An allosteric interleukin-1 receptor modulator mitigates inflammation and photoreceptor toxicity in a model of retinal degeneration.

BACKGROUND: Inflammation and particularly interleukin-1ß (IL-1ß), a pro-inflammatory cytokine highly secreted by activated immune cells during early AMD pathological events, contribute significantly to retinal neurodegeneration. Here, we identify specific cell types that generate IL-1ß and harbor the IL-1 receptor (IL-1R) and pharmacologically validate IL-1ß's contribution to neuro-retinal degeneration using the IL-1R allosteric modulator composed of the amino acid sequence rytvela (as well as the orthosteric antagonist, Kineret) in a model of blue light-induced retinal degeneration. METHODS: Mice were exposed to blue light for 6 h and sacrificed 3 days later. Mice were intraperitoneally injected with rytvela, Kineret, or vehicle twice daily for 3 days. The inflammatory markers F4/80, NLRP3, caspase-1, and IL-1ß were assessed in the retinas. Single-cell RNA sequencing was used to determine the cell-specific expression patterns of retinal Il1b and Il1r1. Macrophage-induced photoreceptor death was assessed ex vivo using retinal explants co-cultured with LPS-activated bone marrow-derived macrophages. Photoreceptor cell death was evaluated by the TUNEL assay. Retinal function was assessed by flash electroretinography. RESULTS: Blue light markedly increased the mononuclear phagocyte recruitment and levels of inflammatory markers associated with photoreceptor death. Co-localization of NLRP3, caspase-1, and IL-1ß with F4/80+ mononuclear phagocytes was clearly detected in the subretinal space, suggesting that these inflammatory cells are the main source of IL-1ß. Single-cell RNA sequencing confirmed the immune-specific expression of Il1b and notably perivascular macrophages in light-challenged mice, while Il1r1 expression was found primarily in astrocytes, bipolar, and vascular cells. Retinal explants co-cultured with LPS/ATP-activated bone marrow-derived macrophages displayed a high number of TUNEL-positive photoreceptors, which was abrogated by rytvela treatment. IL-1R antagonism significantly mitigated the inflammatory response triggered in vivo by blue light exposure, and rytvela was superior to Kineret in preserving photoreceptor density and retinal function. CONCLUSION: These findings substantiate the importance of IL-1ß in neuro-retinal degeneration and revealed specific sources of Il1b from perivascular MPs, with its receptor Ilr1 being separately expressed on surrounding neuro-vascular and astroglial cells. They also validate the efficacy of rytvela-induced IL-1R modulation in suppressing detrimental inflammatory responses and preserving photoreceptor density and function in these conditions, reinforcing the rationale for clinical translation.

The Inability of the Choroid to Revascularize in Oxygen-Induced Retinopathy Results from Increased p53/miR-Let-7b Activity.

Retinopathy of prematurity (ROP) is characterized by an initial retinal avascularization, followed by pathologic neovascularization. Recently, choroidal thinning has also been detected in children formerly diagnosed with ROP; a similar sustained choroidal thinning is observed in ROP models. But the mechanism underlying the lack of choroidal revascularization remains unclear and was investigated in an oxygen-induced retinopathy (OIR) model. In OIR, evidence of senescence was detected, preceded by oxidative stress in the choroid and the retinal pigment epithelium. This was associated with a global reduction of proangiogenic factors, including insulin-like growth factor 1 receptor (Igf1R). Coincidentally, tumor suppressor p53 was highly expressed in the OIR retinae. Curtailing p53 activity resulted in reversal of senescence, normalization of Igf1r expression, and preservation of choroidal integrity. OIR-induced down-regulation of Igf1r was mediated at least partly by miR-let-7b as i) let-7b expression was augmented throughout and beyond the period of oxygen exposure, ii) let-7b directly targeted Igf1r mRNA, and iii) p53 knock-down blunted let-7b expression, restored Igf1r expression, and elicited choroidal revascularization. Finally, restoration of Igf1r expression rescued choroid thickness. Altogether, this study uncovers a significant mechanism for defective choroidal revascularization in OIR, revealing a new role for p53/let-7b/IGF-1R axis in the retina. Future investigations on this (and connected) pathway could further our understanding of other degenerative choroidopathies, such as geographic atrophy.

Retinal neurons curb inflammation and enhance revascularization in ischemic retinopathies via proteinase-activated receptor-2.

Ischemic retinopathies are characterized by sequential vaso-obliteration followed by abnormal intravitreal neovascularization predisposing patients to retinal detachment and blindness. Ischemic retinopathies are associated with robust inflammation that leads to generation of IL-1ß, which causes vascular degeneration and impairs retinal revascularization in part through the liberation of repulsive guidance cue semaphorin 3A (Sema3A). However, retinal revascularization begins as inflammation culminates in ischemic retinopathies. Because inflammation leads to activation of proteases involved in the formation of vasculature, we hypothesized that proteinase-activated receptor (Par)-2 (official name F2rl1) may modulate deleterious effects of IL-1ß. Par2, detected mostly in retinal ganglion cells, was up-regulated in oxygen-induced retinopathy. Surprisingly, oxygen-induced retinopathy-induced vaso-obliteration and neovascularization were unaltered in Par2 knockout mice, suggesting compensatory mechanisms. We therefore conditionally knocked down retinal Par2 with shRNA-Par2-encoded lentivirus. Par2 knockdown interfered with normal revascularization, resulting in pronounced intravitreal neovascularization; conversely, the Par2 agonist peptide (SLIGRL) accelerated normal revascularization. In vitro and in vivo exploration of mechanisms revealed that IL-1ß induced Par2 expression, which in turn down-regulated sequentially IL-1 receptor type I and Sema3A expression through Erk/Jnk-dependent processes. Collectively, our findings unveil an important mechanism by which IL-1ß regulates its own endothelial cytotoxic actions by augmenting neuronal Par2 expression to repress sequentially IL-1 receptor type I and Sema3A expression. Timely activation of Par2 may be a promising therapeutic avenue in ischemic retinopathies.

Microglia/macrophages migrate through retinal epithelium barrier by a transcellular route in diabetic retinopathy: role of PKCζ in the Goto Kakizaki rat model.

Diabetic retinopathy is associated with ocular inflammation, leading to retinal barrier breakdown, macular edema, and visual cell loss. We investigated the molecular mechanisms involved in microglia/macrophages trafficking in the retina and the role of protein kinase Cζ (PKCζ) in this process. Goto Kakizaki (GK) rats, a model for spontaneous type 2 diabetes were studied until 12 months of hyperglycemia. Up to 5 months, sparse microglia/macrophages were detected in the subretinal space, together with numerous pores in retinal pigment epithelial (RPE) cells, allowing inflammatory cell traffic between the retina and choroid. Intercellular adhesion molecule-1 (ICAM-1), caveolin-1 (CAV-1), and PKCζ were identified at the pore border. At 12 months of hyperglycemia, the significant reduction of pores density in RPE cell layer was associated with microglia/macrophages accumulation in the subretinal space together with vacuolization of RPE cells and disorganization of photoreceptors outer segments. The intraocular injection of a PKCζ inhibitor at 12 months reduced iNOS expression in microglia/macrophages and inhibited their migration through the retina, preventing their subretinal accumulation. We show here that a physiological transcellular pathway takes place through RPE cells and contributes to microglia/macrophages retinal trafficking. Chronic hyperglycemia causes alteration of this pathway and subsequent subretinal accumulation of activated microglia/macrophages.

Neovascularization in retinopathy of prematurity: opposing actions of neuronal factors GPR91 and semaphorins 3A.

UNLABELLED: Retinopathy of prematurity (ROP) is a major cause of severe visual deficits in children. This review focuses on the role of newly identified factors from retinal neurons, which through their opposing actions on vascular development contribute to ROP. These hypoxia-generated mediators include the Krebs cycle intermediate, succinate acting via GPR91, and the neuronal guidance molecule Semaphorin 3A. CONCLUSION: Neuron-derived factors guide retinal vascularization and are major contributors to the pathogenesis of ROP.

The Succinate Receptor SUCNR1 Resides at the Endoplasmic Reticulum and Relocates to the Plasma Membrane in Hypoxic Conditions.

The GPCR SUCNR1/GPR91 exerts proangiogenesis upon stimulation with the Krebs cycle metabolite succinate. GPCR signaling depends on the surrounding environment and intracellular localization through location bias. Here, we show by microscopy and by cell fractionation that in neurons, SUCNR1 resides at the endoplasmic reticulum (ER), while being fully functional, as shown by calcium release and the induction of the expression of the proangiogenic gene for VEGFA. ER localization was found to depend upon N-glycosylation, particularly at position N8; the nonglycosylated mutant receptor localizes at the plasma membrane shuttled by RAB11. This SUCNR1 glycosylation is physiologically regulated, so that during hypoxic conditions, SUCNR1 is deglycosylated and relocates to the plasma membrane. Downstream signal transduction of SUCNR1 was found to activate the prostaglandin synthesis pathway through direct interaction with COX-2 at the ER; pharmacologic antagonism of the PGE2 EP4 receptor (localized at the nucleus) was found to prevent VEGFA expression. Concordantly, restoring the expression of SUCNR1 in the retina of SUCNR1-null mice renormalized vascularization; this effect is markedly diminished after transfection of the plasma membrane-localized SUCNR1 N8A mutant, emphasizing that ER localization of the succinate receptor is necessary for proper vascularization. These findings uncover an unprecedented physiologic process where GPCR resides at the ER for signaling function.

Mesenchymal Stromal Cells Promote Retinal Vascular Repair by Modulating Sema3E and IL-17A in a Model of Ischemic Retinopathy.

Ischemic retinopathies (IRs), such as retinopathy of prematurity and diabetic retinopathy, are characterized by an initial phase of microvascular degeneration that results in retinal ischemia, followed by exaggerated pathologic neovascularization (NV). Mesenchymal stromal cells (MSCs) have potent pro-angiogenic and anti-inflammatory properties associated with tissue repair and regeneration, and in this regard exert protection to neurons in ischemic and degenerative conditions; however, the exact mechanisms underlying these functions remain largely unknown. Class III Semaphorins (A-G) are particularly implicated in regulating neural blood supply (as well as neurogenesis) by suppressing angiogenesis and affecting myeloid cell function; this is the case for distinct neuropillin-activating Sema3A as well as PlexinD1-activating Sema3E; but during IR the former Sema3A increases while Sema3E decreases. We investigated whether retinal vascular repair actions of MSCs are exerted by normalizing Semaphorin and downstream cytokines in IR. Intravitreal administration of MSCs or their secretome (MSCs-conditioned media [MSCs-CM]) significantly curtailed vasoobliteration as well as aberrant preretinal NV in a model of oxygen-induced retinopathy (OIR). The vascular repair effects of MSCs-CM in the ischemic retina were associated with restored levels of Sema3E. Vascular benefits of MSCs-CM were reversed by anti-Sema3E; while intravitreal injection of anti-angiogenic recombinant Sema3E (rSema3E) in OIR-subjected mice reproduced effects of MSCs-CM by inhibiting as expected preretinal NV but also by decreasing vasoobliteration. To explain these opposing vascular effects of Sema3E we found in OIR high retinal levels, respectively, of the pro- and anti-angiogenic IL-17A and Sema3A-regulating IL-1ß; IL-17A positively affected expression of IL-1ß. rSema3E decreased concentrations of these myeloid cell-derived pro-inflammatory cytokines in vitro and in vivo. Importantly, IL-17A suppression by MSCs-CM was abrogated by anti-Sema3E neutralizing antibody. Collectively, our findings provide novel evidence by which MSCs inhibit aberrant NV and diminish vasoobliteration (promoting revascularization) in retinopathy by restoring (at least in part) neuronal Sema3E levels that reduce pathological levels of IL-17A (and in turn other proinflammatory factors) in myeloid cells. The ability of MSCs to generate a microenvironment permissive for vascular regeneration by controlling the production of neuronal factors involved in immunomodulatory activities is a promising opportunity for stem cell therapy in ocular degenerative diseases.

AAV-mediated PEX1 gene augmentation improves visual function in the PEX1-Gly844Asp mouse model for mild Zellweger spectrum disorder.

Patients with Zellweger spectrum disorder (ZSD) commonly present with vision loss due to mutations in PEX genes required for peroxisome assembly and function. Here, we evaluate PEX1 retinal gene augmentation therapy in a mouse model of mild ZSD bearing the murine equivalent (PEX1-p[Gly844Asp]) of the most common human mutation. Experimental adeno-associated virus 8.cytomegalovirus.human PEX1.hemagglutinin (AAV8.CMV.HsPEX1.HA) and control AAV8.CMV.EGFP vectors were administered by subretinal injection in contralateral eyes of early (5-week-old)- or later (9-week-old)-stage retinopathy cohorts. HsPEX1.HA protein was expressed in the retina with no gross histologic side effects. Peroxisomal metabolic functions, assessed by retinal C26:0 lysophosphatidylcholine (lyso-PC) levels, were partially normalized after therapeutic vector treatment. Full-field flash electroretinogram (ffERG) analyses at 8 weeks post-injection showed a 2-fold improved retinal response in the therapeutic relative to control vector-injected eyes. ffERG improved by 1.6- to 2.5-fold in the therapeutic vector-injected eyes when each cohort reached 25 weeks of age. At 32 weeks of age, the average ffERG response was double in the therapeutic relative to control vector-injected eyes in both cohorts. Optomotor reflex analyses trended toward improvement. These proof-of-concept studies represent the first application of gene augmentation therapy to treat peroxisome biogenesis disorders and support the potential for retinal gene delivery to improve vision in these patients.

Tyrosine-Protein Phosphatase Non-receptor Type 9 (PTPN9) Negatively Regulates the Paracrine Vasoprotective Activity of Bone-Marrow Derived Pro-angiogenic Cells: Impact on Vascular Degeneration in Oxygen-Induced Retinopathy.

BACKGROUND AND AIM: Insufficient post-ischemic neovascularization is an initial key step in the pathogenesis of Oxygen-Induced Retinopathy (OIR). During neovascularization, pro-angiogenic cells (PACs) are mobilized from the bone marrow and integrate into ischemic tissues to promote angiogenesis. However, the modulation of PAC paracrine activity during OIR and the specific mechanisms involved remain to be explored. Because Tyrosine-protein phosphatase non-receptor type 9 (PTPN9) is reported to be a negative regulator of stem cell differentiation and angiogenesis signaling, we investigated its effect on PAC activity in the context of OIR. METHODS AND RESULTS: In a rat model of OIR, higher levels of PTPN9 in the retina and in bone marrow derived PACs are associated with retinal avascular areas, lower levels of the mobilization factor SDF-1 and decreased number of CD34+/CD117+/CD133+ PACs. PACs exposed ex vivo to hyperoxia display increased PTPN9 expression, which is associated with impaired ability of PAC secretome to promote angiogenesis ex vivo (choroidal vascular sprouting) and in vitro (endothelial cell tubule formation) compared to the secretome of PACs maintained in normoxia. Suppression of PTPN9 (using siRNA) increases VEGF and SDF-1 expression to normalize PAC secretome during hyperoxia, leading to restored angiogenic ability of PAC secretome. Moreover, endothelial cells exposed to the secretome of siPTPN9-treated PACs expressed increased levels of activated form of VEGF receptor 2 (VEGFR2). In the rat model of OIR, intravitreal injection of secretome from siPTPN9-treated PACs significantly reduced retinal vaso-obliteration; this was associated with higher retinal levels of VEGF/SDF-1, and increased recruitment of PACs (CD34+ cells) to the retinal and choroidal vessels. CONCLUSION: Our results suggest that hyperoxia alters the paracrine proangiogenic activity of BM-PACs by inducing PTPN9, which can contribute to impair post-ischemic revascularization in the context of OIR. Targeting PTPN9 restores PAC angiogenic properties, and provide a new target for vessel integrity in ischemic retinopathies.

MicroRNA-96 Promotes Vascular Repair in Oxygen-Induced Retinopathy-A Novel Uncovered Vasoprotective Function.

BACKGROUND AND AIMS: Vascular degeneration is a hallmark in the pathogenesis of oxygen-induced retinopathy (OIR). Dysregulation of microRNAs (miRNAs), key regulators of genes expressions, has been implicated in the regulation of ocular angiogenesis. However, miRNAs specific functions in impaired vascular development during OIR are poorly understood. Herein, we identified miR-96 as one of the most highly expressed miRNAs in the retina and choroid during vascular development and investigated the potential role of miR-96 on microvascular degeneration in a rat OIR model. METHODS AND RESULTS: Next generation sequencing (NGS) and qRT-PCR analysis showed that miR-96 maintain high levels of expression during ocular vascular development. Nevertheless, miR-96 was significantly downregulated in the retina and choroid of OIR rats (80% O2 from P5 to P10) during the phase of microvascular degeneration. Similarly, human retinal microvascular endothelial cells (HRMEC) subjected to hyperoxia (80% O2) showed a significant downregulation of miR-96 evaluated by qPCR. Interestingly, HRMEC supplemented with miR-96 regulated positively the expression of several key angiogenic factors including VEGF and ANG-2. To explore the angiogenic activity of miR-96 on HRMEC, we performed a gain/loss of function study. In a similar way to hyperoxia exposure, we observed a robust angiogenic impairment (tubulogenesis and migration) on HRMEC transfected with an antagomiR-96. Conversely, overexpression of miR-96 stimulated the angiogenic activity of HRMEC and protected against hyperoxia-induced endothelial dysfunction. Finally, we evaluated the potential vasoprotective function of miR-96 in OIR animals. Rat pups intravitreally supplemented with miR-96 mimic (1 mg/kg) displayed a significant preservation of retinal/choroidal microvessels at P10 compared to controls. This result was consistent with the maintenance of physiologic levels of VEGF and ANG-2 in the OIR retina. CONCLUSION: This study demonstrates that miR-96 regulates the expression of angiogenic factors (VEGF/ANG-2) associated to the maintenance of retinal and choroidal microvasculature during physiological and pathological conditions. Intravitreal supplementation of miR-96 mimic could constitute a novel therapeutic strategy to improve vascular repair in OIR and other ischemic retinopathies.

Novel Anti-Interleukin-1ß Therapy Preserves Retinal Integrity: A Longitudinal Investigation Using OCT Imaging and Automated Retinal Segmentation in Small Rodents.

Retinopathy of prematurity (ROP) is the leading cause of blindness in neonates. Inflammation, in particular interleukin-1ß (IL-1ß), is increased in early stages of the disorder, and contributes to inner and outer retinal vasoobliteration in the oxygen-induced retinopathy (OIR) model of ROP. A small peptide antagonist of IL-1 receptor, composed of the amino acid sequence, rytvela, has been shown to exert beneficial anti-inflammatory effects without compromising immunovigilance-related NF-κB in reproductive tissues. We conducted a longitudinal study to determine the efficacy of "rytvela" in preserving the integrity of the retina in OIR model, using optical coherence tomography (OCT) which provides high-resolution cross-sectional imaging of ocular structures in vivo. Sprague-Dawley rats subjected to OIR and treated or not with "rytvela" were compared to IL-1 receptor antagonist (Kineret). OCT imaging and custom automated segmentation algorithm used to measure retinal thickness (RT) were obtained at P14 and P30; gold-standard immunohistochemistry (IHC) was used to confirm retinal anatomical changes. OCT revealed significant retinal thinning in untreated animals by P30, confirmed by IHC; these changes were coherently associated with increased apoptosis. Both rytvela and Kineret subsided apoptosis and preserved RT. As anticipated, Kineret diminished both SAPK/JNK and NF-κB axes, whereas rytvela selectively abated the former which resulted in preserved monocyte phagocytic function. Altogether, OCT imaging with automated segmentation is a reliable non-invasive approach to study longitudinally retinal pathology in small animal models of retinopathy.

MicroRNA expression profile in retina and choroid in oxygen-induced retinopathy model.

BACKGROUND: Ischemic retinopathies (IRs) are leading causes of visual impairment. They are characterized by an initial phase of microvascular degeneration and a second phase of aberrant pre-retinal neovascularization (NV). microRNAs (miRNAs) regulate gene expression, and a number play a role in normal and pathological NV. But, post-transcriptional modulation of miRNAs in the eye during the development of IRs has not been systematically evaluated. AIMS & METHODS: Using Next Generation Sequencing (NGS) we profiled miRNA expression in the retina and choroid during vasodegenerative and NV phases of oxygen-induced retinopathy (OIR). RESULTS: Approximately 20% of total miRNAs exhibited altered expression (up- or down-regulation); 6% of miRNA were found highly expressed in retina and choroid of rats subjected to OIR. During OIR-induced vessel degeneration phase, miR-199a-3p, -199a-5p, -1b, -126a-3p displayed a robust decreased expression (> 85%) in the retina. While in the choroid, miR-152-3p, -142-3p, -148a-3p, -532-3p were upregulated (>200%) and miR-96-5p, -124-3p, -9a-3p, -190b-5p, -181a-1-3p, -9a-5p, -183-5p were downregulated (>70%) compared to controls. During peak pathological NV, miR-30a-5p, -30e-5p and 190b-5p were markedly reduced (>70%), and miR-30e-3p, miR-335, -30b-5p strongly augmented (by up to 300%) in the retina. Whereas in choroid, miR-let-7f-5p, miR-126a-5p and miR-101a-3p were downregulated by (>81%), and miR-125a-5p, let-7e-5p and let-7g-5p were upregulated by (>570%) during NV. Changes in miRNA observed using NGS were validated using qRT-PCR for the 24 most modulated miRNAs. In silico approach to predict miRNA target genes (using algorithms of miRSystem database) identified potential new target genes with pro-inflammatory, apoptotic and angiogenic properties. CONCLUSION: The present study is the first comprehensive description of retinal/choroidal miRNAs profiling in OIR (using NGS technology). Our results provide a valuable framework for the characterization and possible therapeutic potential of specific miRNAs involved in ocular IR-triggered inflammation, angiogenesis and degeneration.

Propranolol Attenuates Proangiogenic Activity of Mononuclear Phagocytes: Implication in Choroidal Neovascularization.

Purpose: Targeting ß-adrenergic receptor signaling with propranolol has emerged as a potential candidate to counteract choroidal neovascularization (CNV). Little is known of its effect on macrophages, which play a critical role in CNV. We investigated the effect of propranolol on angiogenic response of mononuclear phagocytes (MPs). Methods: The angiogenic effect of propranolol was evaluated in laser-induced CNV model. Mice received intraperitoneal injections of propranolol (6 mg/kg/d) or vehicle. CNV area and inflammatory cells were determined respectively by using lectin staining and an anti-IBA-1 antibody on RPE/choroid flat mounts. Inflammatory gene expression was evaluated by quantitative (q) PCR analysis. Mechanisms of propranolol was studied in MP cell lines J774 and RAW264.7 and in primary peritoneal macrophages. Expression of pro- and antiangiogenic mediators was studied. In addition, effects of propranolol treatment of MPs was assessed on choroidal explant. Results: CNV was attenuated by propranolol and concomitantly associated with decreased inflammatory mediators IL-6 and TNFα, albeit with accumulation of (ß-adrenoceptor harboring) MPs in the CNV area. Conditioned media from MPs preincubated with propranolol exerted antiangiogenic effects. Treatment of J774 confirmed the attenuation of inflammatory response to propranolol and increased cleaved caspase-3 on choroidal explant. We found that propranolol increased pigment epithelium-derived factor (PEDF) expression in MPs. Trapping of PEDF with an antibody abrogated antiangiogenic effects of propranolol. PEDF was also detected in CNV-associated MPs. Conclusions: We hereby show that propranolol confers on MPs antiangiogenic properties by increasing PEDF expression, which complements its effects on vascular tissue resulting in inhibition of choroidal vasoproliferation in inflammatory conditions. The study supports possible use of propranolol as a therapeutic modality for CNV.

Immunometabolic modulation of retinal inflammation by CD36 ligand.

In subretinal inflammation, activated mononuclear phagocytes (MP) play a key role in the progression of retinopathies. Little is known about the mechanism involved in the loss of photoreceptors leading to vision impairment. Studying retinal damage induced by photo-oxidative stress, we observed that cluster of differentiation 36 (CD36)-deficient mice featured less subretinal MP accumulation and attenuated photoreceptor degeneration. Moreover, treatment with a CD36-selective azapeptide ligand (MPE-001) reduced subretinal activated MP accumulation in wild type mice and preserved photoreceptor layers and function as assessed by electroretinography in a CD36-dependent manner. The azapeptide modulated the transcriptome of subretinal activated MP by reducing pro-inflammatory markers. In isolated MP, MPE-001 induced dissociation of the CD36-Toll-like receptor 2 (TLR2) oligomeric complex, decreasing nuclear factor-kappa B (NF-κB) and NLR family pyrin domain containing 3 (NLRP3) inflammasome activation. In addition, MPE-001 caused an aerobic metabolic shift in activated MP, involving peroxisome proliferator-activated receptor-γ (PPAR-γ) activation, which in turn mitigated inflammation. Accordingly, PPAR-γ inhibition blocked the cytoprotective effect of MPE-001 on photoreceptor apoptosis elicited by activated MP. By altering activated MP metabolism, MPE-001 decreased immune responses to alleviate subsequent inflammation-dependent neuronal injury characteristic of various vision-threatening retinal disorders.

Azasulfurylpeptide Modulation of CD36-Mediated Inflammation Without Effect on Neovascularization.

Modulation of the cluster of differentiation-36 receptor (CD36) has proven promising for dampening pro-inflammatory macrophage signaling. For example, azapeptides (e.g., 1 and 2) bind CD36 selectively with high affinity, mitigate Toll-like receptor (TLR) agonist-induced overproduction of nitric oxide (NO), and reduce pro-inflammatory cytokine and chemokine production in macrophages. Moreover, semicarbazides 1 and 2 inhibit microvascular sprouting mediated through CD36 in the choroid explant. Seeking a selective CD36 modulator that mediated inflammation without influencing neovascularization, a set of azasulfurylpeptides (e.g., 3a⁻e) were synthesized in which the semicarbazide was replaced by an N-aminosulfamide residue using a novel solid-phase approach. Notably, azasulfurylpeptide 3c diminished selectively CD36-mediated TLR-2-triggered inflammatory response without affecting neovascularization. Subtle chemical modification at the peptide backbone from a carbonyl to a sulfuryl residue has had a selective effect on biological activity providing a valuable probe for studying CD36 chemical biology.

Mechanisms of macular edema: Beyond the surface.

Macular edema consists of intra- or subretinal fluid accumulation in the macular region. It occurs during the course of numerous retinal disorders and can cause severe impairment of central vision. Major causes of macular edema include diabetes, branch and central retinal vein occlusion, choroidal neovascularization, posterior uveitis, postoperative inflammation and central serous chorioretinopathy. The healthy retina is maintained in a relatively dehydrated, transparent state compatible with optimal light transmission by multiple active and passive systems. Fluid accumulation results from an imbalance between processes governing fluid entry and exit, and is driven by Starling equation when inner or outer blood-retinal barriers are disrupted. The multiple and intricate mechanisms involved in retinal hydro-ionic homeostasis, their molecular and cellular basis, and how their deregulation lead to retinal edema, are addressed in this review. Analyzing the distribution of junction proteins and water channels in the human macula, several hypotheses are raised to explain why edema forms specifically in the macular region. "Pure" clinical phenotypes of macular edema, that result presumably from a single causative mechanism, are detailed. Finally, diabetic macular edema is investigated, as a complex multifactorial pathogenic example. This comprehensive review on the current understanding of macular edema and its mechanisms opens perspectives to identify new preventive and therapeutic strategies for this sight-threatening condition.

Ischemic Retinopathies: Oxidative Stress and Inflammation.

Ischemic retinopathies (IRs), such as retinopathy of prematurity (ROP), diabetic retinopathy (DR), and (in many cases) age-related macular degeneration (AMD), are ocular disorders characterized by an initial phase of microvascular changes that results in ischemia, followed by a second phase of abnormal neovascularization that may culminate into retinal detachment and blindness. IRs are complex retinal conditions in which several factors play a key role during the development of the different pathological stages of the disease. Increasing evidence reveals that oxidative stress and inflammatory processes are important contributors to the pathogenesis of IRs. Despite the beneficial effects of the photocoagulation and anti-VEGF therapy during neovascularization phase, the need to identify novel targets to prevent initial phases of these ocular pathologies is still needed. In this review, we provide an update on the involvement of oxidative stress and inflammation in the progression of IRs and address some therapeutic interventions by using antioxidants and anti-inflammatory agents.

Influences of Histidine-1 and Azaphenylalanine-4 on the Affinity, Anti-inflammatory, and Antiangiogenic Activities of Azapeptide Cluster of Differentiation 36 Receptor Modulators.

Azapeptide analogues of growth hormone releasing peptide-6 (GHRP-6) exhibit promising affinity, selectivity, and modulator activity on the cluster of differentiation 36 receptor (CD36). For example, [A1, azaF4]- and [azaY4]-GHRP-6 (1a and 2b) were previously shown to bind selectively to CD36 and exhibited respectively significant antiangiogenic and slight angiogenic activities in a microvascular sprouting assay using choroid explants. The influences of the 1- and 4-position residues on the affinity, anti-inflammatory, and antiangiogenic activity of these azapeptides have now been studied in detail by the synthesis and analysis of a set of 25 analogues featuring Ala1 or His1 and a variety of aromatic side chains at the aza-amino acid residue in the 4-position. Although their binding affinities differed only by a factor of 17, the analogues exhibited significant differences in ability to modulate production of nitric oxide (NO) in macrophages and choroidal neovascularization.

Nuclear localization of platelet-activating factor receptor controls retinal neovascularization.

Platelet-activating factor (PAF) is a pleiotropic phospholipid with proinflammatory, procoagulant and angiogenic actions on the vasculature. We and others have reported the presence of PAF receptor (Ptafr) at intracellular sites such as the nucleus. However, mechanisms of localization and physiologic functions of intracellular Ptafr remain poorly understood. We hereby identify the importance of C-terminal motif of the receptor and uncover novel roles of Rab11a GTPase and importin-5 in nuclear translocation of Ptafr in primary human retinal microvascular endothelial cells. Nuclear localization of Ptafr is independent of exogenous PAF stimulation as well as intracellular PAF biosynthesis. Moreover, nuclear Ptafr is responsible for the upregulation of unique set of growth factors, including vascular endothelial growth factor, in vitro and ex vivo. We further corroborate the intracrine PAF signaling, resulting in angiogenesis in vivo, using Ptafr antagonists with distinct plasma membrane permeability. Collectively, our findings show that nuclear Ptafr translocates in an agonist-independent manner, and distinctive functions of Ptafr based on its cellular localization point to another dimension needed for pharmacologic selectivity of drugs.

Erratum: Nuclear localization of platelet-activating factor receptor controls retinal neovascularization.

[This corrects the article DOI: 10.1038/celldisc.2016.17.].