Immunobiology of a rationally-designed AAV2 capsid following intravitreal delivery in mice.

Adeno-associated virus serotype 2 (AAV2) is a viral vector that can be used to deliver therapeutic genes to diseased cells in the retina. One strategy for altering AAV2 vectors involves the mutation of phosphodegron residues, which are thought to be phosphorylated/ubiquitinated in the cytosol, facilitating degradation of the vector and the inhibition of transduction. As such, mutation of phosphodegron residues have been correlated with increased transduction of target cells, however, an assessment of the immunobiology of wild-type and phosphodegron mutant AAV2 vectors following intravitreal (IVT) delivery to immunocompetent animals is lacking in the current literature. In this study, we show that IVT of a triple phosphodegron mutant AAV2 capsid is associated with higher levels of humoral immune activation, infiltration of CD4 and CD8 T-cells into the retina, generation of splenic germinal centre reactions, activation of conventional dendritic cell subsets, and elevated retinal gliosis compared to wild-type AAV2 capsids. However, we did not detect significant changes in electroretinography arising after vector administration. We also demonstrate that the triple AAV2 mutant capsid is less susceptible to neutralisation by soluble heparan sulphate and anti-AAV2 neutralising antibodies, highlighting a possible utility for the vector in terms of circumventing pre-existing humoral immunity. In summary, the present study highlights novel aspects of rationally-designed vector immunobiology, which may be relevant to their application in preclinical and clinical settings.

Intraocular Pressure Spikes Following iStent Inject and the Relationship to Aqueous Outflow in Open Angle Glaucoma.

PRCIS: Adding trabecular bypass surgery (TBS) to phacoemulsification creates unpredictable short-term intraocular pressure (IOP) control that may be undesirable for patients with advanced glaucoma. Aqueous outflow (AO) responses after TBS are complex and probably multifactorial. PURPOSE: To assess IOP spikes in patients with open angle glaucoma up to 1 month after iStent inject and their relationship to AO patterns measured by hemoglobin video imaging (HVI). PARTICIPANTS AND METHODS: We studied IOP for 4 weeks after TBS with iStent inject in 105 consecutive eyes with open angle glaucoma (6 TBS only and 99 combined with phacoemulsification). The change in IOP after surgery at each time point was compared with baseline measurements and the prior postoperative visit. IOP-lowering medications were stopped on the day of surgery in all patients. A smaller pilot study of 20 eyes (TBS only = 6 and combined = 14) underwent concurrent HVI to observe and quantify perioperative AO. Aqueous column cross-sectional area (AqCA) of one nasal and one temporal aqueous vein was calculated at each time point, and qualitative observations were documented. An additional 5 eyes were studied after phacoemulsification only. RESULTS: Mean IOP for the entire cohort (preoperative 17.3 ± 5.6 mm Hg) was lowest the day after TBS (13.1 ± 5.0 mm Hg) and peaked at 1 week (17.2 ± 8.0 mm Hg), before stabilizing by 4 weeks (15.2 ± 5.2 mm Hg; P < 0.00001). The same IOP pattern was seen when separating the group into a larger cohort without HVI (respectively 15.9 ± 3.2 mm Hg, 12.8 ± 4.9 mm Hg, 16.4 ± 7.4 mm Hg, and 14.1 ± 4.1 mm Hg; N = 85, P < 0.00001) and the smaller HVI pilot study (respectively 21.4 ± 9.9 mm Hg, 14.2 ± 4.9 mm Hg, 20.2 ± 9.7 mm Hg, and 18.9 ± 7.6 mm Hg; N = 20, P < 0.001). More than 30% IOP elevation above baseline occurred in 13.3% of the entire cohort at 1 week after surgery. This increased to 46.7% when IOP was compared with 1 day after surgery. Inconsistent AqCA values and patterns of aqueous flow were demonstrated after TBS. AqCA after phacoemulsification alone was maintained or increased within 1 week in all 5 eyes. CONCLUSION: After iStent inject surgery in patients with open angle glaucoma, intraocular spikes were most commonly seen at 1 week. AO patterns were variable and additional studies are needed to understand the pathophysiology underlying IOP responses after this procedure.

Improving adeno-associated viral (AAV) vector-mediated transgene expression in retinal ganglion cells: comparison of five promoters.

Recombinant adeno-associated viral vectors (AAVs) are an effective system for gene transfer. AAV serotype 2 (AAV2) is commonly used to deliver transgenes to retinal ganglion cells (RGCs) via intravitreal injection. The AAV serotype however is not the only factor contributing to the effectiveness of gene therapies. Promoters influence the strength and cell-selectivity of transgene expression. This study compares five promoters designed to maximise AAV2 cargo space for gene delivery: chicken ß-actin (CBA), cytomegalovirus (CMV), short CMV early enhancer/chicken ß-actin/short ß-globulin intron (sCAG), mouse phosphoglycerate kinase (PGK), and human synapsin (SYN). The promoters driving enhanced green fluorescent protein (eGFP) were examined in adult C57BL/6J mice eyes and tissues of the visual system. eGFP expression was strongest in the retina, optic nerves and brain when driven by the sCAG and SYN promoters. CBA, CMV, and PGK had moderate expression by comparison. The SYN promoter had almost exclusive transgene expression in RGCs. The PGK promoter had predominant expression in both RGCs and AII amacrine cells. The ubiquitous CBA, CMV, and sCAG promoters expressed eGFP in a variety of cell types across multiple retinal layers including Müller glia and astrocytes. We also found that these promoters could transduce human retina ex vivo, although expression was predominantly in glial cells due to low RGC viability. Taken together, this promoter comparison study contributes to optimising AAV-mediated transduction in the retina, and could be valuable for research in ocular disorders, particularly those with large or complex genetic cargos.

Impact of AmaTea® Max on physiological measures and gaming performance in active gamers: A placebo-controlled, double-blind, randomized study.

Background and Aim: The activity of "gaming" has increased greatly in popularity in recent years, with many gamers using nutritional supplements to aid mood and gaming performance. We evaluated the impact of AmaTea® Max (referred to as AmaTea® throughout; a patented dietary supplement consisting of a blend of caffeine and polyphenol antioxidants), compared to both caffeine and a placebo, on gaming and cognitive performance in active gamers. Methods: Subjects reported to the lab on three occasions, separated by approximately 1 week. On each day, they had baseline measurements taken and then played the game Fortnite for four 1-h periods. Measures of cognitive performance, gaming performance, heart rate and blood pressure (BP), and blood cortisol were measured before and at selected times following gameplay. Results: Neither caffeine nor AmaTea® impacted gaming or cognitive performance in a statistically significant manner. However, a trend (P=0.075) was noted for the condition effect for kills/match, with values 21% higher for AmaTea® (1.84) compared to placebo (1.51), and 12% higher for AmaTea® compared to caffeine (1.63). Subjective mood was relatively unaffected, although a condition effect was noted for jittery (P=0.05), with values lower for placebo than for caffeine (P=0.02). BP was minimally elevated with both AmaTea® and caffeine, while cortisol followed the normal diurnal variation and was lower for placebo than AmaTea® and caffeine. Conclusion: AmaTea® modestly increased kills/match during gameplay. It is possible that a different gaming stimulus, varied time of gameplay, or different dosage of the supplement may have yielded different results. Relevance for Subjects: Active gamers who seek to use a dietary supplement for purposes of gaming performance may benefit slightly from ingestion of AmaTea® before gameplay while experiencing greater vigor and lower fatigue as compared to placebo.

The Effects of Trabecular Bypass Surgery on Conventional Aqueous Outflow, Visualized by Hemoglobin Video Imaging.

PRECIS: Hemoglobin Video Imaging (HVI) provides a noninvasive method to quantify aqueous outflow (AO) perioperatively. Trabecular bypass surgery (TBS) is able to improve, and in some cases re-establish, conventional AO. PURPOSE: The purpose of this study was to use HVI to illustrate and quantify effects of TBS on AO through the episcleral venous system. DESIGN: This is a prospective observational cohort study. PARTICIPANTS: Patients were recruited from Sydney Eye Hospital, Australia. The study included 29 eyes from 25 patients, 15 with glaucoma and 14 normal controls. TBS (iStent Inject) was performed on 14 glaucomatous eyes (9 combined phacoemulsification/TBS and 5 standalone TBS). Cataract surgery alone was performed on the remaining eye from the glaucoma group and 2 eyes from the control group. METHODS: We used HVI, a novel clinic-based tool, to visualize and quantify AO perioperatively during routine follow-up to 6 months. Angiographic blood flow patterns were observed within prominent aqueous veins on the nasal and temporal ocular surface. Aqueous column cross-section area (AqCA) was compared before and after surgery. MAIN OUTCOME MEASURES: AqCA, number of aqueous veins, intraocular pressure (IOP) before and after surgery, and number of IOP-lowering medications. RESULTS: Patients with glaucoma had reduced AqCA compared with normal controls (P=0.00001). TBS increased AqCA in 13 eyes at 1 month (n=14; P<0.002), suggesting improved AO. This effect was maintained at 6 months in 7 eyes (n=9, P≤0.05). All patients with unrecordable AO before surgery (n=3; 2 standalone TBS, 1 combined cataract/TBS) established measurable flow after TBS. IOP and/or medication burden became reduced in every patient undergoing TBS. Cataract surgery alone (n=3) increased AqCA in nasal and temporal vessels at 4 weeks after surgery. CONCLUSIONS: HVI provides a safe method for detecting and monitoring AO perioperatively in an outpatient setting. Improvement of AO into the episcleral venous system is expected after TBS and can be visualized with HVI. TBS is able to improve, and in some cases re-establish, conventional AO. Cataract surgery may augment this. Some aqueous veins were first seen after TBS and these patients had unstable postoperative IOP control, which possibly suggests reorganization of aqueous homeostatic mechanisms. HVI may confirm adequacy of surgery during short-term follow-up, but further work is required to assess the potential of HVI to predict surgical outcomes and assist with personalized treatment decisions.

Hemoglobin Video Imaging Provides Novel In Vivo High-Resolution Imaging and Quantification of Human Aqueous Outflow in Patients with Glaucoma.

Purpose: Noninvasive, detailed measurement of the dynamics of human aqueous outflow is difficult to achieve with currently available clinical tools. We used hemoglobin video imaging (HVI) to develop a technique to image and quantify human aqueous outflow noninvasively and in real time. Design: A prospective observational study to describe characteristics of aqueous veins and a pilot prospective interventional feasibility study to develop quantification parameters. Participants: Patients were recruited from the Cambridge University Hospitals NHS Foundation Trust Glaucoma clinic. The observational study included 30 eyes, and the pilot interventional feasibility study was performed on 8 eyes undergoing selective laser trabeculoplasty (SLT). Our SLT protocol also included the installation of pilocarpine and apraclonidine eye drops. Methods: Participants underwent HVI alongside their usual clinic visit. Main Outcome Measures: The change in cross-sectional area (CSA) of the aqueous column within episcleral veins was correlated with intraocular pressure (IOP) reduction and change in visual field mean deviation (MD) before and after intervention. Fluctuations in contrast and pixel intensity of red blood cells in an aqueous vein were calculated to compare the flow rate before and after intervention using autocorrelation analysis. Results: Hemoglobin video imaging enables the direct observation of aqueous flow into the vascular system. Aqueous is seen to centralize within a laminar venous column. Flow is pulsatile, and fluctuations of flow through globe pressure or compression of the aqueous vein are observed. There was a significant increase in the aqueous column after the administration of our SLT protocol (n = 13; P < 0.05). This correlated with the degree of IOP reduction (n = 13; Pearson's correlation coefficient 0.7; P = 0.007) and the improvement in MD observed postintervention (n = 8; Pearson's correlation coefficient 0.75; P = 0.03). Autocorrelation analysis demonstrated a faster rate of decay in an aqueous vein after intervention, indicating an increase in flow rate. Conclusions: Hemoglobin video imaging can be incorporated into a routine clinic slit-lamp examination to allow a detailed assessment and quantification of aqueous outflow in real time. It has the potential to be used to help target therapeutic interventions to improve aqueous outflow and further advance our understanding of aqueous outflow dysregulation in the pathogenesis of glaucoma.

Angiopoietins in Diabetic Retinopathy: Current Understanding and Therapeutic Potential.

Diabetic retinopathy (DR) is the commonest cause of blindness in the working-age population of the developed world. The molecular pathophysiology of DR is complex, and a complete spatiotemporal model of the disease is still being elucidated. Recently, a role for angiopoietin (Ang) proteins in the pathophysiology of DR has been proposed by several research groups, and several aspects of Ang signalling are being explored as novel therapeutic strategies. Here, we review the role of the Ang proteins in two important forms of DR, diabetic macular oedema and proliferative diabetic retinopathy. The function of the Ang proteins in regulating blood vessel permeability and neovascularisation is discussed, and we also evaluate recent preclinical and clinical studies highlighting the potential benefits of modulating Ang signalling as a treatment for DR.

Diabetic retinopathy: a complex pathophysiology requiring novel therapeutic strategies.

INTRODUCTION: Diabetic retinopathy (DR) is the leading cause of vision loss in the working age population of the developed world. DR encompasses a complex pathology, and one that is reflected in the variety of currently available treatments, which include laser photocoagulation, glucocorticoids, vitrectomy and agents which neutralize vascular endothelial growth factor (VEGF). Whilst these options demonstrate modest clinical benefits, none is yet to fully attenuate clinical progression or reverse damage to the retina. This has led to an interest in developing novel therapies for the condition, such as mediators of angiopoietin signaling axes, immunosuppressants, nonsteroidal anti-inflammatory drugs (NSAIDs), oxidative stress inhibitors and vitriol viscosity inhibitors. Further, preclinical research suggests that gene therapy treatment for DR could provide significant benefits over existing treatments options. AREAS COVERED: Here we review the pathophysiology of DR and provide an overview of currently available treatments. We then outline recent advances made towards improved patient outcomes and highlight the potential of the gene therapy paradigm to revolutionize DR management. EXPERT OPINION: Whilst significant progress has been made towards our understanding of DR, further research is required to enable the development of a detailed spatiotemporal model of the disease. In addition, we hope that improvements in our knowledge of the condition facilitate therapeutic innovations that continue to address unmet medical need and improve patient outcomes, with a focus on the development of targeted medicines.

Authentic tart cherry juice reduces markers of inflammation in overweight and obese subjects: a randomized, crossover pilot study.

Subclinical inflammation is frequently noted in chronic diseases such as diabetes, cardiovascular disease (CVD) and obesity. Accumulating epidemiological evidence demonstrates that diets rich in vegetables and fruits, e.g., cherries, may significantly reduce the risk of chronic disease, in part, via antioxidant and anti-inflammatory activities. In this randomized, placebo-controlled crossover study, we recruited 10 at-risk participants (38.1 ± 12.5 years; 8 females, 2 males) with BMI >25.0 kg m-2 (32.2 ± 4.6 kg m-2; 5 obese, 5 overweight) to consume 240 mL (8 ounces) daily of either 100% tart cherry juice (TCJ) or an alternate placebo beverage, for 4 weeks with a 2-week intervening washout period before switching to the alternate beverage for four weeks. Fasting blood samples were collected at the beginning and end of each arm for measurement of biomarkers of inflammation. The erythrocyte sedimentation rate (ESR), an indicator of chronic inflammation, was significantly (p < 0.05) lower with TCJ than with the placebo beverage, which increased ESR by 19%. Mean baseline hsCRP, an indicator of acute inflammation, was 7.0 ± 5.2 mg L-1 and consumption of TCJ did not affect hsCRP levels. The chemokine MCP-1 and cytokine TNF-alpha were lower in participants after consuming TCJ compared to those consuming the placebo beverage. Plasma IL-6 and IL-l0 were not different between treatments. Collectively, the data suggest that authentic 100% TCJ may reduce biomarkers of inflammation often noted in chronic disease and may be a preferable dietary selection compared to artificially flavored beverages with added sugars.

Neuroprotection of retinal ganglion cells by a novel gene therapy construct that achieves sustained enhancement of brain-derived neurotrophic factor/tropomyosin-related kinase receptor-B signaling.

Previous studies have demonstrated that intravitreal delivery of brain-derived neurotrophic factor (BDNF) by injection of recombinant protein or by gene therapy can alleviate retinal ganglion cell (RGC) loss after optic nerve injury. BDNF gene therapy can improve RGC survival in experimental models of glaucoma, the leading cause of irreversible blindness worldwide. However, the therapeutic efficacy of BDNF supplementation alone is time limited at least in part due to BDNF receptor downregulation. Tropomyosin-related receptor kinase-B (TrkB) downregulation has been reported in many neurological diseases including glaucoma, potentially limiting the effect of sustained or repeated BDNF delivery.Here, we characterize a novel adeno-associated virus (AAV) gene therapy (AAV2 TrkB-2A-mBDNF) that not only increases BDNF production but also improves long-term neuroprotective signaling by increasing expression of the BDNF receptor (TrkB) within the inner retina. This approach leads to significant and sustained elevation of survival signaling pathways ERK and AKT within RGCs over 6 months and avoids the receptor downregulation which we observe with treatment with AAV2 BDNF alone. We validate the neuroprotective efficacy of AAV2 TrkB-2A-mBDNF in a mouse model of optic nerve injury, where it outperforms conventional AAV2 BDNF or AAV2 TrkB therapy, before showing powerful proof of concept neuroprotection of RGCs and axons in a rat model of chronic intraocular pressure (IOP) elevation. We also show that there are no adverse effects of the vector on retinal structure or function as assessed by histology and electroretinography in young or aged animals. Further studies are underway to explore the potential of this vector as a candidate for progression into clinical studies to protect RGCs in patients with glaucoma and progressive visual loss despite conventional IOP-lowering treatment.

Efficacy and Safety of the Ab-interno Xen Gel Stent After Failed Trabeculectomy.

AIMS: To assess the efficacy and safety of the Xen gel stent in reducing intraocular pressure (IOP) in eyes with prior failed trabeculectomy and to determine the frequency of complications and further intervention. METHODS: Retrospective case note review of all patients with prior trabeculectomy undergoing Xen surgery across 5 centers from August 2015 to May 2017. RESULTS: In total, 17 surgeries were reviewed. IOP reduced from 21.5 (±2.4) mm Hg preoperatively to 13.6 (±3.4) mm Hg at month 12 (P<0.05). Medication usage reduced from 2.8 (±0.6) preoperatively to 1.0 (±1.3) at month 12 (P<0.05). Adverse events included: numerical hypotony (IOP<6 mm Hg) in 4 cases (23.5%) that all resolved spontaneously, IOP spike of ≥30 mm Hg in 2 (11.8%) cases and transient occlusion of the implant by iris in 1 (5.9%) case. Secondary filtration surgery (Baerveldt tube implantation) was required in 2 (11.8%) cases. Postoperative bleb intervention was required in 9 cases (52.9%), usually in the first month after surgery. CONCLUSIONS: Xen reduces IOP and number of medications in eyes with failed trabeculectomy. Detailed preoperative conjunctival assessment and targeted stent placement is required. Prospective data and follow-up beyond 12 months are required but Xen seems a viable, effective, and safe option after failed trabeculectomy.

Factors predicting the success of trabeculectomy bleb enhancement with needling.

BACKGROUND: Bleb needling is widely used to restore flow and lower intraocular pressure (IOP) in a failing trabeculectomy. We aimed to measure the safety and efficacy of needling in a large cohort and identify factors that were associated with success and failure. METHODS: This retrospective audit included all patients who underwent needling at Addenbrooke's Hospital, Cambridge over a 10-year period. Data were available on 91 patients (98% of patients identified), including 191 needlings on 96 eyes. Success was defined as IOP below 21 mm Hg or 16 mm Hg or 13 mm Hg consistently, without reoperation or glaucoma medication. Risk factors for failure were assessed by Cox proportional hazard regression and Kaplan-Meier curves. RESULTS: Success defined as IOP <16 mm Hg was 66.6% at 12 months and 53% at 3 years and success defined as IOP <21 mm Hg was 77.1% at 12 months and 73.1% at 3 years. Failure after needling was most common in the first 6 months. Factors that predicted failure were flat or fibrotic blebs (non-functional) and no longer injected, while success was predicted by achieving a low IOP immediately after needling. No significant complications were identified. CONCLUSION: Needling was most successful soon after trabeculectomy, but resuscitation of a long-failed trabeculectomy had lower likelihood of success. The safety and efficacy compare favourably with alternative treatment approaches.

Design of a Novel Gene Therapy Construct to Achieve Sustained Brain-Derived Neurotrophic Factor Signaling in Neurons.

Brain-derived neurotrophic factor (BDNF) acting through the tropomyosin-related receptor-B (TrkB) is an important signaling system for the maintenance and survival of neurons. Gene therapy using either recombinant adeno-associated virus (AAV) or lentiviral vectors can provide sustained delivery of BDNF to tissues where reduced BDNF signaling is hypothesized to contribute to disease pathophysiology. However, elevation in BDNF at target sites has been shown to lead to a downregulation of TrkB receptors, thereby reducing the effect of chronic BDNF delivery over time. A novel gene sequence has been designed coding both the ligand (BDNF) and the TrkB receptor in a single transgene separated by a short viral-2A sequence. The single transgene is efficiently processed intracellularly in vitro and in vivo to yield the two mature proteins, which are then independently transported to their final cellular locations: TrkB receptors to the cell surface, and BDNF contained within secretory vesicles. To accommodate the coding sequences of both BDNF and TrkB receptors within the narrow confines of the AAV vectors (4.7 kb pairs), the coding region for the pro-domain of BDNF was removed and the signal peptide sequence modified to improve production, intracellular transport, and secretion of mature BDNF (mBDNF). Intracellular processing and efficacy was shown in HEK293 cells and SH-SY5Y neuroblastoma cells using plasmid DNA and after incorporating the TrkB-2A-mBDNF into an AAV2 vector. Increased BDNF/TrkB-mediated intracellular signaling pathways were observed after AAV2 vector transfection while increased TrkB phosphorylation could be detected in combination with neuroprotection from hydrogen peroxide-induced oxidative stress. Correct processing was also shown in vivo in mouse retinal ganglion cells after AAV2 vector administration to the eye. This novel construct is currently being investigated for its efficacy in animal models to determine its potential to progress to human clinical studies in the future.

Neuroprotective Effects of Human Mesenchymal Stem Cells and Platelet-Derived Growth Factor on Human Retinal Ganglion Cells.

Optic neuropathies such as glaucoma occur when retinal ganglion cells (RGCs) in the eye are injured. Strong evidence suggests mesenchymal stem cells (MSCs) could be a potential therapy to protect RGCs; however, little is known regarding their effect on the human retina. We, therefore, investigated if human MSCs (hMSCs), or platelet-derived growth factor (PDGF) as produced by hMSC, could delay RGC death in a human retinal explant model of optic nerve injury. Our results showed hMSCs and the secreted growth factor PDGF-AB could substantially reduce human RGC loss and apoptosis following axotomy. The neuroprotective pathways AKT, ERK, and STAT3 were activated in the retina shortly after treatments with labeling seen in the RGC layer. A dose dependent protective effect of PDGF-AB was observed in human retinal explants but protection was not as substantial as that achieved by culturing hMSCs on the retina surface which resulted in RGC cell counts similar to those immediately post dissection. These results demonstrate that hMSCs and PDGF have strong neuroprotective action on human RGCs and may offer a translatable, therapeutic strategy to reduce degenerative visual loss. Stem Cells 2018;36:65-78.

Tablets at the bedside - iPad-based visual field test used in the diagnosis of Intrasellar Haemangiopericytoma: a case report.

BACKGROUND: In the assessment of a pituitary mass, objective visual field testing represents a valuable means of evaluating mass effect, and thus in deciding whether surgical management is warranted. CASE PRESENTATION: In this vignette, we describe a 73 year-old lady who presented with a three-week history of frontal headache, and 'blurriness' in the left side of her vision, due to a WHO grade III anaplastic haemangiopericytoma compressing the optic chiasm. We report how timely investigations, including an iPad-based visual field test (Melbourne Rapid Field, (MRF)) conducted at the bedside aided swift and appropriate management of the patient. CONCLUSIONS: We envisage such a test having a role in assessing bed-bound patients in hospital where access to formal visual field testing is difficult, or indeed in rapid testing of visual fields at the bedside to screen for post-operative complications, such as haematoma.

Surgical outcomes of trabeculectomy and glaucoma drainage implant for uveitic glaucoma and relationship with uveitis activity.

IMPORTANCE: This study provides ophthalmologists who manage uveitic glaucoma with important information on factors that can affect the success of surgical management of this challenging disease. BACKGROUND: This study examines surgical outcomes of trabeculectomy and glaucoma device implant (GDI) surgery for uveitic glaucoma, in particular the effect of uveitis activity on surgical outcomes. DESIGN: Retrospective chart review at a tertiary institution. SAMPLES: Eighty-two cases with uveitic glaucoma (54 trabeculectomies and 28 (GDI) surgeries) performed between 1 December 2006 and 30 November 2014. METHODS: Associations of factors with surgical outcomes were examined using univariate and multivariate analysis. MAIN OUTCOME MEASURES: Surgical outcomes as defined in Guidelines from World Glaucoma Association. RESULTS: Average follow up was 26.4 ± 21.5 months. Overall qualified success rate of the trabeculectomies was not statistically different from GDI, being 67% and 75%, respectively (P = 0.60). Primary and secondary GDI operations showed similar success rates. The most common postoperative complication was hypotony (~30%). Active uveitis at the time of operation was higher in trabeculectomy compared with GDI group (35% vs. 14%). Active uveitis at the time of surgery did not significantly increase risk of failure for trabeculectomies. Recurrence of uveitis was significantly associated with surgical failure in trabeculectomy group (odds ratio 4.8, P = 0.02) but not in GDI group. CONCLUSIONS AND RELEVANCE: Surgical success rate of GDI was not significantly different from trabeculectomy for uveitic glaucoma in this study. Regular monitoring, early and prolonged intensive treatment of ocular inflammation is important for surgical success particularly following trabeculectomy.

Retinal regeneration mechanisms linked to multiple cancer molecules: A therapeutic conundrum.

Over the last decade, a large number of research articles have been published demonstrating regeneration and/or neuroprotection of retinal ganglion cells following manipulation of specific genetic and molecular targets. Interestingly, of the targets that have been identified to promote repair following visual system damage, many are genes known to be mutated in different types of cancer. This review explores recent literature on the potential for modulating cancer genes as a therapeutic strategy for visual system repair and looks at the potential clinical challenges associated with implementing this type of therapy. We also discuss signalling mechanisms that have been implicated in cancer and consider how similar mechanisms may improve axonal regeneration in the optic nerve.

Primary open-angle glaucoma.

Glaucoma is an optic neuropathy that is characterized by the progressive degeneration of the optic nerve, leading to visual impairment. Glaucoma is the main cause of irreversible blindness worldwide, but typically remains asymptomatic until very severe. Open-angle glaucoma comprises the majority of cases in the United States and western Europe, of which, primary open-angle glaucoma (POAG) is the most common type. By contrast, in China and other Asian countries, angle-closure glaucoma is highly prevalent. These two types of glaucoma are characterized based on the anatomic configuration of the aqueous humour outflow pathway. The pathophysiology of POAG is not well understood, but it is an optic neuropathy that is thought to be associated with intraocular pressure (IOP)-related damage to the optic nerve head and resultant loss of retinal ganglion cells (RGCs). POAG is generally diagnosed during routine eye examination, which includes fundoscopic evaluation and visual field assessment (using perimetry). An increase in IOP, measured by tonometry, is not essential for diagnosis. Management of POAG includes topical drug therapies and surgery to reduce IOP, although new therapies targeting neuroprotection of RGCs and axonal regeneration are under development.

A Method for the Isolation and Culture of Adult Rat Retinal Pigment Epithelial (RPE) Cells to Study Retinal Diseases.

Diseases such as age-related macular degeneration (AMD) affect the retinal pigment epithelium (RPE) and lead to the death of the epithelial cells and ultimately blindness. RPE transplantation is currently a major focus of eye research and clinical trials using human stem cell-derived RPE cells are ongoing. However, it remains to be established to which extent the source of RPE cells for transplantation affects their therapeutic efficacy and this needs to be explored in animal models. Autotransplantation of RPE cells has attractions as a therapy, but existing protocols to isolate adult RPE cells from rodents are technically difficult, time-consuming, have a low yield and are not optimized for long-term cell culturing. Here, we report a newly devised protocol which facilitates reliable and simple isolation and culture of RPE cells from adult rats. Incubation of a whole rat eyeball in 20 U/ml papain solution for 50 min yielded 4 × 10(4) viable RPE cells. These cells were hexagonal and pigmented upon culture. Using immunostaining, we demonstrated that the cells expressed RPE cell-specific marker proteins including cytokeratin 18 and RPE65, similar to RPE cells in vivo. Additionally, the cells were able to produce and secrete Bruch's membrane matrix components similar to in vivo situation. Similarly, the cultured RPE cells adhered to isolated Bruch's membrane as has previously been reported. Therefore, the protocol described in this article provides an efficient method for the rapid and easy isolation of high quantities of adult rat RPE cells. This provides a reliable platform for studying the therapeutic targets, testing the effects of drugs in a preclinical setup and to perform in vitro and in vivo transplantation experiments to study retinal diseases.

The Capacity of Eye Care Services for Patients with Glaucoma in Botswana.

PURPOSE: To evaluate current delivery of glaucoma care in Botswana; in particular, the service infrastructure available and glaucoma-related workload. METHODS: A multi-center cross-sectional study was undertaken comprising government eye care institutions and ophthalmic personnel across Botswana. Data on human resources, equipment types and numbers, diagnostic criteria routinely used, treatments routinely provided, and new and repeat glaucoma consultations were obtained through quantitative and qualitative surveys. RESULTS: In 27 government eye care institutions there were two general ophthalmologists, neither of whom had a subspecialty interest in glaucoma, 64 ophthalmic nurses, two optometrists, one low vision therapist, one refractionist, and two equipment technicians. Only 8.5% of available ophthalmic human resources were taken up with provision of glaucoma care. About 1/3 of hospitals did not have tonometers, most primary hospitals lacked slit lamp biomicroscopes and most hospitals lacked sensitive diagnostic equipment. A diagnosis of glaucoma was made by either an ophthalmic nurse or an ophthalmologist, but only 10% of institutions could meet recommendations for follow-up assessment. Topical glaucoma medications were prescribed by almost all hospital clinics, usually by ophthalmic nurses. Drug choices were largely determined by local availability. Glaucoma surgery accounted for 0.8% of total eye operations. Glaucoma patients took up 8.5% of total clinic visits. The total number of glaucoma visits was highest in the two hospitals with ophthalmologists. New glaucoma cases took up 10.3% of total glaucoma visits. CONCLUSION: This study highlights the challenges faced in caring for glaucoma patients in Botswana; in particular, lack of professional human resources, equipment and availability of effective treatments.