Calpain inhibitors reduce retinal hypoxia in ischemic retinopathy by improving neovascular architecture and functional perfusion

In ischemic retinopathies, underlying hypoxia drives abnormal neovascularization that damages retina and causes blindness. The abnormal neovasculature is tortuous and leaky and fails to alleviate hypoxia, resulting in more pathological neovascularization and retinal damage. With an established model of ischemic retinopathy we found that calpain inhibitors, when administered in moderation, reduced architectural abnormalities, reduced vascular leakage, and most importantly reduced retinal hypoxia. Mechanistically, these calpain inhibitors improved stability and organization of the actin cytoskeleton in retinal endothelial cells undergoing capillary morphogenesis in vitro, and they similarly improved organization of actin cables within new blood vessels in vivo. Hypoxia induced calpain activity in retinal endothelial cells and severely disrupted the actin cytoskeleton, whereas calpain inhibitors preserved actin cables under hypoxic conditions. Collectively, these findings support the hypothesis that hyper-activation of calpains by hypoxia contributes to disruption of the retinal endothelial cell cytoskeleton, resulting in formation of neovessels that are defective both architecturally and functionally. Modest suppression of calpain activity with calpain inhibitors restores cytoskeletal architecture and promotes formation of a functional neovasculature, thereby reducing underlying hypoxia. In sharp contrast to “anti-angiogenesis” strategies that cannot restore normoxia and may aggravate hypoxia, the therapeutic strategy described here does not inhibit neovascularization. Instead, by improving the function of neovascularization to reduce underlying hypoxia, moderate calpain inhibition offers a method for alleviating retinal ischemia, thereby suggesting a new treatment paradigm based on improvement rather than inhibition of new blood vessel growth.     

Correlations between cutaneous and retinal microaneurysms in diabetes mellitus (preliminary note)

A group of diabetic patients was submitted to a capillaroscopic examination at the level of the peri-ungual vallum and to a fluoroangiographic examination of the retina, for the evaluation of possible correlations between vessel conditions in to various districts. The Authors remarked that the subjects affected by proliferative diabetic retinopathy presented a greater dilatation of the ansae at cutaneous level, with haemorrhagic extravasations in 70% of the cases. On the contrary, on the subjects with nonproliferative diabetic retinopathy numerous aneurysmal "buttons" were noted at cutaneous level, both apically and laterally along the vascular ansa.     

Blood vessel occlusion by Cryptococcus neoformans is a mechanism for haemorrhagic dissemination of infection

Meningitis caused by infectious pathogens is associated with vessel damage and infarct formation, however the physiological cause is often unknown. Cryptococcus neoformans is a human fungal pathogen and causative agent of cryptococcal meningitis, where vascular events are observed in up to 30% of patients, predominantly in severe infection. Therefore, we aimed to investigate how infection may lead to vessel damage and associated pathogen dissemination using a zebrafish model that permitted noninvasive in vivo imaging. We find that cryptococcal cells become trapped within the vasculature (dependent on their size) and proliferate there resulting in vasodilation. Localised cryptococcal growth, originating from a small number of cryptococcal cells in the vasculature was associated with sites of dissemination and simultaneously with loss of blood vessel integrity. Using a cell-cell junction tension reporter we identified dissemination from intact blood vessels and where vessel rupture occurred. Finally, we manipulated blood vessel tension via cell junctions and found increased tension resulted in increased dissemination. Our data suggest that global vascular vasodilation occurs following infection, resulting in increased vessel tension which subsequently increases dissemination events, representing a positive feedback loop. Thus, we identify a mechanism for blood vessel damage during cryptococcal infection that may represent a cause of vascular damage and cortical infarction during cryptococcal meningitis.     

A two-way communication between microglial cells and angiogenic sprouts regulates angiogenesis in aortic ring cultures

Background

Myeloid cells have been associated with physiological and pathological angiogenesis, but their exact functions in these processes remain poorly defined. Monocyte-derived tissue macrophages of the CNS, or microglial cells, invade the mammalian retina before it becomes vascularized. Recent studies correlate the presence of microglia in the developing CNS with vascular network formation, but it is not clear whether the effect is directly caused by microglia and their contact with the endothelium.

Methodology/Principal Findings

We combined in vivo studies of the developing mouse retina with in vitro studies using the aortic ring model to address the role of microglia in developmental angiogenesis. Our in vivo analyses are consistent with previous findings that microglia are present at sites of endothelial tip-cell anastomosis, and genetic ablation of microglia caused a sparser vascular network associated with reduced number of filopodia-bearing sprouts. Addition of microglia in the aortic ring model was sufficient to stimulate vessel sprouting. The effect was independent of physical contact between microglia and endothelial cells, and could be partly mimicked using microglial cell-conditioned medium. Addition of VEGF-A promoted angiogenic sprouts of different morphology in comparison with the microglial cells, and inhibition of VEGF-A did not affect the microglia-induced angiogenic response, arguing that the proangiogenic factor(s) released by microglia is distinct from VEGF-A. Finally, microglia exhibited oriented migration towards the vessels in the aortic ring cultures.

Conclusions/Significance

Microglia stimulate vessel sprouting in the aortic ring cultures via a soluble microglial-derived product(s), rather than direct contact with endothelial cells. The observed migration of microglia towards the growing sprouts suggests that their position near endothelial tip-cells could result from attractive cues secreted by the vessels. Our data reveals a two-way communication between microglia and vessels that depends on soluble factors and should extend the understanding of how microglia promote vascular network formation.     

Reduced Levels of Brain Derived Neurotrophic Factor (BDNF) in the Serum of Diabetic Retinopathy Patients and in the Retina of Diabetic Rats

Diabetic retinopathy (DR) is widely recognized as a neurovascular disease. Retina, being a neuronal tissue of the eye, produces neurotrophic factors for its maintenance. However, diabetes dysregulates their levels and thereby may damage the retina. Among neurotrophins, brain derived neurotrophic factor (BDNF) is the most abundant in the retina. In this study, we investigated the level of BDNF in the serum of patients with DR and also in the serum and retina of streptozotocin-induced diabetic rats. The level of BDNF was significantly decreased in the serum of proliferative diabetic retinopathy patients as compared to that of non-diabetic healthy controls (25.5 ± 8.5–10.0 ± 8.1 ng/ml, p < 0.001) as well as compared to that of diabetic patients with no retinopathy (21.8 ± 4.7–10.0 ± 8.1 ng/ml, p < 0.001), as measured by ELISA techniques. The levels of BDNF in the serum and retina of diabetic rats were also significantly reduced compared to that of non-diabetic controls (p < 0.05). In addition, the expression level of tropomyosin-related kinase B (TrkB) was significantly decreased in diabetic rat retina compared to that of non-diabetic controls as determined by Western blotting technique. Caspase-3 activity was increased in diabetic rat retina after 3 weeks of diabetes and remained elevated until 10 weeks, which negatively correlated with the level of BDNF (r = ?0.544, p = 0.013). Our results indicate that reduced levels of BDNF in diabetes may cause apoptosis and neurodegeneration early in diabetic retina, which may lead to neuro-vascular damage later in DR.     

Alteration of developmental and pathological retinal angiogenesis in angptl4-deficient mice

Proper vessel maturation, remodeling of endothelial junctions, and recruitment of perivascular cells is crucial for establishing and maintaining vessel functions. In proliferative retinopathies, hypoxia-induced angiogenesis is associated with disruption of the vascular barrier, edema, and vision loss. Therefore, identifying factors that regulate vascular maturation is critical to target pathological angiogenesis. Given the conflicting role of angiopoietin-like-4 (ANGPTL4) reported in the current literature using gain of function systems both in vitro and in vivo, the goal of this study was to characterize angiogenesis, focusing on perinatal retinal vascularization and pathological circumstances in angpl4-deficient mice. We report altered organization of endothelial junctions and pericyte coverage, both leading to impaired angiogenesis and increased vascular leakage that were eventually caught up, suggesting a delay in vessel maturation. In a model of oxygen-induced retinopathy, pathological neovascularization, which results from tissue hypoxia, was also strongly inhibited in angptl4-deficient mice. This study therefore shows that ANGPTL4 tunes endothelial cell junction organization and pericyte coverage and controls vascular permeability and angiogenesis, both during development and in pathological conditions.     

Increased Intraocular Insulin-like Growth Factor-I Triggers Blood-Retinal Barrier Breakdown

Blood-retinal barrier (BRB) breakdown is a key event in diabetic retinopathy and other ocular disorders that leads to increased retinal vascular permeability. This causes edema and tissue damage resulting in visual impairment. Insulin-like growth factor-I (IGF-I) is involved in these processes, although the relative contribution of increased systemic versus intraocular IGF-I remains controversial. Here, to elucidate the role of this factor in BRB breakdown, transgenic mice with either local or systemic elevations of IGF-I have been examined. High intraocular IGF-I, resulting from overexpression of IGF-I in the retina, increased IGF-I receptor content and signaling and led to accumulation of vascular endothelial growth factor. This was parallel to up-regulation of vascular Intercellular adhesion molecule I and retinal infiltration by bone marrow-derived microglial cells. These alterations resulted in increased vessel paracellular permeability to both low and high molecular weight compounds in IGF-I-overexpressing retinas and agreed with the loss of vascular tight junction integrity observed by electron microscopy and the altered junctional protein content. In contrast, mice with chronically elevated serum IGF-I did not show alterations in the retinal vasculature structure and permeability, indicating that circulating IGF-I cannot initiate BRB breakdown. Consistent with a key role of IGF-I signaling in retinal diseases, a strong up-regulation of the IGF-I receptor in human retinas with marked gliosis was also observed. Thus, this study demonstrates that intraocular IGF-I, but not systemic IGF-I, is sufficient to trigger processes leading to BRB breakdown and increased retinal vascular permeability. Therefore, therapeutic interventions designed to counteract local IGF-I effects may prove successful to prevent BRB disruption.The BRB⁵ is a selective diffusion barrier that isolates the retina from the blood, maintaining the appropriate milieu for optimal retinal function and excluding potentially harmful stimuli, therefore acting as a critical protective barrier. The BRB consists of outer and inner components. The outer BRB is formed by the retinal pigmentary epithelium, which separates photoreceptors from choroidal permeable vessels. The inner BRB is determined by the presence of tight junctions (TJs) between the endothelial cells of retinal vessels, which limits paracellular flux. In addition, retinal vessels are partially sheathed by glial end-foot processes. Although not considered direct components of the inner BRB, glial cells could play a key role in its formation, maintenance, and breakdown (1). The disruption of the BRB is an important feature not only of non-proliferative and proliferative diabetic retinopathy but also of other diverse ocular disorders (2, 3). Increased vascular permeability results in extravasation of plasma components leading to edema. If the accumulation of fluids threatens the macula it poses a serious risk to visual function. Indeed, macular edema is a leading cause of visual loss among diabetic patients (3).IGF-I has been associated with the pathogenesis of BRB breakdown. Although most studies report an increase of intraocular IGF-I levels in diabetic patients (4, 5), the source of IGF-I is not clear (6, 7), and the relative contribution of local versus serum IGF-I in initiating ocular pathology is unknown. IGF-I is a potent inducer of vascular endothelial growth factor (VEGF) (8), a pro-angiogenic factor that increases vessel permeability (1). We and others have shown in animal models that IGF-I participates in the pathophysiology of diabetic retinopathy by inducing retinal VEGF expression (9, 10). In diabetic retinopathy there is a correlation between VEGF vitreous levels and macular edema (11). Aqueous humor levels of VEGF are also higher in diabetic patients with macular edema, and levels correlate with disease severity (12, 13). Moreover, the early BRB breakdown observed in experimental diabetes coincides with an increase in VEGF (14).To discern the contribution of intraocular versus circulating IGF-I in triggering VEGF production and BRB disruption, in this study we have examined the retinas of two transgenic animal models with elevated IGF-I levels either locally or in the serum. We have found that only IGF-I generated within the eye may trigger the breakdown of the BRB in mice, whereas increased circulating IGF-I did not alter retinal vascular permeability. Therefore, therapeutic interventions designed to counteract local IGF-I effects may prove successful to prevent BRB disruption.     

Effects of seed mass on seedling success in Artocarpus heterophyllus L., a tropical tree species of north-east India

I examined the effects of seed mass on performance of seedlings of Artocarpus heterophyllus L. (Moraceae), a large evergreen late successional shade-tolerant tree species in three contrasting light conditions. Seed mass varied many fold from 1.5 to 14 g in A. heterophyllus. Germination and germination time showed a significant correlation with seed mass. Germination differed significantly among three light regimes (50%, 25% and 3%). Seed mass and light level significantly affected seedling survival. The seedlings that emerged from large seeds survived better than those from small seeds under all light regimes. Survival of seedlings was maximum in 25% light regime for all seed mass classes but did not differ significantly from that at 50% light regime. Survival was significantly lower in 3% light as compared to 50% and 25% light regimes. Seedling vigor (expressed in terms of seedling height, leaf area and dry weight) was also significantly affected by seed mass and light regimes. Seedlings that emerged from larger seeds and grew under 50% light regime produced the heaviest seedlings, while those resulting from smaller seeds and grown under 3% light regime produced the lightest seedlings. Resprouting capacity of seedlings after clipping was significantly affected by seed mass and light regime. Seedlings emerging from larger seeds were capable of resprouting several times successively. Resprouting was more pronounced under 50% and 25% light regimes as compared to 3% light. Success of A. heterophyllus regeneration appears to be regulated by an interactive effect of seed mass and light regime.     

Astrocyte invasion and vasculogenesis in the developing ferret retina

We studied the time course of astrocyte invasion and blood vessel formation in the developing ferret retina using glial fibrillary acidic protein (GFAP)-immunohistochemistry for astrocytes and isolectin B4 histochemistry for blood vessels. As in other mammals, strongly GFAP positive astrocytes invade the ferret retina from the optic nerve. At birth, strongly GFAP positive astrocytes have reached about 22% of the distance between optic disc and outer retinal edge whereas weakly GFAP positive processes already extend to the edge of the retina. At postnatal days P30–P37 about 82% of the distance between optic disc and outer retinal edge and in the adult 88% of this distance is covered with strongly labelled astrocytes. Superficial blood vessels form from the optic disc. They reach up to about 24% of the retinal radius at birth and grow radially across the retina during further development. At P30–P37, the whole retina is covered with superficial blood vessels. The deep vascular layer forms later (around P30) through sprouting from superficial vessels. The radial pattern of astrocyte and vessel growth from the optic disc is not affected by the formation of the area centralis and visual streak.     

Bottomland hardwood forest species responses to flooding regimes along an urbanization gradient

Urbanization alters stream hydrology, hence flooding frequency and duration in floodplain wetlands. Potential impacts include shifts in species composition and survival, making restoration and selection of wetland species difficult. Cephalanthus occidentalis, Fraxinus pennsylvanica, and Quercus shumardii seedlings were subjected to experimental flooding regimes typical of floodplain forests in rural and urban settings. Treatments included a rural flood regime with three 7-day floods, an Urban-short flood regime with six 4-day floods, and an Urban-long flood regime with six 10-day floods over a growing season. Specific responses, measured by stem length, leaf area, and leaf, stem, and root biomass, varied between species from different wetland indicator classes. C. occidentalis, a wetland obligate, was well adapted to both urban flooding regimes, whereas productivity of F. pennsylvanica, a facultative wetland species, and Q. shumardii, a facultative species, was significantly reduced by the Urban-long treatment. Growth rates also varied over time, indicating the importance of temporal flooding patterns on species productivity. Because urban flooding regimes directly and selectively alter species productivity, proper restoration methods in urbanizing environments should include species selection based on current and potential future hydrologic conditions and use of reference standards from reference sites subjected to similar urban hydrologic regimes.     

At last the organ grinder...

Abstract

A prominent and early feature of the retinopathy of diabetes mellitus is a diffuse increase in vascular permeability. As the disease develops, the development of frank macular oedema may result in vision loss. That reactive oxygen species production is likely to be elevated in the retina, and that certain regions of the retina are enriched in substrates for lipid peroxidation, may create an environment susceptible to oxidative damage. This may be more so in the diabetic retina, where hyperglycaemia may lead to elevated oxidant production by a number of mechanisms, including the production of oxidants by vascular endothelium and leukocytes. There is substantial evidence from animal and clinical studies for both impaired antioxidant defences and increased oxidative damage in the retinae of diabetic subjects that have been, in the case of animal studies, reversible with antioxidant supplementation. Whether oxidative damage has a causative role in the pathology of diabetic retinopathy, and thus whether antioxidants can prevent or correct any retinal damage, has not been established, nor has the specific nature of any damaging species been characterised.     

A quantitative histochemical study of the microvasculature of irradiated skin

Short-term X-ray damage to the microvasculature of the skin of newborn rats has been quantitated using Horseradish Peroxidase as a tracer. Image analysis of thick sections on which peroxidase was demonstrated histochemically revealed a radioinduced increase in vascular volume coupled with a decrease in vascular length and an altered frequency distribution of blood vessel calibers which resulted in early telangiectasia. The results afforded by direct counting of peroxidase positive macrophagic cells and microphotometric evaluation of peroxidase present in the connective tissue indicate a progressive increase in capillary permeability as a function of dose and time post-irradiation. The accuracy with which the affected region of blood vessels coincided with the area exposed to the beam favours the hypothesis of direct damage to the vessel wall as a major cause of radioinduced lesion.     

Regenerative Therapeutic Potential of Adipose Stromal Cells in Early Stage Diabetic Retinopathy

Diabetic retinopathy (DR) is the leading cause of blindness in working-age adults. Early stage DR involves inflammation, vascular leakage, apoptosis of vascular cells and neurodegeneration. In this study, we hypothesized that cells derived from the stromal fraction of adipose tissue (ASC) could therapeutically rescue early stage DR features. Streptozotocin (STZ) induced diabetic athymic nude rats received single intravitreal injection of human ASC into one eye and saline into the other eye. Two months post onset of diabetes, administration of ASC significantly improved “b” wave amplitude (as measured by electroretinogram) within 1–3 weeks of injection compared to saline treated diabetic eyes. Subsequently, retinal histopathological evaluation revealed a significant decrease in vascular leakage and apoptotic cells around the retinal vessels in the diabetic eyes that received ASC compared to the eyes that received saline injection. In addition, molecular analyses have shown down-regulation in inflammatory gene expression in diabetic retina that received ASC compared to eyes that received saline. Interestingly, ASC were found to be localized near retinal vessels at higher densities than seen in age matched non-diabetic retina that received ASC. In vitro, ASC displayed sustained proliferation and decreased apoptosis under hyperglycemic stress. In addition, ASC in co-culture with retinal endothelial cells enhance endothelial survival and collaborate to form vascular networks. Taken together, our findings suggest that ASC are able to rescue the neural retina from hyperglycemia-induced degeneration, resulting in importantly improved visual function. Our pre-clinical studies support the translational development of adipose stem cell-based therapy for DR to address both retinal capillary and neurodegeneration.     

Diversity and Above-Ground Biomass Patterns of Vascular Flora Induced by Flooding in the Drawdown Area of China's Three Gorges Reservoir

Hydrological alternation can dramatically influence riparian environments and shape riparian vegetation zonation. However, it was difficult to predict the status in the drawdown area of the Three Gorges Reservoir (TGR), because the hydrological regime created by the dam involves both short periods of summer flooding and long-term winter impoundment for half a year. In order to examine the effects of hydrological alternation on plant diversity and biomass in the drawdown area of TGR, twelve sites distributed along the length of the drawdown area of TGR were chosen to explore the lateral pattern of plant diversity and above-ground biomass at the ends of growing seasons in 2009 and 2010. We recorded 175 vascular plant species in 2009 and 127 in 2010, indicating that a significant loss of vascular flora in the drawdown area of TGR resulted from the new hydrological regimes. Cynodon dactylon and Cyperus rotundus had high tolerance to short periods of summer flooding and long-term winter flooding. Almost half of the remnant species were annuals. Species richness, Shannon-Wiener Index and above-ground biomass of vegetation exhibited an increasing pattern along the elevation gradient, being greater at higher elevations subjected to lower submergence stress. Plant diversity, above-ground biomass and species distribution were significantly influenced by the duration of submergence relative to elevation in both summer and previous winter. Several million tonnes of vegetation would be accumulated on the drawdown area of TGR in every summer and some adverse environmental problems may be introduced when it was submerged in winter. We conclude that vascular flora biodiversity in the drawdown area of TGR has dramatically declined after the impoundment to full capacity. The new hydrological condition, characterized by long-term winter flooding and short periods of summer flooding, determined vegetation biodiversity and above-ground biomass patterns along the elevation gradient in the drawdown area.     

Connexin hemichannel induced vascular leak suggests a new paradigm for cancer therapy

It is 40 years since cancer growth was correlated with neovascularisation. Anti-angiogenic drugs remain at the forefront of cancer investigations but progress has been disappointing and unexpected toxicities are emerging. Gap junction channels are implicated in lesion spread following injury, with channel blockers shown to improve healing; in particular preventing vascular disruption and/or restoring vascular integrity. Here we briefly review connexin roles in vascular leak and endothelial cell death that occurs following acute wounds and during chronic disease, and how connexin channel regulation has been used to ameliorate vascular disruption. We then review chronic inflammatory disorders and trauma in the eye, concluding that vascular disruption under these conditions mimics that seen in tumours, and can be prevented with connexin hemichannel modulation. We apply this knowledge to tumour vessel biology, proposing that contrary to current opinion, these data suggest a need to protect, maintain and/or restore cancer vasculature. This may lead to reduced tumour hypoxia, promote the survival of normal cells, and enable improved therapeutic delivery or more effective radiation therapy.     

Crosstalk between Nitrite,Myoglobin and Reactive Oxygen Species to Regulate Vasodilation under Hypoxia

The systemic response to decreasing oxygen levels is hypoxic vasodilation. While this mechanism has been known for more than a century, the underlying cellular events have remained incompletely understood. Nitrite signaling is critically involved in vessel relaxation under hypoxia. This can be attributed to the presence of myoglobin in the vessel wall together with other potential nitrite reductases, which generate nitric oxide, one of the most potent vasodilatory signaling molecules. Questions remain relating to the precise concentration of nitrite and the exact dose-response relations between nitrite and myoglobin under hypoxia. It is furthermore unclear whether regulatory mechanisms exist which balance this interaction. Nitrite tissue levels were similar across all species investigated. We then investigated the exact fractional myoglobin desaturation in an ex vivo approach when gassing with 1% oxygen. Within a short time frame myoglobin desaturated to 58±12%. Given that myoglobin significantly contributes to nitrite reduction under hypoxia, dose-response experiments using physiological to pharmacological nitrite concentrations were conducted. Along all concentrations, abrogation of myoglobin in mice impaired vasodilation. As reactive oxygen species may counteract the vasodilatory response, we used superoxide dismutase and its mimic tempol as well as catalase and ebselen to reduce the levels of reactive oxygen species during hypoxic vasodilation. Incubation of tempol in conjunction with catalase alone and catalase/ebselen increased the vasodilatory response to nitrite. Our study shows that modest hypoxia leads to a significant nitrite-dependent vessel relaxation. This requires the presence of vascular myoglobin for both physiological and pharmacological nitrite levels. Reactive oxygen species, in turn, modulate this vasodilation response.     

Environmental Enrichment Protects the Retina from Early Diabetic Damage in Adult Rats

Diabetic retinopathy is a leading cause of reduced visual acuity and acquired blindness. Available treatments are not completely effective. We analyzed the effect of environmental enrichment on retinal damage induced by experimental diabetes in adult Wistar rats. Diabetes was induced by an intraperitoneal injection of streptozotocin. Three days after vehicle or streptozotocin injection, animals were housed in enriched environment or remained in a standard environment. Retinal function (electroretinogram, and oscillatory potentials), retinal morphology, blood-retinal barrier integrity, synaptophysin, astrocyte and Müller cell glial fibrillary acidic protein, vascular endothelial growth factor, tumor necrosis factor-α, and brain-derived neurotrophic factor levels, as well as lipid peroxidation were assessed in retina from diabetic animals housed in standard or enriched environment. Environmental enrichment preserved scotopic electroretinogram a-wave, b-wave and oscillatory potential amplitude, avoided albumin-Evan''s blue leakage, prevented the decrease in retinal synaptophysin and astrocyte glial fibrillary acidic protein levels, the increase in Müller cell glial fibrillary acidic protein, vascular endothelial growth factor and tumor necrosis factor-α levels, as well as oxidative stress induced by diabetes. In addition, enriched environment prevented the decrease in retinal brain-derived neurotrophic factor levels induced by experimental diabetes. When environmental enrichment started 7 weeks after diabetes onset, retinal function was significantly preserved. These results indicate that enriched environment could attenuate the early diabetic damage in the retina from adult rats.     

Hypoxia-Induced Retinal Neovascularization in Zebrafish Embryos: A Potential Model of Retinopathy of Prematurity

Retinopathy of prematurity, formerly known as a retrolental fibroplasia, is a leading cause of infantile blindness worldwide. Retinopathy of prematurity is caused by the failure of central retinal vessels to reach the retinal periphery, creating a nonperfused peripheral retina, resulting in retinal hypoxia, neovascularization, vitreous hemorrhage, vitreoretinal fibrosis, and loss of vision. We established a potential retinopathy of prematurity model by using a green fluorescent vascular endothelium zebrafish transgenic line treated with cobalt chloride (a hypoxia-inducing agent), followed by GS4012 (a vascular endothelial growth factor inducer) at 24 hours postfertilization, and observed that the number of vascular branches and sprouts significantly increased in the central retinal vascular trunks 2–4 days after treatment. We created an angiography method by using tetramethylrhodamine dextran, which exhibited severe vascular leakage through the vessel wall into the surrounding retinal tissues. The quantification of mRNA extracted from the heads of the larvae by using real-time quantitative polymerase chain reaction revealed a twofold increase in vegfaa and vegfr2 expression compared with the control group, indicating increased vascular endothelial growth factor signaling in the hypoxic condition. In addition, we demonstrated that the hypoxic insult could be effectively rescued by several antivascular endothelial growth factor agents such as SU5416, bevacizumab, and ranibizumab. In conclusion, we provide a simple, highly reproducible, and clinically relevant retinopathy of prematurity model based on zebrafish embryos; this model may serve as a useful platform for clarifying the mechanisms of human retinopathy of prematurity and its progression.     

Angiotensin and diabetic retinopathy

Diabetic retinopathy develops in patients with both type 1 and type 2 diabetes and is the major cause of vision loss and blindness in the working population. In diabetes, damage to the retina occurs in the vasculature, neurons and glia resulting in pathological angiogenesis, vascular leakage and a loss in retinal function. The renin-angiotensin system is a causative factor in diabetic microvascular complications inducing a variety of tissue responses including vasoconstriction, inflammation, oxidative stress, cell hypertrophy and proliferation, angiogenesis and fibrosis. All components of the renin-angiotensin system including the angiotensin type 1 and angiotensin type 2 receptors have been identified in the retina of humans and rodents. There is evidence from both clinical and experimental models of diabetic retinopathy and hypoxic-induced retinal angiogenesis that the renin-angiotensin system is up-regulated. In these situations, retinal dysfunction has been linked to angiotensin-mediated induction of growth factors including vascular endothelial growth factor, platelet-derived growth factor and connective tissue growth factor. Evidence to date indicates that blockade of the renin-angiotensin system can confer retinoprotection in experimental models of diabetic retinopathy and ischemic retinopathy. This review examines the role of the renin-angiotensin system in diabetic retinopathy and the potential of its blockade as a treatment strategy for this vision-threatening disease.     

Laminins and retinal vascular development

The mechanisms controlling vascular development, both normal and pathological, are not yet fully understood. Many diseases, including cancer and diabetic retinopathy, involve abnormal blood vessel formation. Therefore, increasing knowledge of these mechanisms may help develop novel therapeutic targets. The identification of novel proteins or cells involved in this process would be particularly useful. The retina is an ideal model for studying vascular development because it is easy to access, particularly in rodents where this process occurs post-natally. Recent studies have suggested potential roles for laminin chains in vascular development of the retina. This review will provide an overview of these studies, demonstrating the importance of further research into the involvement of laminins in retinal blood vessel formation.