論文 - 栗原　俊英

Hypoxia-Inducible Factor (HIF)/Vascular Endothelial Growth Factor (VEGF) Signaling in the Retina.

栗原 俊英

Adv Exp Med Biol. （SPRINGER-VERLAG BERLIN）  801   275 - 281 2014年

研究論文（学術雑誌）, 査読有り,  ISSN  0065-2598

　概要を見る

Over a span of two decades, it has become increasingly clear that vascular endothelial growth factor (VEGF) plays an important role in the pathogenesis of retinal diseases including age-related macular degeneration (AMD) and diabetic retinopathy (DR). Based on these observations, anti-VEGF therapies are being developed and approved for clinical use in the treatment of neovascular eye diseases. Hypoxia-inducible factors (HIFs) are transcriptional factors that are stabilized and activated under hypoxic conditions and induce expression of gene products, including VEGF, that are required for cell survival under hypoxia. Here we discuss recent findings from our lab and others that define roles of the HIF-VEGF axis in the retina.

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Utilizing Stem Cell-Derived RPE Cells as A Therapeutic Intervention for Age-Related Macular Degeneration.

栗原 俊英

Adv Exp Med Biol. （SPRINGER-VERLAG BERLIN）  801   323 - 329 2014年

研究論文（学術雑誌）, 査読有り,  ISSN  0065-2598

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Degeneration or dysfunction of the retinal pigment epithelium (RPE) can induce secondary photoreceptor atrophy and catastrophic vision loss in patients with agerelated macular degeneration (AMD). AMD is the leading cause of vision loss in the elderly in industrialized countries and no cure exists for the "dry" or atrophic form to date. However, recent pre-clinical data from several groups suggests that embryonic stem cell-derived RPE cell transplantation may prevent photoreceptor degeneration in animal models of RPE degeneration. Another approach may be to derive RPE cells from autologous induced pluripotent stem cells (iPSCs) reprogrammed from dermal tissue. However, the safety of this approach has been questioned on several levels. In this chapter we will summarize work reported by several groups, including our own, that clearly demonstrate that transplanted RPE cells can provide anatomical and functional photoreceptor rescue in animal models of retinal degeneration. We will also discuss some of the prevailing concerns and challenges associated with this technique.

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Ras pathway inhibition prevents neovascularization by repressing endothelial cell sprouting.

栗原 俊英

J Clin Invest. （AMER SOC CLINICAL INVESTIGATION INC）  123 （ 11 ） 4900 - 4908 2013年11月

研究論文（学術雑誌）, 査読有り,  ISSN  0021-9738

　概要を見る

Vascular networks develop from a growing vascular front that responds to VEGF and other guidance cues. Angiogenesis is required for normal tissue function, but, under conditions of stress, inappropriate vascularization can lead to disease. Therefore, inhibition of angiogenic sprouting may prevent neovascularization in patients with blinding neovascular eye diseases, including macular degeneration. VEGF antagonists have therapeutic benefits but also can elicit off-target effects. Here, we found that the Ras pathway, which functions downstream of a wide range of cytokines including VEGF, is active in the growing vascular front of developing and pathological vascular networks. The endogenous Ras inhibitor p120RasGAP was expressed predominately in quiescent VEGF-insensitive endothelial cells and was ectopically downregulated in multiple neovascular models. MicroRNA-132 negatively regulated p120RasGAP expression. Experimental delivery of alpha-miR-132 to developing mouse eyes disrupted tip cell Ras activity and prevented angiogenic sprouting. This strategy prevented ocular neovascularization in multiple rodent models even more potently than the VEGF antagonist, VEGF-trap. Targeting microRNA-132 as a therapeutic strategy may prove useful for treating multiple neovascular diseases of the eye and for preventing vision loss regardless of the neovascular stimulus.

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VEGF antagonism and age-related macular degeneration: too much of a good thing?

Kurihara T, Westenskow PD, Aguilar E, Friedlander M†

Exp Rev Ophthalmol. 8 （ 2 ） 103 - 105 2013年04月

筆頭著者

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Targeted deletion of Vegfa in adult mice induces vision loss.

栗原 俊英

J Clin Invest. （AMER SOC CLINICAL INVESTIGATION INC）  122 （ 11 ） 4213 - 4217 2012年11月

研究論文（学術雑誌）, 査読有り,  ISSN  0021-9738

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Current therapies directed at controlling vascular abnormalities in cancers and neovascular eye diseases target VEGF and can slow the progression of these diseases. While the critical role of VEGF in development has been well described, the function of locally synthesized VEGF in the adult eye is incompletely understood. Here, we show that conditionally knocking out Vegfa in adult mouse retinal pigmented epithelial (RPE) cells, which regulate retinal homeostasis, rapidly leads to vision loss and ablation of the choriocapillaris, the major blood supply for the outer retina and photoreceptor cells. This deletion also caused rapid dysfunction of cone photoreceptors, the cells responsible for fine visual acuity and color vision. Furthermore, Vegfa deletion showed significant downregulation of multiple angiogenic genes in both physiological and pathological states, whereas the deletion of the upstream regulatory transcriptional factors HIFs did not affect the physiological expressions of angiogenic genes. These results suggest that endogenous VEGF provides critical trophic support necessary for retinal function. Targeting factors upstream of VEGF, such as HIFs, may be therapeutically advantageous compared with more potent and selective VEGF antagonists, which may have more off-target inhibitory trophic effects.

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Using Flow Cytometry to Compare the Dynamics of Photoreceptor Outer Segment Phagocytosis in iPS-Derived RPE Cells.

栗原 俊英

Invest Ophthalmol Vis Sci. （ASSOC RESEARCH VISION OPHTHALMOLOGY INC）  53 （ 10 ） 6282 - 6290 2012年09月

研究論文（学術雑誌）, 査読有り,  ISSN  0146-0404

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PURPOSE. Retinal pigment epithelium (RPE) autologous grafts can be readily derived from induced pluripotent stem (iPS) cells. It is critical to stringently characterize iPS-RPE using standardized and quantifiable methods to be confident that they are safe and adequate replacements for diseased RPE before utilizing them in clinical settings. One important and required function is that the iPS-RPE phagocytose photoreceptor outer segments (POS).
METHODS. We developed a flow cytometry-based assay to monitor binding and internalization of FITC labeled POS by ARPE-19, human fetal RPE (hfRPE), and two types of iPS-RPE. Expression and density of alpha(v)beta(5) integrin, CD36, and MerTK receptors, which are required for phagocytosis, were compared.
RESULTS. Trypsinization of treated RPE cells results in the release of bound POS. The number of freed POS, the percentage of cells that internalized POS, the brightness of the FITC signal from the cells, and the surface density of the phagocytosis receptors on single RPE cells were measured using flow cytometry. These assays reveal that receptor density is dynamic during differentiation and this can affect the binding and internalization dynamics of the RPE cells. Highly differentiated iPS-RPE phagocytose POS more efficiently than hfRPE.
CONCLUSIONS. Caution should be exercised to not use RPE grafts until demonstrating that they are fully functional. The density of the phagocytosis receptors is dynamic and may be used as a predictor for how well the iPS-RPE cells will function in vivo. The phagocytosis dynamics observed between iPS-RPE and primary RPE is very encouraging and adds to mounting evidence that iPS-RPE may be a viable replacement for dysfunctional or dying RPE in human patients. (Invest Ophthalmol Vis Sci. 2012;53:6282-6290) DOI:10.1167/iovs.12-9721

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Biological role of lutein in the light-induced retinal degeneration.

栗原 俊英

J Nutr Biochem. （ELSEVIER SCIENCE INC）  23 （ 5 ） 423 - 429 2012年05月

研究論文（学術雑誌）, 査読有り,  ISSN  0955-2863

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Lutein, a xanthophyll of a carotenoid, is anticipated as a therapeutic product to prevent human eye diseases. However, its biological mechanism is still unclear. Here, we show the molecular mechanism of lutein's effect to reduce photodamage of the retina. We analyzed the light-exposed retinas of Balb/c mice given lutein-supplemented or normal diet. Visual function was measured by electroretinogram, and histological changes were observed. Immunohistochemical and immunoblot analyses were performed to analyze molecular mechanism. The reactive oxygen species induced in the retina was evaluated by fluorescent probes. In the mice after light exposure, reduction of a-wave and b-wave amplitudes in electroretinogram, indicating visual impairment, and thinning of the photoreceptor cell layer owing to apoptosis were both attenuated by lutein diet. Interestingly, gamma-H2AX, a marker for double-strand breaks (DSBs) in DNA, was up-regulated in the photoreceptor cells after light exposure, but this increase was attenuated by lutein diet, suggesting that DSBs caused by photodamage contributed to the photoreceptor cell death and that this change was suppressed by lutein. Moreover, the expression of eyes absent (EYA), which promotes DNA repair and cell survival, was significantly up-regulated with lutein diet in the light-exposed retina. Therefore, lutein induced EYA for DNA repair, which could suppress DNA damage and photoreceptor cell apoptosis. Lutein reduced light-induced oxidative stress in the retina, which might contribute to promote DNA repair. The lutein-supplemented diet attenuated light-induced visual impairment by protecting the photoreceptor cells' DNA. (C) 2012 Elsevier Inc. All rights reserved.

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Renin-Angiotensin System Hyperactivation Can Induce Inflammation and Retinal Neural Dysfunction.

Kurihara T†, Ozawa Y, Ishida S, Okano H, Tsubota K

Int J Inflam. 2012   581695 2012年03月

筆頭著者, 責任著者, 査読有り

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The formation of an angiogenic astrocyte template is regulated by the neuroretina in a HIF-1-dependent manner.

栗原 俊英

Dev Biol. （ACADEMIC PRESS INC ELSEVIER SCIENCE）  363 （ 1 ） 106 - 114 2012年03月

研究論文（学術雑誌）, 査読有り,  ISSN  0012-1606

　概要を見る

The vascular and nervous systems display a high degree of cross-talk and depend on each other functionally. In the vascularization of the central nervous system, astrocytes have been thought to sense tissue oxygen levels in hypoxia-inducible factors (HIFs)-dependent manner and control the vascular growth into the hypoxic area by secreting VEGF. However, recent genetic evidences demonstrate that not only astrocyte HIFs but also astrocyte VEGF expression is dispensable for developmental angiogenesis of the retina. This study demonstrates that hypoxia-inducible factor 1 alpha subunit (HIF-1 alpha), a key transcription factor involved in cellular responses to hypoxia, is most abundantly expressed in the neuroretina, especially retinal progenitor cells (RPCs). A neuroretina-specific knockout of HIF-1 alpha (alpha Cre(+)Hif1 alpha(flox/flox)) showed impaired vascular development characterized by decreased tip cell filopodia and reduced vessel branching. The astrocyte network was hypoplastic in alpha Cre(+)Hif1 alpha(flox/flox) mice. Mechanistically, platelet-derived growth factor A (PDGF-A), a mitogen for astrocytes, was downregulated in the neuroretina of alpha Cre(+)Hif1 alpha(flox/flox) mice. Supplementing PDGF-A restored reduced astrocytic and vascular density in alpha Cre(+)Hif1 alpha(flox/flox) mice. Our data demonstrates that the neuroretina but not astrocytes acts as a primary oxygen sensor which ultimately controls the retinal vascular development by regulating an angiogenic astrocyte template. (C) 2011 Elsevier Inc. All rights reserved.

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Generation of Retinal Pigment Epithelial Cells from Small Molecules and OCT4 Reprogrammed Human Induced Pluripotent Stem Cells.

栗原 俊英

Stem Cells Transl Med. （ALPHAMED PRESS）  1 （ 2 ） 96 - 109 2012年02月

研究論文（学術雑誌）, 査読有り,  ISSN  2157-6564

　概要を見る

Autologous retinal pigment epithelium (RPE) grafts derived from induced pluripotent stem cells (iPSCs) may be used to cure blinding diseases in which RPE dysfunction results in photoreceptor degeneration. Four-, two-, and one-factor-derived iPSCs (4F-, 2F-, and 1F-iPSC5, respectively) were differentiated into fully functional cuboidal pigmented cells in polarized monolayers that express RPE-specific markers. 1F-iPSC-RPE (1F-iPS-RPE) strongly resembles primary human fetal RPE (hfRPE) based on proteomic and untargeted metabolomic analyses, and using novel in vivo imaging technology coupled with electroretinography, we demonstrated that 1F-iPS-RPE mediate anatomical and functional rescue of photoreceptors after transplantation in an animal model of RPE-mediated retinal degeneration. 1F-iPS-RPE cells were injected subretinally as a suspension and formed a monolayer dispersed between host RPE cells. Furthermore, 1F-iPS-RPE do not simply provide trophic support to rescue photoreceptors as previously speculated but actually phagocytose photoreceptor outer segments in vivo and maintain visual cycling. Thus, 1F-iPS-RPE grafts may be superior to conventional iPS-RPE for clinical use because 1F-iPS-RPE closely resemble hfRPE, mediate anatomical and functional photoreceptor rescue in vivo, and are generated using a reduced number of potentially oncogenic reprogramming factors. STEM CELLS TRANSLATIONAL MEDICINE 2012;1:96-109

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Roles of AMP-Activated Protein Kinase in Diabetes-Induced Retinal Inflammation.

栗原 俊英

Invest Ophthalmol Vis Sci. （ASSOC RESEARCH VISION OPHTHALMOLOGY INC）  52 （ 12 ） 9142 - 9148 2011年11月

研究論文（学術雑誌）, 査読有り,  ISSN  0146-0404

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PURPOSE. AMP-activated protein kinase (AMPK) is a sensor of cellular energy status. The purpose of the present study was to elucidate the roles of AMPK in the pathogenesis of diabetic retinopathy using the known AMPK activators resveratrol and AICAR (5-aminoimidazole-4-carboxamide ribonucleoside) in a mouse model.
METHODS. C57BL/6 mice with streptozotocin-induced diabetes were treated with resveratrol orally at 50 mg/kg for 7 days or with AICAR intraperitoneally at 100 mg/kg 24 hours before death. Retinal protein levels of phosphorylated and total AMPK, phosphorylated nuclear factor (NF)-kappa B p65, intercellular adhesion molecule (ICAM)-1, and vascular endothelial growth factor (VEGF) were evaluated by Western blot analysis or enzyme-linked immunosorbent assay. Retinal activity of sirtuin (SIRT) 1 was measured by deacetylase fluorometric assay. Leukocyte adhesion to the retinal vasculature was examined with a concanavalin A lectin perfusion-labeling technique.
RESULTS. Induction of diabetes in mice led to retinal AMPK dephosphorylation, which was significantly reversed by either resveratrol or AICAR. Either resveratrol or AICAR significantly reversed SIRT1 deactivation and NF-kappa B phosphorylation, both of which were induced in the diabetic retina. Administration of resveratrol to diabetic mice significantly reduced diabetes-induced retinal leukocyte adhesion, together with retinal expression of ICAM-1 and VEGF.
CONCLUSIONS. The present findings reveal that diabetes-induced retinal inflammation stems from downregulation of the AMPK pathway, leading subsequently to SIRT1 deactivation and NF-kappa B activation. The data also suggest the potential use of the AMPK activator resveratrol as a therapeutic agent for diabetic retinopathy. (Invest Ophthalmol Vis Sci. 2011;52:9142-9148) DOI: 10.1167/iovs.11-8041

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Astrocyte pVHL and HIF-α isoforms are required for embryonic-to-adult vascular transition in the eye.

栗原 俊英

J Cell Biol. （4）  195 （ 4 ） 689 - 701 2011年11月

研究論文（学術雑誌）, 査読有り,  ISSN  0021-9525

　概要を見る

Successful transition from embryonic to adult circulation is critical for survival of mammalian organisms. This shift occurs in the central cardiovascular circulation and in the eye as oxygen tension increases. However, its regulation is not well understood. We have used combinatorial gene deletion and overexpression assays to assess the effect of astrocyte-targeted deletion of von Hippel-Lindau tumor suppressor (Vhl), hypoxia-inducible factor-alpha s (Hif-alpha s), and Vegf on the normal regression of the hyaloidal vessels, the fetal ocular circulation system. Astrocytic Vhl deletion induced accelerated hyaloidal regression and subsequent massive secondary outgrowth. Combinatorial gene deletion involving Vhl, Hif-alpha s, and Vegf genes revealed that HIF-2 alpha/vascular endothelial growth factor signaling induces secondary outgrowth in Vhl mutants. Conversely, HIF-1 alpha regulated macrophage migration inhibitory factor and promoted macrophage infiltration that accelerates hyaloidal vessel regression. The phenotype observed in Vhl mutants strongly resembles human persistent hyperplastic primary vitreous cases and may provide insights into vascular remodeling mechanisms in other systems.

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Retinal Ganglion Cell Loss in Superoxide Dismutase 1 Deficiency.

栗原 俊英

Invest Ophthalmol Vis Sci. （ASSOC RESEARCH VISION OPHTHALMOLOGY INC）  52 （ 7 ） 4143 - 4150 2011年06月

研究論文（学術雑誌）, 査読有り,  ISSN  0146-0404

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PURPOSE. To investigate the influence of deficiency in superoxide dismutase (SOD) 1, a major antioxidative enzyme, on retinal ganglion cells (RGCs).
METHODS. In the SOD1 total knockout (SOD1-deficient) mice, the level of superoxide anion was measured using dihydroethidium. The number of RGCs was counted in both the retinal sections and the flat-mount retinas after retrograde labeling. Thickness of nerve fiber layer (NFL) was measured in the sections, and the amount of neurofilament protein was measured by immunoblot analysis. Pattern electroretinogram (ERG), which reflects the function of retinal ganglion cells, dark-adapted ERG, and cone ERG were performed. The intraocular pressure (IOP) was measured with an induction-impact tonometer. The levels of SOD-1 and -2 were measured by ELISA, in the serum of 47 newly diagnosed consecutive normal tension glaucoma (NTG) patients and 44 consecutive control subjects.
RESULTS. The level of superoxide anion in the RGC layer was significantly higher in 24-week-old SOD1-deficient mice than in wild-type mice. The RGC number was significantly reduced in 24-week-old SOD1-deficient mice, although they were not in 8-week-old mice. The NFL thickness and neurofilament protein were reduced in 24-week-old SOD1-deficient mice. The amplitude of pattern ERG was significantly reduced, although dark-adapted and cone ERGs showed no impairment, in 24-week-old SOD1-deficient mice. The IOP level was not changed in the SOD1-deficient mice. The serum level of SOD1, but not SOD2, was significantly lower in the NTG patients than in the healthy controls.
CONCLUSIONS. SOD1 deficiency causes RGC vulnerability, which may be involved in the underlying condition of NTG. (Invest Ophthalmol Vis Sci. 2011;52:4143-4150) DOI:10.1167/iovs.106294

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Hydrogen and N-Acetyl-L-Cysteine Rescue Oxidative Stress-Induced Angiogenesis in a Mouse Corneal Alkali-Burn Model.

栗原 俊英

Invest Ophthalmol Vis Sci. （ASSOC RESEARCH VISION OPHTHALMOLOGY INC）  52 （ 1 ） 427 - 433 2011年01月

研究論文（学術雑誌）, 査読有り,  ISSN  0146-0404

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PURPOSE. To investigate the role of reactive oxygen species (ROS) as the prime initiators of the angiogenic response after alkali injury of the cornea and observe the effects of antioxidants in preventing angiogenesis.
METHODS. The corneal epithelia of SOD-1-deficient mice or wild-type (WT) mice were removed after application of 0.15 N NaOH to establish the animal model of alkali burn. ROS production was semiquantitatively measured by dihydroethidium (DHE) fluorescence. Angiogenesis was visualized by CD31 immunohistochemistry. The effects of the specific NF-kappa B inhibitor DHMEQ, the antioxidant N-acetyl-L-cysteine (NAC), and hydrogen (H(2)) solution were observed.
RESULTS. ROS production in the cornea was enhanced immediately after alkali injury, as shown by increased DHE fluorescence (P < 0.01). NF-kappa B activation and the upregulation of vascular endothelial growth factor (VEGF) and monocyte chemoattractant protein-1 (MCP-1) were significantly enhanced (P < 0.01), leading to a significantly larger area of angiogenesis. Angiogenesis in SOD-1(-/-) mice corneas were significantly higher in WT mice (P < 0.01), confirming the role of ROS. Pretreatment with the specific NF-kappa B inhibitor DHMEQ or the antioxidant NAC significantly reduced corneal angiogenesis by downregulating the NF-kappa B pathway (P < 0.01) in both WT and SOD-1(-/-) mice. Furthermore, we showed that irrigation of the cornea with hydrogen (H 2) solution significantly reduced angiogenesis after alkali-burn injury (P < 0.01).
CONCLUSIONS. Immediate antioxidant therapy with H(2)-enriched irrigation solution is a new potent treatment of angiogenesis in cornea to prevent blindness caused by alkali burn. (Invest Ophthalmol Vis Sci. 2011;52:427-433) DOI:10.1167/iovs.106167

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Neural Degeneration in the Retina of the Streptozotocin-Induced Type 1 Diabetes Model.

栗原 俊英

J Diabetes Res. （HINDAWI PUBLISHING CORPORATION）  2011   1433 - 1439 2011年

研究論文（学術雑誌）, 査読有り,  ISSN  1687-5214

　概要を見る

Diabetic retinopathy, a vision-threatening disease, has been regarded as a vascular disorder. However, impaired oscillatory potentials (OPs) in the electroretinogram (ERG) and visual dysfunction are recorded before severe vascular lesions appear. Here, we review the molecular mechanisms underlying the retinal neural degeneration observed in the streptozotocin-(STZ-) induced type 1 diabetes model. The renin-angiotensin system (RAS) and reactive oxygen species (ROS) both cause OP impairment and reduced levels of synaptophysin, a synaptic vesicle protein for neurotransmitter release, most likely through excessive protein degradation by the ubiquitin-proteasome system. ROS also decrease brain-derived neurotrophic factor (BDNF) and inner retinal neuronal cells. The influence of both RAS and ROS on synaptophysin suggests that RAS-ROS crosstalk occurs in the diabetic retina. Therefore, suppressors of RAS or ROS, such as angiotensin II type 1 receptor blockers or the antioxidant lutein, respectively, are potential candidates for neuroprotective and preventive therapies to improve the visual prognosis.

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Regulation of Posttranscriptional Modification as a Possible Therapeutic Approach for Retinal Neuroprotection.

栗原 俊英

J Ophthalmol. （HINDAWI LTD）  2011   663 - 665 2011年

研究論文（学術雑誌）, 査読有り,  ISSN  2090-004X

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Understanding pathogenesis at the molecular level is the first step toward developing new therapeutic approaches. Here, we review the molecular mechanisms of visual dysfunction in two common diseases, innate chorioretinal inflammation and diabetic retinopathy, and the role of the ubiquitin-proteasome system (UPS) in both processes. In innate chorioretinal inflammation, interleukin-6 family ligands induce STAT3 activation in photoreceptors, which causes UPS-mediated excessive degradation of the visual substance, rhodopsin. In diabetic retinopathy, angiotensin II type 1 receptor (AT1R) signaling activates ERK in the inner layers of the retina, causing UPS-mediated excessive degradation of the synaptic vesicle protein, synaptophysin. This latter effect may decrease synaptic activity, in turn adversely affecting neuronal survival. Both mechanisms involve increased UPS activity and the subsequent excessive degradation of a protein required for visual function. Finally, we review the therapeutic potential of regulating the UPS to protect tissue function, citing examples from clinical applications in other medical fields.

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Resveratrol prevents light-induced retinal degeneration via suppressing activator protein-1 activation.

栗原 俊英

Am J Pathol. （AMER SOC INVESTIGATIVE PATHOLOGY, INC）  177 （ 4 ） 1725 - 1731 2010年10月

研究論文（学術雑誌）, 査読有り,  ISSN  0002-9440

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Light damage to the retina accelerates retinal degeneration in human diseases and rodent models. Recently, the polyphenolic phytoalexin resveratrol has been shown to exert various bioactivities in addition to its classical antioxidant property. In the present study, we investigated the effect of resveratrol on light-induced retinal degeneration together with its underlying molecular mechanisms. BALB/c mice with light exposure (5000-lux white light for 3 hours) were orally pretreated with resveratrol at a dose of 50 mg/kg for 5 days. Retinal damage was evaluated by TdT-mediated dUTP nick-end labeling, outer nuclear layer morphometry, and electroretinography. Administration of resveratrol to mice with light exposure led to a significant suppression of light-induced pathological parameters, including TdT-mediated dUTP nick-end labeling-positive retinal cells, outer nuclear layer thinning, and electroretinography changes. To clarify the underlying molecular mechanisms, the nuclear translocation of activator protein-1 subunit c-fos was evaluated by enzyme-linked immunosorbent assay, and the retinal activity of sirtuin 1 was measured by deacetylase fluorometric assay. Retinal activator protein-1 activation, up-regulated following light exposure, was significantly reduced by application of resveratrol. In parallel, retinal sirtuin 1 activity, reduced in animals with light damage, was significantly augmented by resveratrol treatment. Our data suggest the potential use of resveratrol as a therapeutic agent to prevent retinal degeneration related to light damage. (Am J Pathol 2010, 1 77:1725-1731; DOI: 10.2353/ajpath.2010.100098)

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Astrocyte Hypoxic Response Is Essential for Pathological But Not Developmental Angiogenesis of the Retina.

栗原 俊英

Glia. （WILEY-LISS）  58 （ 10 ） 1177 - 1185 2010年08月

研究論文（学術雑誌）, 査読有り,  ISSN  0894-1491

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Vascular/parenchymal crosstalk is increasingly recognized as important in the development and maintenance of healthy vascularized tissues. The retina is an excellent model in which to study the role of cell type-specific contributions to the process of blood vessel and neuronal growth. During retinal vascular development, glial cells such as astrocytes provide the template over which endothelial cells migrate to form the retinal vascular network, and hypoxia-regulated vascular endothelial growth factor (VEGF) has been demonstrated to play a critical role in this process as well as pathological neovascularization. To investigate the nature of cell-specific contributions to this process, we deleted VEGF and its upstream regulators, the hypoxia-inducible transcription factors HIF-1 alpha and HIF-2 alpha, and the negative regulator of HIF alpha, von Hippel Lindau protein (VHL), in astrocytes. We found that loss of hypoxic response and VEGF production in astrocytes does not impair normal development of retinal vasculature, indicating that astrocyte-derived VEGF is not essential for this process. In contrast, using a model of oxygen-induced ischemic retinopathy, we show that astrocyte-derived VEGF is essential for hypoxia-induced neovascularization. Thus, we demonstrate that astrocytes in the retina have highly divergent roles during developmental, physiological angiogenesis, and ischemia-driven, pathological neovascularization. (C) 2010 Wiley-Liss, Inc.

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von Hippel-Lindau protein regulates transition from the fetal to the adult circulatory system in retina.

栗原 俊英

Development. （COMPANY OF BIOLOGISTS LTD）  137 （ 9 ） 1563 - 1571 2010年05月

研究論文（学術雑誌）, 査読有り,  ISSN  0950-1991

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In early neonates, the fetal circulatory system undergoes dramatic transition to the adult circulatory system. Normally, embryonic connecting vessels, such as the ductus arteriosus and the foramen ovale, close and regress. In the neonatal retina, hyaloid vessels maintaining blood flow in the embryonic retina regress, and retinal vessels take over to form the adult-type circulatory system. This process is regulated by a programmed cell death switch mediated by macrophages via Wnt and angiopoietin 2 pathways. In this study, we seek other mechanisms that regulate this process, and focus on the dramatic change in oxygen environment at the point of birth. The von Hippel-Lindau tumor suppressor protein (pVHL) is a substrate recognition component of an E3-ubiquitin ligase that rapidly destabilizes hypoxia-inducible factor alpha s (HIF-alpha s) under normoxic, but not hypoxic, conditions. To examine the role of oxygen-sensing mechanisms in retinal circulatory system transition, we generated retina-specific conditional-knockout mice for VHL (Vhl(alpha-CreKO) mice). These mice exhibit arrested transition from the fetal to the adult circulatory system, persistence of hyaloid vessels and poorly formed retinal vessels. These defects are suppressed by intraocular injection of FLT1-Fc protein [a vascular endothelial growth factor (VEGF) receptor-1 (FLT1)/Fc chimeric protein that can bind VEGF and inhibit its activity], or by inactivating the HIF-1 alpha gene. Our results suggest that not only macrophages but also tissue oxygen-sensing mechanisms regulate the transition from the fetal to the adult circulatory system in the retina.

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Neurodegenerative influence of oxidative stress in the retina of a murine model of diabetes.

Sasaki M, Ozawa Y†, Kurihara T, Kubota S, Yuki K, Noda K, Kobayashi S, Ishida S, Tsubota K

Diabetologia. （SPRINGER）  53 （ 5 ） 971 - 979 2010年05月

査読有り,  ISSN  0012-186X

　概要を見る

Diabetic retinopathy is a progressive neurodegenerative disease, but the underlying mechanism is still obscure. Here, we focused on oxidative stress in the retina, and analysed its influence on retinal neurodegeneration, using an antioxidant, lutein.
C57BL/6 mice with streptozotocin-induced diabetes were constantly fed either a lutein-supplemented diet or a control diet from the onset of diabetes, and their metabolic data were recorded. In 1-month-diabetic mice, reactive oxygen species (ROS) in the retina were measured using dihydroethidium and visual function was evaluated by electroretinograms. Levels of activated extracellular signal-regulated kinase (ERK), synaptophysin and brain-derived neurotrophic factor (BDNF) were also measured by immunoblotting in the retina of 1-month-diabetic mice. In the retinal sections of 4-month-diabetic mice, histological changes, cleaved caspase-3 and TUNEL staining were analysed.
Lutein did not affect the metabolic status of the diabetic mice, but it prevented ROS generation in the retina and the visual impairment induced by diabetes. ERK activation, the subsequent synaptophysin reduction, and the BDNF depletion in the diabetic retina were all prevented by lutein. Later, in 4-month-diabetic mice, a decrease in the thickness of the inner plexiform and nuclear layers, and ganglion cell number, together with increase in cleaved caspase-3- and TUNEL-positive cells, were avoided in the retina of lutein-fed mice.
The results indicated that local oxidative stress that has a neurodegenerative influence in the diabetic retina is prevented by constant intake of a lutein-supplemented diet. The antioxidant, lutein may be a potential therapeutic approach to protect visual function in diabetes.

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