Insulin-like growth factor-2 regulates basal retinal insulin receptor activity.

The retinal insulin receptor (IR) exhibits basal kinase activity equivalent to that of the liver of fed animals, but unlike the liver, does not fluctuate with feeding and fasting; it also declines rapidly after the onset of insulin-deficient diabetes. The ligand(s) that determine basal IR activity in the retina has not been identified. Using a highly sensitive insulin assay, we found that retinal insulin concentrations remain constant in fed versus fasted rats and in diabetic versus control rats; vitreous fluid insulin levels were undetectable. Neutralizing antibodies against insulin-like growth factor 2 (IGF-2), but not insulin-like growth factor 1 (IGF-1) or insulin, decreased IR kinase activity in normal rat retinas, and depletion of IGF-2 from serum specifically reduced IR phosphorylation in retinal cells. Immunoprecipitation studies demonstrated that IGF-2 induced greater phosphorylation of the retinal IR than the IGF-1 receptor. Retinal IGF-2 mRNA content was 10-fold higher in adults than pups and orders of magnitude higher than in liver. Diabetes reduced retinal IGF-2, but not IGF-1 or IR, mRNA levels, and reduced IGF-2 and IGF-1 content in vitreous fluid. Finally, intravitreal administration of IGF-2 (mature and pro-forms) increased retinal IR and Akt kinase activity in diabetic rats. Collectively, these data reveal that IGF-2 is the primary ligand that defines basal retinal IR activity and suggest that reduced ocular IGF-2 may contribute to reduced IR activity in response to diabetes. These findings may have importance for understanding the regulation of metabolic and prosurvival signaling in the retina.

CLINICAL EVALUATION OF THE RAPID ACCESS VITREAL INJECTION GUIDE: A Handheld Instrument for Assisting Intravitreal Injections.

PURPOSE: The Rapid Access Vitreal Injection (RAVI) guide combines the function of an eyelid speculum and measuring caliper into a single instrument for assisting intravitreal injections. This study clinically evaluated the RAVI guide with respect to patient acceptance, complication rates, and operative goals. METHODS: A prospective study was performed on 54 patients undergoing intravitreal injections using the RAVI guide (n = 32) or the speculum/caliper (n = 22). Device-related pain was assessed using the Wong-Baker scoring system, scaled from 0 (no pain) to 10 (agonizing pain). RESULTS: Mean device-related pain score did not differ significantly between the 2 groups, with scores of 0.6 and 0.7 for the RAVI guide and speculum groups, respectively. The rate of significant pain (score of ≥2) was twice as high in the speculum group (7 of 22, 32%) compared with the RAVI guide group (5 of 32, 16%), but this difference was not statistically significant (P = 0.19, Fisher's exact test). Operative goals of avoiding needle touch to lashes/lids and guiding needle insertion to the intended site were achieved in all patients. CONCLUSION: The RAVI guide appeared equivalent to the eyelid speculum in achieving operative goals, with similarly low pain scores. It has the potential for facilitating efficient, accurate, and safe intravitreal injections.

The retinal proteome in experimental diabetic retinopathy: up-regulation of crystallins and reversal by systemic and periocular insulin.

Diabetic retinopathy is the leading cause of blindness in working age persons. Targeted studies have uncovered several components of the pathophysiology of the disease without unveiling the basic mechanisms. This study describes the use of complementary proteomic and genomic discovery methods that revealed that the proteins of the crystallin superfamily are increased dramatically in early diabetic retinopathy. Orthogonal methods confirmed that the amplitude of the up-regulation is greater than other changes described so far in diabetic retinopathy. A detailed time course study during diabetes showed differential up-regulation of the different isoforms of the crystallins superfamily. alpha- and beta-crystallins were regulated primarily at the translation level, whereas gamma-crystallins were also regulated transcriptionally. We also demonstrated cell-specific patterns of expression of the different crystallins in normal and diabetic rat retinas. In addition, systemic and periocular insulin treatments restored retinal crystallin protein expression during diabetes, indicating effects of phosphoinositide 3-kinase/Akt activity. Altogether this work shows the importance of proteomics discovery methods coupled with targeted approaches to unveil new disease mechanistic details and therapeutic targets.

Neuroprotection for diabetic retinopathy.

Diabetic retinopathy (DR) is a neurodegenerative and microvascular disease resulting in functional and structural changes of all cell types in the retina. Several mechanisms for neuroretinal homeostasis, including the blood-retinal barrier, normal metabolite delivery into the retina, and the effect of neurotrophins for the retina, are impaired in DR. However, it is still not clear which components are most important for the development of DR and which may be most useful as therapeutic targets. In this chapter, we summarize the evidence for the neurodegeneration in DR and review normal mechanisms for maintenance of postmitotic cells in the retina and alterations in normal maintenance pathways in DR with emphasis on 'neuroprotection'. Finally, we discuss current neuroprotective strategies and future directions for the treatment of DR.

Diabetes reduces basal retinal insulin receptor signaling: reversal with systemic and local insulin.

Diabetic retinopathy is characterized by early onset of neuronal cell death. We previously showed that insulin mediates a prosurvival pathway in retinal neurons and that normal retina expresses a highly active basal insulin receptor/Akt signaling pathway that is stable throughout feeding and fasting. Using the streptozotocin-induced diabetic rat model, we tested the hypothesis that diabetes diminishes basal retinal insulin receptor signaling concomitantly with increased diabetes-induced retinal apoptosis. The expression, phosphorylation status, and/or kinase activity of the insulin receptor and downstream signaling proteins were investigated in retinas of age-matched control, diabetic, and insulin-treated diabetic rats. Four weeks of diabetes reduced basal insulin receptor kinase, insulin receptor substrate (IRS)-1/2-associated phosphatidylinositol 3-kinase, and Akt kinase activity without altering insulin receptor or IRS-1/2 expression or tyrosine phosphorylation. After 12 weeks of diabetes, constitutive insulin receptor autophosphorylation and IRS-2 expression were reduced, without changes in p42/p44 mitogen-activated protein kinase or IRS-1. Sustained systemic insulin treatment of diabetic rats prevented loss of insulin receptor and Akt kinase activity, and acute intravitreal insulin administration restored insulin receptor kinase activity. Insulin treatment restored insulin receptor-beta autophosphorylation in rat retinas maintained ex vivo, demonstrating functional receptors and suggesting loss of ligand as a cause for reduced retinal insulin receptor/Akt pathway activity. These results demonstrate that diabetes progressively impairs the constitutive retinal insulin receptor signaling pathway through Akt and suggests that loss of this survival pathway may contribute to the initial stages of diabetic retinopathy.

Loss of cholinergic and dopaminergic amacrine cells in streptozotocin-diabetic rat and Ins2Akita-diabetic mouse retinas.

PURPOSE: To identify amacrine cells that are vulnerable to degeneration during the early stages of diabetes. METHODS: Whole retinas from streptozotocin (STZ)-diabetic rats and Ins2(Akita) mice were fixed in paraformaldehyde. Apoptotic cells in the retina were quantified using terminal dUTP nick-end labeling (TUNEL) and active caspase-3 (CM-1) immunohistochemistry. Immunohistochemical markers for choline acetyltransferase (ChAT) and tyrosine hyroxylase (TH) were also used to quantify populations of amacrine cells in the Ins2Akita mouse retinas. RESULTS: The number of TUNEL-positive nuclei increased from 29+/-4 in controls to 72+/-9 in the STZ-diabetic rat retinas after only 2 weeks of diabetes. In rats, CM-1-immunoreactive (IR) cells were found primarily in the inner nuclear and ganglion cell layers after 2, 8, and 16 weeks of diabetes. At each end point, the number of CM-1-IR cells in the retina was elevated by diabetes. Approximately 2% to 6% of the CM-1-IR cells in the inner nuclear layer (INL) were double-labeled for TH immunoreactivity. After 6 months of diabetes in the Ins2Akita mouse, the morphology of the labeled ChAT-IR and TH-IR amacrine cell somas and dendrites appeared normal. A quantitative analysis revealed a 20% decrease in the number of cholinergic and a 16% decrease in dopaminergic amacrine cells in the diabetic mouse retinas, compared with the nondiabetic control. CONCLUSIONS: Dopaminergic and cholinergic amacrine cells are lost during the early stages of retinal neuropathy in diabetes. Loss of these neurons may play a critical role in the development of visual deficits in diabetes.

Sutureless levator plication by conjunctival route: a new technique.

The experiences of sutureless levator plication by conjunctival route surgery are described in 80 primary operations performed for all grades of congenital ptosis in the past 2 years. The surgical steps, postoperative care and postoperative complications are reviewed.

Acute exudative polymorphous vitelliform maculopathy in a patient with Lyme disease.

Acute exudative polymorphous vitelli-form maculopathy (AEPVM) is a rare condition of unclear etiology that has been seen in association with respiratory and viral infections. It has also been reported as a paraneoplastic phenomenon in older individuals. The authors report the first case of AEPVM associated with Lyme disease with over 3.5 years of follow-up. Multimodality serial imaging suggested the lesions began as multiple serous detachments followed by accumulation of photoreceptor outer segments in the subretinal space that gradually resolved over time and gave rise to the characteristic fundus findings at various stages.

Ocular hypertension following intravitreal anti-vascular endothelial growth factor agents.

Age-related macular degeneration (AMD) is the leading cause of severe vision loss in adults over the age of 65 years. The advent of anti-vascular endothelial growth factor (anti-VEGF) intravitreal injections has revolutionized the management of exudative AMD. However, multiple case series of sustained elevated intraocular pressure (IOP) after intravitreal injections of anti-VEGF agents have been reported. Sustained elevated IOP has been reported with all anti-VEGF agents being used in ophthalmology and even in patients without any prior history of glaucoma. No clear correlations to injection frequency or patient characteristics have emerged from the multiple reports so far, but it appears that patients with pre-existing glaucoma or ocular hypertension and those receiving a greater number of injections with shorter injection intervals may be at a higher risk for developing ocular hypertension related to anti-VEGF agents. Until future studies elucidate the pathophysiology of sustained IOP following anti-VEGF injections, it is prudent to recognize the possibility of elevations in IOP in association with anti-VEGF therapy. Treating physicians should look for subtle optic nerve head changes and IOP measurements suspicious for glaucoma and have a low threshold for treating elevated IOP if the patient is likely to require multiple intravitreal anti-VEGF injections. Ocular hypertension following anti-VEGF injections appears to be amenable to anti-glaucoma treatment and every effort should be made to preserve the peripheral vision in these patients where central vision is already threatened by exudative AMD.

Differential roles of hyperglycemia and hypoinsulinemia in diabetes induced retinal cell death: evidence for retinal insulin resistance.

Diabetes pathology derives from the combination of hyperglycemia and hypoinsulinemia or insulin resistance leading to diabetic complications including diabetic neuropathy, nephropathy and retinopathy. Diabetic retinopathy is characterized by numerous retinal defects affecting the vasculature and the neuro-retina, but the relative contributions of the loss of retinal insulin signaling and hyperglycemia have never been directly compared. In this study we tested the hypothesis that increased retinal insulin signaling and glycemic normalization would exert differential effects on retinal cell survival and retinal physiology during diabetes. We have demonstrated in this study that both subconjunctival insulin administration and systemic glycemic reduction using the sodium-glucose linked transporter inhibitor phloridzin affected the regulation of retinal cell survival in diabetic rats. Both treatments partially restored the retinal insulin signaling without increasing plasma insulin levels. Retinal transcriptomic and histological analysis also clearly demonstrated that local administration of insulin and systemic glycemia normalization use different pathways to counteract the effects of diabetes on the retina. While local insulin primarily affected inflammation-associated pathways, systemic glycemic control affected pathways involved in the regulation of cell signaling and metabolism. These results suggest that hyperglycemia induces resistance to growth factor action in the retina and clearly demonstrate that both restoration of glycemic control and retinal insulin signaling can act through different pathways to both normalize diabetes-induced retinal abnormality and prevent vision loss.

Subconjunctivally implantable hydrogels with degradable and thermoresponsive properties for sustained release of insulin to the retina.

The objective of this work is to develop subconjunctivally implantable, biodegradable hydrogels for sustained release of intact insulin to the retina to prevent and treat retinal neurovascular degeneration such as diabetic retinopathy. The hydrogels are synthesized by UV photopolymerization of N-isopropylacrylamide (NIPAAm) monomer and a dextran macromer containing multiple hydrolytically degradable oligolactate-(2-hydroxyetheyl methacrylate) units (Dex-lactateHEMA) in 25:75 (v:v) ethanol:water mixture solvent. Insulin is loaded into the hydrogels during the synthesis process with loading efficiency up to 98%. The hydrogels can release biologically active insulin in vitro for at least one week and the release kinetics can be modulated by varying the ratio between NIPAAm and Dex-lactateHEMA and altering the physical size of the hydrogels. The hydrogels are not toxic to R28 retinal neuron cells in culture medium with 100% cell viability. The hydrogels can be implanted under the conjunctiva without causing adverse effects to the retina based on hematoxylin and eosin stain, immunostaining for microglial cell activation, and electroretinography. These subconjunctivally implantable hydrogels have potential for long-term periocular delivery of insulin or other drugs to treat diabetic retinopathy and other retinal diseases.

Sutureless levator plication by conjunctival route: a new technique.

The experiences of sutureless levator plication by conjunctival route surgery are described in 80 primary operations performed for all grades of congenital ptosis in the past 2 years. The surgical steps, postoperative care and postoperative complications are reviewed.