Breakdown of the blood-retinal barrier in S-antigen-induced uveoretinitis in rats

Experimental autoimmune uveoretinitis was induced in Lewis rats by footpad injection of purified, bovine retinal soluble antigen (S-antigen) with complete Freund's adjuvant. Control animals received adjuvant only. At the peak of ocular inflammation the animals received an intravenous injection of the tracer horseradish peroxidase, and the eyes were removed and processed for peroxidase cytochemistry. A breakdown of the blood-retinal barrier occurred at the level of the retinal venules. Peroxidase reaction product was demonstrable in the interendothelial tight junctions and in the perivascular spaces. Heavy infiltration of inflammatory cells was also found in and around the venular wall. The endothelial cells of venules and capillaries were hypertrophied and showed swollen endoplasmic reticulum and Golgi apparatus as well as increased numbers of ribosomes and cytoplasmic fibrils; pericytes and smooth-muscle cells showed a similar synthetic phenotype and the basal lamina of these cells was thickened.Offprint requests to: E. Essner     

Histopathology of commotio retinae.

The pathogenesis of commotio retinae in animal models has included extracellular edema, intracellular edema, and photoreceptor outer segment disruption. Histopathologic findings obtained within 24 hours of blunt trauma in a human eye with clinically-observed commotio retinae revealed photoreceptor outer segment disruption and damage to the retinal pigment epithelium. The major site of injury in commotio retinae seems to be at the level of the photoreceptor outer segment-retinal pigment epithelium junction.     

Breakdown of the blood-retinal barrier in a model of retinal neovascularization

Breakdown in the blood-retinal barrier occurs in retinal neovascularization in a number of diseases. To study the anatomic basis of this breakdown, we examined retinal neovascularization induced by injection of 250,000 homologous fibroblasts into the vitreous cavity of pigmented rabbits. Neovascularization is evident by electron microscopy in this model 3 days after fibroblast injection. Fluorescein angiography followed by intravenous horseradish peroxidase (HRP) injection was performed prior to enucleation on 2, 3, 5, 7, and 14 days after fibroblast injection. Fluorescein leakage from retinal vessels occurs early (at day 1) and persists as the neovascularization progresses. The leakage in the early stages is concentrated near puckers from the medullary wings. In the later stages, fluorescein leakage is most prominent in the developing tips of the new vessels. Horseradish peroxidase was not observed to leak from the lumen of new vessels. "Gaps" or separations in the endothelial cell junctions were not observed in developing vessels. The breakdown of the blood-retinal barrier in this model of retinal neovascularization is therefore selective, (ie, fluorescein leaks but not HRP) and it is not due to gaps or fenestrations between endothelial cells in developing vessels.     

Model experiments of the outer blood-retinal barrier in vitro]

Because of the difficulty in conducting experiments on the outer blood-retinal barrier in vivo, we developed an in vitro model. Bovine retinal pigment epithelial cells were grown on semipermeable membranes, enabling separate manipulation of the apical and basal medium. As a parameter of barrier function, we measured the transepithelial resistance (TER). Barrier function was also tested with fluorescein. The transepithelial resistance increased under optimal culture conditions, in confluent cultures, by 200 omega and there was no fluorescein leakage. After exposure to trypsin in Ca/Mg-less medium or EDTA or after application of argon laser, we were able to induce a breakdown of the TER and fluorescein leakage. This happened immediately after laser exposure, 1 min after EDTA, and 4 min after trypsin application. We observed no morphological differences after breakdown of the barrier function on the intercellular connections compared to normal confluent cultures following EDTA or trypsin exposure. In all experiments there was a recovery of barrier function after returning the cells to control conditions. These first results demonstrate that our in vitro model is a sensitive method for investigating barrier function in retinal pigment epithelium in cell culture.     

Reduced photoreceptor outer segment layer thickness in mild commotio retinae without ellipsoid zone disruption

Graefe's Archive for Clinical and Experimental Ophthalmology - To quantitatively investigate the reflectivity and structure of the outer retinal layers on spectral-domain optical coherence...     

Breakdown of the blood-retinal barrier in multiple sclerosis measured by vitreous fluorophotometry

Periphlebitis retinae in multiple sclerosis appears as transitory cellular infiltrations around veins in an otherwise normal retina. Similar cellular infiltrations have been found around veins in the central nervous system. In the present study the blood-retinal barrier has been investigated by vitreous fluorophotometry. Eight multiple sclerosis patients with actual periphlebitis retinae and 9 patients with previous but not active periphlebitis retinae were included in this study. Abnormal leakage of fluorescein was manifest in the group of multiple sclerosis patients with periphlebitis retinae. Permeability (1.8 +/- 0.2 X 10(-7) cm/sec; mean +/- SEM) but not in the control group as a whole permeability (1.3 +/- 0.1 X 10(-7) cm/sec; mean +/- SEM) compared to 17 normals (permeability 1.1 +/- 0.005). It is thus concluded that breakdown of the blood-retinal barrier may be transitory when connected with periphlebitis retinae in multiple sclerosis.     

Intrachoroidal dye leakage in indocyanine green fundus angiography after experimental commotio retinae.

Dye leakage in sodium fluorescein (FLUO) fundus angiography indicates damage to the blood-retinal barrier. However, dye leakage in indocyanine green (ICG) fundus angiography does not mean the same, because of the larger molecular size of the dye and impermeability of the choroidal vessels to it. The possibility of dye leakage in ICG angiography has not yet been revealed in an experimental study in which the blood-retinal barrier is undamaged. We report here that intrachoroidal dye leakage may occur in ICG angiography in an experimental model of the traumatic retinal opacity of the rabbit eye, even when the blood-retinal barrier is undamaged. This mechanism of dye leakage in ICG angiography is quite different from the leakage of FLUO angiography. Pathological choroidal vessels with increased permeability, such as choroidal neovascularization under the retinal pigment epithelium, can be observed using ICG angiography.     

视网膜震荡伤眼的多焦视网膜电图改变

目的 观察视网膜震荡伤眼的黄斑区视 网膜功能，评价多焦视网膜电图（mf-ERG）对视网膜震荡伤的诊断价值。 方法 采用德国Roland公司生产的RETI scan 3.15系统对31例单眼视网膜震荡伤患者的伤 眼（伤眼组）和对侧健眼（对侧健眼组）进行检测。记录61个部位反应，比较分析视网膜后 极部不同区域的N₁(第1个负波)、P₁波(第1个正波)平均反应振幅密度。 结果 伤眼组N₁波1～4环平均反应振幅密度、P₁波1～5环平均反应振幅密度明显低 于对侧健眼组。 结论 视网膜震荡伤眼病变区域N₁、P₁波平均反应 振幅密度下降；mf-ERG能对视网膜震荡伤患眼局部视功能进行定量定位测定。（中华眼底病杂志，2004，20：226-228）     

Choroidal alterations during the clinical course of commotio retinae

Graefe's Archive for Clinical and Experimental Ophthalmology - To analyse the alterations in the choroidal structure of eyes with commotio retinae due to blunt-force trauma. This retrospective...     

Influence of vital dyes on the function of the outer blood-retinal barrier in vitro

BACKGROUND: The use of vital dyes in "chromovitrectomy" allows the easier removal of less recognizable structures like epiretinal membranes (EM) or the inner limiting membrane (ILM). In recent years numerous studies demonstrated the use of indocyanine green (ICG), trypan blue (TB) and patent blue (PB) for this indication. Reports of the possible risk of these dyes, i. e. especially ICG, in respect of reduced visual acuity results, possible visual field defects or alterations of the retinal pigment epithelium (RPE) resulted in limitations in their application. MATERIAL AND METHODS: Human RPE cells and choroidal endothelial cells were cultured in monolayers on semipermeable membranes representing an in vitro model of the outer blood-retinal barrier. By measurement of the transepithelial electrical resistance (TER) the stable barrier function was determined. Two different models representing an air-filled and a fluid-filled eye were tested on the one hand by addition of the dye to the culture medium and, on the other, by direct application on the cell monolayer. In these two models ICG (5 mg/ml, 0.5 mg/ml, 0.125 mg/ml), TB (1.5 mg/ml, 0.15 mg/ml) and PB (2.4 mg/ml, 0.24 mg/ml) were applied for three minutes and the influence on the barrier function was determined. RPE cell growth was also tested in these two models after the application of ICG, TB and PB. Finally, monolayers of RPE cells were evaluated by transmission electron microscopy (TEM). RESULTS: After application of TB, PB and the lowest concentration of ICG of 0.125 mg/ml, the TER remained stable in both models. In contrast, ICG in a concentration of 5 mg/ml and 0.5 mg/ml caused a significant TER decrease in the model of the air-filled eye, whereas no influence on the function of the outer blood-retinal barrier was noted in the model of the fluid-filled eye. RPE cell growth rates were not influenced by the addition of the vital dyes, with the exception of ICG in a concentration of 5 mg/ml in the model of the air-filled eye, resulting in a temporary reduction of the cell count. In good correspondence to these results also in TEM intercellular blisters were noted after application of 5 mg/mL ICG for 3 minutes in the model of the air-filled eye. However, damage to the RPE cells themselves was not obvious. No pathological changes in the TEM were noted after application of TB and PB. CONCLUSIONS: The use of PB and TB at the posterior eye segment seems to be safe concerning damage to the PRE and its barrier function. In contrast, ICG in higher concentrations and with longer application times may cause a toxic effect on RPE morphology and function.     

Enzymatic digestion increases permeability of the outer blood-retinal barrier for high-molecular-weight substances

Background: The purpose of the study was to investigate whether lysosomal enzymes can participate in damaging the outer blood-retinal barrier and to examine the role of glycosaminoglycans in maintaining the barrier function for high-molecular-weight substances. Methods: The ciliary artery was cannulated in freshly enucleated pig eyes. Perfusion was performed with buffer (controls), with heparinase (substrate: heparan sulfate), or with lysosomal enzymes freshly prepared from pig retinal pigment epithelium at 36° C, followed by perfusion with the tracer native ferritin (NF) or the marker cationized ferritin (CF). The eyes were examined by electron microscopy. Results: In controls treated with buffer alone, NF was found in high concentration in the lumina of the choroidal capillaries; however, little NF was found in Bruch's membrane (BsM). The tracer did not penetrate to any extent beyond BsM. In eyes digested with heparinase or lysosomal enzymes, significantly higher numbers of tracer molecules were found in BsM. Furthermore, NF penetrated BsM and was apparent in the subretinal space and also inside retinal pigment epithelial cells, probably due to pinocytosis. Conclusions: The results indicate that heparan sulfate proteoglycan is important for the maintenance of the outer blood-retinal barrier and that lysosomal proteases may participate in damaging this barrier, causing increased permeability to high-molecular-weight substances.Presented in part at the Association for Research in Vision and Ophthalmology Annual Meeting, Sarasota, Florida, 28 April 1991     

Repair of outer blood-retinal barrier after severe ocular blunt trauma in rabbits

Retinal contusion is a leading cause of visual loss in ocular blunt trauma. However, its pathogenesis remains controversial. We established a rabbit model of severe retinal contusion with energy of about 2.87 J. Typical retinal edema and sometimes subretinal hemorrhage reproducibly occurred at the posterior pole after injury. These subsided 1 week later with depigmentation in the lesion. Histopathological examination revealed severe damage of the outer layer of retina, for example, disruption of retinal pigment epithelium (RPE) and photoreceptors. Electroretinography showed a decrease in the b wave by 38–47% in amplitudes (P<0.01) during the first 3 days and then returned, although not to normal level. To investigate the damage and repair of blood-retinal barrier (BRB), 5 ml 2% lanthanum solution (La) was injected via the common carotid artery 1–2 min before enucleation. La diffused in the interphotoreceptor space through the damaged junctions of RPE 1 h-3 days after injury. La also reached the nuclei level of photoreceptors up to 14 days after injury. Although a glial scar with scattered RPE cells attached to Bruch's membrane in the severely damage area, no La diffusion was found in the retina 4 weeks after trauma. These results showed incomplete repair of outer BRB after severe blunt trauma.     

Glucose-induced activation of glucose uptake in cells from the inner and outer blood-retinal barrier

PURPOSE: The purpose of this study was to elucidate in vitro the effect of elevated glucose on glucose uptake in the cells comprising the inner and outer blood-retinal barriers: human retinal pigment epithelial (hRPE) and human retinal vascular endothelial (hRVE) cells. METHODS: Primary cultures of hRPE and hRVE cells grown in 5.5 or 22 mM glucose or in 22 mM mannitol were used to measure the rates of glucose uptake with [(3)H]-3-O-methyl glucose as a tracer. GLUT1 expression was measured by Northern and western blot analyses. Cellular fractionation was performed by differential centrifugation. GLUT1 overexpression was accomplished by adenoviral transduction. RESULTS: Increasing media glucose from 5.5 to 22 mM for 30 minutes caused a 1.9-fold increase in the V(max) of glucose uptake in hRPE cells and a 2.5-fold increase in hRVE cells. These increases were nonosmotic and glucose specific, in that the exposure to 22 mM mannitol did not affect the V(max) of glucose uptake. mRNA, total protein expression, and translocation of GLUT1, the glucose transporter predominantly expressed in hRPE and hRVE cells, were not affected by 22 mM glucose for up to 48 hours. High-glucose effects on V(max) were abolished with 10 microg/mL of the microtubule assembly inhibitor nocodazole. hRPE cells transduced to overexpress GLUT1 showed a 1.5-fold increase in the V(max) for glucose uptake versus control-transduced cells. However, the magnitude of glucose-induced increase in glucose uptake was the same in GLUT1- and control-transduced cells. CONCLUSIONS: High glucose induced 1.9- and 2.5-fold increases in the V(max) of glucose uptake in hRPE and hRVE cells, respectively. These increases were not due to an increase in GLUT1 expression. The increases were dependent on microtubule integrity, but not on GLUT1 translocation. The mechanism of the increases is unknown. GLUT1 regulating protein(s) and/or novel glucose transporter(s) may be involved in the regulation of glucose uptake by glucose in the cells that comprise the blood-retinal barrier.     

视网膜震荡伤患者黄斑光敏感度的变化

目的:探讨视网膜震荡伤后黄斑光敏感度在病程中的变化。方法:用Humphrey-750型自动视野分析仪检测视网膜震荡伤后黄斑光敏感度的变化,光敏感度以dB(分贝)表示,dB值越高,表示光敏感度越高,光阈值越低;dB值越低,表示光敏感度越低,光阈值越高。结果以SPSS软件进行分析。结果:视网膜震荡伤组治疗前、后黄斑光敏感度均降低,治疗后平均黄斑光敏感度有所提高,但仍比正常人的平均黄斑光敏感度低。结论:黄斑光敏感度检测可做为视网膜震荡伤后评价视功能恢复情况的一个可靠并且敏感的指标。