C.J. Barry

Expression of cell adhesion molecules and vascular endothelial growth factor in experimental choroidal neovascularisation in the rat.

AIMS To investigate the longevity and reproducibility of choroidal neovascularisation (CNV) induced by krypton laser photocoagulation in the rat. The presence of cell adhesion molecules (CAMs) and vascular endothelial growth factor (VEGF) during the development of CNV was also studied. METHODS 67 pigmented rats underwent retinal photocoagulation by krypton laser. The eyes were examined by either single or serial fluorescein angiography at 3 days, 1, 2–3, 4–5, 7–8, and 12 weeks post photocoagulation. The expression of CAMs (ICAM-1, E-selectin, and CD44) and VEGF post photocoagulation was studied by immunohistochemistry. RESULTS CNV related fluorescein leakage appeared in 46.4% of 766 laser spots delivered to the 58 eyes that were tested at 2–3 weeks post treatment. The ratio of hyperfluorescent laser sites did not change significantly at 8 weeks post laser. The number of leaky spots was independent of the total number of lesions delivered to each eye (at 2–3 weeks post laser 10–15 spots/eye: 44% and 25–30 spots/eye: 49%; t=0.7673; p=0.3903). Nine eyes were followed by serial angiography between 2 and 12 weeks. The laser spots with fluorescein leakage at 2 weeks (51.5%) remained leaky at 12 weeks (51.5%). Histopathologically, macrophage accumulation peaked at 5 days and CNV was firstly observed at 1 week post photocoagulation. ICAM-1, E-selectin, CD44, and VEGF were maximally induced at 3–5 days post laser photocoagulation, and were localised to RPE, choroidal vascular endothelial, and inflammatory cells. VEGF was also detected in intravascular leucocytes at the sites of laser lesions. CONCLUSIONS These studies demonstrated that krypton laser photocoagulation can be successfully used to produce lesions similar to those of human CNV. The response induced remained present for an extended period of time (12 weeks), thus offering a potential model to screen candidate CNV inhibitory agents. In addition, it is proposed that the expression of ICAM-1, E-selectin, CD44, and VEGF before new vessel formation might be linked to the initiation of CNV.

Correlation of Histologic and Clinical Images to Determine the Diagnostic Value of Fluorescein Angiography for Studying Retinal Capillary Detail

PURPOSEnTo delineate morphometric and quantitative features of the capillary image derived from high-resolution fundus fluorescein angiography (FFA) and consequently determine the diagnostic value of FFA for studying the retinal capillary circulation.nnnMETHODSnRetinal capillary images obtained from healthy young subjects using high-resolution FFA were compared with confocal scanning laser microscopic capillary images derived from the retinas of age-matched human donors. Confocal microscopic images were acquired from retinal flatmount tissue after central retinal artery cannulation, perfusion fixation, and antibody labeling. Capillary images from equivalent retinal regions were morphologically and quantitatively analyzed in both groups.nnnRESULTSnTen human subjects (mean age, 27.4 years) were used for FFA studies, and five cadaveric eyes (mean donor age, 26.5 years) were used for histologic studies. In histologic specimens the density of the superficial capillary network was significantly greater than that of the deep capillary network. Despite use of a healthy young population, only 30% of high-resolution FFA studies provided clear capillary images. The configuration of the capillary network in FFA images was comparable to the superficial capillary network in confocal microscope images; however, the density of the capillary network in FFA images was consistently lower than that of histologic images.nnnCONCLUSIONSnFFA provides incomplete morphologic information about the superficial capillary network and even less information about the deep capillary network. Caution should, therefore, be exercised when using FFA data to extrapolate information about microvascular histopathologic processes. The usefulness of newer technology for studying retinal capillary detail should be investigated.

Fluorescein angiography and adverse drug reactions revisited: the Lions Eye experience

Background:u2002 The last major survey of adverse reactions to intravenous fluorescein angiography was performed more than 20u2003years ago. There have been two recent fatalities involving intravenous fluorescein in Australia. It is important to review the current incidence of adverse reactions and latest literature on the pathogenesis, prophylaxis and alternatives to intravenous fluorescein angiography.

The distribution of angioarchitectural changes within the vicinity of the arteriovenous crossing in branch retinal vein occlusion.

OBJECTIVEnBranch retinal vein occlusions (BRVOs) are known to occur most commonly in the vicinity of arteriovenous (A/V) crossings. The authors aimed to identify types of venous wall abnormalities in BRVO and document their position in relation to the A/V crossing.nnnDESIGNnA retrospective review of the color photographs and fluorescein angiograms from the most recent 110 patients with first- or second-order BRVO was performed.nnnMAIN OUTCOME MEASURESnThe films were examined for the presence of angioarchitectural changes of specified type within one-quarter disk diameter of the A/V crossing involved in the BRVO. The specific changes noted were fluorescein leakage, presumed thrombi, and flow abnormalities, which were recorded along with their position in relation to the A/V crossing.nnnRESULTSnOf the 110 patients diagnosed with BRVO, 59 had photography of satisfactory quality. Forty-one (70%) of these 59 patients had venous lesions, of which significantly more (chi-square -5.74, P < 0.02) were downstream (56%) than upstream (12%) from the A/V crossing. Thirty-two percent were upstream and downstream. Of the hemodynamic changes seen, 49% had late venous phase leakage of fluorescein, 85% had abnormal flow, and 7% had presumed thrombi. All thrombi seen were downstream.nnnCONCLUSIONSnVenous lesions in the vicinity of the A/V crossing commonly are seen in BRVO, most of which occur downstream. This suggests that the venous narrowing at the crossing may induce downstream hemodynamic changes predisposing to endothelial damage and thrombus generation.

EVALUATION OF ADENO-ASSOCIATED VIRUS-MEDIATED GENE TRANSFER INTO THE RAT RETINA BY CLINICAL FLUORESCENCE PHOTOGRAPHY

The purpose of this study was to evaluate recombinant adeno-associated virus (AAV) as an in vivo gene transfer vector for the retina and to explore the possibility of monitoring the expression of green fluorescent protein (GFP) using a noninvasive method. Rats were injected subretinally with rAAV-gfp or rAAV-lacZ. Strong expression of the reporter gene in a circular area surrounding the injection site was observed in retinal whole mounts and tissue sections. Higher magnification revealed that cells demonstrating high levels of green fluorescence were hexagonal in shape, indicating they were retinal pigment epithelium (RPE) cells. Histological observation of retinal sections demonstrated that recombinant AAV specifically transduced RPE cells. Ten animals were injected with rAAV-gfp for longitudinal studies and the fluorescence was monitored by retinal fluorescence photography. The GFP signal was detected in 100% of the animals as early as 2 weeks postinjection and remained present throughout the experimental period of 4 months. After 2 weeks, a gradual increase in the number of transduced cells occurred before reaching maximal levels of GFP expression at 8 weeks. This was followed by a small decrease over 4 weeks before reaching stable expression at 16 weeks. Our results demonstrated that rAAV efficiently transduces rat RPE cells and that retinal fluorescence photography is suitable for monitoring GFP expression. By using this noninvasive technique, we demonstrated that repetitive measurements of GFP expression in vivo in the rAAV-gfp-transduced retina are possible. This study demonstrated that retinal fluorescence photography is a potent tool for studying AAV-mediated gene delivery in the retina.

Value of retinal vein pulsation characteristics in predicting increased optic disc excavation

Background: Retinal vein pulsation is often absent in glaucoma, but can be induced by applying a graded ophthalmodynamometric force (ODF) to the eye, which is elevated in glaucoma. Aim: To assess whether ODF has a predictive value in determining glaucoma progression. Methods: 75 patients with glaucoma and suspected glaucoma were examined prospectively in 1996, and then re-examined at a mean of 82 months later. All subjects had intraocular pressure, visual fields, stereo optic disc photography and ODF measured on their initial visit. When venous pulsation was spontaneous, the ODF was said to be 0 g. At re-examination, central corneal thickness and blood pressure were also measured. Initial and subsequent optic disc photographs were compared and graded into those that had increased excavation and those that had remained stable. The relationship between increased excavation (recorded as a binary response) and the measured variables was modelled using a multiple mixed effects logistic regression. Results: ODF at the initial visit was strongly predictive of increased excavation (pu200a=u200a0.004, odds ratio 1.16/g, range 0–60 g), with greater predictive value in women than in men (pu200a=u200a0.004). Visual field mean deviation was predictive of increased excavation (pu200a=u200a0.044), as was optic nerve haemorrhage in association with older age (pu200a=u200a0.038). Central corneal thickness was not significantly predictive of increased excavation (pu200a=u200a0.074) after having adjusted for other variables. Conclusion: ODF measurement seems to be strongly predictive of the patient’s risk for increased optic disc excavation. This suggests that ODF measurement may have predictive value in assessing the likelihood of glaucoma progression.

Generation of transgenic mice with mild and severe retinal neovascularisation

Aim: To generate a mouse model for slow progressive retinal neovascularisation through vascular endothelial growth factor (VEGF) upregulation. Methods: Transgenic mice were generated via microinjection of a DNA construct containing the human VEGF165 (hVEGF) gene driven by a truncated mouse rhodopsin promoter. Mouse eyes were characterised clinically and histologically and ocular hVEGF levels assayed by ELISA. Results: One transgenic line expressing low hVEGF levels showed mild clinical changes such as focal fluorescein leakage, microaneurysms, venous tortuosity, capillary non-perfusion and minor neovascularisation, which remained stable up to 3 months postnatal. Histologically, there were some disturbance and thinning of inner and outer nuclear layers, with occasional focal areas of neovascularisation. By contrast, three other lines expressing high hVEGF levels presented with concomitantly severe phenotypes. In addition to the above, clinical features included extensive neovascularisation, haemorrhage, and retinal detachment; histologically, focal to extensive areas of neovascularisation associated with retinal folds, cell loss in the inner and outer nuclear layers, and partial retinal detachment were common. Conclusions: The authors generated four hVEGF overexpressing transgenic mouse lines with phenotypes ranging from mild to severe neovascularisation. These models are a valuable research tool to study excess VEGF related molecular and cellular changes and provide additional opportunities to test anti-angiogenic therapies.

Ophthalmic imaging today: an ophthalmic photographer's viewpoint – a review

Ophthalmic imaging has changed dramatically since the 1960s with increasingly complex technologies now available. Arguably, the greatest changes have been the development of the digital camera and the speed, processing power and storage of electronic data. Already, ophthalmic practices in many major institutions overseas have paperless medium storage and electronically generated reporting from all equipment that use a computer interface. It is hard to remember the widespread use of photographic film with its attendant costs, or even to remember the days before optical coherence tomography (OCT). These latest technical improvements in ophthalmic imaging are now standard in large Australian institutions and becoming more widespread in smaller private practices. The technicians that operate and maintain this ever‐increasing plethora of gadgetry have seen their work practices change from the darkroom to the complexities of data‐based imaging and storage. It is a fitting time to examine the contemporary state of ophthalmic imaging and what lies on the horizon as we move towards 2020.

Telemedicine Screening of Diabetic Retinopathy Using a Hand-Held Fundus Camera

The objective was to evaluate digital images of the retina from a handheld fundus camera (Nidek NM-100) for suitability in telemedicine screening of diabetic retinopathy. A handheld fundus camera (Nidek) and a standard fundus camera (Zeiss) were used to photograph 49 eyes from 25 consecutive patients attending our diabetic clinic. One patient had cataracts, making it impossible to get a quality image of one of the eyes (retina). The Nidek images were digitized, compressed, and stored in a Fujix DF-10M digitizer supplied with the camera. The digital images and the photographs were presented separately in a random order to three ophthalmologists. The quality of the images was ranked as good, acceptable or unacceptable for diabetic retinopathy diagnosis. The images were also evaluated for the presence of microaneurysms, blot hemorrhages, exudates, fibrous tissue, previous photocoagulation, and new vessel formation. kappa Values were computed for agreement between the photographs and digital images. Overall agreement between the photographs and digital images was poor (kappa < 0.30). On average, only 24% of the digital images were graded as being good quality and 56% as having an acceptable quality. However, 93% of the photographs were graded as good-quality images for diagnosis. The results indicate that the digital images from the handheld fundus camera may not be suitable for diagnosis of diabetic retinopathy. The images shown on the liquid crystal display (LCD) screen of the camera were of good quality. However, the images produced by the digitizer (Fujix DF-10M) attached to the camera were not as good as the images shown on the LCD screen. A better digitizing system may produce better quality images from the Nidek camera.

Evaluation of a portable fundus camera for use in the teleophthalmologic diagnosis of glaucoma.

PURPOSEnThe digital images of the optic disk from a portable fundus camera were evaluated for suitability in teleophthalmologic screening for glaucoma.nnnMETHODSnFifty-one eyes of 27 consecutive patients from our glaucoma clinic were dilated and photographed with a Zeiss FF retinal camera (Carl Zeiss, Oberkochen, Germany) and a portable Nidek NM-100 (Nidek, Tokyo, Japan) fundus camera. Digital images from the portable fundus camera were digitized, compressed and stored in a Fujix DF-10M (Fuji, Tokyo, Japan) digitizer. Lossy compressed digital images and photographs from the Zeiss camera were presented separately in random order to three ophthalmologists for estimation of vertical cup:disk ratios (VCDR) and to evaluate image quality as good, acceptable, or unacceptable for screening glaucoma. Gold standard VCDRs were measured from monoscopic photographic slides obtained using the Zeiss camera by a fourth ophthalmologist.nnnRESULTSnMeasurement of agreement (Kappa values) between estimated VCDR of digital images and photographs by the three ophthalmologists were 0.52, 0.38, and 0.50 respectively. Agreement between gold standard and estimated VCDR from photographs were 0.87, 0.45, and 0.84 respectively (specificity between 79% and 97%, sensitivity between 70% and 95%). Kappa values obtained between gold standard and estimated VCDR from digital images were 0.52, 0.49, and 0.49, respectively (specificity between 68% and 79%, sensitivity between 67% and 87%.nnnCONCLUSIONnModerate to good agreement indicates that the digital images from the portable fundus camera may be suitable for optic disk assessment in the current configuration. This easy to use Nidek hand-held camera could be a viable instrument for teleophthalmology if a better digitizing system is incorporated to improve the quality of the images.