a-murine-model-ischemic-retinal-injury-induced-transient-bilateral

A Murine Model of Ischemic Retinal Injury Induced by Transient Bilateral Common Carotid Artery Occlusion

Diverse vascular diseases such as diabetic retinopathy, occlusion of retinal veins or arteries and ocular ischemic syndrome can lead to retinal ischemia. ...

Keio University School of Medicine

Tokyo Animal Eye Clinic

Prevalence of myopia is increasing worldwide. Outdoor activity is one of the most important environmental factors for myopia control. Here we show that violet light (VL, 360-400nm wavelength) suppresses myopia progression. First, we confirmed that VL suppressed the axial length (AL) elongation in the chick myopia model. Expression microarray analyses revealed that myopia suppressive gene EGR1 was upregulated by VL exposure. VL exposure induced significantly higher upregulation of EGR1 in chick chorioretinal tissues than blue light under the same conditions. Next, we conducted clinical research retrospectively to compare the AL elongation among myopic children who wore eyeglasses (VL blocked) and two types of contact lenses (partially VL blocked and VL transmitting). The data showed the VL transmitting contact lenses suppressed myopia progression most. These results suggest that VL is one of the important outdoor environmental factors for myopia control. Since VL is apt to be excluded from our modern society due to the excessive UV protection, VL exposure can be a preventive strategy against myopia progression.

Violet Light Exposure Can Be a Preventive Strategy Against Myopia Progression.

Age-related macular degeneration and retinitis pigmentosa are leading causes of blindness and share a pathological feature, which is photoreceptor degeneration. To date, the lack of a potential treatment to prevent such diseases has raised great concern. Photoreceptor degeneration can be accelerated by excessive light exposure via an inflammatory response; therefore, anti-inflammatory agents would be candidates to prevent the progress of photoreceptor degeneration. We previously reported that a lactic acid bacterium,  KW3110 ( KW3110), activated macrophages suppressing inflammation in mice and humans. Recently, we also showed that intake of  KW3110 could mitigate visual display terminal (VDT) load-induced ocular disorders in humans. However, the biological mechanism of  KW3110 to retain visual function remains unclear. In this study, we found that  KW3110 activated M2 macrophages inducing anti-inflammatory cytokine interleukin-10 (IL-10) production in vitro using bone marrow-derived M2 macrophages. We also show that IL-10 gene expression was significantly increased in the intestinal immune tissues 6 h after oral administration of  KW3110 in vivo. Furthermore, we demonstrated that intake of  KW3110 suppressed inflammation and photoreceptor degeneration in a murine model of light-induced retinopathy. These results suggest that  KW3110 may have a preventive effect against degrative retinal diseases.

 KW3110 Prevents Blue Light-Induced Inflammation and Degeneration in the Retina.

To investigate meibomian gland (MG) morphology by noncontact infrared meibography in Shih Tzu dogs with or without keratoconjunctivitis sicca (KCS).

Assessment of meibomian gland morphology by noncontact infrared meibography in Shih Tzu dogs with or without keratoconjunctivitis sicca.

Neovascular retinal diseases are the leading causes of blindness in advanced countries. To date, anti-VEGF (vascular endothelial growth factor) drugs are clinically effective and widely used for these diseases. However, recent animal and clinical studies reported that potent and long-term VEGF antagonism may induce chorioretinal atrophy. Thus, physiological amount of VEGF is required for the homeostasis in the retina. Hypoxia-inducible factors (HIFs) are transcription factors located upstream of VEGF. We hypothesized that ectopically stabilized HIFs induce pathological amount of VEGF involved with retinal neovascularization. Therefore, HIF inhibition could be an alternative therapeutic candidate targeting the pathological amount of VEGF while holding a physiological amount of VEGF. To test this hypothesis, topotecan and doxorubicin, HIF inhibitors with different mechanisms were administered to the murine oxygen-induced retinopathy (OIR) model. We found that both topotecan and doxorubicin significantly prevented pathological but not physiological neovascularization in OIR. Furthermore, impaired visual function observed in OIR can also be suppressed by administering topotecan. These data suggested that HIF inhibition may be effective for pathological angiogenesis and neurodegeneration of the retina.

Pharmacological HIF inhibition prevents retinal neovascularization with improved visual function in a murine oxygen-induced retinopathy model.

Sphingosine-1-phosphate (S1P) is a lysophospholipid mediator, promoting angiogenesis and inflammation via interactions with its receptors (S1P1-5), but the receptors and signaling pathways responsible for the progression of choroidal neovascularization (CNV) remain unknown. We investigated the roles of S1P/S1P receptors in RPE cells. ARPE-19 cells were treated with S1P dissolved in carrier proteins of albumin or apolipoprotein M (ApoM). The mRNA expression levels of interleukin-8 (IL-8), C-C motif chemokine ligand 2 (CCL2), and vascular endothelial growth factor (VEGF) were evaluated using quantitative real-time polymerase chain reaction. The protein level of hypoxia-inducible factor (HIF)-1α was assessed via enzyme-linked immunosorbent assay. HIF transcriptional activity was evaluated with a dual-reporter luciferase assay. Cellular barrier integrity was evaluated using transepithelial electrical resistance and the FITC-dextran permeability assay. The suppressive effect of an S1P antagonist on CNV progression was investigated with a laser-induced CNV model in mice. The increase in expression of IL-8, CCL2, and VEGF due to albumin-bound S1P was significantly mitigated by an S1P2 antagonist. The expression of HIF-1α significantly decreased with inhibition of S1P2 and S1P3. In addition, albumin-bound S1P disrupted the barrier integrity of retinal pigment epithelial cells via S1P2, whereas integrity was strengthened by ApoM-bound S1P. CNV lesions were significantly reduced in the mouse model with intravitreal injection of S1P2 antagonist. This study demonstrated that S1P significantly promotes angiogenesis, inflammation, and barrier integrity, which was attenuated by inhibition of S1P2 or S1P3, suggesting that regulation of S1P2 and S1P3 is a novel therapeutic target for CNV.

The role of sphingosine 1-phosphate receptors on retinal pigment epithelial cells barrier function and angiogenic effects.

Age-related macular degeneration (AMD) is the leading cause of blindness and can be classified into two types called atrophic AMD (dry AMD) and neovascular AMD (wet AMD). Dry AMD is characterized by cellular degeneration of the retinal pigment epithelium, choriocapillaris, and photoreceptors. Wet AMD is characterized by the invasion of abnormal vessels from the choroid. Although anti-vascular endothelial growth factor (VEGF) therapy has a potent therapeutic effect against the disease, there is a possibility of chorio-retinal atrophy and adverse systemic events due to long-term robust VEGF antagonism. We focused on hypoxia-inducible factor (HIF) regulation of VEGF transcription, and report the suppressive effects of HIF inhibition against ocular phenotypes in animal models. Many of the known HIF inhibitors are categorized as anti-cancer drugs, and their systemic side effects are cause for concern in clinical use. In this study, we explored food ingredients that have HIF inhibitory effects and verified their effects in an animal model of AMD.

Therapeutic Effect of  Extract and Hydroxycitric Acid Inhibiting Hypoxia-Inducible Factor in a Murine Model of Age-Related Macular Degeneration.

To evaluate electroretinogram (ERG) responses under anesthesia with midazolam, medetomidine, and butorphanol tartrate (MMB) combination compared with pentobarbital sodium and ketamine/xylazine (KX).

Effect of midazolam, medetomidine, and butorphanol tartrate combination anesthetic on electroretinograms of mice.

Lactoferrin, a type of glycoprotein, is contained in exocrine fluids such as tears, breast milk, sweat, and saliva, and is known to have anti-microbial, antioxidant, and anti-cancer effects. In the ophthalmological field, topical administration of lactoferrin has been reported to have a therapeutic effect in a murine dry eye model. Hypoxia-inducible factor (HIF) regulates various gene expressions under hypoxia, including vascular endothelial growth factor (VEGF), and is considered as an alternative target for neovascular ocular diseases such as age-related macular degeneration (AMD). We previously screened natural products and identified lactoferrin as a novel HIF inhibitor. In this study, we confirmed that lactoferrin has an HIF inhibitory effect and a therapeutic effect in a murine model of neovascular AMD.

Lactoferrin Has a Therapeutic Effect  HIF Inhibition in a Murine Model of Choroidal Neovascularization.

Neovascular retinal degenerative diseases are the leading causes of blindness in developed countries. Anti-vascular endothelial growth factor (VEGF) therapy is commonly used to treat these diseases currently. However, recent reports indicate that long term suppression of VEGF in the eye is associated with chorioretinal atrophy. Therefore, a physiological amount of VEGF is required for retinal homeostasis. Hypoxia-inducible factor (HIF) is a transcriptional factor upstream of VEGF. We previously reported that HIF regulated pathological angiogenesis in the retina of murine models of oxygen-induced retinopathy and laser-induced choroidal neovascularization. Most of the known HIF inhibitors are anti-cancer agents which may have systemic adverse effects in for clinical use; thus, there is a need for safer and less invasive HIF inhibitors. In this study, we screened marine products, especially fish ingredients, and found that six species of fish had HIF inhibitory effects. Among them, administration of  ingredients significantly suppressed retinal neovascular tufts by inhibiting HIF expression in a murine oxygen-induced retinopathy model. These results indicate that particular fish ingredients can act as anti-angiogenic agents in retinal neovascularization diseases.

Hypoxia-Inducible Factor Inhibitors Derived from Marine Products Suppress a Murine Model of Neovascular Retinopathy.

Diabetic retinopathy (DR) is one of the leading causes of blindness globally. Retinal neuronal abnormalities occur in the early stage in DR. Therefore, maintaining retinal neuronal activity in DR may prevent vision loss. Previously, pemafibrate, a novel selective peroxisome proliferator-activated receptor alpha modulator, was suggested as a promising drug in hypertriglyceridemia. However, the role of pemafibrate remains obscure in DR. Therefore, we aimed to unravel systemic and retinal changes by pemafibrate in diabetes. Adult mice were intraperitoneally injected with streptozotocin (STZ) to induce diabetes. After STZ injection, diet supplemented with pemafibrate was given to STZ-induced diabetic mice for 12 weeks. During the experiment period, body weight and blood glucose levels were examined. Electroretinography was performed to check the retinal neural function. After sacrifice, the retina, liver, and blood samples were subjected to molecular analyses. We found pemafibrate mildly improved blood glucose level as well as lipid metabolism, boosted liver function, increased serum fibroblast growth factor21 level, restored retinal functional deficits, and increased retinal synaptophysin protein expression in STZ-induced diabetic mice. Our present data suggest a promising pemafibrate therapy for the prevention of early DR by improving systemic metabolism and protecting retinal function.

Pemafibrate Protects Against Retinal Dysfunction in a Murine Model of Diabetic Retinopathy.

To compare the superficial punctate fluorescein staining in dogs with and without aqueous tear deficiency.

Punctate fluorescein staining scores in dogs with or without aqueous tear deficiency.

Neovascular retinal degeneration is a leading cause of blindness in advanced countries. Anti-vascular endothelial growth factor (VEGF) drugs have been used for neovascular retinal diseases; however, anti-VEGF drugs may cause the development of chorioretinal atrophy in chronic therapy as they affect the physiological amount of VEGF needed for retinal homeostasis. Hypoxia-inducible factor (HIF) is a transcription factor inducing VEGF expression under hypoxic and other stress conditions. Previously, we demonstrated that HIF was involved with pathological retinal angiogenesis in murine models of oxygen-induced retinopathy (OIR), and pharmacological HIF inhibition prevented retinal neovascularization by reducing an ectopic amount of VEGF. Along with this, we attempted to find novel effective HIF inhibitors. Compounds originally isolated from mushroom-forming fungi were screened for prospective HIF inhibitors utilizing cell lines of 3T3, ARPE-19 and 661W. A murine OIR model was used to examine the anti-angiogenic effects of the compounds. As a result, 2-azahypoxanthine (AHX) showed an inhibitory effect on HIF activation and suppressed  mRNA upregulation under CoCl-induced pseudo-hypoxic conditions. Oral administration of AHX significantly suppressed retinal neovascular tufts in the OIR model. These data suggest that AHX could be a promising anti-angiogenic agent in retinal neovascularization by inhibiting HIF activation.

Article	Year
Violet Light Exposure Can Be a Preventive Strategy Against Myopia Progression. EBioMedicine\| PubMed ID: 28063778	2017
KW3110 Prevents Blue Light-Induced Inflammation and Degeneration in the Retina. Nutrients\| PubMed ID: 30558320	2018
Assessment of meibomian gland morphology by noncontact infrared meibography in Shih Tzu dogs with or without keratoconjunctivitis sicca. Veterinary ophthalmology\| PubMed ID: 30715776	2019
Pharmacological HIF inhibition prevents retinal neovascularization with improved visual function in a murine oxygen-induced retinopathy model. Neurochemistry international\| PubMed ID: 30872009	2019
The role of sphingosine 1-phosphate receptors on retinal pigment epithelial cells barrier function and angiogenic effects. Prostaglandins & other lipid mediators\| PubMed ID: 31415870	2019
Therapeutic Effect of Extract and Hydroxycitric Acid Inhibiting Hypoxia-Inducible Factor in a Murine Model of Age-Related Macular Degeneration. International journal of molecular sciences\| PubMed ID: 31614647	2019
Effect of midazolam, medetomidine, and butorphanol tartrate combination anesthetic on electroretinograms of mice. Molecular vision\| PubMed ID: 31741653	2019
Lactoferrin Has a Therapeutic Effect HIF Inhibition in a Murine Model of Choroidal Neovascularization. Frontiers in pharmacology\| PubMed ID: 32180725	2020
Hypoxia-Inducible Factor Inhibitors Derived from Marine Products Suppress a Murine Model of Neovascular Retinopathy. Nutrients\| PubMed ID: 32290307	2020
Pemafibrate Protects Against Retinal Dysfunction in a Murine Model of Diabetic Retinopathy. International journal of molecular sciences\| PubMed ID: 32872333	2020
Punctate fluorescein staining scores in dogs with or without aqueous tear deficiency. Veterinary ophthalmology\| PubMed ID: 32961030	2020
A Fairy Chemical Suppresses Retinal Angiogenesis as a HIF Inhibitor. Biomolecules\| PubMed ID: 33020402	2020

Yukihiro Miwa

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