Model Of Retinopathy Research Articles

Corticosteroids reduce pathological angiogenesis yet compromise reparative vascular remodeling in a model of retinopathy

Tissue inflammation is often broadly associated with cellular damage, yet sterile inflammation also plays critical roles in beneficial tissue remodeling. In the central nervous system, this is observed through a predominantly innate immune response in retinal vascular diseases such as age-related macular degeneration, diabetic retinopathy, and retinopathy of prematurity. Here, we set out to elucidate the dynamics of the immune response during progression and regression of pathological neovascularization in retinopathy. In a mouse model of oxygen-induced retinopathy, we report that dexamethasone, a broad-spectrum corticosteroid, suppresses initial formation of pathological preretinal neovascularization in early stages of disease, yet blunts reparative inflammation by impairing distinct myeloid cell populations, and hence reduces beneficial vascular remodeling in later stages of disease. Using genetic depletion of distinct components of the innate immune response, we demonstrate that CX3C chemokine receptor 1-expressing microglia contribute to angiogenesis. Conversely, myeloid cells expressing lysozyme 2 are recruited to sites of damaged blood vessels and pathological neovascularization where they partake in a reparative process that ultimately restores circulatory homeostasis to the retina. Hence, the Janus-faced properties of anti-inflammatory drugs should be considered, particularly in stages associated with persistent neovascularization.

Evaluation of the effect of melatonin treatment on telomere length of the retinal pigment epithelium in streptozotocin-induced diabetic rat model

ObjectivesWe aimed to investigate the effect of diabetic retinopathy and melatonin treatment on the relative telomer lengths (RTL) in retinal pigment epithelium (RPE) cells in a streptozotocin-induced diabetic rat model.BackgroundTL can be used to evaluate diabetes mellitus, its complications, and the effectiveness of its treatment. However, TL assessment has not been performed in retinal cells in a diabetic retinopathy model until now.MethodsForty Sprague-Dawley male rats were randomly divided into four groups. The experimental groups were: Control Group (C): non- diabetic rats; Diabetes Mellitus Group (DM): rats induced to diabetes without treatment; Melatonin and Diabetes Mellitus Group (Mel + DM): rats induced to diabetes and after confirmation, treated with melatonin; Melatonin Group (Mel): rats were not induced to diabetes, treated with melatonin. Diabetes was induced by intraperitoneal administration of streptozotocin solution after 12 h food fasting. For eight weeks after the diabetes was induced, melatonin was administered via subcutaneous injection at a dose of 10 mg / kg. RTLs were measured by qPCR method with modifications. The comparison of averaged data among groups was performed using least significant difference (LSD) and Kruskal – Wallis Test and One way ANOVA test.ResultsRTL was significantly similar in control and melatonin group. RTL was thinnest in DM group, in addition melatonin treatment significantly prevented the RTL shortening in DM + Mel group (p = 0.031).ConclusionWe demonstrated that diabetic retinopathy led to the shortening of RTL in RPE cells in rats and melatonin treatment prevents this shortening.

Development of an Open-Source Dataset of Flat-Mounted Images for the Murine Oxygen-Induced Retinopathy Model of Ischemic Retinopathy.

To describe an open-source dataset of flat-mounted retinal images and vessel segmentations from mice subject to the oxygen-induced retinopathy (OIR) model. Flat-mounted retinal images from mice killed at postnatal days 12 (P12), P17, and P25 used in prior OIR studies were compiled. Mice subjected to normoxic conditions were killed at P12, P17, and P25, and their retinas were flat-mounted for imaging. Major blood vessels from the OIR images were manually segmented by four graders (JSC, HKR, KBL, JM), with cross-validation performed to ensure similar grading. Overall, 1170 images were included in this dataset. Of these images, 111 were of normoxic mice retina, and 1048 were mice subject to OIR. The majority of images from OIR mice were obtained at P17. The 50 images obtained from an external dataset, OIRSeg, did not have age labels. All images were manually segmented and used in the training or testing of a previously published deep learning algorithm. This is the first open-source dataset of original and segmented flat-mounted retinal images. The dataset has potential applications for expanding the development of generalizable and larger-scale artificial intelligence and analyses for OIR. This dataset is published online and publicly available at dx.doi.org/10.6084/m9.figshare.23690973. This open access dataset serves as a source of raw data for future research involving big data and artificial intelligence research concerning oxygen-induced retinopathy.

Automated Early Diabetic Retinopathy Detection Using a Deep Hybrid Model

Recently, the primary reason for blindness in adults has been diabetic retinopathy (DR) disease. Therefore, there is an increasing demand for a real-time efficient classification and detection system for diabetic retinopathy (DR) to overcome fast-growing disease (DR). We introduced a novel deep hybrid model for auto-mated diabetic retinopathy (DR) disease recognition and classification. Our model leverages the power of CNN architectures: Inception V3 and VGG16 models by combining their strengths to cater to exact requirements. VGG16 model efficiently captures fine features and wide-ranging features such as textures and edges, crucial for classifying initial signs of DR. Similarly, Inception V3’s architecture is proficient at detecting multiscale patterns, providing an extensive setting for shaping the occurrence of more complex DR severity stages. Our deep hybrid model allows the extraction of various appearance features in retinal images, which can better assist the classification and detection of DR. Our proposed model evaluated on diverse datasets, including EyePACS1 and APTOS2019, demonstrating confident performance of 99.63% accuracy in classifying the DR severity levels on EyePACS1 dataset, while 98.70% accuracy on the APTOS2019 dataset, indicating that our proposed deep hybrid model well distinguished different stages and highly efficient in DR detection. This model helps clinicians and medical experts to classify and identify diabetic retinopathy DR stages and severity levels early. This automatic system helps to manage and treat the patient more effectively and introduces timely treatment.

Monoclonal antibodies that block Roundabout 1 and 2 signaling target pathological ocular neovascularization through myeloid cells.

Roundabout (ROBO) 1 and 2 are transmembrane receptors that bind secreted SLIT ligands through their extracellular domains (ECDs) and signal through their cytoplasmic domains to modulate the cytoskeleton and regulate cell migration, adhesion, and proliferation. SLIT-ROBO signaling regulates pathological ocular neovascularization, which is a major cause of vision loss worldwide, but pharmacological tools to prevent SLIT-ROBO signaling are lacking. Here, we developed human monoclonal antibodies (mAbs) against the ROBO1 and ROBO2 ECDs. One antibody that inhibited in vitro SLIT2 signaling through ROBO1 and ROBO2 (anti-ROBO1/2) also reduced ocular neovascularization in oxygen-induced retinopathy (OIR) and laser-induced corneal neovascularization (CNV) mouse models in vivo. Single-cell RNA sequencing of OIR retinas revealed that antibody treatment affected several cell types relevant to physiological and pathological angiogenesis, including endothelial cells, pericytes, and a heterogeneous population of myeloid cells. mAb treatment improved blood-retina barrier integrity and prevented pathological pericyte activation in OIR. SLIT-ROBO signaling inhibition prevented pathological activation of myeloid cells and increased neuroprotective myeloid populations normally seen in the developing retina. Microglia/infiltrating macrophage-specific ablation of Robo1 and Robo2 or knockout of the downstream effector phosphatidylinositol 3-kinase (Pik3cg) encoding PI3Kγ in both OIR and CNV models phenocopied anti-ROBO1/2 treatment, further demonstrating the key role of myeloid cells as drivers of ocular neovascular diseases. ROBO1/2 blocking antibodies may thus provide a promising strategy to combat inflammation in blinding eye diseases.

Single-Cell Multiomics Profiling Reveals Heterogeneity of Müller Cells in the Oxygen-Induced Retinopathy Model.

Retinal neovascularization poses heightened risks of vision loss and blindness. Despite its clinical significance, the molecular mechanisms underlying the pathogenesis of retinal neovascularization remain elusive. This study utilized single-cell multiomics profiling in an oxygen-induced retinopathy (OIR) model to comprehensively investigate the intricate molecular landscape of retinal neovascularization. Mice were exposed to hyperoxia to induce the OIR model, and retinas were isolated for nucleus isolation. The cellular landscape of the single-nucleus suspensions was extensively characterized through single-cell multiomics sequencing. Single-cell data were integrated with genome-wide association study (GWAS) data to identify correlations between ocular cell types and diabetic retinopathy. Cell communication analysis among cells was conducted to unravel crucial ligand-receptor signals. Trajectory analysis and dynamic characterization of Müller cells were performed, followed by integration with human retinal data for pathway analysis. The multiomics dataset revealed six major ocular cell classes, with Müller cells/astrocytes showing significant associations with proliferative diabetic retinopathy (PDR). Cell communication analysis highlighted pathways that are associated with vascular proliferation and neurodevelopment, such as Vegfa-Vegfr2, Igf1-Igf1r, Nrxn3-Nlgn1, and Efna5-Epha4. Trajectory analysis identified a subset of Müller cells expressing genes linked to photoreceptor degeneration. Multiomics data integration further unveiled positively regulated genes in OIR Müller cells/astrocytes associated with axon development and neurotransmitter transmission. This study significantly advances our understanding of the intricate cellular and molecular mechanisms underlying retinal neovascularization, emphasizing the pivotal role of Müller cells. The identified pathways provide valuable insights into potential therapeutic targets for PDR, offering promising directions for further research and clinical interventions.

Capsiate Improves Glucose Metabolism by Improving Insulin Sensitivity in Diabetic Retinopathy Mice

Purpose Capsiate (cap) is a metabolite that affects a number of biological processes, and diabetic retinopathy (DR) is now known to be the primary cause of end-stage eye illness. Methods In order to examine the effects of the cap intervention on body weight, nutritional intake, changes in body weight composition, glucose metabolism levels, retinopathy, and oxidative stress levels, we proposed using a mouse model of diabetic retinopathy caused by STZ. Results Our findings demonstrated that, in addition to increasing lean body mass and lowering fat body mass content, cap intervention significantly improved body weight and dietary consumption in STZ mice. Additionally, our results on glucose metabolism revealed that cap had a significant impact on insulin resistance and the stabilization of OGTT levels. In conclusion, we examined the levels of oxidative stress and retinopathy. We discovered that the cap intervention greatly reduced the levels of MDA and significantly improved the levels of VEGF and retinopathy. In contrast, the STZ group’s levels of SOD, CAT, and GSH were significantly higher. Conclusions According to our research, the Cap intervention improved the damage caused by diabetic retinopathy by reversing the levels of oxidative stress and the disrupted state of glucose metabolism, which in turn decreased the levels of VEGF.

METTL14-mediated m6A methylation regulates pathological retinal neovascularization by targeting autophagy

Pathological retinal neovascularization (RNV) is a prevalent characteristic of various ocular diseases, including proliferative diabetic retinopathy (PDR), retinopathy of prematurity (ROP), and retinal vein occlusion (RVO). While the importance of N6-methyladenosine (m6A) modification in diverse disease contexts is well-established, its functional role in pathological RNV remains unclear. Herein, we investigated the involvement of m6A modification and its core methyltransferase, METTL14, in a model of oxygen-induced retinopathy (OIR) to elucidate their contribution to retinal angiogenesis. In this study, we observed heightened levels of m6A modification and elevated expression of METTL14 in the OIR model, suggesting their potential implication in pathological RNV. Employing targeted knockdown of METTL14, we revealed that its depletion activated autophagy flux in human retinal vascular endothelial cells (HRVECs), consequently inhibiting the angiogenic capacity of endothelial cells. Mechanistically, we demonstrated that METTL14 exerts its regulatory influence on autophagy flux by modulating the stability of ATG7, a pivotal protein involved in autophagy. Specifically, METTL14 knockdown led to increased ATG7 expression at both mRNA and protein levels, accompanied by reduced m6A methylation of ATG7 mRNA and enhanced mRNA stability. Moreover, silencing of ATG7 counteracted the effects of METTL14 knockdown on endothelial cell functions, emphasizing ATG7 as a downstream target of METTL14-mediated autophagy in HRVECs. After all, our findings provide valuable insights into the pathogenesis of retinal pathological angiogenesis and potential therapeutic targets for the treatment of ocular neovascular diseases.

Paquinimod attenuates retinal injuries by suppressing the S100A9/TLR4 signaling in an experimental model of diabetic retinopathy

Diabetic retinopathy (DR), the most common ocular complication of diabetes mellitus (DM), has exhibited an increase in incidence over the past decade. S100 calcium-binding protein A9 (S100A9) plays a significant role in inflammation and cancer. Toll-like receptor 4 (TLR4), a transmembrane receptor, initiates signaling cascades upon ligand binding. S100A9 activates TLR4, and their involvement in various diseases is well-established. We found elevated S100A9/TLR4 pathway proteins in the vitreous of DR patients. Bioinformatics analysis revealed differential gene expression related to this pathway. These proteins were also detected in diabetic rat retinas and induced structural damage. Paquinimod, an S100A9 inhibitor, decreased pathway protein expression and reduced retinal damage. Our study validates the S100A9/TLR4 pathway in diabetic retinas and suggests its potential as a therapeutic target for DR. Targeting S100A9 could offer a novel approach to prevention and treatment.

Effects of mesenchymal stromal cells and human recombinant Nerve Growth Factor delivered by bioengineered human corneal lenticule on an innovative model of diabetic retinopathy.

Diabetic retinopathy (DR) is a microvascular complication of diabetes in which neurodegeneration has been recently identified as a driving force. In the last years, mesenchymal stromal cells (MSCs) and neurotrophins like Nerve Growth Factor (NGF), have garnered significant attention as innovative therapeutic approaches targeting DR-associated neurodegeneration. However, delivering neurotrophic factors directly in the eye remains a challenge. Hence, this study evaluated the effects of MSCs from human amniotic fluids (hAFSCs) and recombinant human NGF (rhNGF) delivered by human corneal lenticule (hCL) on a high glucose (HG) induced ex vivo model simulating the molecular mechanisms driving DR. Porcine neuroretinal explants exposed to HG (25 mM for four days) were used to mimic DR ex vivo. hCLs collected from donors undergoing refractive surgery were decellularized using 0.1% sodium dodecyl sulfate and then bioengineered with hAFSCs, microparticles loaded with rhNGF (rhNGF-PLGA-MPs), or both simultaneously. Immunofluorescence (IF) and scanning electron microscopy (SEM) analyses were performed to confirm the hCLs bioengineering process. To assess the effects of hAFSCs and rhNGF, bioengineered hCLs were co-cultured with HG-treated neuroretinal explants and following four days RT-PCR and cytokine array experiments for inflammatory, oxidative, apoptotic, angiogenic and retinal cells markers were performed. Data revealed that HG-treated neuroretinal explants exhibit a characteristic DR-phenotype, including increased level of NF-kB, NOS2, NRF2 GFAP, VEGFA, Bax/Bcl2 ratio and decreased expression of TUBB3 and Rho. Then, the feasibility to bioengineer decellularized hCLs with hAFSCs and rhNGF was demonstrated. Interestingly, co-culturing hAFSCs- and rhNGF- bioengineered hCLs with HG-treated neuroretinal explants for four days significantly reduced the expression of inflammatory, oxidative, apoptotic, angiogenic and increased retinal markers. Overall, we found for the first time that hAFSCs and rhNGF were able to modulate the molecular mechanisms involved in DR and that bioengineered hCLs represents a promising ocular drug delivery system of hAFSCs and rhNGF for eye diseases treatment. In addition, results demonstrated that porcine neuroretinal explants treated with HG is a useful model to reproduce ex vivo the DR pathophysiology.

Phlebotomy-induced anemia reduces oxygen-induced retinopathy severity and dampens retinal developmental transcriptomic pathways in rats.

Phlebotomy-induced-anemia (PIA), which induces tissue hypoxia and angiogenesis, occurs universally among infants at risk for severe retinopathy of prematurity (ROP). We hypothesized that PIA exacerbates pathologic retinal neovascularization in ROP. We induced PIA to a hematocrit of 18% among rats undergoing the established 50/10 oxygen-induced retinopathy (OIR) model. Rats were euthanized at P15 and P20, during the avascular and neovascular phases of OIR, respectively. Retinal vascular morphometry, cytokine/chemokine concentrations, transcriptomes, and mRNA expression of angiogenic and iron-deficiency markers were compared to non-PIA controls. In OIR, PIA decreased percent avascular area at P15 by 35%, percent neovascular area at P20 by 42%, and select pro-inflammatory cytokine/chemokine concentrations at both time points. At P20, PIA increased mRNA expression of angiopoietin 2/ vascular endothelial growth factor-A 2-fold and transferrin and transferrin receptor 5-fold. RNA sequencing showed dampened pathways of angiogenesis, inflammation, and neural development in anemic OIR females. Contrary to our hypothesis, PIA decreased OIR severity and retinal cytokine and chemokine levels and dampened transcriptomic pathways central to retinal vascular and neural development in neonatal rats. These data suggest PIA provides a protective effect from OIR. Further investigation into the functional effect of these molecular changes is warranted. This is the first preclinical study to investigate the impact of neonatal anemia on oxygen-induced retinopathy (OIR) outcomes. This study adds to the literature that anemia decreases neovascularization, decreases cytokine and chemokine levels, and dampens angiogenic and neural transcriptomic pathways in the rat 50/10 OIR model. The study identifies a sex-specific transcriptomic response to anemia in the 50/10 OIR model, with females primarily impacted.

Heterotypic macrophages/microglia differentially contribute to retinal ischaemia and neovascularisation.

Diabetic retinopathy is characterised by neuroinflammation that drives neuronal and vascular degenerative pathology, which in many individuals can lead to retinal ischaemia and neovascularisation. Infiltrating macrophages and activated retina-resident microglia have been implicated in the progression of diabetic retinopathy, although the distinct roles of these immune cells remain ill-defined. Our aim was to clarify the distinct roles of macrophages/microglia in the pathogenesis of proliferative ischaemic retinopathies. Murine oxygen-induced retinopathy is commonly used as a model of ischaemia-induced proliferative diabetic retinopathy (PDR). We evaluated the phenotype macrophages/microglia by immunostaining, quantitative real-time RT-PCR (qRT-PCR), flow cytometry and scRNA-seq analysis. In clinical imaging studies of diabetic retinopathy, we used optical coherence tomography (OCT) and OCT angiography. Immunostaining, qRT-PCR and flow cytometry showed expression levels of M1-like macrophages/microglia markers (CD80, CD68 and nitric oxide synthase 2) and M2-like macrophages/microglia markers (CD206, CD163 and macrophage scavenger receptor 1) were upregulated in areas of retinal ischaemia and around neo-vessels, respectively. scRNA-seq analysis of the ischaemic retina revealed distinct ischaemia-related clusters of macrophages/microglia that express M1 markers as well as C-C chemokine receptor 2. Inhibition of Rho-kinase (ROCK) suppressed CCL2 expression and reduced CCR2-positive M1-like macrophages/microglia in areas of ischaemia. Furthermore, the area of retinal ischaemia was reduced by suppressing blood macrophage infiltration not only by ROCK inhibitor and monocyte chemoattractant protein-1 antibody but also by GdCl3. Clinical imaging studies of diabetic retinopathy using OCT indicated potential involvement of macrophages/microglia represented by hyperreflective foci in areas of reduced perfusion. These results collectively indicated that heterotypic macrophages/microglia differentially contribute to retinal ischaemia and neovascularisation in retinal vascular diseases including diabetic retinopathy. This adds important new information that could provide a basis for a more targeted, cell-specific therapeutic approach to prevent progression to sight-threatening PDR.

DDLA: a double deep latent autoencoder for diabetic retinopathy diagnose based on continuous glucose sensors.

The current diagnosis of diabetic retinopathy is based on fundus images and clinical experience. However, considering the ineffectiveness and non-portability of medical devices, we aimed to develop a diagnostic model for diabetic retinopathy based on glucose series data from the wearable continuous glucose monitoring system. Therefore, this study developed a novel method, i.e., double deep latent autoencoder, for exploring glycemic variability influence from multi-day glucose data for diabetic retinopathy. Specifically, the model proposed in this research could encode continuous glucose sensor data with non-continuous and variable length via the integration of a data reorganization module and a novel encoding module with fragmented-missing-wise objective function. Additionally, the model implements a double deep autoencoder, which integrated convolutional neural network, long short-term memory, to jointly capturing the inter-day and intra-day glucose latent features from glucose series. The effectiveness of the proposed model is evaluated through a cross-validation method to clinical datasets of 765 type 2 diabetes patients. The proposed method achieves the highest accuracy value (0.89), precision value (0.88), and F1 score (0.73). The results suggest that our model can be used to remotely diagnose and screen for diabetic retinopathy by learning potential features of glucose series data collected by wearable continuous glucose monitoring systems.

TRNA-derived fragment tRF-30 propels diabetes-induced retinal microvascular complications by regulating STAT3 signaling.

Transfer RNA-derived fragments (tRFs) represent a novel class of non-coding RNA transcripts that possess specific biological functions. However, the involvement of tRFs in retinal microvascular diseases remains poorly understood. In this study, we aimed to reveal whether modulation of tRF-30 expression could attenuate pathological retinal neovascular diseases. Our findings demonstrate a significant upregulation of tRF-30 expression levels in both in vivo models of diabetic retinopathy (DR) and in vitro endothelial sprouting models. Conversely, inhibition of tRF-30 expression suppressed the formation of abnormal neovascularization in the retina in vivo, while reducing the proliferation and migration activity of retinal vascular endothelial cells in vitro. We also found that tRF-30 modulates retinal neovascularization through the tRF-30/TRIB3/signal transducer and activated transcription 3 signaling pathway. Furthermore, we validated a significant upregulation of tRF-30 expression levels in the vitreous humor of DR patients, with high levels of both validity and specificity in diagnostic testing. Collectively, our findings highlight a pro-angiogenic role for tRF-30 in DR. Intervening in the tRF-30 signaling pathway may represent a promising prevention and treatment strategy for retinal angiogenesis.

Enhancing Gpx1 palmitoylation to inhibit angiogenesis by targeting PPT1

The significance of protein S-palmitoylation in angiogenesis has been largely overlooked, leaving various aspects unexplored. Recent identification of Gpx1 as a palmitoylated protein has generated interest in exploring its potential involvement in novel pathological mechanisms related to angiogenesis. In this study, we demonstrate that Gpx1 undergoes palmitoylation at cysteine-76 and -113, with PPT1 playing a crucial role in modulating the depalmitoylation of Gpx1. Furthermore, we find that PPT1-regulated depalmitoylation negatively impacts Gpx1 protein stability. Interestingly, inhibiting Gpx1 palmitoylation, either through expression of a non-palmitoylated Gpx1 mutant or by expressing PPT1, significantly enhances neovascular angiogenesis. Conversely, in PPT1-deficient mice, angiogenesis is notably attenuated compared to wild-type mice in an Oxygen-Induced Retinopathy (OIR) model, which mimics pathological angiogenesis. Physiologically, under hypoxic conditions, Gpx1 palmitoylation levels are drastically reduced, suggesting that increasing Gpx1 palmitoylation may have beneficial effects. Indeed, enhancing Gpx1 palmitoylation by inhibiting PPT1 with DC661 effectively suppresses retinal angiogenesis in the OIR disease model. Overall, our findings highlight the pivotal role of protein palmitoylation in angiogenesis and propose a novel mechanism whereby the PPT1-Gpx1 axis modulates angiogenesis, thereby providing a potential therapeutic strategy for targeting PPT1 to combat angiogenesis.

A Nanogel Formulation of Anti-VEGF Peptide for Ocular Neovascularization Treatment.

Age-related macular degeneration (AMD) is an eye disorder that can lead to visual impairment in elder patients, and current treatments include repeated injections of monoclonal antibody-based antivascular endothelial growth factor (anti-VEGF) agents. This study investigates the potential of a nanoformulation of a peptide anti-VEGF molecule for neovascular AMD. Anti-VEGF peptide HRHTKQRHTALH (HRH), which has high affinity to VEGF-Fc receptor, was used as the bioactive agent to control neovascularization of the retina. The nanoformulation consisting of hyaluronic acid nanogel was generated by incorporating divinyl sulfone and cholesterol to increase the stability and control the size of the nanodrug. The encapsulation efficacy of nanogel was 65%, and drug release was 34.72% at the end of 192 h. Obtained nanogels were efficiently internalized in 15 min by human umbilical vascular endothelial cells (HUVECs) and ARPE-19 cells, and results indicate that nanoformulation is not toxic to ARPE-19 cells, whereas it inhibits HUVEC proliferation owing to anti-VEGF peptide in the nanogel structure. In the coculture experiment in which retinal penetration was modeled, it was observed that the nanogel reached HUVECs and negatively affected their proliferation without disturbing the monolayer of ARPE-19 cells. In vivo experiments with chick chorioallantoic membrane revealed that nanogel formulation has higher antiangiogenesis activity compared to free HRH. Additionally, in an oxygen-induced retinopathy model, the excessive growth of blood vessels was notably suppressed in mice treated with HRH-loaded nanogel. This research indicates that nanogels formulated in this study are promising candidates as a topical treatment for AMD.

LncRNA-MM2P regulates retinal neovascularization through M2 macrophage polarization

The study aims to investigate the effects and potential mechanisms of lncRNA-MM2P on retinal neovascularization in a mouse model of oxygen-induced retinopathy (OIR). The OIR model was established in C57BL/6J mice. RAW264.7 cell line and bone marrow-derived macrophages (BMDMs) from mice were used for in vitro studies. RT-qPCR was used to analyze the expressions of lncRNA and mRNAs. The protein expression levels were determined by western blotting. The size of avascular areas and neovascular tufts were assessed based on isolectin B4 immunofluorescence staining images. The human retinal endothelial cells (HRECs) were used to evaluate the proliferation, migration, and tube formation of endothelial cells. The expression of lncRNA-MM2P was significantly upregulated from P17 to P25 in OIR retinas. Knockdown of lncRNA-MM2P levels in vivo led to a significant reduction in the neovascular tufts and avascular areas in the retinas of OIR mice. Knockdown of lncRNA-MM2P levels in vitro suppressed the expression of M2 markers in macrophages. Moreover, we found a significant inhibition of avascular areas and neovascular tufts in OIR mice injected intravitreally with M2 macrophages treated by shRNA-MM2P. The cellular functions of proliferation, migration, and tube formation were significantly attenuated in HRECs cultured with a supernatant of shRNA-MM2P-treated M2 macrophages. Our results indicate that lncRNA-MM2P regulates retinal neovascularization by inducing M2 polarization of macrophages in OIR mice. Therefore, lncRNA-MM2P may be a potential molecular target for immunoregulation of retinal neovascularization.

Reduction of pathological retinal neovascularization, vessel obliteration, and artery tortuosity by PEDF protein in an oxygen-induced ischemic retinopathy rat model.

Retinopathy of prematurity (ROP) is a severe retinal disease in premature infants characterized by pathological neovascularization, obliteration of retinal vessels and increased vessel tortuosity. Currently, there are no completely satisfactory treatments for ROP. Pigment epithelium-derived factor (PEDF), a potent inhibitor of angiogenesis, appears late in gestation and its deficiency may be linked to development of ROP. This study investigates the preclinical efficacy of PEDF protein alone or in combination with VEGF antagonists for treating ROP. The safety of PEDF protein in the rat eye was assessed using functional invivo measurements and histology. The efficacy of intravitreal injections (IVI) of various treatments was evaluated in a rat oxygen-induced retinopathy (OIR) model using invivo imaging and flatmount analyses. No functional or histological side-effects were found in rat eyes after intravitreal PEDF protein injection. PEDF protein alone or combined with anti-VEGF drugs significantly reduced pathological neovascularization and vessel obliteration, comparable to the effects of anti-VEGF drugs alone. Regarding arterial tortuosity, treatment with a combination of PEDF, and VEGF antagonist was more effective than treatment with anti-VEGF alone. IVI of PEDF protein is safe. PEDF protein alone or combined with VEGF antagonists shows similar efficacy in reducing pathological neovascularization and vessel obliteration as anti-VEGF agents. Furthermore, only treatments involving PEDF protein, alone or with VEGF antagonists, significantly improved the quality of retinal vasculature. Thus, PEDF protein alone or combined with anti-VEGF agents presents a promising alternative to current anti-VEGF treatments for ROP.

Endothelial c-Src Mediates Neovascular Tuft Formation in Oxygen-Induced Retinopathy

Vascular retinopathy, characterised by abnormal blood vessel growth in the retina, frequently results in vision impairment or loss. Neovascular tufts, a distinctive pathological feature of this condition, are highly leaky blood vessel structures, exacerbating secondary complications. Despite their clinical significance, the mechanisms underlying tuft development are not fully elucidated, posing challenges for effective management and treatment of vascular retinopathy. This study investigates the role of c-Src in neovascular tuft formation. Although c-Src has been acknowledged as a pivotal regulator in developmental angiogenesis within the retinal vasculature, its specific role in governing pathological retinal angiogenesis remains to be fully understood.The Oxygen-Induced Retinopathy (OIR) model was used for neovascular tufts formation in both Cre-mediated vascular specific c-Src knockout mice and wildtype littermates. High-resolution imaging and analysis of isolated retinas was conducted. c-Src depletion demonstrated a significant reduction in neovascular tufts within the OIR model. This decrease in tuft formation was observed independently of any alterations in cell death, cell proliferation or cell adhesion and the absence of c-Src did not impact tuft pericyte coverage and junctional morphology. These findings underscore the critical role of c-Src in the pathogenesis of neovascular tufts in vascular retinopathy. Understanding the molecular mechanisms involving c-Src may offer valuable insights for the development of targeted therapies aimed at mitigating vision-threatening complications associated with retinopathy.

Agonism of β3-Adrenoceptors Inhibits Pathological Retinal Angiogenesis in the Model of Oxygen-Induced Retinopathy.

In response to hypoxia, sympathetic fibers to the retina activate β-adrenoceptors (β-ARs) that play an important role in the regulation of vascular and neuronal functions. We investigated the role of β3-AR using the mouse model of oxygen-induced retinopathy (OIR). Mouse pups were exposed to 75% oxygen at postnatal day 7 (PD7) followed by a return to room air at PD12. The β3-AR preferential agonist BRL37344 was subcutaneously administered once daily at different times after the return to room air. At PD17, the OIR mice underwent flash and pattern electroretinogram. After sacrifice, retinal wholemounts were used for vessel staining or immunohistochemistry for astrocytes, Müller cells, or retinal ganglion cells (RGCs). In retinal homogenates, the levels of markers associated with neovascularization (NV), the blood-retinal barrier (BRB), or astrocytes were determined by western blot, and quantitative reverse-transcription polymerase chain reaction was used to assess β3-AR messenger. Administration of the β3-AR antagonist SR59230A was performed to verify BRL37344 selectivity. β3-AR expression is upregulated in response to hypoxia, but its increase is prevented by BRL37344, which counteracts NV by inhibiting the pro-angiogenic pathway, activating the anti-angiogenic pathway, recovering BRB-associated markers, triggering nitric oxide production, and favoring revascularization of the central retina through recovered density of astrocytes that ultimately counteracts NV in the midperiphery. Vasculature rescue prevents dysfunctional retinal activity and counteracts OIR-associated retinal ganglion cell loss. β3-AR has emerged as a crucial intermediary in hypoxia-dependent NV, suggesting a role of β3-AR agonists in the treatment of proliferative retinopathies.