CCL2/CCR2 and CX3CL1/CX3CR1 chemokine axes and their possible involvement in age-related macular degeneration

The causes of age-related macular degeneration (AMD) are not well understood. Due to demographic shifts in the industrialized world a growing number of people will develop AMD in the coming decades. To develop treatments it is essential to characterize the disease's pathogenic process. Over the past few years, numerous studies have focused on the role of chemotactic cytokines, also known as chemokines. Certain chemokines, such as CCL2 and CX3CL1, appear to be crucial in subretinal microglia and macrophage accumulation observed in AMD, and participate in the development of retinal degeneration as well as in choroidal neovascularization. This paper reviews the possible implications of CCL2 and CX3CL1 signaling in AMD. Expression patterns, single nucleotide polymorphisms (SNPs) association studies, chemokine and chemokine receptor knockout models are discussed. Future AMD treatments could target chemokines and/or their receptors

Thinning of the RPE and choroid associated with T lymphocyte recruitment in aged and light-challenged mice

International audienceThe choroidal vasculature is essential when it comes to bringing oxygen and nutrients to the functioning retina and evacuating debris resulting from the normal visual cycle. Choroidal thinning is a common feature in many human eye diseases, including high myopia [1,2] and retinitis pigmentosa [3,4], and has been reproducibly observed with age [5-7]. However, the association between choroidal thinning and age-related macular degeneration (AMD) remains controversial. Some authors have reported the loss of choriocapillaries in eyes with exudative AMD [8], and choroidal thinning has been detected in some studies [9-11]. Choroidal thinning has also been associated with geographic atrophy (GA), the dry form of late AMD [12-15]. A morphometric analysis by Ramrattan et al. more than two decades ago showed a decrease in choriocapillary density and diameter with age and in GA, but choroidal thinning was only significant with age [6]. Moreover, it has been reported that the choriocapillaries and choroid are thinner in areas where the RPE has degenerated [8]. However, all studies agree that aging is associated with significant choroidal thinning [16-18]. The exact mechanisms behind choroidal thinning with age or disease are not clear. The RPE is a monolayer of pigmented cells situated between photoreceptors and Bruch's membrane; its plays an essential role in the visual cycle. RPE65, which is also called 11-cis retinol isomerase and is strongly expressed in the RPE, participates in the production of 11-cis retinal [19], which is essential for photoreceptor function [20]. Mutations in the RPE65 gene cause progressive photoreceptor degeneration [21,22] and adult RPE65 −/

Effects of triamcinolone acetonide on vessels of the posterior segment of the eye

PURPOSE: This study investigates the effects of triamcinolone acetonide (TA) on retinal endothelial cells in vitro and explores the potential vascular toxic effect of TA injected into the vitreous cavity of rats in vivo. METHODS: Subconfluent endothelial cells were treated with either 0.1 mg/ml or 1 mg/ml TA in 1% ethanol. Control cells were either untreated or exposed to 1% ethanol. Cell viability was evaluated at 24 h, 72 h, and five days using the tetrazolium 3-(4,5-dimethylthiazol-2-yl)-2,5 phenyltetrazolium bromide test (MTT) and lactate dehydrogenase (LDH) assays. Cell proliferation was evaluated by 5-bromo-2-deoxyuridine (BrdU) test. Apoptosis was evaluated by terminal deoxynucleotidyl transferase dUTP nick end labeling assay (TUNEL assay), annexin-binding, and caspase 3 activation. Caspase-independent cell deaths were investigated by immunohistochemistry using antibodies against apoptosis inducing factor (AIF), cytochrome C, microtubule-associated protein (MAP)-light chain 3 (MAP-LC3), and Leukocyte Elastase Inhibitor/Leukocyte Elastase Inhibitor-derived DNase II (LEI/L-DNase II). In vivo, semithin and ultrathin structure analysis and vascular casts were performed to examine TA-induced changes of the choroidal vasculature. In addition, outer segments phagocytosis assay on primary retinal pigment epithelium (RPE) cells was performed to assess cyclooxygenase (COX-2) and vascular endothelial growth factor (VEGF) mRNAs upregulation with or without TA. RESULTS: The inhibitory effect of TA on cell proliferation could not explain the significant reduction in cell viability. Indeed, TA induced a time-dependent reduction of bovine retinal endothelial cells viability. Annexin-binding positive cells were observed. Cytochrome C was not released from mitochondria. L-DNase II was found translocated to the nucleus, meaning that LEI was changed into L-DNase II. AIF was found nuclearized in some cells. LC3 labeling showed the absence of autophagic vesicles. No autophagy or caspase dependent apoptosis was identified. At 1 mg/ml TA induced necrosis while exposure to lower concentrations for 3 to 5 days induced caspase independent apoptosis involving AIF and LEI/L-DNase II. In vivo, semithin and ultrathin structure analysis and vascular casts revealed that TA mostly affected the choroidal vasculature with a reduction of choroidal thickness and increased the avascular areas of the choriocapillaries. Experiments performed on primary RPE cells showed that TA downregulates the basal expression of COX-2 and VEGF and inhibits the outer segments (OS)-dependent COX-2 induction but not the OS-dependent VEGF induction. CONCLUSIONS: This study demonstrates for the first time that glucocorticoids exert direct toxic effect on endothelial cells through caspase-independent cell death mechanisms. The choroidal changes observed after TA intravitreous injection may have important implications regarding the safety profile of TA use in human eyes

Rôles des chimiokines dans le développement de la dégénérescence maculaire liée à l’âge

International audienceRole of chemokines in the development of age-related macular degeneration. Age-related macular degeneration (AMD) is the main cause of irreversible blindness in industrialized nations. Recent research has emphasized the importance of inflam-matory processes in pathogenesis of this disease. Chemotactic cytokines also named chemokines are important mediators of inflammation and might have a role in development of this disease. They appear to be crucial in the subretinal microglia / macrophage accumulation observed in AMD and may participate in the development of retinal degeneration and in choroidal neovascularization. This paper reviews the possible implication of chemokines in the development of AMD.La dégénérescence maculaire liéeà l'âge (DMLA) est la principale cause de cécité irréversible dans les pays industrialisés. Lesétudes récentes mettent en exergue l'importance des processus inflammatoires dans le développement de la maladie. Les cytokines chimiotactiques, dénommées chimiokines, qui apparaissent comme des médiateurs importants de l'inflammation, pourraient jouer un rôle dans le développement de la DMLA. Plus particulièrement, elles semblent indispensables dans le processus d'accumulation des microglies/macrophages dans l'espace sous-rétinien observé au cours de la DMLA. Elles pourraient par conséquent partici-per au développement de la dégénérescence rétinienne et de la néovascularisation choroïdienne. Dans cette revue, nous décrirons l'implication des chimiokines et de leurs récepteurs dans le développement de la DMLA

APOE Isoforms Control Pathogenic Subretinal Inflammation in Age-Related Macular Degeneration

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CD36 plays an important role in the clearance of oxLDL and associated age-dependent sub-retinal deposits

Age-related macular degeneration (AMD) represents the major cause of vision loss in industrialized nations. Laminar deposits in Bruch's membrane (BM) are among the first prominent histopathologic features, along with drusen formation, and have been found to contain oxidized lipids. Increases in concentrations of oxidized LDL (oxLDL) in plasma are observed with age and high fat high (HFHC) cholesterol diet. CD36 is the principal receptor implicated in uptake of oxLDL, and is expressed in the retinal pigment epithelium (RPE). We determined if CD36 participates in oxLDL uptake in RPE and correspondingly in clearance of sub-retinal deposits. Uptake of oxLDL by RPE in vitro and in vivo was CD36-dependent. CD36 deficiency in mice resulted in age-associated accumulation of oxLDL and sub-retinal BM thickening, despite fed a regular diet. Conversely, treatment of HFHC-fed ApoE null mice with a CD36 agonist, EP80317 (300 μg/kg/day), markedly diminished thickening of BM, and partially preserved (in part) photoreceptor function. In conclusion, our data uncover a new role for CD36 in the clearance of oxidized lipids from BM and in the prevention of age-dependent sub-retinal laminar deposits

Deletion of the myeloid endothelin-B receptor confers long-term protection from angiotensin II-mediated renal, retinal & vascular injury

International audienceThe endothelin system may be an important player in hypertensive end-organ injury as endothelin-1 increases blood pressure and is pro-inflammatory. The immune system is emerging as an important regulator of blood pressure and we have shown that the early hypertensive response to angiotensin-II infusion was amplified in mice deficient of myeloid endothelin-B (ETB) receptors (LysM-CreEdnrblox/lox). Hypothesizing that these mice would display enhanced organ injury, we gave angiotensin-II to LysM-CreEdnrblox/lox and littermate controls (Ednrblox/lox) for six weeks. Unexpectedly, LysM-CreEdnrblox/lox mice were significantly protected from organ injury, with less proteinuria, glomerulosclerosis and inflammation of the kidney compared to controls. In the eye, LysM-CreEdnrblox/lox mice had fewer retinal hemorrhages, less microglial activation and less vessel rarefaction. Cardiac remodeling and dysfunction were similar in both groups at week six but LysM-CreEdnrblox/lox mice had better endothelial function. Although blood pressure was initially higher in LysM-CreEdnrblox/lox mice, this was not sustained. A natriuretic switch at about two weeks, due to enhanced ETB signaling in the kidney, induced a hypertensive reversal. By week six, blood pressure was lower in LysM-CreEdnrblox/lox mice than in controls. At six weeks, macrophages from LysM-CreEdnrblox/lox mice were more anti-inflammatory and had greater phagocytic ability compared to the macrophages of Ednrblox/lox mice. Thus, myeloid cell ETB receptor signaling drives this injury both through amplifying hypertension and by inflammatory polarization of macrophages

FoxP3 expression by retinal pigment epithelial cells: transcription factor with potential relevance for the pathology of age-related macular degeneration

Background: Forkhead-Box-Protein P3 (FoxP3) is a transcription factor and marker of regulatory T cells, converting naive T cells into Tregs that can downregulate the effector function of other T cells. We previously detected the expression of FoxP3 in retinal pigment epithelial (RPE) cells, forming the outer blood-retina barrier of the immune privileged eye. Methods: We investigated the expression, subcellular localization, and phosphorylation of FoxP3 in RPE cells in vivo and in vitro after treatment with various stressors including age, retinal laser burn, autoimmune inflammation, exposure to cigarette smoke, in addition of IL-1 beta and mechanical cell monolayer destruction. Eye tissue from humans, mouse models of retinal degeneration and rats, and ARPE-19, a human RPE cell line for in vitro experiments, underwent immunohistochemical, immunofluorescence staining, and PCR or immunoblot analysis to determine the intracellular localization and phosphorylation of FoxP3. Cytokine expression of stressed cultured RPE cells was investigated by multiplex bead analysis. Depletion of the FoxP3 gene was performed with CRISPR/Cas9 editing. Results: RPE in vivo displayed increased nuclear FoxP3-expression with increases in age and inflammation, long-term exposure of mice to cigarette smoke, or after laser burn injury. The human RPE cell line ARPE-19 constitutively expressed nuclear FoxP3 under non-confluent culture conditions, representing a regulatory phenotype under chronic stress. Confluently grown cells expressed cytosolic FoxP3 that was translocated to the nucleus after treatment with IL-1 beta to imitate activated macrophages or after mechanical destruction of the monolayer. Moreover, with depletion of FoxP3, but not of a control gene, by CRISPR/Cas9 gene editing decreased stress resistance of RPE cells. Conclusion: Our data suggest that FoxP3 is upregulated by age and under cellular stress and might be important for RPE function

CD36 Deficiency Leads to Choroidal Involution via COX2 Down-Regulation in Rodents

Florian Sennelaub and colleagues show that CD36 deficiency leads to choroidal involution, a key feature of "dry" age-related macular degeneration, via COX-2 down-regulation in the retinal pigment epithelium