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Tübingen, Germany

Arango-Gonzalez B.,Center for Ophthalmology
PloS one | Year: 2012

To explore the effect of ketamine-xylazine anesthesia on light-induced retinal degeneration in rats. Rats were anesthetized with ketamine and xylazine (100 and 5 mg, respectively) for 1 h, followed by a recovery phase of 2 h before exposure to 16,000 lux of environmental illumination for 2 h. Functional assessment by electroretinography (ERG) and morphological assessment by in vivo imaging (optical coherence tomography), histology (hematoxylin/eosin staining, TUNEL assay) and immunohistochemistry (GFAP and rhodopsin staining) were performed at baseline (ERG), 36 h, 7 d and 14 d post-treatment. Non-anesthetized animals treated with light damage served as controls. Ketamine-xylazine pre-treatment preserved retinal function and protected against light-induced retinal degeneration. In vivo retinal imaging demonstrated a significant increase of outer nuclear layer (ONL) thickness in the non-anesthetized group at 36 h (p<0.01) and significant reduction one week (p<0.01) after light damage. In contrast, ketamine-xylazine pre-treated animals showed no significant alteration of total retinal or ONL thickness at either time point (p>0.05), indicating a stabilizing and/or protective effect with regard to phototoxicity. Histology confirmed light-induced photoreceptor cell death and Müller cells gliosis in non-anesthetized rats, especially in the superior hemiretina, while ketamine-xylazine treated rats showed reduced photoreceptor cell death (TUNEL staining: p<0.001 after 7 d), thicker ONL and longer IS/OS. Fourteen days after light damage, a reduction of standard flash induced a-wave amplitudes and a-wave slopes (p = 0.01) and significant alterations in parameters of the scotopic sensitivity function (e.g. Vmax of the Naka Rushton fit p = 0.03) were observed in non-treated vs. ketamine-xylazine treated animals. Our results suggest that pre-treatment with ketamine-xylazine anesthesia protects retinas against light damage, reducing photoreceptor cell death. These data support the notion that anesthesia with ketamine-xylazine provides neuroprotective effects in light-induced cell damage.

Julien S.,Center for Ophthalmology | Schraermeyer U.,Center for Ophthalmology
Neurobiology of Aging | Year: 2012

Lipofuscin is a cytologic hallmark of aging in metabolically active postmitotic cells including neurons, cardiac muscle cells, and the retinal pigment epithelium (RPE). High levels of lipofuscin are involved in the pathogenesis of age-related macular degeneration (AMD), the main cause of blindness in the elderly population in the western world. Degradation and exocytosis of lipofuscin by RPE cells have not been observed in vivo until now, and no drug is known to eliminate the intracellular amount of lipofuscin. Here, we show that in monkeys treated with a small molecule belonging to the tetrahydropyridoethers class (n = 36 of 48 monkeys), RPE cells significantly release lipofuscin. In 4 eyes, macrophages were detected which had taken up lipofuscin. They were located between the Bruch's membrane and the RPE, and in the choroid. The quantification of pigment granules was performed by transmission electron microscopy. Our findings open the way to develop therapeutic strategies to remove lipofuscin from RPE cells, which may have implications for the treatment of age-related macular degeneration in which lipofuscin accumulation in cells is a causative factor. © 2012 Elsevier Inc.

Schraermeyer U.,Center for Ophthalmology | Julien S.,Center for Ophthalmology
Graefe's Archive for Clinical and Experimental Ophthalmology | Year: 2012

Background In this study, the effect of intravitreal injection of bevacizumab on choroidal blood vessels was examined in primate eyes. Methods Four Cynomolgus monkeys received an intravitreal injection of 1.25 mg bevacizumab. The eyes were enucleated on days 1, 4, 7 and 14. For each animal, one eye was embedded in paraffin whereas the other eye was embedded for electron microscopy. Seven untreated or PBS (phosphate buffered saline)-injected monkeys were used as controls. Results Thrombotic microangiopathy was found in the choriocapillaris and choroidal vessels of all eight injected eyes. Acute microangiopathy was characterized ultrastructurally as swelling of the endothelium, loss of fenestrations and complete collapse of the capillaries, and was commonly observed in bevacizumab-treated eyes. Quantitative analysis showed reduction of the lumina of the choriocapillaris in the eyes of three of the monkeys. Bevacizumab was frequently localized inside the blood vessels, often filling the entire breadth of the vessels, and formed clusters with blood cells. Death of photoreceptors occurred in two monkeys. Conclusions This study indicate that intravitreal injection of bevacizumab in monkeys induces activation of platelets, degranulation of thrombocytes and neutrophils, formation of immune complexes, thrombotic microangiopathy and alteration of the blood flow in choroidal vessels. © Springer-Verlag 2012.

Schraermeyer U.,Center for Ophthalmology | Julien S.,Center for Ophthalmology
Expert Opinion on Biological Therapy | Year: 2013

Objective: Due to its low price, bevacizumab, which binds vascular endothelial growth factor, is currently used off-label for the treatment of over 50 different eye diseases and has been adopted worldwide despite the absence of serious preclinical data. This study examines the effects of intravitreal bevacizumab on monkey eyes with particular focus on choroidal and retinal vessels. Methods: Cynomolgus monkeys received an intravitreal injection of 1.25 mg bevacizumab with or without 125I labeling. The eyes were enucleated between 1 and 14 days after injection and were investigated by electron microscopy, immunocytochemistry, histochemistry or autoradiography. Untreated and phosphate buffered saline (PBS)-injected monkeys were used as controls. Results: Bevacizumab locally accumulated at high concentration within individual blood vessels. It formed electron-dense deposits inside retinal veins and between red and white blood cells, activated thrombocytes and induced retinal vein thrombosis. Retinal cells like Müller cells, astrocytes and microglia were also activated. High amounts of bevacizumab were found in retinal and choroidal vessels which may interfere with blood flow. Conclusions: The deposits on the retinal vein walls may provide a mechanistic basis for the observed retinal blood flow alterations after bevacizumab treatment in patients. © 2013 Informa UK, Ltd.

Kustermann S.,Center for Ophthalmology | Hildebrandt H.,Hannover Medical School | Bolz S.,Center for Ophthalmology | Dengler K.,Center for Ophthalmology | And 2 more authors.
Journal of Comparative Neurology | Year: 2010

Polysialic acid (polySia) is a posttranslational modification of the neural cell adhesion molecule NCAM, which in the vertebrate brain is dynamically regulated during development and crucially involved in developmental and adult neurogenesis. In the fish retina, new neurons are persistently generated, but the possible contribution of polySia has not yet been addressed. Here we used immunohistochemistry with NCAM- and polySia-specific antibodies to study spatiotemporal expression patterns of NCAM and polySia in the developing and mature zebrafish retina. As early as 2.3 days postfertilization (dpf), NCAM but not poly-Sia was detected on cell somata and fibers of the developing retina. At 4.3 dpf polySia immunoreactivity first appeared in the ventral retina and was localized to the nascent outer nuclear layer (ONL). In mature zebrafish, polySia immunoreactivity in the ONL extended to the entire retina. Colocalization with rhodopsin-EGFP in transgenic zebrafish or the Müller glia-specific protein cellular retinaldehyde-binding protein (CRALBP) revealed that polySia immunoreactivity was confined to the compartment of radial Müller glia processes crossing the ONL and to a small band of processes positioned proximal to the horizontal cell layer of the mature retina. As shown by 5-bromo-2-deoxyuridine (BrdU) labeling, both newly generated rod precursors within the mature ONL and precursors of the marginal zone were polySia-negative. Thus, polySia-negative rod precursors of the mature zebrafish retina face a polySia-NCAM-positive microenvironment presented by radial Müller glia. In view of the prominent role of polySia in other neurogenic systems, this pattern indicates that polySia provides environmental cues that are relevant for the generation of new rods. © 2009 Wiley-Liss, Inc.

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