Microperimetry and Structural Risk Factors on OCT in Intermediate Age-Related Macular Degeneration.

PURPOSE: To determine the relationship between structural biomarkers on OCT that increase the risk of disease progression and microperimetric retinal sensitivity in patients with intermediate age-related macular degeneration (iAMD). DESIGN: Prospective cross-sectional, observational study. PARTICIPANTS: Forty-five eyes of 23 patients with iAMD. METHODS: Patients underwent OCT and microperimetry. OCT scans were evaluated for the risk factors intraretinal hyperreflective foci (HRF), hyporeflectivity within drusenoid lesions (HRDL), subretinal drusenoid deposits, double-layer sign (DLS), and drusen volume. Microperimetric retinal sensitivity was analyzed with a 33-point grid covering the macula. With a novel method of determining what part of the retina corresponded to each microperimetry point, a Voronoi diagram was constructed, dividing the macula in cells consisting of the region nearer to each point than any other. The Voronoi diagram was superimposed on the OCT, making it possible to determine the point-to-point location of the OCT risk factors. Univariable and multivariable linear mixed-effect models were used for analysis. MAIN OUTCOME MEASURES: Association between microperimetric retinal sensitivity and OCT risk factors at individual measuring points. RESULTS: One thousand four hundred seventy-nine points of retinal sensitivity and corresponding structural area on OCT were included in this study. Retinal sensitivity was significantly decreased with presence of the OCT risk factors HRF, HRDL, DLS, and drusen volume (all P < 0.001) when analyzed with the univariable linear mixed-effect model. The multivariable model showed a significant decrease of retinal sensitivity with presence of HRF (P < 0.001), DLS (P = 0.025), and greater drusen volume (P < 0.001). CONCLUSIONS: Presence of HRF, DLS, and greater drusen volume, all of which increase the risk of disease progression, is significantly and independently associated with decreased microperimetric retinal sensitivity in patients with iAMD. FINANCIAL DISCLOSURE(S): Proprietary or commercial disclosure may be found in the Footnotes and Disclosures at the end of this article.

Complement factor H Y402H polymorphism results in diminishing CD4+ T cells and increasing C-reactive protein in plasma.

Age-related macular degeneration (AMD) is a common cause of visual loss among the elderly. Genetic variants in the gene encoding complement factor H (CFH) have been identified as an AMD susceptibility gene, however, the mechanistic link is debated. Here, we investigated the link between the CFH Y402H genotype and low-grade inflammation. We recruited 153 healthy individuals, 84 participants with dry stages of AMD, and 148 participants with neovascular AMD. All participants were subjected to detailed retinal examination, and interview regarding comorbidities and lifestyle. Blood samples were analyzed for level of C-Reactive Protein (CRP), white blood cell differential count, and stained with fluorescent antibodies to differentiate CD4+ and CD8+ T cells. CFH Y402H genotyping was performed using an allele-specific polymerase chain reaction genotyping assay. Splenocytes from young and aged wild type and Cfh null mutant C57BL/6J mice were examined for CD4+ and CD8+ T cells. Healthy individuals with the CFH Y402H at-risk polymorphism HH had higher levels of CRP and lower proportions of CD4+ T cells compared to persons with the YH or YY polymorphism (P = 0.037, Chi-square). Healthy individuals with the HH polymorphism displayed lower proportions of CD4+ T cells with ageing (P < 0.01, one-way ANOVA), whereas both young and aged Cfh null mutant mice displayed lower proportions of CD4+ T cells (P < 0.001 and P < 0.05; unpaired t test). Participants with dry AMD and the HH polymorphism had similarly lower proportions of CD4+ T cells (P = 0.024, one-way ANOVA), but no difference in CRP-levels. In the neovascular stage of AMD, there was no difference in proportion of CD4+ cells or CRP levels according to genotype. The risk-associated CFH genotype is associated with an age-related decrease in proportion of CD4+ T cells and increased levels of CRP in healthy individuals. This indicates that decreased complement regulation results in extensive changes in innate and adaptive immune compartments that precede development of AMD.

Driving vision in patients with neovascular AMD in anti-VEGF treatment.

PURPOSE: To report real-world experiences on driving vision in patients with neovascular age-related macular degeneration (AMD) undergoing intravitreal anti-VEGF treatment. METHODS: Retrospective cohort study of treatment-naïve patients with neovascular AMD commenced in anti-VEGF treatment (n = 416) and followed for 4 years in a pro re nata treatment regimen. Monocular best-corrected visual acuity (BCVA) measured using ETDRS was performed on the treatment eye at all visits and on the fellow eye at baseline, every 6 months, and upon any patient-reported change in vision. Driving vision was defined as BCVA in the best-seeing eye of ≥70 ETDRS letters (equivalent to ≥0.5 Snellen) corresponding to the minimum BCVA required in many countries. RESULTS: Driving vision was present in 280 patients (67%) and was sustained in 86%, 74%, 65% and 59% of the patients at 1, 2, 3 and 4 years, respectively. Lower BCVA in the best-seeing eye predicted loss of driving vision. In patients without driving vision at baseline, driving vision was regained in 29%, 36%, 39% and 41% of the patients at 1, 2, 3 and 4 years, respectively; but only 35% sustained driving vision after the first year. Lower age and higher BCVA in best-seeing eye predicted regain of driving vision. CONCLUSION: Driving vision can be sustained in the majority of the patients if they have driving vision at baseline. This study provides important prognostic information for patients with neovascular AMD.

Chemokine Profile and the Alterations in CCR5-CCL5 Axis in Geographic Atrophy Secondary to Age-Related Macular Degeneration.

Purpose: Geographic atrophy (GA) secondary to age-related macular degeneration (AMD) is a progressive disease with no treatment option. Previous studies show chemokine-mediated recruitment of immune cells in the retina, and therefore we investigated systemic levels of chemokines and chemokine receptors in patients with GA. Methods: This observational prospective study was conducted at a single center. We included 122 participants with no immune disease: 41 participants with GA and no choroidal neovascularization, 51 patients with neovascular AMD, and 30 healthy control individuals. Flow cytometric analysis was used to detect expression level of C-C chemokine receptor (CCR)1, CCR2, CCR3, CCR5, and C-X-C motif chemokine receptor (CXCR)3 on peripheral blood mononuclear cells (CD14+ monocytes, CD4+ T cells, CD8+ T cells). Plasma levels of C-C motif ligand (CCL)11, C-X-C motif chemokine (CXCL)10, and CCL5 were measured by specific immunoassays. Enlargement rate of GA lesion was measured from autofluorescence images. Results: Participants with GA have a specific chemokine profile with a higher expression of CCR5 than healthy controls in peripheral blood mononuclear cells, and a higher plasma levels of CCL-5. Further, GA was associated with higher monocytic expression of CCR2 than in neovascular AMD. We found that a high expression level of CCR5 on CD8+ T cells was associated with slower enlargement rate of atrophic lesion. Conclusions: The study showed an association between systemic chemokine profile and GA formation. Further studies are needed to fully elucidate the possible role of systemic chemokine regulation in mediating pathogenesis of GA.

Association of CD11b+ Monocytes and Anti-Vascular Endothelial Growth Factor Injections in Treatment of Neovascular Age-Related Macular Degeneration and Polypoidal Choroidal Vasculopathy.

Importance: CD11b+ immune cells have been implicated in the formation of choroidal neovascularization in experimental studies on animals and disease-association studies on humans. However, the clinical importance of such observations remains unknown. Objective: To investigate whether the proportion of CD11b+ circulating monocytes is associated with the number of anti-vascular endothelial growth factor (anti-VEGF) injections in neovascular age-related macular degeneration (AMD) and polypoidal choroidal vasculopathy (PCV). Design, Setting, and Participants: These observational cohort studies collected data from January 1, 2010, through December 31, 2013, and from January 1, 2015, through December 31, 2018. Fresh venous blood samples were acquired for flow cytometric immune studies in patients with neovascular AMD or PCV receiving treatment with aflibercept or ranibizumab as needed for 36 months. Patients (n = 81) without immune diseases were consecutively recruited from a single center in Denmark. Exposures: Proportion of CD11b+ circulating monocytes. Main Outcomes and Measures: The estimation of the number of intravitreal anti-VEGF injections given at 12, 24, and 36 months by the proportion of CD11b+ circulating monocytes and the correlation between these values. The angiogenic role of CD11b+ circulating monocytes was further evaluated by investigating the expression of the known proangiogenic receptor CCR2. Results: Eighty-one patients were included in the analysis (54% women; mean [SD] age, 76 [7] years). The proportion of CD11b+ monocytes at baseline positively estimated the future number of anti-VEGF injections at 12 (ρ = 0.77; 95% CI, 0.35-0.93; P = .004), 24 (ρ = 0.82; 95% CI, 0.44-0.95; P = .002), and 36 (ρ = 0.78; 95% CI, 0.34-0.94; P = .005) months. This association was also found retrospectively in a larger sample of patients with neovascular AMD at 12 (ρ = 0.46; 95% CI, 0.16-0.68; P = .004), 24 (ρ = 0.49; 95% CI, 0.20-0.70; P = .002), and 36 (ρ = 0.65; 95% CI, 0.41-0.80; P < .001) months and patients with PCV at 12 (ρ = 0.27; 95% CI, -0.28 to 0.68; P = .30), 24 (ρ = 0.60; 95% CI, 0.12-0.85; P = .02), and 36 (ρ = 0.70; 95% CI, 0.27-0.90; P = .005) months, suggesting that this association is not specific to AMD but rather reflects VEGF activity in neovascularization. CD11b+ monocytes highly coexpressed CCR2, an important monocytic marker of proangiogenic activity. Conclusions and Relevance: Results of this study demonstrated that the proportion of circulating CD11b+ monocytes estimated and correlated with the number of anti-VEGF injections in patients with neovascular AMD and PCV. Additional longitudinal studies are needed to determine whether these findings have clinical relevance to influence treatment algorithms or provide novel targets for medical therapy.

Systemic Levels of Interleukin-6 Correlate With Progression Rate of Geographic Atrophy Secondary to Age-Related Macular Degeneration.

Purpose: Geographic atrophy (GA) is a clinical phenotype of late age-related macular degeneration (AMD) with no current treatment available. In this study, we investigated markers of chronic inflammation in plasma of patients with GA and how these relate to progression rate. Methods: We prospectively included 42 patients with GA, 41 patients with neovascular AMD, and 27 healthy controls. We quantified levels of interleukin (IL)-1ß, IL-6, IL-8, tumor necrosis factor (TNF) receptor 2, and C-reactive protein (CRP). We adapted an inflammation summary score to cluster conceptually related markers of chronic inflammation. Enlargement rate of the atrophic lesion was measured from fundus autofluorescence images performed at baseline and after 1 year. Results: Patients with GA showed an increase in proinflammatory markers of IL-6 (P = 0.009), TNF receptor 2 (P = 0.013), and CRP (P = 0.017) compared to healthy controls. We found that IL-8 levels were markedly higher in patients with GA when compared to patients with neovascular AMD (P = 0.013). The inflammation summary score was high in patients with neovascular AMD (P = 0.024), but even higher in patients with GA (<0.001), when compared to healthy controls. GA enlargement was measured in 36 patients, who completed follow-up. Plasma levels of IL-6 had a moderate but significant correlation with GA enlargement rate (R2 = 0.23, P = 0.0035). Conclusions: Markers of chronic inflammation strongly associates with presence of GA secondary to AMD. Plasma IL-6 possesses predictive ability of progression and constitutes the first known plasma biomarker of disease activity in GA. These findings shed light into a poorly understood clinical phenotype of AMD and highlights the important role of chronic inflammation in GA.

Patients with a fast progression profile in geographic atrophy have increased CD200 expression on circulating monocytes.

IMPORTANCE: Geographic atrophy (GA) is a progressing atrophy of the neuroretina with no treatment option. BACKGROUND: Age-related malfunction of retinal microglia amplifies response towards age-related tissue stress in age-related macular degeneration. Here, we investigated monocyte CD200 expression - the circulating middleman negotiating retinal microglial activity - in a poorly understood subtype of age-related macular degeneration. DESIGN: Prospective case-control study. PARTICIPANTS: Forty-six patients with GA and 26 healthy controls were included. METHODS: All participants were subjected to a structured interview and detailed retinal examination. Controls were recruited from patient's spouses accompanying them in the clinic to match the groups best possibly. Participants had no history of immune disorders or cancer, and did not receive any immune-modulating medication. Patients did not have any history or sign of choroidal neovascularization in either eye. Fresh drawn blood was stained with monoclonal antibodies and prepared for flow cytometry to evaluate CD200 expression in monocytes and their functional subsets. MAIN OUTCOME MEASURES: The percentage of CD200+ monocytes in patients and controls. RESULTS: We found that monocytes were more CD200 positive in patients with GA compared to healthy age-matched controls. Then, we explored the potential relationship between CD200 expression and important fundus autofluorescence patterns that predict disease progression. Patients with a high risk of progression (patients with high degree of hyperautofluorescence) had distinctly increased CD200 expression compared to other patients with GA. CONCLUSIONS AND RELEVANCE: Our data reveals that abnormal monocytic CD200 expression is present in GA, and in particular among those identified as fast progressors.

Extended HLA-G haplotypes in patients with age-related macular degeneration.

The study aims to determine if genetic polymorphisms in the human leukocyte antigen (HLA)-G gene are associated with age-related macular degeneration (AMD). HLA-G is important for immunological tolerance, and it is also known to have angiogenic effects. Polymorphisms in the 5'-upstream regulatory region (URR) and 3'-untranslated region (UTR) of HLA-G have been associated with a number of diseases, especially with respect to a 14 bp insertion/deletion (ins/del) polymorphism in the 3'UTR. Full gene sequencing was performed on a cohort of 146 AMD patients and 63 healthy controls aged 60 years or older and HLA-G haplotypes were determined. Analyses were performed on a publicly available gene expression dataset from the NCBI GEO database (accession number GSE29801) from which expression data for HLA-G, -C and -A were extracted. Analysis of the GEO dataset showed that both HLA-G and -C was expressed in the back of the eye and that expression was upregulated in the macular area of AMD. No differences were observed between patients and controls when analysing the distribution of haplotypes in the HLA-G promoter, coding region, 3'UTR or the 14 bp ins/del polymorphism of the 3'UTR. The increased expression of HLA-G in the macula of AMD patients indicates a role of HLA-G in the micro environment as part of the AMD pathogenesis. This is supported by the expression of HLA-C, which has previously been shown to play a role in AMD. The HLA-G haplotype distribution did not display any differences between AMD patients and controls. This article is protected by copyright. All rights reserved.

Serum carotenoids and macular pigment optical density in patients with intestinal resections and healthy subjects: an exploratory study.

Reduced absorption capacity in patients with intestinal resections (IR) could result in malabsorption of fat-soluble components like carotenoids, which are of clinical interest in relation to visual health. In this case cohort, we investigated the association between IR and serum lutein, zeaxanthin, ß-carotene and macular pigment optical density, when compared with healthy controls. Ten patients with IR and twelve healthy controls were included in the study. Baseline characteristics were comparable between groups, except for higher serum TAG (P < 0·05) and shorter bowel length (P < 0·0001) in the group with IR. Serum lutein, zeaxanthin, ß-carotene and macular pigment optical density were >15 % lower in the patient group compared with healthy controls (P < 0·05, adjusted for age) and, in the case of serum lutein and zeaxanthin, also for dietary intake of carotenoids. Results suggest that for a test of macular carotenoid supplementation, subjects with a potentially clinically significant carotenoid deficit could be recruited among patients with IR.

Systemic frequencies of T helper 1 and T helper 17 cells in patients with age-related macular degeneration: A case-control study.

Age-related macular degeneration (AMD) is a degenerative disease of the retina and a leading cause of irreversible vision loss. We investigated the systemic differences in the frequency of T helper (Th) 1 and Th17 cells in patients with non-exudative and exudative AMD and compared to age-matched controls. Flow cytometry was used to determine the systemic frequency of Th1 (CD4+CXCR3+IL12RB2+) and Th17 (CD4+CCR6+IL23R+) cells, and percentage of CD4+ T-cells expressing CXCR3, IL12RB2, CCR6, IL23R, and co-expressing CXCR3 and CCR6. The frequency of Th1 cells and CXCR3+ CD4+ T-cells was lower in patients with exudative AMD. A significant age-dependent decrement in Th1 was observed in controls, but not in non-exudative or exudative AMD. This may be related to the CXCR3+ CD4+ T-cells, which showed similar pattern in controls, but not in non-exudative or exudative AMD. No significant group differences were observed for the frequency of Th17 cells. Correlation networks found several differences between controls and AMD. These data suggests the involvement of the adaptive immune system in AMD and supports the notion of AMD as a systemic disease. Our observations warrant further investigation into the role of the adaptive immune system in the pathogenesis of AMD.

Systemic and Ocular Long Pentraxin 3 in Patients with Age-Related Macular Degeneration.

Age-related macular degeneration (AMD) has been associated with both systemic and ocular alterations of the immune system. In particular dysfunction of complement factor H (CFH), a soluble regulator of the alternative pathway of the complement system, has been implicated in AMD pathogenesis. One of the ligands for CFH is long pentraxin 3 (PTX3), which is produced locally in the retinal pigment epithelium (RPE). To test the hypothesis that PTX3 is relevant to retinal immunohomeostasis and may be associated with AMD pathogenesis, we measured plasma PTX3 protein concentration and analyzed the RPE/choroid PTX3 gene expression in patients with AMD. To measure the ability of RPE cells to secrete PTX3 in vitro, polarized ARPE-19 cells were treated with activated T cells or cytokines (interferon (IFN)-gamma and/or tumor necrosis factor (TNF)-alpha) from the basolateral side; then PTX3 protein concentration in supernatants and PTX3 gene expression in tissue lysates were quantified. Plasma levels of PTX3 were generally low and did not significantly differ between patients and controls (P=0.307). No statistically significant difference was observed between dry and exudative AMD nor was there any correlation with hsCRP or CFH genotype. The gene expression of PTX3 increased in RPE/choroid with age (P=0.0098 macular; P=0.003 extramacular), but did not differ between aged controls and AMD patients. In vitro, ARPE-19 cells increased expression of the PTX3 gene as well PTX3 apical secretions after stimulation with TNF-alpha or activated T cells (P<0.01). These findings indicate that PTX3 expressed in the eye cannot be detected systemically and systemic PTX3 may have little or no impact on disease progression, but our findings do not exclude that locally produced PTX3 produced in the posterior segment of the eye may be part of the AMD immunopathogenesis.

The association between neovascular age-related macular degeneration and regulatory T cells in peripheral blood.

PURPOSE: To investigate regulatory T cells (Tregs) and subsets of the Treg population in patients with neovascular age-related macular degeneration (AMD). PATIENTS AND METHODS: Twenty-one neovascular AMD cases and 12 age-matched controls without retinal pathology were selected. Patients were recruited from our outpatient retinal clinic. Control individuals were typically spouses. The diagnosis of neovascular AMD was confirmed using fluorescein and indocyaningreen angiography. Fresh venous blood was analyzed by flow cytometry using fluorochrome-conjugated antibodies to the Treg surface antigens CD4, CD25, CD127, CD45RA, and CD31. Main outcome measures were the percentage of CD25(high)CD127(low) Tregs, the percentage of CD45RA(+) naïve Tregs, and the percentage of CD31(+) recent thymic emigrant Tregs. RESULTS: Comparing patients with neovascular AMD to controls, no significant differences were found in the percentages of CD4(+) lymphocytes, CD25(high)CD127(low) Tregs, CD45RA(+) naïve Tregs, or CD31(+) recent thymic emigrant Tregs. CONCLUSION: Our data does not indicate an altered state of systemic Treg cells in neovascular AMD.

Early and exudative age-related macular degeneration is associated with increased plasma levels of soluble TNF receptor II.

PURPOSE: We have recently identified homeostatic alterations in the circulating T cells of patients with age-related macular degeneration (AMD). In cultures of retinal pigment epithelial cells, we have demonstrated that T-cell-derived cytokines induced the upregulation of complement, chemokines and other proteins implicated in AMD pathogenesis. The purpose of this study was to test whether increased plasma levels of cytokines were present in patients with AMD. METHODS: We conducted a case-control study. Age-related macular degeneration status was assessed using standardized multimodal imaging techniques. Plasma was isolated from freshly drawn peripheral venous blood samples and analysed for interleukin (IL)15, IL18, interferon (IFN)γ, soluble tumour necrosis factor (TNF) receptor II (sTNFRII) and complement factor H (CFH) Y402H genotype. RESULTS: We included 136 individuals with early or late forms of AMD and 74 controls. Significantly increased levels of sTNFRII were observed in patients with early or exudative AMD (p < 0.01). After adjusting for CFH Y402H genotype, age, sex and smoking history, the level of sTNFRII remained a significant predictor for prevalence of AMD with odds ratios at 3.0 in the middle and 3.6 in the highest tertiles. Levels of IL15, IL18 and IFNγ were low and not associated with AMD. CONCLUSIONS: Increased plasma level of sTNFRII is found to be associated with AMD. The data supports the observations of low-grade, systemic inflammatory alterations in patients with AMD. However, it remains to be determined whether increased levels of TNFα can be found, which directly reflects an increased activity of macrophages and T cells.

CX3CL1/CX3CR1 and CCL2/CCR2 chemokine/chemokine receptor complex in patients with AMD.

PURPOSE: The chemokine receptors CX3CR1 and CCR2 have been implicated in the development of age-related macular degeneration (AMD). The evidence is mainly derived from experimental cell studies and murine models of AMD. The purpose of this study was to investigate the association between expression of CX3CR1 and CCR2 on different leukocyte subsets and AMD. Furthermore we measured the plasma levels of ligands CX3CL1 and CCL2. METHODS: Patients attending our department were asked to participate in the study. The diagnosis of AMD was based on clinical examination and multimodal imaging techniques. Chemokine plasma level and chemokine receptor expression were measured by flow-cytometry. RESULTS: A total of 150 participants were included. We found a significantly lower expression of CX3CR1 on CD8+ T cells in the neovascular AMD group compared to the control group (p = 0.04). We found a significant positive correlation between CCR2 and CX3CR1 expression on CD8+ cells (r = 0.727, p = 0.0001). We found no difference in plasma levels of CX3CL1 and CCL2 among the groups. CONCLUSIONS: Our results show a down regulation of CX3CR1 on CD8+ cells; this correlated to a low expression of CCR2 on CD8+ cells. Further studies are needed to elucidate the possible role of this cell type in AMD development.

Dysregulation of CXCR3 expression on peripheral blood leukocytes in patients with neovascular age-related macular degeneration.

PURPOSE: The chemokine receptor CXCR3 has been strongly related to inhibition of angiogenesis. The purpose of this study was to investigate the association between expression of CXCR3 on peripheral blood leukocytes and age-related wet macular degeneration. Furthermore, we measured the plasma concentration of chemokines CXCL9 to -11. METHODS: The study group consisted of patients with age-related macular degeneration (AMD) attending our department. Patients referred for reasons other than AMD were enrolled as control subjects. The expression of CXCR3 on T cells and the plasma concentration of CXCL9 to -11 were measured using flow cytometry. RESULTS: We looked at all CD8(+) T cells expressing CXCR3 and found a significantly lower percentage of these cells in the neovascular AMD group compared to the age-matched control group (P = 0.05). When dividing the CD8(+) cells into functional groups according to their expression of CXCR3, we found a significantly lower percentage of CD8(+) CXCR3(high) cells in the group with neovascular AMD compared to the control group (P = 0.038). We found a lower percentage of CD4(+)CD69(+)CXCR3(+) T cells in the group of patients with neovascular AMD when compared to the age-matched control group (P = 0.052). CONCLUSIONS: Our results point toward a systemic dysregulation of CXCR3 in patients with neovascular AMD. Since there is evidence to suggest that CXCR3 is able to alter the response of VEGF, the primary driver of choroidal neovascularization (CNV) formation, low levels of CXCR3 could potentially drive some patients toward a more angiogenic profile leading to CNV formation and growth. CXCR3-enhancing molecules could therefore be a possible target for treatment of AMD.

Blood expression levels of chemokine receptor CCR3 and chemokine CCL11 in age-related macular degeneration: a case-control study.

BACKGROUND: Dysregulation of the CCR3/CCL11 pathway has been implicated in the pathogenesis of choroidal neovascularisation, a common feature of late age-related macular degeneration (AMD). The aim of this study was to investigate the expression of CCR3 and its ligand CCL11 in peripheral blood in patients with neovascular AMD. METHODS: Patients with neovascular AMD and healthy controls were included. Blood samples were obtained and prepared for flow cytometry to investigate the expression of CCR3. Levels of CCL11 were measured in plasma using Cytometric Bead Array. Differences between the groups were tested using Kruskal-Wallis test and Mann-Whitney U test. RESULTS: Patients (n = 83) with neovascular AMD and healthy control persons (n = 114) were included in the study. No significant difference in the expression of CCR3 was found on CD9+ granulocytes when comparing patients suffering from neovascular AMD with any of the control groups. We did not find any alteration in CCL11 levels in patients among the age matched groups. There was no correlation between expression of CCR3/CCL11 and clinical response to treatment with anti-vascular endothelial growth factor (VEGF). CONCLUSION: Our results do not suggest a systemic alteration of the CCR3/CCL11 receptor/ligand complex in patients with neovascular AMD.

In patients with neovascular age-related macular degeneration, physical activity may influence C-reactive protein levels.

PURPOSE: Association of neovascular age-related macular degeneration (AMD) with C-reactive protein (CRP) was previously reported, indicating a relation to systemic low-grade inflammation. However, visual impairment limits physical activity, and physical activity modulates CRP levels. Here, we investigated the impact of physical activity on CRP levels in patients with neovascular AMD and control individuals. SUBJECTS AND METHODS: We recruited participants from our outpatient AMD program, and control individuals from non-AMD patients, visitors, and department staff. After initial screening of 191 individuals, we included 98 patients with neovascular AMD and 77 controls. All were screened using digital fundus photography and optical coherence tomography, and interviewed about medical history and physical activity. Venous blood samples were obtained for high-sensitivity CRP. RESULTS: Physically active individuals had lower CRP than physically inactive individuals (P=0.003), and physical activity was associated with lower CRP in patients (P=0.038) and controls (P=0.031). Patients and controls did not differ in percentage physically active (P=0.807) or in overall CRP levels (P=0.394). The independent contribution of physical activity on CRP was confirmed in a multiple regression analysis (P=0.009), in which the presence of neovascular AMD did not contribute significantly (P=0.913). CONCLUSION: Our findings suggest that elevated CRP levels in patients with neovascular AMD are at least partly explained by physical inactivity. Future studies of systemic inflammation among the visually impaired should include disease-related implications, such as the impact of physical activity.

The association between plasma 25-hydroxyvitamin D and subgroups in age-related macular degeneration: a cross-sectional study.

OBJECTIVES: To evaluate potential differences in plasma 25-hydroxyvitamin in subtypes of age-related macular degeneration (AMD), and in patients in Clinical Age-Related Maculopathy Staging (CARMS) group 5 with or without subretinal fibrosis. METHODS: This single-center cross-sectional study included 178 participants during a period of 20 months. Ninety-five patients belonged to CARMS 5; twelve belonged to CARMS 4; twenty-two belonged to CARMS 2 or 3; and 49 individuals did not have AMD (CARMS 1). Following a structured interview, a detailed bilateral retinal examination was performed and participants were allocated to their respective subgroups in accordance with the Clinical Age-Related Maculopathy Staging system. Plasma 25-hydroxyvitamin D2 and D3 were analyzed using liquid chromatography-tandem mass spectrometry. Genomic DNA was extracted from leukocytes and genotyped for single nucleotide polymorphisms (SNPs) in the vitamin D metabolism. Differences in plasma 25-hydroxyvitamin D were determined in the subgroups as well as between patients in CARMS 5 with or without subretinal fibrosis. RESULTS: Plasma 25-hydroxyvitamin D was comparable in patients across CARMS groups 1 to 5 (p = 0.83). In CARMS 5, the presence of subretinal fibrosis was associated with significantly lower concentrations of 25-hydroxyvitamin D as compared to the absence of subretinal fibrosis (47.2 versus 75.6 nmol/L, p<0.001). Patients in CARMS 5 with subretinal fibrosis were more likely to have insufficient levels of 25-hydroxyvitamin D compared to patients without subretinal fibrosis (p = 0.006). No association was found between the SNPs rs10877012, rs2228570, rs4588, or rs7041 and AMD subgroups or plasma 25-hydroxyvitamin levels. CONCLUSIONS: This study suggests that the presence of subretinal fibrosis in patients belonging to CARMS 5 may be associated with a poor vitamin D status. Our observations warrant further investigation into the role of vitamin D in the development of subretinal fibrosis.

Age-related macular degeneration is associated with increased proportion of CD56(+) T cells in peripheral blood.

PURPOSE: To examine the association between age-related changes in the T-cell compartment and prevalence of age-related macular degeneration (AMD). DESIGN: Case-control study. PARTICIPANTS: A total of 117 AMD cases and 106 controls were included prospectively. METHODS: Fresh-drawn peripheral blood samples were processed for flow cytometric analysis of T-cell populations. Plasma samples were analyzed for anti-cytomegalovirus (CMV) immunoglobulin (Ig)G and complement factor H (CFH) Y402H genotype. The diagnosis of AMD was made according to the Clinical Age-Related Maculopathy Staging System. MAIN OUTCOME MEASURES: Association between frequency of aged T cells and prevalence of AMD. RESULTS: The prevalence of AMD was associated with distinct age-related changes in the T-cell compartment. Specifically, the patients with AMD had an increased frequency of CD28(-) T cells that expressed the CD56 surface marker (patients, 34.9% vs. aged controls, 25.8%; P = 0.002). Participants in the highest tertile of CD56(+) CD28(-) T cells had an odds ratio (OR) for the presence of AMD of 3.2 (95% confidence interval [CI], 1.2-8.8) after adjustment for CFH genotype, anti-CMV IgG positivity, age, sex, and smoking history. The adjusted OR of the presence of AMD for persons having at least 1 CFH H402 risk allele increased from 3.5 (95% CI, 1.5-8.1) to 13.3 (95% CI, 3.3-53.6) for persons with at least 1 CFH H402 risk allele and above the median level of CD56(+) CD28(-) T cells. CONCLUSIONS: We found increased levels of circulating aged CD56(+) CD28(-) T cells in patients with AMD. Although this supports the notion of AMD as a systemic disease, it also suggests that the adaptive immune system is implicated in its pathogenesis.

Increased expression of CD200 on circulating CD11b+ monocytes in patients with neovascular age-related macular degeneration.

OBJECTIVE: Dysregulation of retinal microglial activity has been implicated in the pathogenesis of neovascular age-related macular degeneration. Microglia activity can be regulated through the membrane protein CD200 and its corresponding receptor, the CD200 receptor (CD200R). Because both the ligand and the receptor are expressed on a broad spectrum of cell types, we set out to study the expression of CD200 and CD200R on CD11b+ monocytes, granulocytes, and subsets of T lymphocytes. DESIGN: Prospective, case-control study. PARTICIPANTS: The study population consisted of 62 patients with neovascular age-related macular degeneration (AMD) and 44 age-matched controls without AMD. METHODS: The participants were aged 60 years or older, had no history of immune dysfunction or cancer, and were not receiving immune-modulating therapy. All participants were subjected to a structured interview, and detailed retinal imaging was performed: fundus autofluorescence imaging, digital color fundoscopy, and spectral-domain optical coherence tomography. Fluorescein and indocyanine green angiography were performed in patients with suspected neovascular AMD. Visual acuity was measured in both eyes. Fresh venous blood was obtained and stained with monoclonal antibodies and analyzed using flow cytometry within 6 hours of phlebotomy. MAIN OUTCOME MEASURES: The percentage of CD11b+ monocytes, granulocytes, and CD4+/CD8+ T lymphocytes positive for CD200 or CD200R in patients and controls, respectively. RESULTS: Patients with neovascular AMD had a higher percentage of CD11b+CD200+ monocytes and CD200+ monocytes compared with controls. Multiple regression analysis revealed that the intergroup differences observed were independent of age. Moreover, an age-related increment in CD200 expression on monocytes was observed in controls with healthy eyes, but not in patients with neovascular AMD. We did not find any differences in CD200 and CD200R expression between patients with subretinal fibrosis and patients without subretinal fibrosis. CONCLUSIONS: The surface expression of CD200 on circulating CD11b+ monocytes was found to be increased in patients with neovascular AMD compared with controls with healthy eyes. This novel finding supports the notion that altered regulation of the inflammatory response plays an integral role in the pathogenesis of AMD. FINANCIAL DISCLOSURE(S): The author(s) have no proprietary or commercial interest in any materials discussed in this article.