Treatment Of Proliferative Retinopathy Research Articles

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

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

Tractional Retinal Detachment: Prevalence and Causes in Nigerians.

To determine the causes of tractional retinal detachment (TRD) in Nigerians. A prospective, multicentre study evaluating eyes diagnosed to have TRD. History, clinical examination (including visual acuity, intraocular pressure measurement, anterior segment examination and dilated fundoscopy) and systemic evaluation (including previous diagnosis of diabetes, hypertension, sickle-cell disease and others) were performed in TRD eyes out of a cohort of retinal detachment eyes. The prevalence of TRD of the 237 patients diagnosed with RD within a one-year study period was 25.7% (61 patients). Eighty eyes were diagnosed with TRD. Thirty-eight eyes of nineteen patients (31%) were bilateral, and 42 (69%) were unilateral. There were 38 male patients (62.3%) and 23 female patients (37.7%). The mean age was 52.3 ± 12.7 years (11-69 years). 88.5% of all TRD patients had an associated systemic disease, diabetes being the most common disease in 88.8% of them. Proliferative diabetic retinopathy was the most common cause of TRD (77.5%) and the most common cause of bilateral TRD. Both trauma and proliferative sickle-cell retinopathy occurred in 3.8% of the eyes. 68.8% of TRD eyes were blind at the presentation. However, the causes of TRD did not show any significant association with blindness (P = 0.819). Proliferative diabetic retinopathy poses a significant threat to vision, being the most common cause of TRD. Early detection and treatment of proliferative retinopathy in diabetes and sickle-cell disease, and trauma prevention will significantly reduce the burden of blindness due to TRD.

Single-cell transcriptome analyses reveal microglia types associated with proliferative retinopathy.

Pathological angiogenesis is a major cause of irreversible blindness in individuals of all age groups with proliferative retinopathy (PR). Mononuclear phagocytes (MPs) within neovascular areas contribute to aberrant retinal angiogenesis. Due to their cellular heterogeneity, defining the roles of MP subsets in PR onset and progression has been challenging. Here, we aimed to investigate the heterogeneity of microglia associated with neovascularization and to characterize the transcriptional profiles and metabolic pathways of proangiogenic microglia in a mouse model of oxygen-induced PR (OIR). Using transcriptional single-cell sorting, we comprehensively mapped all microglia populations in retinas of room air (RA) and OIR mice. We have unveiled several unique types of PR-associated microglia (PRAM) and identified markers, signaling pathways, and regulons associated with these cells. Among these microglia subpopulations, we found a highly proliferative microglia subset with high self-renewal capacity and a hypermetabolic microglia subset that expresses high levels of activating microglia markers, glycolytic enzymes, and proangiogenic Igf1. IHC staining shows that these PRAM were spatially located within or around neovascular tufts. These unique types of microglia have the potential to promote retinal angiogenesis, which may have important implications for future treatment of PR and other pathological ocular angiogenesis-related diseases.

Activated \u03b12-Macroglobulin Regulates LRP1 Levels at the Plasma Membrane through the Activation of a Rab10-dependent Exocytic Pathway in Retinal M\xfcller Glial Cells

Activated α2-macroglobulin (α2M*) and its receptor, low-density lipoprotein receptor-related protein 1 (LRP1), have been linked to proliferative retinal diseases. In Müller glial cells (MGCs), the α2M*/LRP1 interaction induces cell signaling, cell migration, and extracellular matrix remodeling, processes closely associated with proliferative disorders. However, the mechanism whereby α2M* and LRP1 participate in the aforementioned pathologies remains incompletely elucidated. Here, we investigate whether α2M* regulates both the intracellular distribution and sorting of LRP1 to the plasma membrane (PM) and how this regulation is involved in the cell migration of MGCs. Using a human Müller glial-derived cell line, MIO-M1, we demonstrate that the α2M*/LRP1 complex is internalized and rapidly reaches early endosomes. Afterward, α2M* is routed to degradative compartments, while LRP1 is accumulated at the PM through a Rab10-dependent exocytic pathway regulated by PI3K/Akt. Interestingly, Rab10 knockdown reduces both LRP1 accumulation at the PM and cell migration of MIO-M1 cells induced by α2M*. Given the importance of MGCs in the maintenance of retinal homeostasis, unravelling this molecular mechanism can potentially provide new therapeutic targets for the treatment of proliferative retinopathies.

Endothelial adenosine A2a receptor-mediated glycolysis is essential for pathological retinal angiogenesis

Adenosine/adenosine receptor-mediated signaling has been implicated in the development of various ischemic diseases, including ischemic retinopathies. Here, we show that the adenosine A2a receptor (ADORA2A) promotes hypoxia-inducible transcription factor-1 (HIF-1)-dependent endothelial cell glycolysis, which is crucial for pathological angiogenesis in proliferative retinopathies. Adora2a expression is markedly increased in the retina of mice with oxygen-induced retinopathy (OIR). Endothelial cell-specific, but not macrophage-specific Adora2a deletion decreases key glycolytic enzymes and reduces pathological neovascularization in the OIR mice. In human primary retinal microvascular endothelial cells, hypoxia induces the expression of ADORA2A by activating HIF-2α. ADORA2A knockdown decreases hypoxia-induced glycolytic enzyme expression, glycolytic flux, and endothelial cell proliferation, sprouting and tubule formation. Mechanistically, ADORA2A activation promotes the transcriptional induction of glycolytic enzymes via ERK- and Akt-dependent translational activation of HIF-1α protein. Taken together, these findings advance translation of ADORA2A as a therapeutic target in the treatment of proliferative retinopathies and other diseases dependent on pathological angiogenesis.

Abstract 177: Adenosine Receptor 2A-mediated Endothelial Glycolysis Critically Contributes to Pathological Angiogenesis of Retina

Pathological proliferative retinopathy is the most common cause of blindness in patients of all ages worldwide. Investigation of the molecular mechanisms underlying pathological retina neovascularization is critical to the development of new therapeutic strategies. In the current study, we investigated the effect of macrophage or endothelial cell adenosine receptor 2A (A 2A R) deletion in pathological retinal vascular growth, and further explored the potential mechanism. Our results showed that: (1) A 2A R expression was localized to retinal vessels and was markedly induced in pathological neovascularization tufts in a mouse model of oxygen-induced proliferative retinopathy (OIR) (Figure A); (2) endothelial cell, but not macrophage, -specific A 2A R deletion significantly decreased pathological neovascularization in the mouse OIR model (Figure B);. The levels of glycolytic enzymes, angiogenic factors and lactate secretion were markedly reduced in retina of endothelial cell-specific A 2A R knockout mice; (3) in human retinal microvascular endothelial cells (HRMECs), hypoxia markedly induced the expression of A 2A R, but not A 1 R, A 2B R and A 3 R, by activating HIF-2α. Deletion or blockade of A 2A R by small interfering RNA (siRNA) or antagonist ZM241385 decreased hypoxia or adenosine-induced glycolytic enzyme expression, lactate secretion and cell proliferation via suppression of the cAMP/PKA/CREB/HIF-1α pathway; (4) blockade of glycolysis by 3PO or 2-DG abolished the adenosine-induced HRMEC proliferation under hypoxic conditions. Taken together, we have demonstrated a critical role of endothelial cell A 2A R in promoting pathological vascular development and revealed a novel glycolysis-driven mechanism for endothelial cell A 2A R-mediated pathological angiogenesis in proliferative retinopathy. Thus, A 2A R is a promising therapeutic target in the treatment of proliferative retinopathy.

Acetyl-11-keto-β-boswellic acid reduces retinal angiogenesis in a mouse model of oxygen-induced retinopathy

Retinal diseases characterized by pathologic retinal angiogenesis are the leading causes of blindness worldwide. Although therapies directed toward vascular endothelial growth factor (VEGF) represent a significant step forward in the treatment of proliferative retinopathies, further improvements are needed. In the last few years, an intense research activity has focused around the use of herbal and traditional natural medicines as an alternative for slowing down the progression of proliferative retinopathies. In the present study, we investigated the antiangiogenic effects of acetyl-11-keto-β-boswellic acid (AKBA), one of the active principles derived from the plant Boswellia serrata, used in Ayurvedic systems of medicine. We studied the antiangiogenic properties of AKBA using the mouse model of oxygen-induced retinopathy (OIR), which mimics the neovascular response seen in human retinopathy of prematurity. We first evaluated the effects of subcutaneously administered AKBA on the expression/activity of proteins which are known to play a role in the OIR model. In the retina, AKBA increased expression and activity of Src homology region 2 domain-containing phosphatase 1 and reduced the phosphorylation of the transcription factor signal transducer and activator of transcription 3 (STAT3) as well as VEGF expression and VEGF receptor (VEGFR)-2 phosphorylation. Likely as a result of these effects, AKBA significantly reduced retinal neovascularization in OIR mice without affecting retinal cell survival and retinal function. Using retinal explants cultured in hypoxia and an activator of STAT3 phosphorylation, we showed that the AKBA-induced inhibition of VEGFR-2 phosphorylation is likely to be mediated by a mechanism depending on an SHP-1/STAT3/VEGF axis. In the OIR model, neovascularization results from the activation of retinal endothelial cells, therefore we evaluated whether AKBA affected the angiogenic response of human retinal microvascular endothelial cells (HRMECs). We observed that AKBA reduced proliferation, migration and tube formation in HRMECs stimulated with exogenous VEGF, while it reduced migration and tube formation in untreated HRMECs. Taken together, our results demonstrate the antiangiogenic effects of AKBA in a model of pathologic neovascularization, providing a rationale for further investigation of AKBA as a promising therapeutic agent to reduce the impact of proliferative retinopathies.

Antiangiogenic Effectiveness of the Urokinase Receptor-Derived Peptide UPARANT in a Model of Oxygen-Induced Retinopathy.

Pharmacologic control of neovascularization is a promising approach for the treatment of retinal angiogenesis. UPARANT, an inhibitor of the urokinase-type plasminogen activator receptor (uPAR), inhibits VEGF-driven angiogenesis in vitro and in vivo. This study investigates for the first time the effectiveness of UPARANT in counteracting pathologic neovascularization in the retina. Murine retinal fragments and a mouse model of oxygen-induced retinopathy (OIR) were used. In mice with OIR, UPARANT-treated retinas were analyzed for avascular area and neovascular tuft formation. Levels of transcription and proangiogenic factors were determined. UPARANT effects on the blood-retinal barrier (BRB), visual function, retinal cytoarchitecture, and inflammatory markers were also assessed. Human umbilical vein endothelial cells (HUVECs) and chick embryo chorioallantoic membrane (CAM) in which angiogenesis was induced by the vitreous fluid from patients with proliferative diabetic retinopathy (PDR) were also used. UPARANT reduced VEGF-induced angiogenesis in retinal fragments. In mice with OIR, UPARANT decreased neovascular response, VEGF, and VEGF receptor-2 activity. Transcription factors regulating VEGF expression were also reduced. UPARANT restored BRB integrity, recovered visual loss, and reduced levels of inflammatory markers. Restored electroretinogram does not involve any rescue in the retinal cytoarchitecture. Finally, UPARANT blocked PDR vitreous fluid-induced angiogenesis in HUVEC and CAM assays. The finding that UPARANT is effective against neovascularization may help to establish uPAR as a target in the treatment of proliferative retinopathies. The potential application of UPARANT in retinal diseases is further supported by UPARANT capacity to counteract the angiogenic activity of PDR vitreous fluid.

Endothelial-specific deficiency of Junctional Adhesion Molecule-C promotes vessel normalisation in proliferative retinopathy.

In proliferative retinopathies, like proliferative diabetic retinopathy and retinopathy of prematurity (ROP), the hypoxia response is sustained by the failure of the retina to revascularise its ischaemic areas. Non-resolving retina ischaemia/hypoxia results in upregulation of pro-angiogenic factors and pathologic neovascularisation with ectopic, fragile neovessels. Promoting revascularisation of the retinal avascular area could interfere with this vicious cycle and lead to vessel normalisation. Here, we examined the function of endothelial junctional adhesion molecule-C (JAM-C) in the context of ROP. Endothelial-specific JAM-C-deficient (EC-JAM-C KO) mice and littermate JAM-C-proficient (EC-JAM-C WT) mice were subjected to the ROP model. An increase in total retinal vascularisation was found at p17 owing to endothelial JAM-C deficiency, which was the result of enhanced revascularisation and vessel normalisation, thereby leading to significantly reduced avascular area in EC-JAM-C KO mice. In contrast, pathologic neovessel formation was not affected by endothelial JAM-C deficiency. Consistent with improved vessel normalisation, tip cell formation at the interface between vascular and avascular area was higher in EC-JAM-C KO mice, as compared to their littermate controls. Consistently, JAM-C inactivation in endothelial cells resulted in increased spreading on fibronectin and enhanced sprouting in vitro in a manner dependent on β1-integrin and on the activation of the small GTPase RAP1. Together, endothelial deletion of JAM-C promoted endothelial cell sprouting, and consequently vessel normalisation and revascularisation of the hypoxic retina without altering pathologic neovascularisation. Thus, targeting endothelial JAM-C may provide a novel therapeutic strategy for promoting revascularisation and vessel normalisation in the treatment of proliferative retinopathies.

Expression, Localization, and Functional Coupling of the Somatostatin Receptor Subtype 2 in a Mouse Model of Oxygen-Induced Retinopathy

Purpose. In the mouse model of oxygen-induced retinopathy (OIR), somatostatin-14 (SRIF) acting at the SRIF receptor subtype 2 (sst(2)) inhibits angiogenic responses to hypoxia through a downregulation of vascular endothelial growth factor. Information about where SRIF-sst(2) interactions take place is lacking, and downstream effectors mediating SRIF-sst(2) antiangiogenic actions are unknown. Methods. In the OIR model, retinal expression of SRIF was evaluated with RT-PCR and radioimmunoassay. The bindings of [(125)I]LTT-SRIF-28 and [(125)I]Tyr(3)-octreotide were measured in coronal sections of the eye. With Western blot analysis, the authors evaluated the levels of sst(2A) and the expression and activity of the signal transducer and activator of transcription (STAT)3. The analysis of STAT3 was performed in hypoxic mice treated with the sst(2) agonist octreotide or with the sst(2) antagonist D-Tyr(8) cyanamid 154806 (CYN). Retinal localization of sst(2A) was assessed by single and double immunohistochemistry with an endothelial cell marker. Results. In the hypoxic retina, both SRIF and sst(2) levels as well as [(125)I]Tyr(3)-octreotide binding were downregulated. In addition, sst(2A) immunostaining was decreased in the neuroretina but was increased in capillaries. Hypoxia increased both the expression and the activity of STAT3. This increase was inhibited by octreotide but was strengthened by CYN. Conclusions. These data suggest that sst(2) expressed by capillaries may be responsible for the antiangiogenic effects of SRIF and that downstream effectors in this action include the transcription factor STAT3. These results support the possibility of using sst(2)-selective ligands in the treatment of proliferative retinopathies and indicate STAT3 as an additional target for a novel therapeutic approach.

Retinopatia diabética

Diabetic retinopathy is a frequently observed complication in both type 1 and type 2 diabetes, specially in patients with long term disease and poor glicemic control. Irreversible visual loss appears at the final stages of diabetic retinopathy and it is considered one of the most tragic of diabetic complications. It is also considered an important factor of morbidity and has a high economical impact once it is the leading cause of blindness. The pathophysiology of the retinal microvascular alterations is related to the chronic hyperglycemia that leads to the following circulatory disturbances: loss of vascular tonus, increase in vascular permeability, edema and exudation, with vascular obstruction and ischemia that stimulates neovascularization, which may lead to fibrous retraction and vitreous hemorrhages with retinal detachment. Recent studies have indicated that the strict glicemic and blood pressure controls are effective in reducing or blocking the progression of retinopathy. Up to now no pharmacological agents have shown to be effective in preventing or reducing neovascularization and visual loss. Presently, the most effective available treatment for proliferative retinopathy is laser photocoagulation. Further studies are needed to obtain new products and technologies that could effectively prevent or block retinopathy progression.

The GH-IGF-I axis and diabetes complications.

Over 60 yr ago, Houssay demonstrated that removal of the pituitary improved the diabetes of animals and made them particularly sensitive to insulin (Carbohydrate metabolism. BA Houssay. N Engl J Med 1936; 214: 971–975), and Young reported that permanent diabetes could be produced by injection of pituitary extract (Permanent diabetes produced by pituitary (anterior lobe) injections. FG Young. Lancet 1937; ii: 372–374). Rolf Luft and colleagues in Sweden translated the Houssay model to humans by performing hypophysectomy in the hope of ameliorating vascular complications (Hypophysectomy in man. R Luft, H Olivecrona, B Sjogren. Nord Med 1952; 47: 351–354). At the same time, remission of diabetic retinopathy was reported in a woman with long-standing type 1 diabetes when she developed postpartum anterior pituitary insufficiency (Sheehan's syndrome) (The Houssay phenomenon in man: recovery from retinopathy in a case of diabetes with Simmonds' disease. JE Poulsen. Diabetes 1953; 2 : 7–12). In the 1960s and 1970s, there was considerable enthusiasm for pituitary ablation in the treatment of proliferative retinopathy, an approach happily supplanted by retinal photocoagulation.

Corneal sensation in insulin dependent and non-insulin dependent diabetics with proliferative retinopathy.

The corneal sensation of diabetic patients undergoing pan retinal photocoagulation for the treatment of proliferative retinopathy was measured using the Cochet-Bonnet aesthesiometer. Insulin dependent and non-insulin dependent diabetics both showed significantly decreased corneal sensitivity, and there was no significant change following pan-retinal photocoagulation. Insulin dependent diabetics with proliferative retinopathy had been diagnosed for a significantly longer period than non-insulin dependent diabetics, but had better corneal sensitivity than the latter group. Male patients were more likely to have brown irides, whereas there was a preponderance of blue irides amongst the females. Patients with blue irides also had significantly lower sensitivity than patients with brown irides. The results are discussed.