety of a single treatment with intravitreal ranibizumab plus peripheral scatter laser (RaScaL) in patients with diabetic macular edema associated with peripheral retinal nonperfusion on ultrawide-field fluorescein angiography (UWFA). Study Design: A 6-month, randomized, controlled, prospective phase I/II study of 30 treatment-naïve eyes of 22 patients (8 bilateral patients) with visual impairment secondary to diabetic macular edema associated with peripheral nonperfusion on UWFA. Patients were randomized to receive ranibizumab plus UWFA-guided peripheral scatter laser (n = 15) or triamcinolone acetonide plus macular laser (n = 15). Results: At 6 months, the RaScaL group patients had fewer recurrences warranting retreatment (33% vs. 80%, p < 0.003). Mean change in final visual acuity and central foveal thickness were not statistically significant between groups. Conclusion: This pilot study suggests the efficacy, safety and durability of the RaScaL treatment regimen in patients with diabetic macular edema associated with peripheral nonperfusion on UWFA. © 2014 S. Karger AG, Basel.

Ten-year incidence of age-related macular degeneration according to diabetic retinopathy classification among medicare beneficiaries.

PURPOSE: To compare the longitudinal incidence over 10 years of dry and wet age-related macular degeneration (AMD) in a U.S. sample of Medicare beneficiaries with no diabetes mellitus, diabetes mellitus without retinopathy, nonproliferative diabetic retinopathy (NPDR), and proliferative diabetic retinopathy (PDR). METHODS: Using Medicare claims data, the 10-year incidence of dry and wet AMD from 1995 to 2005 in beneficiaries older than 69 years with newly diagnosed diabetes mellitus (n = 6,621), NPDR (n = 1,307), and PDR (n = 327) compared with each other and matched controls without diabetes for each group. RESULTS: After controlling for covariates, newly diagnosed NPDR was associated with significantly increased risk of incident diagnosis of dry AMD (hazard ratio, 1.24; 95% confidence interval: 1.08-1.43) and wet AMD (hazard ratio 1.68; 95% confidence interval: 1.23-2.31). Newly diagnosed PDR was associated with significantly increased risk of wet AMD only (hazard ratio 2.15; 95% confidence interval: 1.07-4.33). Diabetes without retinopathy did not affect risk of dry or wet AMD. There was no difference in risk of wet AMD in PDR compared with NPDR. CONCLUSION: Elderly individuals with NPDR or PDR may be at higher risk of AMD compared to those without diabetes mellitus or diabetic retinopathy.

Costs of newly diagnosed neovascular age-related macular degeneration among medicare beneficiaries, 2004-2008.

PURPOSE: To examine associations between newly diagnosed neovascular age-related macular degeneration and direct medical costs. METHODS: This retrospective observational study matched 23,133 Medicare beneficiaries diagnosed with neovascular age-related macular degeneration between 2004 and 2008 with a control group of 92,532 beneficiaries on the basis of age, sex, and race. The index date for each case-control set corresponded to the first diagnosis for the case. Main outcome measures were total costs per patient and age-related macular degeneration-related costs per case 1 year before and after the index date. RESULTS: Mean cost per case in the year after diagnosis was $12,422, $4,884 higher than the year before diagnosis. Postindex costs were 41% higher for cases than controls after adjustment for preindex costs and comorbid conditions. Age-related macular degeneration-related costs represented 27% of total costs among cases in the postindex period and were 50% higher for patients diagnosed in 2008 than in 2004. This increase was attributable primarily to the introduction of intravitreous injections of vascular endothelial growth factor antagonists. Intravitreous injections averaged $203 for patients diagnosed in 2004 and $2,749 for patients diagnosed in 2008. CONCLUSION: Newly diagnosed neovascular age-related macular degeneration was associated with a substantial increase in total medical costs. Costs increased over time, reflecting growing use of anti-vascular endothelial growth factor therapies.

Angiotensin II-induced MMP-2 activity and MMP-14 and basigin protein expression are mediated via the angiotensin II receptor type 1-mitogen-activated protein kinase 1 pathway in retinal pigment epithelium: implications for age-related macular degeneration.

Accumulation of various lipid-rich extracellular matrix (ECM) deposits under the retinal pigment epithelium (RPE) has been observed in eyes with age-related macular degeneration (AMD). RPE-derived matrix metalloproteinase (MMP)-2, MMP-14, and basigin (BSG) are major enzymes involved in the maintenance of ECM turnover. Hypertension (HTN) is a systemic risk factor for AMD. It has previously been reported that angiotensin II (Ang II), one of the most important hormones associated with HTN, increases MMP-2 activity and its key regulator, MMP-14, in RPE, inducing breakdown of the RPE basement membrane, which may lead to progression of sub-RPE deposits. Ang II exerts most of its actions by activating the mitogen-activated protein kinase (MAPK) signaling pathway. Herein is explored the MAPK signaling pathway as a potential key intracellular modulator of Ang II-induced increase in MMP-2 activity and MMP-14 and BSG protein expression. It was observed that Ang II stimulates phosphorylation of extracellular signal-regulated kinase (ERK) and p38 MAPK in RPE cells and ERK/p38 and Jun N-terminal kinase (JNK) in mice. These effects were mediated by Ang II type 1 receptors. Blockade of ERK or p38 MAPK abrogated the increase in MMP-2 activity and MMP-14 and BSG proteins in ARPE-19 cells. A better understanding of the molecular events by which Ang II induces ECM dysregulation is of critical importance to further define its contribution to the progression of sub-RPE deposits in AMD patients with HTN.

Comparison of color fundus photographs and fundus autofluorescence images in measuring geographic atrophy area.

PURPOSE: To assess the agreement between color fundus photographs (CFP) and fundus autofluorescence (FAF) images when measuring geographic atrophy (GA) area and reproducibility of measurements between graders. Frequency and disagreement types were also determined. METHODS: Eyes with GA secondary to age-related macular degeneration had CFP and FAF imaging on the same day. Seventy-two eyes from 72 patients were included in the analysis. Three graders calculated GA area using digital imaging software. Main outcome measures included agreement between graders for GA area on both FAF and CFP and agreement between both imaging modalities. RESULTS: The intraclass correlation for the 3 graders for FAF images was 0.99 (95% confidence interval, 0.98-0.99). For CFP, it was 0.96 (95% confidence interval, 0.94-0.97). The intraclass correlation between imaging modalities for Graders 1, 2, and 3 were 0.93, 0.85, and 0.87, respectively. Sensitivities to detect involvement of fovea (CFP, 86-97%; FAF, 72-93%) and specificities to detect sparing of fovea (CFP, 74-76%; FAF, 59-88%) overlapped between imaging modalities. CONCLUSION: Both CFP and FAF imaging are reliable for measuring GA area. Interobserver agreement was slightly higher for FAF images. Although the high agreement between modalities suggests that either would be appropriate for measuring GA area, using both may be the best approach for following GA progression.

Phase 1 Clinical Trial of Elamipretide in Dry Age-Related Macular Degeneration and Noncentral Geographic Atrophy: ReCLAIM NCGA Study.

Purpose: Assess the safety, tolerability, and feasibility of subcutaneous administration of the mitochondrial-targeted drug elamipretide in patients with dry age-related macular degeneration (AMD) and noncentral geographic atrophy (NCGA) and to perform exploratory analyses of change in visual function. Design: Phase 1, single-center, open-label, 24-week clinical trial with preplanned NCGA cohort. Participants: Adults ≥ 55 years of age with dry AMD and NCGA. Methods: Participants received subcutaneous elamipretide 40-mg daily; safety and tolerability assessed throughout. Ocular assessments included normal-luminance best-corrected visual acuity (BCVA), low-luminance BCVA (LLBCVA), normal-luminance binocular reading acuity (NLBRA), low-luminance binocular reading acuity (LLBRA), spectral-domain OCT, fundus autofluorescence (FAF), and patient self-reported function by low-luminance questionnaire (LLQ). Main Outcome Measures: Primary end point was safety and tolerability. Prespecified exploratory end-points included changes in BCVA, LLBCVA, NLBRA, LLBRA, geographic atrophy (GA) area, and LLQ. Results: Subcutaneous elamipretide was highly feasible. All participants (n = 19) experienced 1 or more nonocular adverse events (AEs), but all AEs were either mild (73.7%) or moderate (26.3%); no serious AEs were noted. Two participants exited the study because of AEs (conversion to neovascular AMD, n = 1; intolerable injection site reaction, n = 1), 1 participant discontinued because of self-perceived lack of efficacy, and 1 participant chose not to continue with study visits. Among participants completing the study (n = 15), mean ± standard deviation (SD) change in BCVA from baseline to week 24 was +4.6 (5.1) letters (P = 0.0032), while mean change (SD) in LLBCVA was +5.4 ± 7.9 letters (P = 0.0245). Although minimal change in NLBRA occurred, mean ± SD change in LLBCVA was -0.52 ± 0.75 logarithm of the minimum angle of resolution units (P = 0.005). Mean ± SD change in GA area (square root transformation) from baseline to week 24 was 0.14 ± 0.08 mm by FAF and 0.13 ± 0.14 mm by OCT. Improvement was observed in LLQ for dim light reading and general dim light vision. Conclusions: Elamipretide seems to be well tolerated without serious AEs in patients with dry AMD and NCGA. Exploratory analyses demonstrated possible positive effect on visual function, particularly under low luminance. A Phase 2b trial is underway to evaluate elamipretide further in dry AMD and NCGA.

Phase 1 Clinical Trial of Elamipretide in Intermediate Age-Related Macular Degeneration and High-Risk Drusen: ReCLAIM High-Risk Drusen Study.

Purpose: To assess safety, tolerability, and feasibility of subcutaneous administration of the mitochondrial-targeted drug elamipretide in patients with intermediate age-related macular degeneration (AMD) and high-risk drusen (HRD) and to perform exploratory analyses of change in visual function. Design: Phase 1, single-center, open-label, 24-week clinical trial with preplanned HRD cohort. Participants: Adult patients ≥55 years of age with intermediate AMD and HRD. Methods: Participants received subcutaneous elamipretide 40 mg daily, with safety and tolerability assessed throughout the study. Ocular assessments included normal-luminance best-corrected visual acuity (BCVA), low-luminance best-corrected visual acuity (LLVA), normal-luminance binocular reading acuity (NLRA), low-luminance binocular reading acuity (LLRA), spectral-domain OCT, fundus autofluorescence (FAF), mesopic microperimetry, dark adaptation, and low-luminance questionnaire (LLQ). Main Outcome Measures: The primary end point was safety and tolerability. Prespecified exploratory end points included changes from baseline in BCVA, LLVA, NLRA, LLRA, retinal pigment epithelium (RPE)-drusen complex (DC) volume by OCT, FAF, mesopic microperimetry, dark adaptation, and LLQ results. Results: Subcutaneous administration of elamipretide was highly feasible. All participants with HRD (n = 21) experienced 1 or more adverse events (AEs), but all were mild (57%) or moderate (43%), with the most common events related to injection site reactions. No serious systemic AEs occurred. One participant discontinued because of injection site reaction, 1 participant withdrew because they did not wish to continue study visits, and 1 participant withdrew after experiencing transient visual impairment. Among the 18 participants who completed the study, mean change in BCVA from baseline to 24 weeks was +3.6 letters (P = 0.014) and LLVA was +5.6 letters (P = 0.004). Compared with baseline, mean NLRA improved by -0.11 logarithm of the minimum angle of resolution (logMAR) units (P = 0.001), and LLRA by -0.28 logMAR units (P < 0.0001). Significant improvements were found in 6 of 7 subscales of the LLQ (P <0.0015). No significant changes were observed for RPE-DC volume, FAF, mesopic microperimetry, or dark adaptation. Conclusions: Elamipretide appeared to be generally safe and well tolerated in treating intermediate AMD and HRD. Exploratory analyses demonstrate a positive effect on visual function, particularly under low-luminance conditions. Further study of elamipretide for treatment of intermediate AMD with HRD is warranted.

Incidence of Referable Retinal Disease in Diabetic Patients at a Primary Care Practice.

Purpose: This work tests the feasibility of remote ophthalmic imaging to identify referable retinal abnormalities and assesses the effectiveness of color fundus photography (CFP) vs optical coherence tomography (OCT) for this purpose. Methods: This prospective, nonrandomized study included 633 patients with diabetes at Duke Primary Care. Undilated patients underwent screening with CFP and OCT camera (MaestroCare, Topcon). Images were graded independently for interpretability and the presence of predetermined retinal disease. Retinal disease was classified as diabetic retinopathy (DR) referable to a retina specialist or incidental findings referable to either a retina specialist or a general ophthalmologist, depending on severity. Results: Mean (SD) age of screened patients was 66 (13) years, and 49% were women. The average glycated hemoglobin A1c level was 7.6 % (SD, 1.7%), and 30% of the patients were on insulin. The average duration of diabetes was 5.9 (SD, 7.3) years. Remote images from OCT were significantly more interpretable than CFP (98% vs 83%, respectively; P < .001). Referral rates were 9% for DR and 28% for incidental findings. Among patients with DR, OCT and CFP were helpful in 58% and 87% of cases, respectively (P < .001). Conclusions: Remote diagnosis of ophthalmic imaging at the point of service may allow for early identification of retinal disease and timely referral and treatment. Our approach showed that OCT had significantly better interpretability, while CFP was more helpful in identifying DR. These findings may be important when choosing the screening device in a specific context.

Cigarette smoke-related hydroquinone induces filamentous actin reorganization and heat shock protein 27 phosphorylation through p38 and extracellular signal-regulated kinase 1/2 in retinal pigment epithelium: implications for age-related macular degeneration.

Retinal pigment epithelium (RPE)-derived membranous debris named blebs, may accumulate and contribute to sub-RPE deposit formation, which is the earliest sign of age-related macular degeneration (AMD). Oxidative injury to the RPE might play a significant role in AMD. However, the underlying mechanisms are unknown. We previously reported that hydroquinone (HQ), a major pro-oxidant in cigarette smoke, foodstuff, and atmospheric pollutants, induces actin rearrangement and membrane blebbing in RPE cells as well as sub-RPE deposits in mice. Here, we show for the first time that phosphorylated Heat shock protein 27 (Hsp27), a key regulator of actin filaments dynamics, is up-regulated in RPE from patients with AMD. Also, HQ-induced nonlethal oxidative injury led to Hsp27mRNA up-regulation, dimer formation, and Hsp27 phosphorylation in ARPE-19 cells. Furthermore, we found that a cross talk between p38 and extracellular signal-regulated kinase (ERK) mediates HQ-induced Hsp27 phosphorylation and actin aggregate formation, revealing ERK as a novel upstream mediator of Hsp27 phosphorylation. Finally, we demonstrated that Hsp25, p38, and ERK phosphorylation are increased in aging C57BL/6 mice chronically exposed to HQ, whereas Hsp25 expression is decreased. Our data suggest that phosphorylated Hsp27 might be a key mediator in AMD and HQ-induced oxidative injury to the RPE, which may provide helpful insights into the early cellular events associated with actin reorganization and bleb formation involved in sub-RPE deposits formation relevant to the pathogenesis of AMD.

Proteomics characterization of cell membrane blebs in human retinal pigment epithelium cells.

Age-related macular degeneration (AMD) is the leading cause of legal blindness among the elderly population in the industrialized world, affecting about 14 million people in the United States alone. Smoking is a major environmental risk factor for AMD, and hydroquinone is a major component in cigarette smoke. Hydroquinone induces the formation of cell membrane blebs in human retinal pigment epithelium (RPE). Blebs may accumulate and eventually contribute first to sub-RPE deposits and then drusen formation, which is a prominent histopathologic feature in eyes with AMD. As an attempt to better understand the mechanisms involved in early AMD, we sought to investigate the proteomic profile of RPE blebs. Isolated blebs were subjected to SDS-PAGE fractionation, and in-gel trypsin-digested peptides were analyzed by LC-MS/MS that lead to the identification of a total of 314 proteins. Identified proteins were predominantly involved in oxidative phosphorylation, cell junction, focal adhesion, cytoskeleton regulation, and immunogenic processes. Importantly basigin and matrix metalloproteinase-14, key proteins involved in extracellular matrix remodeling, were identified in RPE blebs and shown to be more prevalent in AMD patients. Altogether our findings suggest, for the first time, the potential involvement of RPE blebs in eye disease and shed light on the implication of cell-derived microvesicles in human pathology.

Use of fundus autofluorescence images to predict geographic atrophy progression.

PURPOSE: Fundus autofluorescence imaging has been shown to be helpful in predicting progression of geographic atrophy (GA) secondary to age-related macular degeneration. We assess the ability of fundus autofluorescence imaging to predict rate of GA progression using a simple categorical scheme. METHODS: Subjects with GA secondary to age-related macular degeneration with fundus autofluorescence imaging acquired at least 12 months apart were included. Rim area focal hyperautofluorescence was defined as percentage of the 500-µm-wide margin bordering the GA that contained increased autofluorescence. Rim area focal hyperautofluorescence on baseline fundus autofluorescence images was assessed and categorized depending on the extent of rim area focal hyperautofluorescence (Category 1: ≤33%; Category 2: between 33 and 67%; Category 3: ≥67%). Total GA areas at baseline and follow-up were measured to calculate change in GA progression. RESULTS: Forty-five eyes of 45 subjects were included; average duration of follow-up was 18.5 months. Median growth rates differed among categories of baseline rim area focal hyperautofluorescence (P = 0.01 among Categories 1, 2, and 3; P = 0.008 for Category 1 compared with Category 3, Jonckheere-Terpstra test). CONCLUSION: A simple categorical scheme that stratifies the amount of increased autofluorescence in the 500-µm margin bordering GA may be used to differentiate faster and slower progressors.

Incomplete response to Anti-VEGF therapy in neovascular AMD: Exploring disease mechanisms and therapeutic opportunities.

Intravitreal anti-vascular endothelial growth factor (VEGF) drugs have revolutionized the treatment of neovascular age-related macular degeneration (NVAMD). However, many patients suffer from incomplete response to anti-VEGF therapy (IRT), which is defined as (1) persistent (plasma) fluid exudation; (2) unresolved or new hemorrhage; (3) progressive lesion fibrosis; and/or (4) suboptimal vision recovery. The first three of these collectively comprise the problem of persistent disease activity (PDA) in spite of anti-VEGF therapy. Meanwhile, the problem of suboptimal vision recovery (SVR) is defined as a failure to achieve excellent functional visual acuity of 20/40 or better in spite of sufficient anti-VEGF treatment. Thus, incomplete response to anti-VEGF therapy, and specifically PDA and SVR, represent significant clinical unmet needs. In this review, we will explore PDA and SVR in NVAMD, characterizing the clinical manifestations and exploring the pathobiology of each. We will demonstrate that PDA occurs most frequently in NVAMD patients who develop high-flow CNV lesions with arteriolarization, in contrast to patients with capillary CNV who are highly responsive to anti-VEGF therapy. We will review investigations of experimental CNV and demonstrate that both types of CNV can be modeled in mice. We will present and consider a provocative hypothesis: formation of arteriolar CNV occurs via a distinct pathobiology, termed neovascular remodeling (NVR), wherein blood-derived macrophages infiltrate the incipient CNV lesion, recruit bone marrow-derived mesenchymal precursor cells (MPCs) from the circulation, and activate MPCs to become vascular smooth muscle cells (VSMCs) and myofibroblasts, driving the development of high-flow CNV with arteriolarization and perivascular fibrosis. In considering SVR, we will discuss the concept that limited or poor vision in spite of anti-VEGF may not be caused simply by photoreceptor degeneration but instead may be associated with photoreceptor synaptic dysfunction in the neurosensory retina overlying CNV, triggered by infiltrating blood-derived macrophages and mediated by Müller cell activation Finally, for each of PDA and SVR, we will discuss current approaches to disease management and treatment and consider novel avenues for potential future therapies.