Identification of Structures Labeled by Indocyanine Green in the Rat Choroid and Retina Can Guide Interpretation of Indocyanine Green Angiography.

Purpose: Indocyanine green (ICG) is an albumin and lipoprotein binding dye absorbing in the far red used in angiography to visualize choroidal vessels (ICG angiography [ICGA]). To guide interpretation, ICG transport in the choroid, RPE, and retina of rats was studied. Methods: Two conditions were used: RPE/choroid organoculture, incubated for 45 minutes in DMEM medium, 1% fetal bovine serum containing 0.25 mg/mL ICG and RPE/choroid and neural retina flat-mounts at 1 and 6 hours after intravenous ICG injection. Early and late sequences of ICGA were recorded until 6 hours. Ultra-deep red confocal microscope was used to localize ICG in flat-mounts and immunohistochemistry was performed for caveolin-1, tryptase (mast cell marker), and tubulin ß3 (a nerve marker). Results: In the organoculture, ICG penetrated homogeneously in the cytoplasm and stained the membranes of the RPE. At 1 hour after intravenous injection, ICG appeared in fine granules in RPE, partly labeled with caveolin-1 and decreasing at 6 hours. At 1 hour and 6 hours, ICG was found in the retinal vessels, faintly in the inner retina, and in the photoreceptor outer segments at 6 hours. In the choroid, ICG colocalized with mast cells, immunostained with tryptase, and accumulated along the large tubulin ß3-labeled nerve bundles. The hypothesis was raised on the interpretation of late ICGA infrared photography in case of transthyretin amyloidosis with neuropathy. Conclusions: Beside being a vascular dye, ICG is transported from the vessels to the RPE toward the outer retina. It stains mast cells and large choroidal nerves. These observations could help the analysis of ICGA images.

[Macular edema: Understand mechanisms to develop treatments]. / Les œdèmes maculaires - Mieux comprendre leurs mécanismes pour mieux les traiter.

Macular edema is an increase in volume of the central area of the retina, responsible for visual acuity. Visual symptoms handicap the lives of millions of patients with macular edema secondary to chronic and sometimes acute retinal disease. Proteins that neutralize the vascular endothelial growth factor (VEGF) pathway or glucocorticoids, at the cost of repeated intraocular injections over years, limit visual symptoms. A better understanding of why and how edema forms and how therapeutic molecules exert an anti-edematous effect will help prevent this disabling and blinding retinal complication from occurring.

Title: Les œdèmes maculaires - Mieux comprendre leurs mécanismes pour mieux les traiter. Abstract: L'œdème maculaire est une augmentation de volume de la macula, zone centrale de la rétine, responsable de l'acuité visuelle. Des symptômes visuels handicapent la vie de millions de patients atteints d'œdème maculaire secondaire à une maladie chronique et parfois aiguë de la rétine. Les protéines qui neutralisent la voie du facteur de croissance de l'endothélium vasculaire (VEGF) ou les glucocorticoïdes, au prix d'injections intraoculaires répétées pendant des années, limitent les symptômes visuels. Mieux comprendre pourquoi et comment l'œdème se forme et comment les molécules thérapeutiques exercent un effet anti-œdémateux permettra de mieux prévenir la survenue de cette complication rétinienne handicapante et cécitante.

Mechanisms of FH Protection Against Neovascular AMD.

A common allele (402H) of the complement factor H (FH) gene is the major risk factor for age-related macular degeneration (AMD), the leading cause of blindness in the elderly population. Development and progression of AMD involves vascular and inflammatory components partly by deregulation of the alternative pathway of the complement system (AP). The loss of central vision results from atrophy and/or from abnormal neovascularization arising from the choroid. The functional link between FH, the main inhibitor of AP, and choroidal neovascularization (CNV) in AMD remains unclear. In a murine model of CNV used as a model for neovascular AMD (nAMD), intraocular human recombinant FH (recFH) reduced CNV as efficiently as currently used anti-VEGF (vascular endothelial growth factor) antibody, decreasing deposition of C3 cleavage fragments, membrane attack complex (MAC), and microglia/macrophage recruitment markers in the CNV lesion site. In sharp contrast, recFH carrying the H402 risk variant had no effect on CNV indicating a causal link to disease etiology. Only the recFH NTal region (recFH1-7), containing the CCPs1-4 C3-convertase inhibition domains and the CCP7 binding domain, exerted all differential biological effects. The CTal region (recFH7-20) containing the CCP7 and CCPs19-20 binding domains was antiangiogenic but did not reduce the microglia/macrophage recruitment. The antiangiogenic effect of both recFH1-20 and recFH-CCP7-20 resulted from thrombospondin-1 (TSP-1) upregulation independently of the C3 cleavage fragments generation. This study provides insight on the mechanistic role of FH in nAMD and invites to reconsider its therapeutic potential.

Glial cells of the human fovea.

Purpose: The exact cellular types that form the human fovea remain a subject of debate, and few studies have been conducted on human macula to solve this question. The purpose of this study was to perform immunohistochemistry on fresh human samples to characterize the glial cells that form the human fovea. Methods: Immunohistochemistry was performed using antibodies against proteins expressed in astrocytes or in retinal Müller glial cells or both types of cells on six human macula obtained from eyes enucleated for peripheral intraocular tumors and on two postmortem eyes from healthy donors. The posterior poles of the enucleated eyes were cryosectioned and stained with antibodies against the glial proteins GFAP, vimentin, CRALBP, glutamine synthetase, and connexin 43. Results: A population of cells positive for GFAP and negative for glutamine synthetase and CRALBP that express connexin 43 were identified at the roof of the foveal pit. These cells are distinct from the Müller cone cells described by Yamada and Gass, suggesting that another type of foveal glial cells, most likely astrocytes, are present in the human fovea. Conclusions: This study showed that in humans, astrocytic glial cells cover the foveal pit. Their roles in macula homeostasis and mechanisms of macular diseases disease remain to be determined.

Nogo-A-targeting antibody promotes visual recovery and inhibits neuroinflammation after retinal injury.

N-Methyl-D-aspartate (NMDA)-induced neuronal cell death is involved in a large spectrum of diseases affecting the brain and the retina such as Alzheimer's disease and diabetic retinopathy. Associated neurological impairments may result from the inhibition of neuronal plasticity by Nogo-A. The objective of the current study was to determine the contribution of Nogo-A to NMDA excitotoxicity in the mouse retina. We observed that Nogo-A is upregulated in the mouse vitreous during NMDA-induced inflammation. Intraocular injection of a function-blocking antibody specific to Nogo-A (11C7) was carried out 2 days after NMDA-induced injury. This treatment significantly enhanced visual function recovery in injured animals. Strikingly, the expression of potent pro-inflammatory molecules was downregulated by 11C7, among which TNFα was the most durably decreased cytokine in microglia/macrophages. Additional analyses suggest that TNFα downregulation may stem from cofilin inactivation in microglia/macrophages. 11C7 also limited gliosis presumably via P.Stat3 downregulation. Diabetic retinopathy was associated with increased levels of Nogo-A in the eyes of donors. In summary, our results reveal that Nogo-A-targeting antibody can stimulate visual recovery after retinal injury and that Nogo-A is a potent modulator of excitotoxicity-induced neuroinflammation. These data may be used to design treatments against inflammatory eye diseases.