Retinal neovascularization is a common feature of several ocular neovascular diseases, which are the leading cause of blindness in the world. Current treatments are administered through invasive intravitreal injections, leading to poor patient compliance, serious ocular complications and heavy economic burdens. Thus, an alternative less or non-invasive therapeutic strategy is in demand. Here, a non-invasive oral tyrosine kinase inhibitor, CM082, was evaluated in a retinal neovascularization model induced by hypoxia in zebrafish larvae. We found that CM082 effectively suppressed retinal neovascularization, rescued cell loss in the retinal ganglion cell layer, and rescued the visual function deficiency. Our results elucidated that CM082 mediated its therapeutic efficacy primarily through the inhibition of Vegfr2 phosphorylation. The findings demonstrated that CM082 possessed strong antiangiogenic effects and may serve as a potential treatment for angiogenesis in ocular neovascular diseases.

HbSC disease, a less severe form of sickle cell disease, affects the retina more frequently and patients have higher rates of proliferative retinopathy that can progress to vision loss. This study aimed to identify differences in the expression of endothelial cell-derived molecules associated with the pathophysiology of proliferative sickle cell retinopathy (PSCR). RNAseq was used to compare the gene expression profile of circulating endothelial colony-forming cells from patients with SC hemoglobinopathy and proliferative retinopathy (n = 5), versus SC patients without retinopathy (n = 3). Real-time polymerase chain reaction (qRT-PCR) was used to validate the RNAseq results. A total of 134 differentially expressed genes (DEGs) were found. DEGs were mainly associated with vasodilatation, type I interferon signaling, innate immunity and angiogenesis. Among the DEGs identified, we highlight the most up-regulated genes ROBO1 (log2FoldChange = 4.32, FDR = 1.35E-11) and SLC38A5 (log2FoldChange = 3.36 FDR = 1.59E-07). ROBO1, an axon-guided receptor, promotes endothelial cell migration and contributes to the development of retinal angiogenesis and pathological ocular neovascularization. Endothelial SLC38A5, an amino acid (AA) transporter, regulates developmental and pathological retinal angiogenesis by controlling the uptake of AA nutrient, which may serve as metabolic fuel for the proliferation of endothelial cells (ECs) and consequent promotion of angiogenesis. Our data provide an important step towards elucidating the molecular pathophysiology of PSCR that may explain the differences in ocular manifestations between individuals with hemoglobinopathies and afford insights for new alternative strategies to inhibit pathological angiogenesis.

UNASSIGNED: Retinal neovascularization is a significant feature of advanced age-related macular degeneration (AMD) and a major cause of blindness in patients with AMD. However, the underlying mechanism of this pathological neovascularization remains unknown. Iron metabolism has been implicated in various biological processes. This study was conducted to investigate the effects of iron metabolism on retinal neovascularization in neovascular AMD (nAMD).
UNASSIGNED: C57BL/6J and very low-density lipoprotein receptor (VLDLR) knockout (Vldlr-/-) mice, a murine model of nAMD, were used in this study. Bulk-RNA sequencing was used to identify differentially expressed genes. Western blot analysis was performed to test the expression of proteins. Iron chelator deferiprone (DFP) was administrated to the mice by oral gavage. Fundus fluorescein angiography was used to evaluate retinal vascular leakage. Immunofluorescence staining was used to detect macrophages and iron-related proteins.
UNASSIGNED: RNA sequencing (RNA-seq) results showed altered transferrin expression in the retina and RPE of Vldlr-/- mice. Disrupted iron homeostasis was observed in the retina and RPE of Vldlr-/- mice. DFP mitigated iron overload and significantly reduced retinal neovascularization and vascular leakage. In addition, DFP suppressed the inflammation in Vldlr-/- retinas. The reduced signals of macrophages were observed at sites of neovascularization in the retina and RPE of Vldlr-/- mice after DFP treatment. Further, the IL-6/JAK2/STAT3 signaling pathway was activated in the retina and RPE of Vldlr-/- mice and reversed by DFP treatment.
UNASSIGNED: Disrupted iron metabolism may contribute to retinal neovascularization in nAMD. Restoring iron homeostasis by DFP could be a potential therapeutic approach for nAMD.

UNASSIGNED: To longitudinally investigate the changes in intraretinal microvascular abnormalities (IRMAs) over time, employing swept-source optical coherence tomography angiography in eyes with diabetic retinopathy.
UNASSIGNED: In this retrospective, longitudinal study, we evaluated 12 × 12-mm swept-source optical coherence tomography angiography centered on the macula at baseline and last available follow-up visit for (1) IRMA changes during follow-up, defined as (a) stable, (b) regressed, (c) obliterated, and (d) progressed; and the (2) development of new neovascularization (NV) and their origins. Competing-risk survival analysis was used to assess the factors associated with these changes.
UNASSIGNED: In total, 195 eyes from 131 participants with diabetic retinopathy were included. Stable, regressed, obliterated, and progressed IRMA were observed in 65.1%, 12.8%, 11.3%, and 19% of eyes with diabetic retinopathy, respectively. Anti-VEGF injections during the follow-up periods and a slower increase of foveal avascular zone were associated with IRMA regression (P < 0.001 and P = 0.039). Obliterated IRMA were correlated with previous panretinal photocoagulation (P < 0.001) and a lower deep capillary plexus vessel density at baseline (P = 0.007), as well as with follow-up anti-VEGF injections (P = 0.025). A higher baseline ischemia index (ISI) and panretinal photocoagulation during the follow-up periods were associated with IRMA progression (P = 0.049 and P < 0.001). A faster increase in ISI predicted the development of NV elsewhere (NVE) from veins (P < 0.001). No significant factors were found to be associated with NVE originating from IRMA.
UNASSIGNED: Changes in IRMA closely correlated with the severity of retinal ischemia and treatment. Notably, our study confirmed the potential, yet relatively rare, development of NVE from IRMA in a large cohort; however, the risk factors associated with this transformation require further exploration.

This study aimed to determine whether the state of retinal vascularization after anti-vascular endothelial growth factor (anti-VEGF) injection can help predict the risk of reactivated retinopathy of prematurity (ROP) requiring treatment and whether repeated ranibizumab injection will be effective in such cases. We retrospectively reviewed 24 infants (43 eyes) who received ranibizumab monotherapy between January 2021 and December 2022. All eyes were classified as having non-retreated ROP or retreated ROP. The state of ROP at the time of treatment, the time required for resolution of plus disease, and the extent of vascularization at 4 and 8 weeks after treatment were analyzed. Extent of temporal retinal vascularization was measured with serial fundus images using disc-fovea distance (DF) unit and disc diameter (DD). Reactivated ROP requiring treatment occurred in six infants (25.0%) and ten eyes (23.3%) after ranibizumab treatment. The mean retreatment interval was 9.0 ± 3.3 weeks (range 4-16). In the retreated ROP group, the time required for the resolution of plus disease after primary injection was longer compared to the control group (13.3 days vs 5.2 days), with a mean ROP regression time of 3.4 weeks. All eyes in the retreated ROP showed retinal vascularization < 0.5 DF from the original site at 4 weeks after injection. In 90% of cases with retreated ROP, the extent of vascularization at 8 weeks after injection was within 1 DF from the original ROP site, and all cases showed reactivation in the posterior Zone II area. The extent of retinal neovascularization in the retreated group was an average of 0.7 DD (vs 1.7 DD) and 1.3 DD (vs 3.3 DD) at 4 and 8 weeks after injection, respectively. After ranibizumab retreatment, only one reactivated case with vitreous traction progressed to focal retinal detachment, while all other cases regressed with peripheral vascular development. The continuation of delayed retinal blood vessel development after ≥ 8 weeks may indicate a high likelihood of reactivated ROP requiring treatment. In the absence of vitreous traction, ranibizumab reinjection is likely to be effective in treating reactivated ROP requiring treatment.

UNASSIGNED: Retinopathy of prematurity (ROP) results from postnatal hyperoxia exposure in premature infants and is characterized by aberrant neovascularization of retinal blood vessels. Epithelial membrane protein-2 (EMP2) regulates hypoxia-inducible factor (HIF)-induced vascular endothelial growth factor (VEGF) production in the ARPE-19 cell line and genetic knock-out of Emp2 in a murine oxygen-induced retinopathy (OIR) model attenuates neovascularization. We hypothesize that EMP2 blockade via intravitreal injection protects against neovascularization.
UNASSIGNED: Ex vivo choroid sprouting assay was performed, comparing media and human IgG controls versus anti-EMP2 antibody (Ab) treatment. In vivo, eyes from wild-type (WT) mice exposed to hyperoxia from postnatal (P) days 7 to 12 were treated with P12 intravitreal injections of control IgG or anti-EMP2 Abs. Neovascularization was assessed at P17 by flat mount imaging. Local and systemic effects of anti-EMP2 Ab treatment were assessed.
UNASSIGNED: Choroid sprouts treated with 30 µg/mL of anti-EMP2 Ab demonstrated a 48% reduction in vessel growth compared to control IgG-treated sprouts. Compared to IgG-treated controls, WT OIR mice treated with 4 µg/g of intravitreal anti-EMP2 Ab demonstrated a 42% reduction in neovascularization. They demonstrated down-regulation of retinal gene expression in pathways related to vasculature development and up-regulation in genes related to fatty acid oxidation and tricarboxylic acid cycle respiratory electron transport, compared to controls. Anti-EMP2 Ab-treated OIR mice did not exhibit gross retinal histologic abnormalities, vision transduction abnormalities, or weight loss.
UNASSIGNED: Our results suggest that EMP2 blockade could be a local and specific treatment modality for retinal neovascularization in oxygen-induced retinopathies, without systemic adverse effects.

BACKGROUND: Retinopathy of Prematurity (ROP) is a proliferative retinal vascular disease occurring in the retina of premature infants and is the main cause of childhood blindness. Nowadays anti-VEGF and retinal photocoagulation are mainstream treatments for ROP, but they develop a variety of complications. Hydrogen (H2) is widely considered as a useful neuroprotective and antioxidative therapeutic method for hypoxic-ischemic disease without toxic effects. However, whether H2 provides physiological angiogenesis promotion, neovascularization suppression and glial protection in the progression of ROP is largely unknown.This study aims to investigate the effects of H2 on retinal angiogenesis, neovascularization and neuroglial dysfunction in the retinas of oxygen-induced retinopathy (OIR) mice.
METHODS: In this study, mice that were seven days old and either wild-type (WT) or Nrf2-deficient (Nrf2-/-) were exposed to 75% oxygen for 5 days and then returned to normal air conditions. Different stages of hydrogen gas (H2) inhalation were administered. Vascular obliteration, neovascularization, and blood vessel leakage were analyzed and compared. To count the number of neovascularization endothelial nuclei, routine HE staining of retinal sections was conducted. Immunohistochemistry was performed using DyLight 594 labeled GSL I-isolectin B4 (IB4), as well as primary antibodies against proliferating cell nuclear antigen (PCNA), glial fibrillary acidic protein (GFAP), and Iba-1. Western blots were used to measure the expression of NF-E2-related factor 2 (Nrf2), vascular endothelial growth factor (VEGF), Notch1, Dll4, and HIF-1α. Additionally, the expression of target genes such as NQO1, HO-1, Notch1, Hey1, Hey2, and Dll4 was measured. Human umbilical vein endothelial cells (HUVECs) treated with H2 under hypoxia were used as an in vitro model. RT-PCR was used to evaluate the mRNA expression of Nrf2, Notch/Dll4, and the target genes. The expression of reactive oxygen species (ROS) was observed using immunofluorescence staining.
RESULTS: Our results indicate that 3-4% H2 does not disturb retinal physiological angiogenesis, but ameliorates vaso-obliteration and neovascularization in OIR mice. Moreover, H2 prevents the decreased density and reverses the morphologic and functional changes in retinal astrocytes caused by oxygen-induced injury. In addition, H2 inhalation reduces microglial activation, especially in the area of neovascularization in OIR mice. H2 plays a protective role in vascular regeneration by promoting Nrf2 activation and suppressing the Dll4-induced Notch signaling pathway in vivo. Also, H2 promotes the proliferation of HUVECs under hypoxia by negatively regulating the Dll4/Notch pathway and reducing ROS levels through Nrf2 pathway aligning with our findings in vivo.Moreover, the retinal oxygen-sensing mechanisms (HIF-1α/VEGF) are also involved in hydrogen-mediated retinal revascularization and neovascularization suppression.
CONCLUSIONS: Collectively, our results indicate that H2 could be a promising therapeutic agent for POR treatment and that its beneficial effect in human ROP might involve the activation of the Nrf2-Notch axis as well as HIF-1α/VEGF pathways.

OBJECTIVE: Diabetic retinopathy (DR) is a common complication of diabetes, and recent findings have shown that long noncoding RNAs (lncRNAs) may be involved in its pathogenesis. Through bioinformatics analysis, we found that lncRNA ATP2B2-IT2 may be involved in this process. This study primarily investigated the expression of the lncRNA ATP2B2-IT2 in human retinal microvascular endothelial cells (HRMECs) under high-glucose conditions and its effects on HRMEC proliferation, migration, and neovascularization.
METHODS: We used RT‒PCR to assess the expression levels of lncRNA ATP2B2-IT2 and vascular endothelial growth factor (VEGF) in HRMECs under normal glucose (5.5 mmol/L) and high glucose (30 mmol/L) conditions. HRMECs were subsequently divided into four groups: the normal glucose (NG), high glucose (HG), high glucose with lncRNA ATP2B2-IT2 silencing (HG + si-lncRNA ATP2B2-IT2), and high glucose with silencing control (HG + si-NC) groups. The expression levels of the lncRNA ATP2B2-IT2 and VEGF in each group were determined using RT‒PCR. Thereafter, cell proliferation, migration, and neovascularization were assessed using CCK-8, Transwell, and tube formation assays, respectively.
RESULTS: RT‒PCR revealed that the expression levels of the lncRNA ATP2B2-IT2 and VEGF were greater in the HG group than in the NG group (P < 0.05). After silencing of the lncRNA ATP2B2-IT2, the expression of VEGF decreased significantly (P < 0.05). Subsequent CCK-8, Transwell, and tube formation assays demonstrated that compared to those in the NG group, the HRMECs in the HG group exhibited significantly increased proliferation, migration, and neovascularization (P < 0.05). However, after silencing of the lncRNA ATP2B2-IT2, the proliferation, migration, and neovascularization of HRMECs were significantly decreased in the HG + si-lncRNA ATP2B2-IT2 group compared to those in the HG group (P < 0.05).
CONCLUSIONS: LncRNA ATP2B2-IT2 may promote the proliferation, migration and neovascularization of HRMECs under high-glucose conditions.

Fundus neovascularization diseases are a series of blinding eye diseases that seriously impair vision worldwide. Currently, the means of treating these diseases in clinical practice are continuously evolving and have rapidly revolutionized treatment opinions. However, key issues such as inadequate treatment effectiveness, high rates of recurrence, and poor patient compliance still need to be urgently addressed. Multifunctional nanomedicine can specifically respond to both endogenous and exogenous microenvironments, effectively deliver drugs to specific targets and participate in activities such as biological imaging and the detection of small molecules. Nano-in-micro (NIM) delivery systems such as metal, metal oxide and up-conversion nanoparticles (NPs), quantum dots, and carbon materials, have shown certain advantages in overcoming the presence of physiological barriers within the eyeball and are widely used in the treatment of ophthalmic diseases. Few studies, however, have evaluated the efficacy of NIM delivery systems in treating fundus neovascular diseases (FNDs). The present study describes the main clinical treatment strategies and the adverse events associated with the treatment of FNDs with NIM delivery systems and summarizes the anatomical obstacles that must be overcome. In this review, we wish to highlight the principle of intraocular microenvironment normalization, aiming to provide a more rational approach for designing new NIM delivery systems to treat specific FNDs.

This retrospective observational study aimed to investigate the difference in 4-year outcomes of ranibizumab or aflibercept therapy for macular neovascularization (MNV) with high myopia between pathologic myopia (PM) and non-PM. This study was conducted at Kyoto University Hospital and included consecutive treatment-naïve eyes with active myopic MNV, in which a single intravitreal ranibizumab or aflibercept injection was administered, followed by a pro re nata (PRN) regimen for 4 years. Based on the META-PM study classification, eyes were assigned to the non-PM and PM groups. This study analyzed 118 eyes of 118 patients (non-PM group, 19 eyes; PM group, 99 eyes). Baseline, 1-year, and 2-year best-corrected visual acuity (BCVA) were significantly better in the non-PM group (P = 0.02, 0.01, and 0.02, respectively); however, the 3-year and 4-year BCVA were not. The 4-year BCVA course was similar in both groups. However, the total number of injections over 4 years was significantly higher in the non-PM than in the PM group (4.6 ± 2.6 vs. 2.9 ± 2.6, P = 0.001). Four-year BCVA significantly correlated only with baseline BCVA in both non-PM (P = 0.047, β = 0.46) and PM groups (P < 0.001, β = 0.59). In conclusion, over the 4-year observation period, the BCVA course after anti-VEGF therapy for myopic MNV was similar in the eyes with non-PM and those with PM; however, more additional injections in a PRN regimen were required in the eyes with non-PM compared to those with PM. Thus, more frequent and careful follow-up is required for the eyes with non-PM compared with those with PM to maintain long-term BCVA.