Inflammatory mechanisms contributing to retinal alterations in HIV infection and long-term ART.

People living with HIV (PLWH) may face an increased risk of eye complications associated with ageing, chronic inflammation, and the toxicity arising from long-term antiretroviral therapy (ART). This review aims to understand how inflammatory pathways contribute to retinal alterations observed in PLWH on long-term ART. This review was conducted using four electronic database searches, namely Scopus, Hinari, Google Scholar, and PubMed; from 1996 (when ART became available) until January 2022, without language restriction. Sources from clinical trials, meta-analyses, randomised controlled trials, and systematic reviews were used. Dysregulated para-inflammation (chronic inflammation) damages the blood-retina barrier, resulting in the altered retinal immune privilege and leading to the development of retinal and blood vessel changes. There is an interplay between the effects of the disease versus ART. ART causes mitochondrial toxicity, which affects the retinal ganglion cells and retinal pigment epithelium (RPE) due to oxidative stress. Infection by HIV also affects retinal microglia, which contributes to RPE damage. Both of these mechanisms affect the blood vessels. Assessing the integrity of the inner and outer blood-retina barrier is a pivotal point in pinpointing the pathogenesis of inner retinal alterations. Optical coherence tomography is a valuable tool to assess these changes. There is a paucity of research to understand how these structural changes may affect visual function, such as contrast sensitivity and colour vision.

Neutrophil-inflicted vasculature damage suppresses immune-mediated optic nerve regeneration.

In adult mammals, injured retinal ganglion cells (RGCs) fail to spontaneously regrow severed axons, resulting in permanent visual deficits. Robust axon growth, however, is observed after intra-ocular injection of particulate ß-glucan isolated from yeast. Blood-borne myeloid cells rapidly respond to ß-glucan, releasing numerous pro-regenerative factors. Unfortunately, the pro-regenerative effects are undermined by retinal damage inflicted by an overactive immune system. Here, we demonstrate that protection of the inflamed vasculature promotes immune-mediated RGC regeneration. In the absence of microglia, leakiness of the blood-retina barrier increases, pro-inflammatory neutrophils are elevated, and RGC regeneration is reduced. Functional ablation of the complement receptor 3 (CD11b/integrin-αM), but not the complement components C1q-/- or C3-/-, reduces ocular inflammation, protects the blood-retina barrier, and enhances RGC regeneration. Selective targeting of neutrophils with anti-Ly6G does not increase axogenic neutrophils but protects the blood-retina barrier and enhances RGC regeneration. Together, these findings reveal that protection of the inflamed vasculature promotes neuronal regeneration.

Dissimilar Effect of P-Glycoprotein and Breast Cancer Resistance Protein Inhibition on the Distribution of Erlotinib to the Retina and Brain in Humans and Mice.

P-glycoprotein (P-gp) and breast cancer resistance protein (BCRP) are two ATP-binding cassette efflux transporters that are coexpressed at the human blood-brain barrier (BBB) and blood-retina barrier (BRB). While pharmacological inhibition of P-gp and/or BCRP results in increased brain distribution of dual P-gp/BCRP substrate drugs, such as the tyrosine kinase inhibitor erlotinib, the effect of P-gp and/or BCRP inhibition on the retinal distribution of such drugs has hardly been investigated. In this study, we used positron emission tomography (PET) imaging to assess the effect of transporter inhibition on the distribution of [11C]erlotinib to the human retina and brain. Twenty two healthy volunteers underwent two PET scans after intravenous (i.v.) injection of a microdose (<5 µg) of [11C]erlotinib, a baseline scan, and a second scan either with concurrent i.v. infusion of tariquidar to inhibit P-gp (n = 5) or after oral intake of single ascending doses of erlotinib (300 mg, 650 mg, or 1000 mg, n = 17) to saturate erlotinib transport. In addition, transport of [3H]erlotinib to the retina and brain was assessed in mice by in situ carotid perfusion under various drug transporter inhibition settings. In comparison to the baseline PET scan, coadministration of tariquidar or erlotinib led to a significant decrease of [11C]erlotinib total volume of distribution (VT) in the human retina by -25 ± 8% (p ≤ 0.05) and -41 ± 16% (p ≤ 0.001), respectively. In contrast, erlotinib intake led to a significant increase in [11C]erlotinib VT in the human brain (+20 ± 16%, p ≤ 0.001), while administration of tariquidar did not result in any significant changes. In situ carotid perfusion experiments showed that both P-gp and BCRP significantly limit the distribution of erlotinib to the mouse retina and brain but revealed a similar discordant effect at the mouse BRB and BBB following co-perfusion with tariquidar and erlotinib as in humans. Co-perfusion with prototypical inhibitors of solute carrier transporters did not reveal a significant contribution of organic cation transporters (e.g., OCTs and OCTNs) and organic anion-transporting polypeptides (e.g., OATP2B1) to the retinal and cerebral distribution of erlotinib. In conclusion, we observed a dissimilar effect after P-gp and/or BCRP inhibition on the retinal and cerebral distribution of [11C]erlotinib. The exact mechanism for this discrepancy remains unclear but may be related to the function of an unidentified erlotinib uptake carrier sensitive to tariquidar inhibition at the BRB. Our study highlights the great potential of PET to study drug distribution to the human retina and to assess the functional impact of membrane transporters on ocular drug distribution.

Therapeutic targeting of the complement system in ocular disease.

The complement system is involved in the pathogenesis of several ocular diseases, providing a rationale for the investigation of complement-targeting therapeutics for these conditions. Dry age-related macular degeneration, as characterised by geographic atrophy (GA), is currently the most active area of research for complement-targeting therapeutics, with a complement C3 inhibitor approved in the United States earlier this year marking the first approved therapy for GA. This review discusses the role of complement in ocular disease, provides an overview of the complement-targeting agents currently under development for ocular conditions, and reflects on the lessons that can be learned from the preclinical investigations and clinical trials conducted in this field to date.

Efficacy and Safety of Low-Intensity Pulsed Ultrasound-Induced Blood-Retinal Barrier Opening in Mice.

Systemic drugs can treat various retinal pathologies such as retinal cancers; however, their ocular diffusion may be limited by the blood-retina barrier (BRB). Sonication corresponds to the use of ultrasound (US) to increase the permeability of cell barriers including in the BRB. The objective was to study the efficacy and safety of sonication using microbubble-assisted low-intensity pulsed US in inducing a transient opening of the BRB. The eyes of C57/BL6J mice were sonicated at different acoustic pressures (0.10 to 0.50 MPa). Efficacy analyses consisted of fluorescein angiography (FA) performed at different timepoints and the size of the leaked molecules was assessed using FITC-marked dextrans. Tolerance was assessed by fundus photographs, optical coherence tomography, immunohistochemistry, RT-qPCR, and electroretinograms. Sonication at 0.15 MPa was the most suitable pressure for transient BRB permeabilization without altering the morphology or function of the retina. It did not increase the expression of inflammation or apoptosis markers in the retina, retinal pigment epithelium, or choroid. The dextran assay suggested that drugs up to 150 kDa in size can cross the BRB. Microbubble-assisted sonication at an optimized acoustic pressure of 0.15 MPa provides a non-invasive method to transiently open the BRB, increasing the retinal diffusion of systemic drugs without inducing any noticeable side-effect.

Morphine pharmacokinetics and opioid transporter expression at the blood-retina barrier of male and female mice.

Opioids are effective analgesics for treating moderate to severe pain, however, their use must be weighed against their dangerous side effects. Investigations into opioid pharmacokinetics provide crucial information regarding both on- and off-target drug effects. Our recent work showed that morphine deposits and accumulates in the mouse retina at higher concentrations than in the brain upon chronic systemic exposure. We also found reduced retinal expression of P-glycoprotein (P-gp), a major opioid extruder at the blood-brain barrier (BBB). Here, we systematically interrogated the expression of three putative opioid transporters at the blood-retina barrier (BRB): P-gp, breast cancer resistance protein (Bcrp) and multidrug resistance protein 2 (Mrp2). Using immunohistochemistry, we found robust expression of P-gp and Bcrp, but not Mrp2, at the inner BRB of the mouse retina. Previous studies have suggested that P-gp expression may be regulated by sex hormones. However, upon acute morphine treatment we found no sex differences in morphine deposition levels in the retina or brain, nor on transporter expression in the retinas of males and females with a high or low estrogen:progesterone ratio. Importantly, we found that P-gp, but not Bcrp, expression significantly correlated with morphine concentration in the retina, suggesting P-gp is the predominant opioid transporter at the BRB. In addition, fluorescence extravasation studies revealed that chronic morphine treatment did not alter the permeability of either the BBB or BRB. Together, these data suggest that reduced P-gp expression mediates retinal morphine accumulation upon systemic delivery, and in turn, potential effects on circadian photoentrainment.

Cerebrospinal Fluid Pressure Reduction Induces Glia-Mediated Retinal Inflammation and Leads to Retinal Ganglion Cell Injury in Rats.

Low intracranial pressure (LICP)-induced translaminar cribrosa pressure difference (TLCPD) elevation has been proven as a risk factor in glaucomatous neurodegeneration, whereas the underlying retinal immune features of LICP-induced retinal ganglion cells (RGC) injury remain elusive. Here, we identified the retinal immune characteristics of LICP rats, and minocycline (Mino) treatment was utilized to analyze its inhibitory role in glia-mediated retinal inflammation of LICP rats. The results showed that retrograde axonal transport was decreased in LICP rats without significant RGC loss, indicating the RGC injury was at an early stage before the morphological loss. The activation of retinal microglia and astrocytes with morphologic and M1 or A1-marker alternations was detected in TLCPD elevation rats, the activation level is more dramatic in HIOP rats than in the LICP rats (P<0.05). Besides, we detected reduced retinal tight junction protein expressions, accompanied by specific imbalance patterns of T lymphocytes in the retina of both LICP and HIOP rats (P<0.05). Further Mino treatment showed an effective inhibitory role in glia-driven inflammatory responses in LICP rats, including improving retrograde axonal transport, inhibiting retinal glial activation and proinflammatory subtype polarization, and alleviating the blood-retina barrier compromise. This study identified the glia-mediated retinal inflammation features triggered by LICP stimulus, and Mino application exhibited an effective role in the inhibition of retinal glia-mediated inflammation in LICP-induced TLCPD elevation rats.

Reduced folate carrier 1 is present in retinal microvessels and crucial for the inner blood retinal barrier integrity.

BACKGROUND: Reduced folate carrier 1 (RFC1; SLC19a1) is the main responsible transporter for the B9 family of vitamins named folates, which are essential for normal tissue growth and development. While folate deficiency resulted in retinal vasculopathy, the expression and the role of RFC1 in blood-retinal barrier (BRB) are not well known. METHODS: We used whole mount retinas and trypsin digested microvessel samples of adult mice. To knockdown RFC1, we delivered RFC1-targeted short interfering RNA (RFC1-siRNA) intravitreally; while, to upregulate RFC1 we delivered lentiviral vector overexpressing RFC1. Retinal ischemia was induced 1-h by applying FeCl3 to central retinal artery. We used RT-qPCR and Western blotting to determine RFC1. Endothelium (CD31), pericytes (PDGFR-beta, CD13, NG2), tight-junctions (Occludin, Claudin-5 and ZO-1), main basal membrane protein (Collagen-4), endogenous IgG and RFC1 were determined immunohistochemically. RESULTS: Our analyses on whole mount retinas and trypsin digested microvessel samples of adult mice revealed the presence of RFC1 in the inner BRB and colocalization with endothelial cells and pericytes. Knocking down RFC1 expression via siRNA delivery resulted in the disintegration of tight junction proteins and collagen-4 in twenty-four hours, which was accompanied by significant endogenous IgG extravasation. This indicated the impairment of BRB integrity after an abrupt RFC1 decrease. Furthermore, lentiviral vector-mediated RFC1 overexpression resulted in increased tight junction proteins and collagen-4, confirming the structural role of RFC1 in the inner BRB. Acute retinal ischemia decreased collagen-4 and occludin levels and led to an increase in RFC1. Besides, the pre-ischemic overexpression of RFC1 partially rescued collagen-4 and occludin levels which would be decreased after ischemia. CONCLUSION: In conclusion, our study clarifies the presence of RFC1 protein in the inner BRB, which has recently been defined as hypoxia-immune-related gene in other tissues and offers a novel perspective of retinal RFC1. Hence, other than being a folate carrier, RFC1 is an acute regulator of the inner BRB in healthy and ischemic retinas.

Netrin-1 binding to Unc5B regulates Blood-Retina Barrier integrity.

Background: The blood brain barrier (BBB) preserves neuronal function in the central nervous system (CNS) by tightly controlling metabolite exchanges with the blood. In the eye, the retina is likewise protected by the blood-retina barrier (BRB) to maintain phototransduction. We showed that the secreted guidance cue Netrin-1 regulated BBB integrity, by binding to endothelial Unc5B and regulating canonical ß-catenin dependent expression of BBB gene expression. Objective: Here, we investigated if Netrin-1-binding to endothelial Unc5B also controlled BRB integrity, and if this process involved Norrin/ß-catenin signaling, which is the major known driver of BRB development and maintenance. Methods: We analyzed Tamoxifen-inducible loss- and gain- of-function alleles of Unc5B, Ntn1 and Ctnnb1 in conjunction with tracer injections and biochemical signaling studies. Results: Inducible endothelial Unc5B deletion, and inducible global Ntn1 deletion in postnatal mice reduced phosphorylation of the Norrin receptor LRP5, leading to reduced ß-catenin and LEF1 expression, conversion of retina endothelial cells from a barrier-competent Claudin-5+/PLVAP- state to a Claudin-5-/PLVAP+ leaky phenotype, and extravasation of injected low molecular weight tracers. Inducible Ctnnb1 gain of function rescued vascular leak in Unc5B mutants, and Ntn1 overexpression induced BRB tightening. Unc5B expression in pericytes contributed to BRB permeability, via regulation of endothelial Unc5B. Mechanistically, Netrin-1-Unc5B signaling promoted ß-catenin dependent BRB signaling by enhancing phosphorylation of the Norrin receptor LRP5 via the Discs large homologue 1 (Dlg1) intracellular scaffolding protein. Conclusions: The data identify Netrin1-Unc5B as novel regulators of BRB integrity, with implications for diseases associated with BRB disruption.

Untargeted Analysis of Lipids Containing Very Long Chain Fatty Acids in Retina and Retinal Tight Junctions.

Several recent studies suggest that C24-C38 very long chain fatty acids (VLCFA) play an important role in vision, and decreased levels of retina VLCFA have been associated with vision disorders including the onset and progression of diabetic retinopathy in animal models. Traditional methods for VLCFA analysis lack the sensitivity and specificity needed to enable detailed characterization of VLCFA incorporation into complex lipids in tissues and subcellular components. To assess whether decreased VLCFA in diabetic retina are directly implicated in diabetes-induced breakdown of the blood-retinal barrier, we demonstrated the utility of performing untargeted lipid analysis via Orbitrap high resolution/accurate mass MS and MS/MS-based shotgun lipidomics to identify structural lipids containing VLCFA substituents. This comprehensive and highly sensitive approach to untargeted lipid identification enabled us to characterize low-abundance sphingolipids containing very long chain fatty acids from isolated retinal tight junction complexes, as well as VLCFA-containing phospholipids in retinal tissues. To facilitate future biochemical and physiological studies of the roles of VLCFA in blood-retina barrier integrity and maintenance of vision, this chapter describes steps to isolate tight junction complexes from a cell culture model of the outer blood-retinal barrier and perform untargeted Orbitrap high resolution/accurate mass-based lipid analysis to identify VLCFA in tight junctions and retina tissue.

Morphine Accumulates in the Retina Following Chronic Systemic Administration.

Opioid transport into the central nervous system is crucial for the analgesic efficacy of opioid drugs. Thus, the pharmacokinetics of opioid analgesics such as morphine have been extensively studied in systemic circulation and the brain. While opioid metabolites are routinely detected in the vitreous fluid of the eye during postmortem toxicological analyses, the pharmacokinetics of morphine within the retina of the eye remains largely unexplored. In this study, we measured morphine in mouse retina following systemic exposure. We showed that morphine deposits and persists in the retina long after levels have dropped in the serum. Moreover, we found that morphine concentrations (ng/mg tissue) in the retina exceeded brain morphine concentrations at all time points tested. Perhaps most intriguingly, these data indicate that following chronic systemic exposure, morphine accumulates in the retina, but not in the brain or serum. These results suggest that morphine can accumulate in the retina following chronic use, which could contribute to the deleterious effects of chronic opioid use on both image-forming and non-image-forming visual functions.

Brain and Retinal Organoids for Disease Modeling: The Importance of In Vitro Blood-Brain and Retinal Barriers Studies.

Brain and retinal organoids are functional and dynamic in vitro three-dimensional (3D) structures derived from pluripotent stem cells that spontaneously organize themselves to their in vivo counterparts. Here, we review the main literature data of how these organoids have been developed through different protocols and how they have been technically analyzed. Moreover, this paper reviews recent advances in using organoids to model neurological and retinal diseases, considering their potential for translational applications but also pointing out their limitations. Since the blood-brain barrier (BBB) and blood-retinal barrier (BRB) are understood to play a fundamental role respectively in brain and eye functions, both in health and in disease, we provide an overview of the progress in the development techniques of in vitro models as reliable and predictive screening tools for BBB and BRB-penetrating compounds. Furthermore, we propose potential future directions for brain and retinal organoids, in which dedicated biobanks will represent a novel tool for neuroscience and ophthalmology research.

Ex Vivo and In Vitro Analysis Identify a Detrimental Impact of Neutrophil Extracellular Traps on Eye Structures in Equine Recurrent Uveitis.

Equine recurrent uveitis (ERU) is a common ocular disease of horses and described as a model for human autoimmune uveitis. This immune-mediated, inflammatory condition progressively destroys the eye, ultimately leading to blindness. Genetic and autoimmune factors, next to infections with Leptospira, are discussed as key factors in the pathogenesis. Furthermore, a release of neutrophil extracellular traps (NETs) by activated neutrophils is involved. NETs are composed of decondensed chromatin and proteins that can immobilize invading pathogens. However, if NETs accumulate, they can contribute to detrimental autoimmune processes. Thus, we aimed to investigate the impact of NETs in ERU patients. Therefore, we quantified several NET-markers (cell-free DNA, nucleosomes, citrullinated histone H3, histone-myeloperoxidase complexes, interleukin-17, equine cathelicidin 1 and DNase I activity) and NET-autoantibodies in sera and vitreous body fluids (VBF) of ERU-diseased horses and correlated the data with the disease status (signalment, ERU scores and Leptospira infection status). NET markers were detected to varying degrees in VBF of diseased horses, and partially correlated to disease severity and the presence of Leptospira spp. Cell-free DNA and nucleosomes as NET markers correlate with ERU severity in total and VBF scores, despite the presence of active DNases. Additionally, a significant correlation between fundus affection in the eye and NET autoantibodies was detectable. Therefore, we further investigated the influence of VBF samples from equine patients and isolated NETs on the blood-retina barrier in a cell culture model. VBF of diseased horses significantly induced cytotoxicity in retinal pigment epithelial cells. Moreover, partially digested NETs also resulted in cytotoxic effects. In the presence of lipopolysaccharide (LPS), the main component of the leptospiral surface, both undigested and completely digested NETs were cytotoxic. Correlations between the ERU-scores and Leptospira were also calculated. Detection of leptospiral DNA, and antibody titers of the serovar Grippotyphosa correlated with disease severity. In addition, a correlation between Leptospira and several NET markers was observed in VBF. Altogether, our findings suggest a positive correlation between NET markers with disease severity and involvement of Leptospira in the VBF of ERU-diseased horses, as well as a cytotoxic effect of NETs in eyes.

Blood-retina barrier dysfunction in experimental autoimmune uveitis: the pathogenesis and therapeutic targets.

Experimental autoimmune uveitis (EAU), an animal model of human uveitis, is characterized by infiltration of autoimmune T cells in the uvea as well as in the retina of susceptible animals. EAU is induced by the immunization of uveitogenic antigens, including either retinal soluble-antigen or interphotoreceptor retinoid-binding proteins, in Lewis rats. The pathogenesis of EAU in rats involves the proliferation of autoimmune T cells in peripheral lymphoid tissues and breakdown of the blood-retinal barrier, primarily in the uvea and retina, finally inducing visual dysfunction. In this review, we describe recent EAU studies to facilitate the design of a therapeutic strategy through the interruption of uveitogenic factors during the course of EAU, which will be helpful for controlling human uveitis.

The role of lipopolysaccharides in diabetic retinopathy.

Diabetes mellitus (DM) is a complex metabolic syndrome characterized by hyperglycemia. Diabetic retinopathy (DR) is the most common complication of DM and the leading cause of blindness in the working-age population of the Western world. Lipopolysaccharides (LPS) is an essential ingredient of the outer membrane of gram-negative bacteria, which induces systemic inflammatory responses and cellular apoptotic changes in the host. High-level serum LPS has been found in diabetic patients at the advanced stages, which is mainly due to gut leakage and dysbiosis. In this light, increasing evidence points to a strong correlation between systemic LPS challenge and the progression of DR. Although the underlying molecular mechanisms have not been fully elucidated yet, LPS-related pathobiological events in the retina may contribute to the exacerbation of vasculopathy and neurodegeneration in DR. In this review, we focus on the involvement of LPS in the progression of DR, with emphasis on the blood-retina barrier dysfunction and dysregulated glial activation. Eventually, we summarize the recent advances in the therapeutic strategies for antagonising LPS activity, which may be introduced to DR treatment with promising clinical value.

Adult-induced genetic ablation distinguishes PDGFB roles in blood-brain barrier maintenance and development.

Platelet-derived growth factor B (PDGFB) released from endothelial cells is indispensable for pericyte recruitment during angiogenesis in embryonic and postnatal organ growth. Constitutive genetic loss-of-function of PDGFB leads to pericyte hypoplasia and the formation of a sparse, dilated and venous-shifted brain microvasculature with dysfunctional blood-brain barrier (BBB) in mice, as well as the formation of microvascular calcification in both mice and humans. Endothelial PDGFB is also expressed in the adult quiescent microvasculature, but here its importance is unknown. We show that deletion of Pdgfb in endothelial cells in 2-months-old mice causes a slowly progressing pericyte loss leading, at 12-18 months of age, to ≈50% decrease in endothelial:pericyte cell ratio, ≈60% decrease in pericyte longitudinal capillary coverage and >70% decrease in pericyte marker expression. Similar to constitutive loss of Pdgfb, this correlates with increased BBB permeability. However, in contrast to the constitutive loss of Pdgfb, adult-induced loss does not lead to vessel dilation, impaired arterio-venous zonation or the formation of microvascular calcifications. We conclude that PDFGB expression in quiescent adult microvascular brain endothelium is critical for the maintenance of pericyte coverage and normal BBB function, but that microvessel dilation, rarefaction, arterio-venous skewing and calcification reflect developmental roles of PDGFB.

Ultrasound and Microbubbles for the Treatment of Ocular Diseases: From Preclinical Research towards Clinical Application.

The unique anatomy of the eye and the presence of various biological barriers make efficacious ocular drug delivery challenging, particularly in the treatment of posterior eye diseases. This review focuses on the combination of ultrasound and microbubbles (USMB) as a minimally invasive method to improve the efficacy and targeting of ocular drug delivery. An extensive overview is given of the in vitro and in vivo studies investigating the mechanical effects of ultrasound-driven microbubbles aiming to: (i) temporarily disrupt the blood-retina barrier in order to enhance the delivery of systemically administered drugs into the eye, (ii) induce intracellular uptake of anticancer drugs and macromolecules and (iii) achieve targeted delivery of genes, for the treatment of ocular malignancies and degenerative diseases. Finally, the safety and tolerability aspects of USMB, essential for the translation of USMB to the clinic, are discussed.

Selective Targeting and Tissue Penetration to the Retina by a Systemically Administered Vascular Homing Peptide in Oxygen Induced Retinopathy (OIR).

Pathological angiogenesis is the hallmark of ischemic retinal diseases among them retinopathy of prematurity (ROP) and proliferative diabetic retinopathy (PDR). Oxygen-induced retinopathy (OIR) is a pure hypoxia-driven angiogenesis model and a widely used model for ischemic retinopathies. We explored whether the vascular homing peptide CAR (CARSKNKDC) which recognizes angiogenic blood vessels can be used to target the retina in OIR. We were able to demonstrate that the systemically administered CAR vascular homing peptide homed selectively to the preretinal neovessels in OIR. As a cell and tissue-penetrating peptide, CAR also penetrated into the retina. Hyperoxia used to induce OIR in the retina also causes bronchopulmonary dysplasia in the lungs. We showed that the CAR peptide is not targeted to the lungs in normal mice but is targeted to the lungs after hyperoxia-/hypoxia-treatment of the animals. The site-specific delivery of the CAR peptide to the pathologic retinal vasculature and the penetration of the retinal tissue may offer new opportunities for treating retinopathies more selectively and with less side effects.