Choroidal Neovascularization Formation Research Articles

Morphofunctional Distinction of Retinal Pigment Epithelium in Normal and Age-related Macular Degeneration. Review of the Literature

The retinal pigment epithelium performs a number of highly specialized functions. The relevance of studying morphological changes in the pigment epithelium is associated with an increase in the number of patients with age-related macular degeneration (AMD), polypoid choroidal vasculopathy (PCV), central serous retinopathy and retinitis pigmentosa. The increase in the number of patients with these pathologies is associated both with the general trend of aging of the world’s population and with modern opportunities for mass diagnostics. The pigment epithelium performs a number of functions that ensure the normal functioning of the retina. The main structural unit is a hexagonal epithelial cell with a large number of melanosomes containing the pigment melanin. Pathomorphological changes in the pigment leaf are an important indicator of the development of AMD. In most cases, an elevation or detachment is formed, in which three types are distinguished: druzenoid, fibrovascular (hemorrhagic) and serous. According to a prospective multicenter study, in 19 % of cases, the druzenoid form progresses into atrophy of the pigment epithelium. There are a number of studies that present harbingers of integrity damages. Fibrovascular detachment is the pathomorphological basis for the formation of submacular hemorrhage. The pathogenesis of serous exfoliation in AMD is associated with a range of different mechanisms, ranging from degenerative changes in the Bruch membrane to the formation of choroidal neovascularization. In the scientific publications of a number of authors, information is presented about the complication of detachment in the form of the development of a rupture of the retinal pigment epithelium. The key criterion for a threatening rupture is the height of the pigment leaf detachment. Damage of the integrity of the pigment leaf in the long term is a factor in reducing or complete loss of vision. This is especially characteristic when a gap is formed in the central area of the retina. Also, visual impairment is observed in cases of submacular hemorrhage and the formation of a fibrous scar. An important point, at present, is the development of protocols for the management of patients with ruptures on the background of the underlying disease.

Diagnostic Challenges in Inflammatory Choroidal Neovascularization.

Inflammation plays a key role in the induction of choroidal neovascularization (CNV). Inflammatory choroidal neovascularization (iCNV) is a severe but uncommon complication of both infectious and non-infectious uveitides. It is hypothesized that its pathogenesis is similar to that of wet age-related macular degeneration (AMD), and involves hypoxia as well as the release of vascular endothelial growth factor, stromal cell-derived factor 1-alpha, and other mediators. Inflammatory CNV develops when inflammation or infection directly involves the retinal pigment epithelium (RPE)-Bruch's membrane complex. Inflammation itself can compromise perfusion, generating a gradient of retinal-choroidal hypoxia that additionally promotes the formation of choroidal neovascularization in the course of uveitis. The development of choroidal neovascularization may be a complication, especially in conditions such as punctate inner choroidopathy, multifocal choroiditis, serpiginous choroiditis, and presumed ocular histoplasmosis syndrome. Although the majority of iCNV cases are well defined and appear as the "classic" type (type 2 lesion) on fluorescein angiography, the diagnosis of iCNV is challenging due to difficulties in differentiating between inflammatory choroiditis lesions and choroidal neovascularization. Modern multimodal imaging, particularly the recently introduced technology of optical coherence tomography (OCT) and OCT angiography (noninvasive and rapid imaging modalities), can reveal additional features that aid the diagnosis of iCNV. However, more studies are needed to establish their role in the diagnosis and evaluation of iCNV activity.

Tocilizumab attenuates choroidal neovascularization by regulating macrophage polarization through the IL-6R/STAT3/VEGF pathway

Globally, age-related macular degeneration (AMD) is the leading cause of irreversible visual impairment. Up to 80% of severe vision loss is caused by AMD, which is characterized by the development of choroidal neovascularization (CNV). Uncertainty exists regarding the precise pathophysiological mechanisms of CNV. It has been suggested that the interleukin (IL) IL-6/IL-6R signaling pathway is crucial in the progression of CNV. Tocilizumab (TCZ), a monoclonal antibody, binds to soluble and membrane-bound IL-6R and competitively inhibits IL-6 downstream signaling. Previous research has demonstrated that TCZ promotes several roles related to inflammation and neovascularization. However, the effects of TCZ on CNV and the underlying mechanism are still unknown. This study found that TCZ administration decreased the area and leakage of CNV lesions in the mice model of laser-induced CNV. Additionally, results demonstrated that TCZ promotes the expression of iNOS, CCL-3, CCL-5, TNF-α and inhibits the expression of Arg-1, IL-10, YM-1 and CD206. Furthermore, TCZ treatment inhibited the signal transducer and activator of transcription (STAT) STAT3/vascular endothelial growth factor (VEGF) pathway, which was activated after CNV formation. Colivelin, a STAT3 agonist, reversed the inhibitory effects of TCZ on CNV formation and macrophage polarization. In a mouse model of laser-induced CNV, our findings demonstrated that TCZ attenuated CNV formation and inhibited the leakage of CNV lesions by regulating macrophage polarization via inhibiting the STAT3/VEGF axis. TCZ is the potential therapeutic strategy for CNV.

Treatment Strategies for Anti-VEGF Resistance in Neovascular Age-Related Macular Degeneration by Targeting Arteriolar Choroidal Neovascularization.

Despite extensive use of intravitreal anti-vascular endothelial growth factor (anti-VEGF) biologics for over a decade, neovascular age-related macular degeneration (nAMD) or choroidal neovascularization (CNV) continues to be a major cause of irreversible vision loss in developed countries. Many nAMD patients demonstrate persistent disease activity or experience declining responses over time despite anti-VEGF treatment. The underlying mechanisms of anti-VEGF resistance are poorly understood, and no effective treatment strategies are available to date. Here we review evidence from animal models and clinical studies that supports the roles of neovascular remodeling and arteriolar CNV formation in anti-VEGF resistance. Cholesterol dysregulation, inflammation, and ensuing macrophage activation are critically involved in arteriolar CNV formation and anti-VEGF resistance. Combination therapy by neutralizing VEGF and enhancing cholesterol removal from macrophages is a promising strategy to combat anti-VEGF resistance in CNV.

Topical Ophthalmic Liposomes Dual-Modified with Penetratin and Hyaluronic Acid for the Noninvasive Treatment of Neovascular Age-Related Macular Degeneration.

Since intrinsic ocular barrier limits the intraocular penetration of therapeutic protein through eye drops, repeated intravitreal injections of anti-vascular endothelial growth factor (anti-VEGF) agents are the standard therapy for neovascular age-related macular degeneration (nAMD), which are highly invasive and may cause particular ocular complications, leading to poor patient compliance. Using Penetratin (Pen) as the ocular penetration enhancer and hyaluronic acid (HA) as the retina-targeting ligand, a dual-modified ophthalmic liposome (Penetratin hyaluronic acid-liposome/Conbercept, PenHA-Lip/Conb) eye drop was designed to non-invasively penetrate the ocular barrier and deliver anti-VEGF therapeutic agents to the targeted intraocular tissue. PenHA-Lip effectively penetrates the ocular barrier and targets the retinal pigment epithelium via corneal and non-corneal pathways. After a single topical administration of conbercept-loaded PenHA-Lip (PenHA-Lip/Conb), the intraocular concentration of conbercept peaked at 18.74 ± 1.09 ng/mL at 4 h, which is 11.55-fold higher than unmodified conbercept. In a laser-induced choroidal neovascularization (CNV) mouse model, PenHA-Lip/Conb eye drops three times daily for seven days inhibited CNV formation and progression without any significant tissue toxicity and achieved an equivalent effect to a single intravitreal conbercept injection. PenHA-Lip efficiently and safely delivered conbercept to the posterior eye segment and may be a promising noninvasive therapeutic option for nAMD.

Splenic monocytes drive pathogenic subretinal inflammation in age-related macular degeneration

Age-related macular degeneration (AMD) is invariably associated with the chronic accumulation of activated mononuclear phagocytes in the subretinal space. The mononuclear phagocytes are composed of microglial cells but also of monocyte-derived cells, which promote photoreceptor degeneration and choroidal neovascularization. Infiltrating blood monocytes can originate directly from bone marrow, but also from a splenic reservoir, where bone marrow monocytes develop into angiotensin II receptor (ATR1)+ splenic monocytes. The involvement of splenic monocytes in neurodegenerative diseases such as AMD is not well understood. Using acute inflammatory and well-phenotyped AMD models, we demonstrate that angiotensin II mobilizes ATR1+ splenic monocytes, which we show are defined by a transcriptional signature using single-cell RNA sequencing and differ functionally from bone marrow monocytes. Splenic monocytes participate in the chorio-retinal infiltration and their inhibition by ATR1 antagonist and splenectomy reduces the subretinal mononuclear phagocyte accumulation and pathological choroidal neovascularization formation. In aged AMD-risk ApoE2-expressing mice, a chronic AMD model, ATR1 antagonist and splenectomy also inhibit the chronic retinal inflammation and associated cone degeneration that characterizes these mice. Our observation of elevated levels of plasma angiotensin II in AMD patients, suggests that similar events take place in clinical disease and argue for the therapeutic potential of ATR1 antagonists to inhibit splenic monocytes for the treatment of blinding AMD.

Intravenous 3K3A-Activated Protein C Inhibits Murine Ocular Inflammation and Suppresses Ocular Choroidal Neovascularization

Introduction: Choroidal neovascularization (CNV) is characterized by the growth of newly formed abnormal blood vessels from the choroid extending into the neurosensory retina, leading to vision loss. CNV is a common cause of visual impairment in neovascular age-related macular degeneration (nAMD) patients. Current treatments failed to stop the continuity of the neurodegenerative process. Despite the urgency, no neuroprotective agent has been found yet to treat AMD and other neurodegenerative ocular diseases. 3K3A-activated protein C (APC) is a cell-signaling analog of the endogenous blood serine protease APC. Preclinical studies showed that circulating plasma 3K3A-APC readily crosses the blood-brain barrier (BBB) and reaches neurons due to EPCR-dependent transport. In rodent models of stroke, brain injury, and neurodegenerative disorders, 3K3A-APC exerts neuroprotective, anti-inflammatory, and vasculoprotective activities. Due to its pleiotropic effects, 3K3A-APC has advanced to clinical trial stages for treating acute brain damage and chronic neurodegenerative diseases, which share similarities with retinal neurodegenerative disorders. Notably, the retina is part of the central nervous system (CNS), and the blood-retina barrier shares functional and structural similarities with the BBB. We previously demonstrated that intraocular administration of 3K3A-APC functions as a pleiotropic cytoprotective molecule within the retina, reducing ocular inflammation, suppressing CNV growth, and exhibiting promising therapeutic potential as a neuroprotective agent for treating ocular disorders. Intraocular administration is commonly utilized for delivering therapeutics to treat retinal conditions. However, safety concerns, the burden on the healthcare system, and patient comfort may indicate that systemic administration of 3K3A-APC is a beneficial alternative. Aim: To determine whether intravenously injecting 3K3A-APC into mice tails will cross the blood-retina barrier, induce anti-inflammatory activities in the retina, and suppress CNV. Methods: The efficacy of systemic administration of 3K3A-APC was evaluated using two C57BL/6J murine models: the LPS-induced ocular inflammation and the laser-induced CNV model. CNV was triggered by laser and ocular inflammation was triggered by intravitreal injection of lipopolysaccharide (LPS). Murine recombinant 3K3A-APC was administered systemically via tail injection. Fluorescein isothiocyanate (FITC)-dextran perfusion followed by retinal flatmount staining with anti-CD11b was used to assess inflammatory co-localization of cells to retinal vessels. FITC-dextran perfusion followed by retinal pigment epithelium (RPE)-choroid flatmount was used to assess CNV vascular penetration using 3D confocal imaging. Results: LPS injection triggered robust inflammatory cell infiltration in both the anterior and posterior chambers of the eyes. Systemic administration of 3K3A-APC (0.5 mg/kg, 1 and 4 hours post-LPS) significantly inhibited ocular inflammation. Immunostaining performed 24 hours post LPS revealed that 3K3A-APC reduced the number of CD11b+ monocyte and their extravasation into the retinal parenchyma (Figure). In the laser-induced CNV model, systemic administration of 0.2 mg/kg 3K3A-APC was applied one hour and four days post-laser. Evaluation of retinal specimens ten days post-laser showed that 3K3A-APC treatment significantly suppressed CNV growth, attenuated the abnormal growth of blood vessels from the choroid into the sensory retina, and decreased myeloid cell recruitment to the RPE-choroid area. Conclusion: Our study reveals that systemically administered 3K3A-APC effectively inhibits ocular inflammation and reduces CNV formation. Drawing insights from the neuroprotective activities of 3K3A-APC in the brain and CNS and recognizing the retina, particularly the macula, as an extension of the brain, we propose 3K3A-APC as a potential neuroprotective treatment for AMD and other neurodegenerative retinal pathologies. Given the established safety of systemic 3K3A-APC administration in phase 2 clinical trials for ischemic stroke, our findings support further exploring 3K3A-APC as a novel therapeutic approach in ophthalmology. Further research is warranted to elucidate fully the mechanisms underlying 3K3A-APC's beneficial effects on the retina.

5α-Hydroxycostic acid inhibits choroidal neovascularization in rats through a dual signalling pathway mediated by VEGF and angiopoietin 2

Background5α-Hydroxycostic acid is a eudemane sesquiterpene that is isolated from the natural plant, Laggera alata. It exerts anti-inflammatory and anti-angiogenic effects on human breast cancer cells, but its role and underlying mechanism in choroidal neovascularization (CNV) are still unclear. We conducted a study to verify that 5α-Hydroxycostic acid can inhibit the formation and leakage of CNV, and describe the possible dual pathway by which it exerts its inhibitory effects in this process.MethodsAn in vitro model of choroidal neovascularization was established using VEGF164, while a rat model of choroidal neovascularization was established using a 532 nm laser. In both models, the effects of 5α-Hydroxycostic acid in vivo and in vitro were evaluated to determine its inhibitory effect on abnormal cell proliferation, migration and tubule formation, as well as its effect on pathological changes in choroidal tissues and the area of neovascularization leakage in rats. The levels of components in the VEGF/VEGFR and Ang2/Tie2 signaling pathways were measured in tissues and cells.ResultsIn vitro experiments have shown that 5α-Hydroxycostic acid can inhibit abnormal cell proliferation, migration and angiogenesis. Additionally, 5α-Hydroxycostic acid enhances cell adhesion by inhibiting the phosphorylation pathways of VEGFR2 and Tie2. In vivo experiments demonstrated that 5α-Hydroxycostic acid has a positive therapeutic effect on choroidal neovascularization in rats. It can effectively reduce vascular leakage, consistent with the results of the cell experiments.Conclusion5α-Hydroxycostic acid can inhibit choroidal neovascularization by interfering with the VEGF- and Ang2/Tie2-related pathways, and it may be a good candidate drug for treating CNV.

NR4A1 deletion promotes pro-angiogenic polarization of macrophages derived from classical monocytes in a mouse model of neovascular age-related macular degeneration

BackgroundNeovascular age-related macular degeneration causes vision loss from destructive angiogenesis, termed choroidal neovascularization (CNV). Cx3cr1−/− mice display alterations in non-classical monocytes and microglia with increased CNV size, suggesting that non-classical monocytes may inhibit CNV formation. NR4A1 is a transcription factor that is necessary for maturation of non-classical monocytes from classical monocytes. While Nr4a1−/− mice are deficient in non-classical monocytes, results are confounded by macrophage hyper-activation. Nr4a1se2/se2 mice lack a transcriptional activator, resulting in non-classical monocyte loss without macrophage hyper-activation.Main bodyWe subjected Nr4a1−/− and Nr4a1se2/se2 mice to the laser-induced CNV model and performed multi-parameter flow cytometry. We found that both models lack non-classical monocytes, but only Nr4a1−/− mice displayed increased CNV area. Additionally, CD11c+ macrophages were increased in Nr4a1−/− mice. Single-cell transcriptomic analysis uncovered that CD11c+ macrophages were enriched from Nr4a1−/− mice and expressed a pro-angiogenic transcriptomic profile that was disparate from prior reports of macrophage hyper-activation.ConclusionsThese results suggest that non-classical monocytes are dispensable during CNV, and NR4A1 deficiency results in increased recruitment of pro-angiogenic macrophages.

Dynamic changes in macrophage morphology during the progression of choroidal neovascularization in a laser-induced choroidal neovascularization mouse model

BackgroundNeovascular age-related macular degeneration (AMD) is responsible for the majority of severe vision loss cases and is mainly caused by choroidal neovascularization (CNV). This condition persists or recurs in a subset of patients and regresses after 5 or more years of anti-vascular endothelial growth factor (VEGF) treatment. The precise mechanisms of CNV continue to be elucidated. According to our previous studies, macrophages play a critical role in CNV. Herein, we aimed to determine the morphological changes in macrophages in CNV to help us understand the dynamic changes.MethodsMice were subjected to laser injury to induce CNV, and lesion expansion and macrophage transformation were examined by immunofluorescence and confocal analysis. Several strategies were used to verify the dynamic changes in macrophages. Immunofluorescence and confocal assays were performed on choroidal flat mounts to evaluate the morphology and phenotype of macrophages in different CNV phases, and the results were further verified by western blotting and RT–PCR.ResultsThe location of infiltrated macrophages changed after laser injury in the CNV mouse model, and macrophage morphology also dynamically changed. Branching macrophages gradually shifted to become round with the progression of CNV, which was certified to be an M2 phenotypic shift.ConclusionsDynamic changes in macrophage morphology were observed during CNV formation, and the round-shaped M2 phenotype could promote neovascularization. In general, the changes in morphology we observed in this study can help us to understand the critical role of macrophages in CNV progression and exploit a potential treatment option for CNV indicated by a shift in macrophage polarity.

Honokiol Attenuates Choroidal Neovascularization by Inhibiting the Hypoxia-Inducible Factor-α/Vascular Endothelial Growth Factor Axis via Nuclear Transcription Factor-Kappa B Activation

Purpose Honokiol is a lignan isolated from Magnolia officinalis and exhibits anti-angiogenic properties. This study was conducted to investigate the role of honokiol in choroidal neovascularization. Methods C57BL/6 mice were treated with honokiol at 10–20 mg/kg by daily intraperitoneal injection from day 1 to 6 after laser photocoagulation. ARPE-19 cells were cultured under hypoxic conditions with or without the presence of honokiol. After laser photocoagulation and honokiol treatment, hematoxylin and eosin staining, immunofluorescence and fundus fluorescein angiography were used to analyze the effect of honokiol on choroidal neovascularization formation. Quantitative real-time PCR, western blot, enzyme-linked immunosorbent assay, immunofluorescence, luciferase assay, and chromatin immunoprecipitation were performed to explore the mechanism of honokiol in the pathological process of choroidal neovascularization. Finally, the role of honokiol on the human choroidal vascular endothelial cells was detected by using 5-ethynyl-20-deoxyuridine assay, Transwell and Tube formation assays. Results The results of hematoxylin and eosin staining and immunofluorescence suggested that honokiol reduced the thickness, length, and area of choroidal neovascularization lesions in laser-induced choroidal neovascularization mouse model. Fundus fluorescein angiography showed that choroidal neovascularization leakage was reduced in honokiol group and the concentration of 20 mg/kg showed better effects. Mechanism studies have shown that honokiol exerted inhibitory effects on choroidal neovascularization by inactivating hypoxia-inducible factor-1α/vascular endothelial growth factor axis through the nuclear transcription factor-kappa B signaling pathway. The same results were obtained in ARPE-19 cells under hypoxic conditions. Furthermore, the conditional medium of retinal pigmented epithelial cells promoted the proliferation, migration, and tube formation of human choroidal vascular endothelial cells, while honokiol reversed these. Conclusion We demonstrated that honokiol attenuated choroidal neovascularization formation by inactivating the hypoxia-inducible factor-1α/vascular endothelial growth factor axis through nuclear transcription factor-kappa B signaling pathway.

Neutrophil extracellular traps boost laser-induced mouse choroidal neovascularization through the activation of the choroidal endothelial cell TLR4/HIF-1α pathway.

Choroidal neovascularization (CNV) is characterized by the infiltration of immune cells, particularly neutrophils. Neutrophil extracellular trap (NET) facilitates the angiogenesis of pulmonary endothelial cells via activating Toll-like receptor 4 (TLR4). TLR4 promotes the expression of transcription factor hypoxia inducible factor-1α (HIF-1α), which promotes inflammation and angiogenesis via the up-regulation of metalloproteinase-9 (MMP-9) and interleukin-1β (IL-1β). In the present study, we aimed to identify the formation of NET and its role in CNV. Our results showed that NET levels were increased in a mouse laser-induced CNV model via oxidative stress, whereas the inhibition of NET alleviated CNV. In vitro, NET activated the TLR4/HIF-1α pathway in human choroidal endothelial cells (HCECs). Additionally, NET increased the transcription and expression of MMP-9 and IL-1β in HCECs via activating the TLR4/HIF-1α pathway. Meanwhile, NET promoted the inflammatory response accompanied by the proliferation, migration and tube formation of HCECs in a MMP-9- and IL-1β-dependent manner. In conclusion, NET was up-regulated in CNV and promoted the formation of CNV via activating the TLR4/HIF-1α pathway in choroidal endothelial cells. Our data uncovered the novel role of NET in promoting the formation of CNV. The underlying mechanism of NET could be targeted to delay the process of CNV.

High-Fat Diet Alters Acylcarnitine Metabolism of the Retina and Retinal Pigment Epithelium/Choroidal Tissues in Laser-Induced Choroidal Neovascularization Rat Models.

Choroidal neovascularization (CNV) is age-related macular degeneration's (AMD) main pathological change. High-fat diet (HFD) is associated with a form of CNV; however, the specific mechanism is unclear. Mitochondrial dysfunction, characterized by abnormal acylcarnitine, occurs during metabolic screening of serum or other body tissues in AMD. This study investigates HFD's role in retinal and retinal pigment epithelium (RPE)/choroidal acylcarnitine metabolism in CNV formation. Chow diet and HFD-BN rats are laser-treated to induce CNV. Acylcarnitine species are quantitatively characterized by ultrahigh-performance liquid chromatography-tandem mass spectrometry. Optical coherence tomography and fundus fluorescein angiography evaluate CNV severity. HFD promotes weight gain, dyslipidemia, and CNV formation. In CNV rats, few medium-chain fatty acids (MCFAs) acylcarnitine in the RPE/choroid are initially affected. When an HFD is administered to these, even MCFA acylcarnitine in the RPE/choroid is found to decline. However, in the retina, odd acylcarnitines are increased, revealing "an opposite" change within the RPE/choroid, accompanied by influencing glycolytic key enzymes. The HFD+CNV group incorporated fewer long-chain acylcarnitines, like C18:2, into the retina than controls. HFD hastens choroidal neovascularization. The study comprehensively documented acylcarnitine profiles in a CNV rat model. Acylcarnitine's odd-even and carbon-chain length properties may guidefuture therapeutics.

MicroRNA-376b-3p Suppresses Choroidal Neovascularization by Regulating Glutaminolysis in Endothelial Cells.

Choroidal neovascularization (CNV) is a common pathological change of various ocular diseases that causes serious damage to central vision. Accumulated evidence shows that microRNAs (miRNAs) are closely related with the regulation of endothelial metabolism, which plays crucial roles in angiogenesis. Here, we investigate the molecular mechanism underlying the regulation of endothelial glutamine metabolism by miR-376b-3p in the progression of CNV. Human retinal microvascular endothelial cells (HRMECs) were transfected with control or miR-376b-3p mimics, and the expression of glutaminase 1 (GLS1), a rate-limiting enzyme in glutaminolysis, was detected by real-time PCR or Western blotting. The biological function and glutamine metabolism of transfected HRMECs were measured by related kits. Luciferase reporter assays were used to validate the CCAAT/enhancer-binding protein beta (CEBPB) was a target of miR-376b-3p. Chromatin immunoprecipitation and RNA immunoprecipitation assays were performed to verify the binding of CEBPB on the promoter region of GLS1. Fundus fluorescein angiography and immunofluorescence detected the effect of miR-376b-3p agomir on rat laser-induced CNV. The expression of miR-376b-3p was decreased, whereas GLS1 expression was increased in the retinal pigment epithelial-choroidal complexes of rats with CNV. HRMECs transfected with miR-376b-3p mimic showed inhibition of CEBPB, resulting in the inactivation of GLS1 transcription and glutaminolysis. Moreover, the miR-376b-3p mimic inhibited proliferation, migration and tube formation but promoted apoptosis in HRMECs, whereas these effects counteracted by α-ketoglutarate supplementation or transfection with CEBPB overexpression plasmid. Finally, the intravitreal administration of the miR-376b-3p agomir restrained CNV formation. Collectively, miR-376b-3p is a suppressor of glutamine metabolism in endothelial cells that could be expected to become a therapeutic target for the treatment of CNV-related diseases.

“One stone and two birds” strategy to treat neovascular age-related macular degeneration by a novel retinoid drug, EYE-101

Choroidal neovascularization (CNV) is a typical pathological feature of neovascular age-related macular degeneration and has become a major cause of vision loss in the elderly. Current therapies require repeated intraocular injections of anti-VEGF drugs by inhibiting endothelial angiogenic effects, which is painful and may cause adverse effects on normal vascular and neuronal functions. Herein, we designed a novel retinoid drug, EYE-101, determined its therapeutic effects on CNV, and clarified the anti-angiogenic mechanism. The results show that administration of EYE-101 did not cause obvious cytotoxicity and ocular tissue toxicity at the concentrations less than 5 μM. Topical administration of EYE-101 could reduce choroidal sprouting, suppress laser-induced CNV formation, and decrease pericyte coverages on ocular vessels. Administration of EYE-101 also suppressed endothelial cell proliferation, migration, and tube formation and reduced pericyte proliferation, migration, recruitment towards endothelial cells. EYE-101 exerted its anti-angiogenic effects by targeting endothelial cells and pericytes via antagonizing Wnt/β-catenin signaling and PDGF signaling. Thus, EYE-101 administration may offer an“one stone and two birds” strategy for the prevention and treatment of ocular neovascular disorders.

Subretinal application of cell cultures: Surgical methods and its transition to the use in human

AbstractThe progression of age‐related macular degeneration (AMD) results in irreversible and untreatable impairment of central vision. The formation of the drusen and subsequent atrophy of in the retinal pigmented epithelium (RPE) with development of neovascular membranes remain the typical features of AMD pathogenesis. As the result of the disorganization of the RPE the death photoreceptor cell develops, which is associated with altered RPE phagocytosis, increased RPE permeability, formation of choroidal neovascularization, and involvement of resident and monocytic microglial cells. Nevertheless, the exact cause of the RPE degeneration is still not fully understood. Up to date there is no therapeutic approach, which could reverse the course of AMD progression. Injection of anti‐vascular endothelial growth factors (VEGFs), laser‐photo coagulation and photodynamic therapy of the neovascularisations are being widely used to treat the complications of exudative AMD and therefore to prevent visual loss. Since few decades, the concept of substitutional cell‐based regenerative treatment has been investigated by many groups. It could offer in the near future an alternative approach to restore impaired vision in AMD. Th studies have shown that subretinal implantation of RPE cells seeded on a biodegradable scaffold enhances the integration of the donor cells compared to the injection of cell‐suspension as method of delivery. This could improve the differentiation of donor RPE cells into the host retina and help with the proper cellularization of the RPE monolayer. However, the implantation techniques including surgical procedure with exposion of subretinal space and the development and use of delivery instruments remains challenging. Experimental studies with subretinal RPE transplantation are focused on development of optimal surgical techniques, which could be easily transferred to the application in humans. The cultivation and reproducible viability of hRPEs for transplantation purposes on biodegradable scaffolds for human transplantation shall correspond and be adapted to the instrumentation used during the surgery. Hence, the advances in the methodology and the results of improved approaches shall be investigated further.

GPx4 reduces laser‐induced choroidal neovascularization by inhibiting ferroptosis

AbstractPurpose: To investigate the protective effects of glutathione peroxidase 4 (GPx4) in laser‐Induced age‐related macular degeneration (AMD) animal model.Methods: In this study, Norway brown rats were used to establish a laser‐induced AMD model. To investigate the protective role of GPx4, rats received laser photocoagulation and intravitreal injection of GPx4 (25 μg/eye) or PBS on the same day. The size and vascular leakage of choroidal neovascularization (CNV) were measured on 7 and 14 days after lasing by using OCT and FA. Immunohistochemical staining of the choroidal flat mounts with CD31 were used to assess angiogenesis.Results: Compared with PBS group, intravitreal injection of GPx4 decreased the CNV size of 0.083 ± 0.121, P = 0.07 on day 7 and 0.11 ± 0.135 (p < 0.05) on day 14. By leakage grading, the CNV lesion with pathologically significant leakage (grade 2a and 2b) was reduced in GPx4 group (2%) and increased in PBS group (25.6%) from day 7 to day 14. The immunostaining with CD31 showed smaller lesions in GPx4 group than PBS group.Conclusions: This study showed that GPx4 inhibited CNV formation and providing a novel molecular target against ferroptosis for wet AMD treatment.

Licochalcone A alleviates laser-induced choroidal neovascularization by inhibiting the endothelial-mesenchymal transition via PI3K/AKT signaling pathway

Choroidal neovascularization (CNV) is a hallmark of wet age-related macular degeneration, which severely impairs central vision. Studies have shown that endothelial-mesenchymal transition (EndMT) is involved in the pathogenesis of CNV. Licochalcone A (lico A), a flavonoid extracted from the root of licorice, shows the inhibition on EndMT, but it remains unclear whether it can suppress the formation of CNV. The aim of this study is to investigate the effects of lico A on laser-induced CNV, and EndMT process in vitro and vivo. We established the model of CNV with a krypton laser in Brown-Norway rats and then intraperitoneally injected lico A. Our experimental results demonstrated that the leakage of CNV was relieved, and the area of CNV was reduced in lico A-treated rats. Cell migration and tube formation in oxidized low-density lipoprotein (Ox-LDL)-stimulated HUVECs were inhibited by lico A and promoted by PI3K activator 740Y-P. The protein expressions of snai1 and α-SMA were increased, and CD31 and VE-cadherin were decreased in the model rats of CNV, but partially reversed after treatment with lico A. The expression of CD31 was decreased and α-SMA was increased in OX-LDL-treated HUVECs, which was further strengthened by 740Y-P, while the expression of CD31 was up-regulated and α-SMA was down-regulated in lico A treated HUVECs. Our data revealed that EndMT process was alleviated by lico A. Meanwhile, PI3K/AKT signaling pathway was activated in model rat of CNV and Ox-LDL-stimulated HUVECs, which can be suppressed with treatment of lico A. Our experimental results confirmed for the first time that lico A has the potential to alleviate CNV by inhibiting the endothelial-mesenchymal transition via PI3K/AKT signaling pathway.

PGE2 promotes macrophage recruitment and neovascularization in murine wet-type AMD models

Age-related macular degeneration (AMD), a progressive chronic disease of the central retina, is a leading cause of blindness worldwide. Activated macrophages recruited to the injured eyes greatly contribute to the pathogenesis of choroidal neovascularization (CNV) in exudative AMD (wet AMD). This study describes the effects of cyclooxygenase-2 (COX2)/prostaglandin E2 (PGE2) signalling on the macrophage activation and CNV formation of wet AMD. In a mouse model of laser-induced wet AMD, the mice received an intravitreal injection of celecoxib (a selective COX2 inhibitor). Optical coherence tomography (OCT), fundus fluorescein angiography (FFA), choroidal histology of the CNV lesions, and biochemical markers were assessed. The level of PGE2 expression was high in the laser-induced CNV lesions. Macrophage recruitment and CNV development were significantly less after celecoxib treatment. E-prostanoid1 receptor (EP1R)/protein kinase C (PKC) signalling was involved in M2 macrophage activation and interleukin-10 (IL-10) production of bone marrow-derived macrophages (BMDMs) in vitro. In addition, IL-10 was found to induce the proliferation and migration of human choroidal microvascular endothelial cells (HCECs). Thus, the PGE2/EP1R signalling network serves as a potential therapeutic target for CNV of the wet-type AMD.Graphical AXv86vuL_n_YsWmKFp4z_2Video abstract

Application and mechanism of anti-VEGF drugs in age-related macular degeneration.

Age-related macular degeneration (AMD) is the leading cause of blindness in the elderly. The incidence rate increases with age in people over 50 years of age. With the advent of China’s aging society, the number of patients is increasing year by year. Although researchers have done a lot of basic research and clinical research on the pathogenesis and treatment of AMD in recent years, the pathogenesis of AMD is still controversialdue to the complexity of the disease itself. AMD is the primary cause of blindness in the elderly over 50 years old. It is characterized by the formation of choroidal neovascularization (CNV) and the over secretion of vascular endothelial growth factor (VEGF) as its main mechanism, which can eventually lead to vision loss or blindness. The occurrence and development of AMD is an extremely complex process, in which a large number of regulatory factors and cytokines are involved. Most of the existing treatments are for its concomitant CNV. Targeted VEGF drugs for neovascularization, such as Lucentis and Aflibercept, are the first-line drugs for AMD. Their application has greatly reduced the blinding rate of patients. However, there are still some patients who have no response to treatment or cannot maintain their vision after long-term treatment. Frequent injection also increases the risk of complications and economic burden. In order to further improve the quality of life and long-term prognosis of AMD patients, a variety of new treatmentshave been or will be applied in clinic, including combined treatment with the same or different targets to improve the curative effect, change or simplify the mode of medication, inhibit VEGF receptor tyrosine protein kinase and so on. This article provides a brief review of the research progress of anti-VEGF drugs and their mechanisms for the treatment of AMD, it is expected to provide a better treatment plan for AMD treatment.