Dexamethasone reverses the effects of high glucose on human retinal endothelial cell permeability and proliferation in vitro.

Diabetic macular oedema (DMO), a leading cause of preventable visual loss in the working population, is caused by an increase in microvascular endothelial cell permeability, and its prevalence is on the increase in parallel with the rising worldwide prevalence of diabetes. It is known that retinal vascular leakage in DMO is contributed to by VEGF upregulation as well as non-VEGF dependent inflammatory pathways, and the potential use of anti-inflammatory agents such as the glucocorticoids, including dexamethasone are being extensively studied. However, the mechanisms of action of dexamethasone in DMO reduction are not fully understood. Using human primary retinal endothelial cells (REC) the in vitro effect of dexamethasone in modulating the proliferation, permeability and gene expression of key tight and adheren junction components, and the expression of angiopoietins (Ang) 1 and 2 in high (25 mM) glucose conditions were investigated. High glucose decreased REC proliferation, an effect that was reversed by dexamethasone. High glucose conditions significantly increased REC permeability and decreased claudin-5, occludin and JAM-A gene expression; dexamethasone was effective in partially reversing these changes, restoring EC permeability to the normal or near normal state. High glucose levels resulted in reduction of Ang1 secretion, although Ang2 levels were consistently high. DEX increased Ang1 and decreased Ang2, indicating that the balance of Ang1/Ang2 may be important in determining functional changes in REC under high glucose conditions.

The effect of hyperglycaemia on permeability and the expression of junctional complex molecules in human retinal and choroidal endothelial cells.

Diabetic retinopathy is the leading cause of preventable blindness in the working population and its prevalence continues to increase as the worldwide prevalence of diabetes grows. Diabetic choroidopathy is less well studied and occurs in the late stages of diabetic eye disease. The main cause of visual loss in diabetic eye disease is diabetic macular oedema caused by an increase in microvascular endothelial permeability. Endothelial cell permeability is influenced by multiple factors which have not been fully elucidated, particularly in human models. In addition, the gene and protein expression between retinal and choroidal endothelial cells, even in humans, has been shown to be heterogeneous. The aim of this project was to determine, in vitro, the effect of high glucose (25 mM) on human paracellular permeability in retinal and choroidal endothelial cells. The expression of selected tight junction molecules (Occludin, Claudin-5, JAM-A and JAM-C) and adheren junction (VE-Cadherin) molecules was also compared between retinal and choroidal endothelial cells and with high glucose. High glucose conditions significantly increased the permeability in both retinal and choroidal endothelial cells monolayers although the increase was higher in retinal endothelial cells. Under normal glucose culture conditions microarray analysis determined that occludin and claudin-5 gene expression was higher in retinal endothelial cells than choroidal endothelial cells, and western blotting indicated that claudin-5 protein expression was also higher in retinal endothelial cells whilst JAM-A, and C and VE-Cadherin levels were similar. In retinal endothelial cells exposed to high glucose claudin-5, occludin and JAM-A was found to be reduced, whereas the expression of VE-Cadherin and JAM-C was unchanged when evaluated with western blotting, immunofluorescence and qPCR. None of the proteins were significantly decreased by high glucose in choroidal endothelial cells. The increase in retinal endothelial cell permeability is likely caused by a decrease in selective tight junction protein expression, leading to increased paracellular permeability. This may indicate differences in junctional molecule regulation of permeability in retinal compared to choroidal endothelial cells.

Prevalence of age-related macular degeneration in an elderly UK Caucasian population-The Bridlington Eye Assessment Project: a cross-sectional study.

ImportanceThere is paucity of data on prevalence and disease asymmetry of age-related macular degeneration (AMD), particularly the earlier stages, in the UK population.Objective and PurposeTo determine the prevalence of age-related macular degeneration in an elderly Caucasian UK population.DesignCross-sectional population study, 2002-2006.ParticipantsResidents in the study area of Bridlington aged 65 years and older.MethodsFull-ophthalmic examination was undertaken in 3549 participants, of eligible 6319 Caucasian population (response rate of 56%). Non-stereoscopic Colour fundus photographs (30°) were graded masked using a modified Rotterdam Classification for 3475 (98%) participants with gradable images. Prevalence for different AMD grades were calculated. Demographic details were analysed then integrated with the AMD gradings for full analysis. Prevalence rates for the different AMD Grades were calculated, as well as the age-specific prevalences.ResultsAMD prevalence in the worst eye were 38.5% grade 0, 41.4% grade 1, 12.8% grade 2, 2.8% grade 3, and 4.6% grade 4. Geographic atrophy (grade 4a) occurred in 2.5%, and neovascular AMD (grade 4b) in 1.8%. Prevalence increased with age such that grade 4 (advanced) AMD was 2.2% in the 65-69 years group, 15.8% for the 85-90, and 21.2% for over 90 years. There was significant asymmetry between the two eyes of individuals with advanced AMD (P<0.001), such that vision loss was unilateral. Persons with more advanced AMD grades were more likely to be dissatisfied with their vision.ConclusionsAdvanced AMD occurs more commonly in the UK Caucasian population than previously reported. Significant asymmetry between the two eyes occurs in individuals with unilateral advanced AMD so that visual impairment statistics do not represent true prevalence of advanced AMD. Persons with more advanced AMD were more likely to be dissatisfied with their vision.

The management of retinal vein occlusion: is interventional ophthalmology the way forward?

Retinal vein occlusions (RVO) are the second commonest sight threatening vascular disorder. Despite its frequency treatments for RVO are unsatisfactory and include several that have not been tested by large, well designed, prospective, randomised controlled trials. There is also the lack of long term follow up in many of the available small uncontrolled studies, and the timings of interventions are haphazard. This review aims to evaluate the current knowledge relating to the pathogenesis, suggested treatments for the different types of RVO, and their complications. Isovolaemic haemodilution is of limited benefit and should be avoided in patients with concurrent cardiovascular, renal, or pulmonary morbidity. Evidence to date does not support any therapeutic benefit from radial optic neurotomy, optic nerve decompression, or arteriovenous crossing sheathotomy on its own. Vitrectomy combined with intravenous thrombolysis may offer promise for central RVO. Similarly, vitrectomy combined with arteriovenous sheathotomy intravenous tissue plasminogen activator may offer benefits for branch RVO. RVOs occur at significantly high frequency to allow future prospective randomised controlled studies to be conducted to evaluate the role of different therapeutic modalities singly or in combination.

Effect of age related macular degeneration on the Eger macular stressometer photostress recovery time.

AIM: To assess the repeatability of Eger macular stressometer (EMS) measures of photostress recovery and determine their association with other measures of visual function. METHODS: EMS photostress recovery time was measured in 90 patients with bilateral exudative age related macular degeneration (AMD), 19 with bilateral atrophic AMD and 47 with both forms of the condition (mean age 79 (SD 13) years). Measurements were made on two occasions separated by 1 year. Intrasession repeatability was assessed by repeating the measures after a 10 minute recovery period at the first visit. Distance visual acuity was measured with a logMAR chart, near visual acuity with a MNRead chart at 25 cm, contrast sensitivity with a Pelli-Robson chart, and the presence of central visual disturbance assessed with an Amsler grid. A questionnaire was used to assess self reported difficulties with glare recovery. RESULTS: The average EMS recovery time was 11.0 (SD 8.9) seconds, decreasing by 1.6 (5.2) seconds on repeated measurement (p<0.05). EMS photostress recovery was not correlated with visual function measures or subjective difficulties with lights (p>0.05). EMS photostress recovery time did not predict those whose vision decreased over the following year compared with those among whom it remained stable. CONCLUSIONS: The EMS test is not a useful tool in determining the severity or progression of AMD.

Prevalence of reticular pseudodrusen in newly presenting adult onset foveomacular vitelliform dystrophy.

PurposeTo report the association and prevalence of reticular pseudodrusen (RPD) in eyes with newly presenting adult onset foveomacular vitelliform dystrophy (AFVD). To compare the strength of association with other pathologies resulting from dysfunction of the choroid-Bruch's membrane-retinal pigment epithelium (RPE) complex, including eyes with geographic atrophy (GA) and angioid streaks.MethodsRetrospective single-centre review of all consecutive newly presenting AFVD. Multimodal imaging with spectral domain optical coherence tomography, fundus photographs, red-free/blue light images, and fundus fluorescein angiograms were graded for the presence of RPD. For comparison, all consecutive newly presenting cases of GA and eyes with angioid streaks were studied.ResultsFifteen (15) patients were identified with AFVD (mean age of 77.3 years; 73.3% female). Mean age of patients with AFVD and RPD was 80.5 years (SD 3.7), whereas that of patients with AFVD without RPD was 75.1 years (SD 7.0). This age difference did not reach statistical significance, P=0.1. Six (40%) had identifiable RPD; being a bilateral finding in 100% of patients. No males with AFVD and RPD were identified. A total of 92 eyes presented with GA. Twenty-three (23) of these (25.0%) had RPD. Twelve (12) patients presented with identifiable angioid streaks, with 4 (36.4%) having RPD.ConclusionRPD are a frequent finding in eyes with newly presenting AFVD; not being restricted to AMD, but a finding common among diseases where pathophysiological mechanisms involve damage to Bruch's membrane and the RPE, whether genetic or degenerative. Our study supports the concept that they occur with high but variable frequencies in eyes with various pathologies.

Verteporfin photodynamic therapy of choroidal neovascularization in angioid streaks: one-year results of a prospective case series.

PURPOSE: To report the 12-month results on the use of verteporfin photodynamic therapy (PDT) in the treatment of choroidal neovascularization (CNV) secondary to angioid streaks. STUDY DESIGN: Five-center prospective case series. METHODS: Patients with CNV secondary to angioid streaks who were treated with PDT were recruited and followed up at 3-month intervals for 12 months, with the addition of visits at 1.5 and 4.5 months if deemed appropriate by the investigator. Best-corrected visual acuity (BCVA) was measured at each visit after full refraction or with their current distance spectacles using Early Treatment Diabetic Retinopathy Study logarithm of the minimum angle of resolution charts. Stereoscopic fundus fluorescein angiography was used to determine baseline lesion characteristics and location. RESULTS: Twenty-two patients were recruited (23 eyes, 16 with subfoveal CNV and 7 with juxtafoveal; all classic no occult). Seventeen patients (77%) had angioid streaks secondary to pseudoxanthoma elasticum. In the subfoveal group, median BCVA at baseline was 49 letters (approximate Snellen equivalent, 20/100) and was 46 at 12 months (approximate Snellen equivalent, 20/125). Twelve of 16 eyes (75%) lost fewer than 8 letters, whereas 14 of 16 eyes (88%) lost fewer than 15 letters. The mean CNV greatest linear dimension (GLD) was 2520 microm at baseline. At 12 months, 7 of 16 eyes with subfoveal CNV at baseline were leaking (GLD = 3220 microm; P = 0.62). The mean number of treatments in the first 12 months was 2.9. In the juxtafoveal group, the median BCVA at baseline was 66 letters (approximate Snellen equivalent, 20/50) and was 51 letters at 12 months (approximate Snellen equivalent, 20/100). Two of 7 eyes (29%) gained 8 or more letters at the 12-month examination, whereas 4 of 7 eyes (57%) lost more than 15 letters. The mean CNV GLD at baseline was 1890 microm. At 12 months, 1 of 7 eyes with juxtafoveal CNV at baseline was leaking. Choroidal neovascularization progressed from juxtafoveal to subfoveal location during the follow-up period in 4 of 7 eyes. The mean number of treatments in the first 12 months was 3.4. No side effects were noted in either patient group. CONCLUSIONS: This small series suggests that treatment of CNV secondary to angioid streaks with verteporfin PDT seems to limit visual loss in most patients through the first 12 months of follow-up, particularly in those with subfoveal lesions at baseline.

Isolation, culture, and characterisation of human macular inner choroidal microvascular endothelial cells.

AIM: To develop a method for the reliable isolation of adult human macular inner choroidal endothelial cells (ICECs) and to subsequently characterise them for their expression of a range of endothelial cell associated surface markers. METHOD: Human ICECs were isolated after manual dissection of maculas from fresh human posterior segments. Following enzyme digestion to form a single cell suspension, the ICECs were isolated using anti-CD31 coated Dynabeads. The isolated cells were grown in culture and examined for typical endothelial cell morphology, surface expression of vWf, CD 31, CD 105, VEGF receptors 1 and 2, and expression of E-selectin after stimulation with TNF-alpha. The cells were also examined for their ability to form fenestrations and capillary-like tubes in Matrigel. RESULTS: The method enabled the rapid isolation of viable cells that demonstrated typical endothelial cobblestone morphology in culture. The cells stained positive for CD31, vWf, CD105, VEGF receptors 1 and 2, and E-selectin (after stimulation with TNF-alpha). The cells stained negative for alpha smooth muscle actin and fibroblast surface protein. The cells also developed fenestrations when cultured on fibronectin coated plates and formed capillary-like tubes structures when cultured on Matrigel. CONCLUSIONS: This technique isolates cells from the human macular inner choroid that display features consistent with vascular endothelial cells. These cells could subsequently be used to further the understanding of the pathophysiological mechanisms of diseases of the inner choroid, such as choroidal neovascularisation.

The efficacy of sirolimus in the treatment of patients with refractory uveitis.

AIMS: To determine the efficacy of sirolimus in the treatment of patients with severe non-infectious uveitis. METHODS: Eight patients with severe non-infectious uveitis were recruited to an open study. Inclusion criteria were limited to patients whose disease was not controlled with at least two or more separate steroid sparing immunosuppressants (either because of unacceptable side effects or ineffectiveness of the drug) or who required regular doses of corticosteroids either as high dose systemic or orbital floor injections in order to control their disease. Intraocular inflammation, visual acuity, symptoms, corticosteroid burden, drug toxicity, and side effects were monitored. RESULTS: Sirolimus therapy was effective in five of the eight patients, all of whom had their dose of corticosteroids reduced or discontinued. Treatment in three patients was considered a failure as it caused intolerable side effects and/or failed to control the uveitis. Side effects were common and were typically gastrointestinal or cutaneous in nature. The severity of symptoms was dose dependent in most cases and occurred at trough blood levels above 25 ng/ml. CONCLUSION: Sirolimus is an effective and potent immunosuppressive treatment in the majority of patients with non-infectious uveitis and can reduce the need for long term supplementary corticosteroid therapy. Further studies are required to establish the long term efficacy and safety of sirolimus alone or in combination with other steroid sparing immunosuppressants.

A review of therapies for diabetic macular oedema and rationale for combination therapy.

Diabetic macular oedema (DMO) is responsible for significant visual impairment in diabetic patients. The primary cause of DMO is fluid leakage resulting from increased vascular permeability through contributory anatomical and biochemical changes. These include endothelial cell (EC) death or dysfunction, pericyte loss or dysfunction, thickened basement membrane, loss or dysfunction of glial cells, and loss/change of EC Glycocalyx. The molecular changes include increased reactive oxygen species, pro-inflammatory changes: advanced glycation end products, intracellular adhesion molecule-1, Complement 5-9 deposition and cytokines, which result in increased paracellular permeability, tight junction disruption, and increased transcellular permeability. Laser photocoagulation has been the mainstay of treatment until recently when pharmacological treatments were introduced. The current treatments for DMO target reducing vascular leak in the macula once it has occurred, they do not attempt to treat the underlying pathology. These pharmacological treatments are aimed at antagonising vascular endothelial growth factor (VEGF) or non-VEGF inflammatory pathways, and include intravitreal injections of anti-VEGFs (ranibizumab, aflibercept or bevacizumab) or steroids (fluocinolone, dexamethasone or triamcinolone) as single therapies. The available evidence suggests that each individual treatment modality in DMO does not result in a completely dry macula in most cases. The ideal treatment for DMO should improve vision and improve morphological changes in the macular (eg, reduce macular oedema) for a significant duration, reduced adverse events, reduced treatment burden and costs, and be well tolerated by patients. This review evaluates the individual treatments available as monotherapies, and discusses the rationale and potential for combination therapy in DMO. A comprehensive review of clinical trials related to DMO and their outcomes was completed. Where phase III randomised control trials were available, these were referenced, if not available, phase II trials have been included.