Central Serous Chorioretinopathy: Morphological and Functional Outcome after Subthreshold Thermal Laser Coagulation with a Frequency-Doubled Nd:YAG Continuous-Wave Laser.

PURPOSE: Central serous chorioretinopathy (CSCR) presents itself as a serous detachment of the central neurosensory retina (NR), which may be accompanied by focal detachment of the retinal pigment epithelium (RPE) and changes in the RPE itself. It is often self-limiting; however, if the macular region is affected, visual impairment can be serious. If spontaneous remission does not occur, data on the effectiveness of further treatment options are sparse. We therefore decided to examine the effectiveness of subthreshold laser photocoagulation (ST-LP) on best-corrected visual acuity (BCVA) and subretinal fluid (SRF) resorption. We conducted a retrospective analysis of all patients who underwent ST-LP based on the diagnosis of CSCR in a German university eye hospital from 2009 to 2014. METHODS: The diagnosis of CSCR was based on the following criteria: detachment of the NR and possibly the RPE visible on ophthalmoscopy, evidence of SRF on optical coherence tomography (OCT), visualization of one or more source points typical for CSCR in fluorescein angiography, and exclusion of differential diagnoses. The time between the anamnestic onset of symptomatic complaints and ST-LP was determined as well as BCVA and OCT before ST-LP. ST-LP was performed as a subthreshold thermal laser coagulation with a frequency-doubled Nd:YAG continuous-wave laser. Follow-up examinations were scheduled at 4, 8, and 12 weeks after ST-LP. RESULTS: Fifty-four eyes of 49 patients were included in the study. The median age of patients was 47 years. Eighty-nine percent of the included patients were male. Twenty percent of patients had a first manifestation of CSCR, 69% had a recurrence, and 11% had persistent SRF for >6 months. The median visual acuity rose from 0.30 at baseline (BL) to 0.10 at 4 weeks and 0.00 at 8 weeks, before dropping slightly to 0.05 at 12 weeks. Changes of visual acuity in comparison to BL were statistically significant (p < 0.05). The initial median retinal thickness of 397 µm at BL decreased to 264 µm at 4 weeks, to 236 µm at 8 weeks, and to 239 µm at 12 weeks (decreases to BL all statistically significant p < 0.05). CONCLUSION: In our cohort, we were able to achieve substantial and significant clinical benefit through ST-LP measured by improvement in BCVA. Furthermore, we were also able to demonstrate measurable, significant morphological improvements as decreased retinal thickness and increased resorption of SRF as probable mechanisms explaining clinical improvement of CSCR with ST-LP. The advantage of ST-LP over other methods is the low risk of adverse events and its high availability. Controlled, randomized studies are necessary to confirm the data and demonstrate the effect over a longer period of time.

Optical Coherence Tomography Angiography Findings in Proliferative Diabetic Retinopathy.

PURPOSE: To characterize preretinal neovascularizations (NV) and their corresponding branching routes in proliferative diabetic retinopathy (PDR) with optical coherence tomography angiography (OCTA) and compare the findings with fluorescein angiography (FA). METHODS: In patients with PDR, angiograms were acquired with spectral-domain OCTA (CIRRUS 5000, OCTA AngioPlexTMCarl Zeiss Meditec, Inc.) and FA (Zeiss FF450PlusIR fundus camera or Spectralis HRA-OCT SLO, Heidelberg Engineering Inc.) and were consecutively evaluated. Neovascularization of the disc (NVD) and neovascularization elsewhere (NVE) were analyzed with 6 × 6 and 8 × 8 mm OCTA flow images and B-scans with flow registration. Segmentations of the vitreoretinal interface (VRI) and superficial retina were performed for analysis. Two independent investigators examined OCTA findings and compared them to corresponding FA. RESULTS: Forty-two eyes of 30 patients with PDR were analyzed. A total of 76 NV with their corresponding proliferation routes were visualized and characterized, with 55 (72.4%) proliferating along the posterior hyaloid membrane (PHM), 14 (18.4%) along the epiretinal membrane, and 7 (9.2%) along the fibrovascular membrane. The posterior vitreous was partially detached in 37 of 42 eyes (88.1%), completely detached in 1 of 42 eyes (2.4%), and adherent in 1 of 42 eyes (2.4%). In 38 of 42 cases, OCTA was superior (n = 23) or equivalent (n = 15) to FA in detecting NV and provided a more detailed information of the neovascular vessels. In 4 of 42 study eyes, OCTA was inferior to FA. CONCLUSIONS: OCTA is a useful tool to detect NV in PDR. In comparison to FA, OCTA has the advantages that it is noninvasive and the image capture takes only seconds. We were able to identify all NV and characterize their corresponding proliferation routes in the VRI, the superficial retina slab, or the B-scan with flow registration. Through evading the masking effect of dye leakage in FA, OCTA is capable of better visualization of NV. FA, however, remains essential for the detection of all NV, since OCTA supplies a smaller detection field. Additionally, we identified the PHM as the main proliferating route of diabetic NV (72.4%), marking it as an important structure for sprouting vessels in neoangiogenesis in PDR.

OCT Angiographic Findings in Retinal Angiomatous Proliferation. / OCT-angiografische Befunde bei retinalen angiomatösen Proliferationen.

BACKGROUND: OCT angiography (OCT-A) allows non-invasive blood flow registration of the retina and choroid. In contrast to fluorescein angiography (FA), no dye has to be administered. The OCT-A also provides depth-selective information. OCT-A and FA were compared in patients with neovascular age-related macular degeneration (AMD) with retinal angiomatous proliferation (RAP) stage 1. In stage 1, the neovascularizations are intraretinal. In contrast to the two-dimensional total image of the FA, the OCT-A allows a depth-selective display of the individual retinal layers. In this way, a conclusion can be drawn about the place of origin of the RAP. PATIENTS AND METHODS: Three patients with neovascular AMD and RAP stage 1 were included. They were examined with OCT (ZEISS CIRRUS HD-OCT 5000, Carl Zeiss Meditec, Inc., Dublin, USA), OCT-A (ZEISS AngioPlex OCT-Angiography) as well as FA (HRA2, Heidelberg Engineering) between January 2016 and March 2019. A complete ophthalmological examination was performed. A qualitative analysis of the OCT-A images (3 × 3 and 6 × 6 mm) and the FA images was carried out. Leaks in the FA were compared with the en-face images of the OCT-A followed by a depth-selective assignment using the corresponding B-scans of the OCT-A. RESULTS: It was one woman and two men aged 66 - 89 years. The visual acuity was 0.4 in the first, 0.5 p in the second and 0.8 in the third patient. The diagnosis of RAP stage 1 could be made both in the OCT, the FA and the OCT-A. All patients showed macular edema in the OCT. The FA showed selective hyperfluorescence in the early phase and fluorescein extravasation in the late phase. In OCT-A, the blood flow in all patients could be shown in the hyperreflective structure of the RAP in the B-scan. The first patient showed two RAP lesions in the FA, which were in the deep vascular plexus in the OCT-A. In the second patient, three RAP lesions were found in the FA, and a total of five RAP lesions in the OCT-A. One could be located in the superficial and deep vascular plexus, four in the deep vascular plexus. The third patient showed one RAP lesion in the FA as well as in the OCT-A, which could be assigned to the superficial vascular plexus. CONCLUSION: The OCT-A is well suited for the diagnosis of RAP stage 1. In the present cases, the diagnosis in the OCT-A could be made as clearly as by FA. A major advantage of the OCT-A results from the non-invasive character and the depth selectivity. The RAP 1 lesions could be assigned to both the superficial and the deep vascular plexus. Depth selection is not possible with the FA due to the summary picture.

[Patient's Knowledge after Informed Consent for Intravitreal Injections]. / Intravitreale Injektionen ­ welche Informationen aus dem Aufklärungsgespräch bleiben Patienten im Gedächtnis?

BACKGROUND: Intravitreal treatment (IVT) is one of the most common ophthalmological procedures. Therapeutic effectiveness is however dependent on patient compliance. Unwanted treatment cessation rates are high though. The authors therefore decided to analyse the patient's knowledge and treatment expectations, as discrepancies are known to negatively affect compliance and thus treatment outcomes. PATIENTS AND METHODS: The study was designed as an exploratory survey. In total, 100 patients presenting to an outpatient clinic of a tertiary care centre from October to December 2016 were included. A structured, anonymised questionnaire was handed out, consisting mainly of question items with closed code lists as response domains. Solely descriptive analysis of results was performed. RESULTS: The median age of patients was 73 years. 70% had received more than 3 IVTs in at least one eye. Age-related macula degeneration was the most common underlying cause (52%). 64% expected improvement of visual acuity after IVT. 42% could not name one medication used in their IVT. 55% felt that the information provided during informed consent had been adequate. 69% did not know the post-surgical occurrence of endophthalmitis. Three patients were confident of being able to drive a car directly after IVT. CONCLUSION: Patient's knowledge of their underlying disease, treatment goals and complications rates exhibited some deficiencies. Standardised patient information sheets could be of significant use and were actively suggested by patients to improve the informed consent process.

Transport and fate of aflibercept in VEGF-A165-challenged retinal endothelial cells.

Retinal vessels are at least in part involved in clearing of Fc terminus-containing proteins from the vitreous. In vitro, the Fc fusion protein aflibercept is transported through a monolayer of unchallenged immortalized bovine retinal endothelial cells (iBREC), mediated by the neonatal Fc receptor (FcRn), but part of the Fc fusion protein is also degraded. Aflibercept's target VEGF-A not only enhances the permeability of REC by destabilization of tight junctions (TJs) thereby allowing for paracellular flow, it may also lower the intracellular stability of the Fc fusion protein by changing its binding properties to the FcRn. Therefore, we investigated the transport and fate of aflibercept in VEGF-A165-challenged iBREC. All cell culture media were supplemented with 5% fetal bovine serum (FBS) as its absence results in accumulation of aflibercept in iBREC due to deregulated expression of transport proteins. Early after exposure of a confluent iBREC monolayer cultivated on gold electrodes to 5% FBS, the cell index (CI) - assessed as a measure of barrier function, cell viability and cell adhesion - transiently declined but recovered again within a few hours to high values. These values remained stable for several days associated with a strong expression of the TJ-protein claudin-1, indicative of a functional barrier formed by the iBREC monolayer. Transient changes of the plasma membrane localizations of claudin-5 and vascular endothelial cadherin - both important for regulation of paracellular flow - accompanied the transient reduction of the CI not prevented by VEGF-binding proteins. Treatment of iBREC with 50 ng/ml VEGF-A165 for one day resulted in a strong and persistent decline of the CI associated with a low expression level of the TJ-protein claudin-1; reversion to normal values was complete one day after aflibercept's addition at a final concentration of 250 µg/ml. Expressions of other proteins involved in regulation of paracellular flow or transcellular transport were not significantly changed. More aflibercept passed through the monolayer of iBREC cultivated on permeable membrane inserts pretreated with VEGF-A for one day, but this was not affected by a FcRn-inhibiting antibody. Subcellular localization of aflibercept was hardly changed in VEGF-A-exposed iBREC 3 h after its addition to the cells; inhibition of (non)-lysosomal or proteasomal proteases then only weakly affected the amount of internalized aflibercept. iBREC also internalized VEGF-A which was barely detectable as early as 2 h after addition of aflibercept. In contrast, blocking the tyrosine kinase activity of VEGF receptor(s) did not prevent VEGF-A's uptake. Inhibition of cellular proteases strongly increased the amount of internalized VEGF-A in the absence and presence of the Fc fusion protein. We therefore conclude that a FcRn-mediated transport plays a minor role in aflibercept's passage through a leaky barrier of REC. Even early after addition of aflibercept to VEGF-A-exposed iBREC, the levels of free intracellular VEGF-A are low, as aflibercept likely prevents binding of VEGF-A to its receptor. Interestingly, the growth factor's detrimental effects still persist for nearly one day.

VEGF receptor 2 inhibitor nintedanib completely reverts VEGF-A165-induced disturbances of barriers formed by retinal endothelial cells or long-term cultivated ARPE-19 cells.

Various severe ocular diseases are associated with an elevated intravitreal expression of VEGF-A which increases the permeability of retinal endothelial cells (REC) or retinal pigment epithelial (RPE) cells in vivo and in vitro. Inhibition of VEGF receptor 2 (VEGFR2) is sufficient to completely prevent VEGF-A165-induced dysfunctions of barriers formed by long-term cultivated, immortal human ARPE-19 cells or immortalized bovine retinal endothelial cells (iBREC). Extended exposure to VEGF-A could result in additional activation of other growth factor receptors, potentially promoting synergistic effects of corresponding factors on various cellular processes including angiogenesis. Based on these observations, we investigated whether blocking of VEGFR2 is also sufficient to revert VEGF-A-induced changes of the barriers consisting of iBREC (i.e. inner blood-retina barrier) or ARPE-19 cells (i.e. outer blood-retina barrier) in vitro. Alterations of confluent monolayers' properties induced by treatment with VEGF-A165 for one day followed by addition of small molecule inhibitors of the VEGFR2 were determined by continuous cell index (CI) measurements using the microelectronic biosensor system for cell-based assays xCELLigence. VEGF-A165 induced a long-lasting drop of the otherwise high CI of iBREC accompanied by reduced expression of the tight junction (TJ) protein claudin-1 and subtle changes of the plasma membrane localizations of TJ-protein claudin-5 and of vascular endothelial cadherin. Blocking mainly VEGFR2 with 10 nM nintedanib, 10 nM tivozanib or 500 nM ZM323881 efficiently reverted these changes within one day; higher concentrations of nintedanib or additional inhibition of neuropilin-1 were not superior. Interestingly, the CI of short-term cultivated, confluent ARPE-19 cells slightly increased in the presence of VEGF-A165, but was not changed by nintedanib. In contrast, VEGF-A165 markedly reduced the transepithelial electrical resistance of ARPE-19 cells cultivated on porous membrane inserts for three weeks, which was also accompanied by a significant loss of the then strongly plasma membrane-expressed TJ-protein ZO-1. These alterations were completely reverted within one day by 10 nM nintedanib of which higher concentrations were not superior. None of the inhibitors tested diminished the strong barrier properties of iBREC or long-term cultivated ARPE-19 cells. Taken together, inhibition of VEGFR2 efficiently reverts VEGF-A165-induced barrier disturbances of both cell types forming and regulating the inner and outer blood-retina barrier. As synergistic actions of growth factors seem to play only a minor role in inducing a barrier dysfunction, specific inhibition of VEGFR2 could be an interesting option to treat VEGF-A-induced macular edema without obvious effects on vitality and functions of REC and RPE cells.

Comparison between Findings in Optical Coherence Tomography Angiography and in Fluorescein Angiography in Patients with Diabetic Retinopathy.

OBJECTIVE: To compare typical findings of diabetic retinopathy in optical coherence tomography angiography (OCTA) and fluorescein angiography (FA). SUBJECTS/METHODS: 42 patients were enrolled in this study. We performed FA and obtained en face 3 × 3 mm OCTA images of the macular region. The count of microaneurysms (MAs) and the size of the foveal avascular zone (FAZ) were compared. The assessability of the imaging modalities was graded in each eye. RESULTS: 53 eyes of 42 patients with a mean age of 61 years were included. 36/53 eyes revealed nonproliferative diabetic retinopathy, 17/53 eyes had proliferative diabetic retinopathy. The mean size of the FAZ was 0.39 mm2 in FA and 0.42 mm2 in OCTA. The mean MA count was 14 in FA and 13 in OCTA. The assessability was favorable to OCTA in 38-41/53 eyes regarding the FAZ and favorable to FA in 45-49/53 eyes regarding MAs. CONCLUSION: We found a good agreement for the size of the FAZ and a weak agreement regarding the count of MAs in both imaging modalities. The readers favored OCTA for the assessment of the FAZ and FA for the assessment of MAs. Complementary use of FA and OCTA ensures the best diagnostic approach in patients with diabetic retinopathy.

Mutation-Dependent Pathomechanisms Determine the Phenotype in the Bestrophinopathies.

Best vitelliform macular dystrophy (BD), autosomal dominant vitreoretinochoroidopathy (ADVIRC), and the autosomal recessive bestrophinopathy (ARB), together known as the bestrophinopathies, are caused by mutations in the bestrophin-1 (BEST1) gene affecting anion transport through the plasma membrane of the retinal pigment epithelium (RPE). To date, while no treatment exists a better understanding of BEST1-related pathogenesis may help to define therapeutic targets. Here, we systematically characterize functional consequences of mutant BEST1 in thirteen RPE patient cell lines differentiated from human induced pluripotent stem cells (hiPSCs). Both BD and ARB hiPSC-RPEs display a strong reduction of BEST1-mediated anion transport function compared to control, while ADVIRC mutations trigger an increased anion permeability suggesting a stabilized open state condition of channel gating. Furthermore, BD and ARB hiPSC-RPEs differ by the degree of mutant protein turnover and by the site of subcellular protein quality control with adverse effects on lysosomal pH only in the BD-related cell lines. The latter finding is consistent with an altered processing of catalytic enzymes in the lysosomes. The present study provides a deeper insight into distinct molecular mechanisms of the three bestrophinopathies facilitating functional categorization of the more than 300 known BEST1 mutations that result into the distinct retinal phenotypes.

[The Importance of the Preoperative Information from a Patient's Perspective - a Structured Interview of 100 Ophthalmological University-Based Outpatients]. / Die Erwartungen an das Aufklärungsgespräch aus Patientensicht ­ strukturierte Befragung von 100 Patienten einer ophthalmologischen Universitätsambulanz.

BACKGROUND: The patient's knowledge about their illness, as well as their expectations regarding pre-intervention, consultation and treatment, may differ from the physician's assumptions. Therefore, it is of great importance that the physician can identify misconceptions and missing knowledge and to focus on those points in the preoperative consultation, as well as meeting patient expectations as to the consultation itself. The aim of this study was to identify such expectations and the knowledge gaps of patients scheduled for ophthalmologic treatment. METHOD: An anonymous questionnaire containing predominantly closed questions was handed out to 100 patients in an ophthalmological outpatient clinic of a tertiary care center. Answers were mostly single choice items on a rating scale. RESULTS: 55% of patients had received ophthalmological interventions prior to receiving the questionnaire; 36% received more than two. More than half had not informed themselves about the planned procedure prior to their appointment. They were worried the most about complications (59%) and least about the anaesthesia (29%). When asked, patients attributed the highest priority to provision of information regarding complications and most often requested information on implications of the planned surgery on daily activities. CONCLUSION: Roughly half of the patients came without having informed themselves prior to the consultation. A comprehensive explanation with regard to success rates and possible post-surgical impairments appears to be essential. Possibilities of new media, such as the internet, surprisingly do not seem to be of importance to patients in this context.

[Invasive Eyelid Infections by ß-Hemolytic Streptococci of Serogroup A]. / Invasive Lidinfektionen durch ß-hämolysierende Streptokokken der Serogruppe A.

BACKGROUND: Invasive soft tissue infections by Streptococcus pyogenes are rapidly progressive and potentially life-threatening infectious diseases. These can also affect the eyelid. Aggressive virulence factors and the synthesis of exotoxins can lead to complications, such as periorbital necrotizing fasciitis (PONF) and streptococcal toxic shock syndrome (STSS). The clinical picture is characterized by four patients with invasive eyelid infections. MATERIALS AND METHODS: Photographic documentation, radiological imaging, laboratory and smear diagnostics and intravenous antibiotic therapy were performed on all patients according to the recommendations of the German Robert Koch Institute and the local infectiology board. RESULTS: In all patients, Streptococcus pyogenes was culturally detected in a direct swab. The antibiogram showed sensitivity to the common intravenous antibiotics. The time interval between symptom onset and presentation at the clinic was between two days and one week. All patients had high systemic inflammatory parameters on admission: Pat. 1: CRP 259 mg/l, leukocytes 20.1 giga/l; Pat. 2: CRP 375 mg/l, leukocytes 15.6 giga/l; Pat. 3: CRP 378 mg/l, leukocytes 38.7 giga/l; Pat. 4: CRP 483 mg/l, leukocytes 1.7 giga/l; normal values: CRP < 5 mg/l, leucocytes 4.4 - 11.3 giga/l. In Pat. 2 and 3, a periorbital necrotizing fasciitis was diagnosed due to rapidly progressing necrosis in the area of cutis and subcutis and systemic toxicity. Pat. 3 and 4 met the diagnostic criteria of STSS. Pat. 2, 3 and 4 had to be relocated to an intermediate or intensive care unit with sepsis, despite immediate intravenous antibiotic therapy. Patient 3 underwent surgical debridement during the stay in the intensive care unit. Thanks to interdisciplinary management (ophthalmology, infectiology, ear, nose and throat medicine, internal medicine and intensive care medicine), all patients were finally discharged from our inpatient treatment in a significantly improved general condition. CONCLUSION: Invasive streptococcal infections represent a challenge in the daily routine of an ophthalmologist. Interdisciplinary management and immediate onset of high-dose intravenous antibiotic therapy are crucial for successful treatment.

[Successful Systemic Lysis Therapy in Acute Retinal Arterial Occlusions]. / Erfolgreiche systemische Lysetherapie bei akuten retinalen Arterienverschlüssen.

BACKGROUND: Retinal artery occlusion leads to dramatic and irreversible vision loss. There is currently no evidence-based standard therapy. According to the German guidelines on retinal artery occlusions, intravenous fibrinolysis therapy can be performed up to a time window of 4 h 30 min. METHODS: Two patients were treated accordingly. RESULTS: In patient 1, systemic lysis therapy was used in branch retinal artery occlusion (BRAO) of the inferior temporal retinal artery with macular involvement 4 h 15 min after symptom onset. Immediately after the therapy, the patient reported significant improvement in symptoms. Three months after therapy, retinal function was good, but with subtle atrophy of the inner neurosensory retina. Patient 2, 2 h 30 min after onset of symptoms of the inferior temporal BRAO, the patient experienced further deterioration, with clinical signs of a central retinal artery occlusion (CRAO). Visual acuity deteriorated to light perception. Emergency intravenous lysis therapy, administered 3 h later, gave an improvement in visual acuity with preservation of the inferior visual field. In both patients, a marked improvement in visual acuity was observed immediately after the lysis therapy: Patient 1: right eye, best corrected visual acuity (BCVA) initial 0.5, BCVA 3 days after lysis therapy 1.0, no defects in Goldmann visual field. Patient 2: left eye, BCVA initial 0.4, then sudden deterioration to light perception, BCVA 1 month after lysis therapy 0.6, persisting visual field defects in the superior hemisphere with preservation of the inferior visual field. CONCLUSIONS: Two patients with acute retinal artery occlusion were treated successfully with systemic intravenous fibrinolysis.

Fate of the Fc fusion protein aflibercept in retinal endothelial cells: competition of recycling and degradation.

PURPOSE: Intravitreal injection of the VEGF-binding protein aflibercept is widely used to treat various ocular diseases. In vitro, immortalized bovine retinal endothelial cells (iBREC) take up and transport aflibercept through the cell layer in a serum-dependent manner, likely mediated through the neonatal Fc receptor (FcRn), but degradation of the Fc domain-containing protein might be a competing intracellular process. Therefore, aflibercept's associations with proteins either involved in FcRn-mediated transport or in the lysosomal pathway were studied. METHODS: Confluent iBREC pre-cultivated with or without FBS were exposed for 4 h to in vivo achievable 250 µg/ml aflibercept, before cells were harvested for immunofluorescence staining or preparation of protein extracts. Intracellular localization of aflibercept and putative co-localizations with proteins involved in transport of IgG/FcRn complexes, i.e., endosomal Rab4 and Rab11, components of the cytoskeleton, motor proteins, or with marker proteins characteristic of multivesicular bodies or lysosomes were assessed by co-immunofluorescence stainings. Amounts of expressed endogenous proteins and of internalized aflibercept were determined by Western blot analyses. RESULTS: Aflibercept-specific perinuclear staining overlapped with that of the motor protein dynein whereas double staining with an anti-kinesin antibody resulted in a patchy pattern. In addition, aflibercept was typically present close to microtubules and often co-localized with α-tubulin. Rab4 and Rab11 stainings partly overlapped with the perinuclear staining of aflibercept whereas co-localization with Rab7 (in late endosomes/lysosomes) was only rarely seen. Interestingly, aflibercept but not the IgG bevacizumab broadly co-localized with the cation-independent mannose 6-phosphate receptor characteristic of multivesicular endosomes. In accordance with partial degradation beside transcytosis, the amount of intracellular aflibercept increased when cells were treated with protease inhibitors MG-132 or MG-101. Serum-deprived iBREC expressed less Rab11 and dynein but slightly more Rab4. CONCLUSION: After uptake by iBREC, aflibercept is present in organelles associated with FcRn-mediated transport, but part of the protein is subject to degradation. Transport inhibition of aflibercept during cultivation without FBS is likely a consequence of an attenuated exocytosis due to decreased expression of Rab11.

[Visualization of Retinal Neovascularization with Optical Coherence Tomography Angiography in Comparison with Fluorescein Angiography]. / Darstellung von retinalen Neovaskularisationen mit der optischen Kohärenztomografie-Angiografie im Vergleich mit der Fluoreszeinangiografie.

OBJECTIVE: Comparison of retinal neovascularizations of the disc (NVD) and elsewhere (NVE) in optical coherence tomography angiography (OCTA) and fluorescein angiography (FA) in patients with proliferative diabetic retinopathy. MATERIALS AND METHODS: 15 consecutive patients were included in this study. All patients received an OCTA with a 3 × 3 mm scan of the region of interest with the ZEISS OCT Cirrus 5000 with the AngioPlex module. The size of the neovascularization (NV) was determined manually in OCTA and FA and compared between the two methods. RESULTS: 20 eyes of 15 patients with proliferative diabetic retinopathy with an average age of 57 years were included. The mean size of NVDs was 3.44 mm2 in OCTA and 3.75 mm2 in FA, the mean size of NVDs was 1.06 mm2 in OCTA and 1.54 mm2 in FA. Taking into account a generally larger area measured in the FA, the two methods showed good overall agreement. CONCLUSION: There was a good agreement for the size of the NVs in both methods. OCTA can be used as a simple and non-invasive method to visualize retinal neovascularizations.

[Comparison Between Automated and Manual Measurement of Foveal Avascular Zone Retinopathy in Optical Coherence Tomography]. / Evaluation der automatisierten Messung von ischämischen Veränderungen der Makula bei diabetischer Makulopathie.

OBJECTIVE: A comparison between automated and manual measurements of a foveal avascular zone in optical coherence tomography angiography (OCTA) in patients with diabetic retinopathy (DR). MATERIAL AND METHODS: Consecutive patients with non-proliferative DR were included in this study. All patients received an OCTA, with a 3 × 3 mm scan of the macular region taken with the Zeiss OCT CIRRUS 5000 with the AngioPlex module. The size of the foveal avascular zone (FAZ) was determined both manually and with the help of the automated measurement metrics. Next, the measurements obtained using manual and automated methods were compared. In addition, the circularity index determined in metrics was examined for correlations with the size and area of the FAZ. RESULTS: Thirty-four eyes from 28 patients with non-proliferative diabetic retinopathy with a mean age of 63 years were included. The mean size of the foveal avascular zone was 0.34 ± 0.12 mm2 (0.08 - 0.65 mm2) for manual evaluation and 0.23 ± 0.11 mm2 (range 0.03 - 0.49 mm2) in metrics. The circularity index in metrics averaged 0.58 and showed a statistically significant correlation with the size of the manually measured FAZ. CONCLUSION: There was a comparable result for the size of the FAZ in both measurement methods. Automated measurements with metrics can reliably represent changes in the FAZ for most patients, based on the calculated area, as well as on the circumference and the circularity index.

[Retinal Artery Occlusions]. / Arterielle Gefäßverschlüsse der Retina.

Retinal artery occlusions are acute vascular diseases. Very often they are caused by an embolisation. Rarely a thrombosis is caused by an arteriitis. In the acute phase diagnosis in most of the cases can be made by ophthalmoscopy due to the whitish retinal edema and in central retinal artery occlusion the cherry red spot. Difficulties in making the diagnosis can occur in the chronic stage, especially after reperfusion of the retinal vessels, when the fundus colour returned to normal again. Several diagnostic procedures help to make the diagnosis. Unfortunately there is no generally accepted therapeutic approach for retinal artery occlusions. The visual prognosis often is poor. It is very important to rule out underlying diseases because the patients have an increased risk for stroke or cardiovascular events.

[Retinal Vein Occlusions]. / Venöse Gefäßverschlüsse der Retina.

Retinal vein occlusions (RVO) are common retinal diseases. They are caused by an obstruction of retinal veins. RVO often result in visual deterioration and, in rare cases, can lead to blindness, but also be asymptomatic. The prognosis is various and depends on the extent, the localization of the occlusion and the retinal perfusion, especially in the macular area. The etiology often is a thrombosis caused by vascular sclerosis, rarely a vasculitis. Because often an underlying disease is present the medical clarification is necessary. The ophthalmological treatment of the macular edema includes intravitreal injection of VEGF inhibitors and steroids. Laser photocoagulation is being proposed as second line treatment. Neovascular complications like preretinal neovascularizations with and without vitreous hemorrhage are treated with laser or vitrectomy. Iris neovascularization can also occur and is treated with laser photocoagulation or cyclophotocoagulation.

Inhibition of Single Routes of Intracellular Signaling is Not Sufficient to Neutralize the Biphasic Disturbance of a Retinal Endothelial Cell Barrier Induced by VEGF-A165.

BACKGROUND/AIMS: Hallmark of diabetic macular edema is the enhanced permeability of retinal endothelial cells (REC) induced by vascular endothelial growth factor (VEGF-A165), which acts through activating specific receptors. To improve the predictability of inhibitors' potentials to block harmful effects of VEGF-A165, we investigated if its signaling pathways triggered in REC are redundant. METHODS: Immortalized bovine REC monolayers were treated with inhibitors specific for various protein kinases in combination with VEGF-A165. Permeability was monitored continuously by measurements of the cell index (CI) to reveal even subtle and transient changes. Expression of tight junction (TJ) proteins was determined as additional indicator of barrier stability. RESULTS: After a sharp but transient CI drop caused by VEGF-A165 early after its addition, further exposure resulted in a continuous CI decline over several days associated with loss of TJ protein claudin-1. Both phases were blocked by inhibition of VEGF receptor 2. Tested inhibitors of intracellular kinases had a limited or no effect, or were efficient only in certain phases of exposure to VEGF-A165, e.g. inhibiting protein kinase C only prevented the early response. High concentrations of some inhibitors even resulted in VEGF-independent barrier destabilization. CONCLUSIONS: Specific kinase inhibitors differently affect VEGF-A165-triggered processes in distinct phases of its action. VEGF-A165-initiated signaling is redundant and blocking of key proteins of single pathways is not sufficient to suppress REC barrier breakdown.

Neonatal Fc receptor FcRn is involved in intracellular transport of the Fc fusion protein aflibercept and its transition through retinal endothelial cells.

Retinal endothelial cells (REC) likely contribute to the clearance of intravitreally injected IgG. Because this is of high relevance to the pharmacokinetic assessment of the widely used therapeutic Fc fusion protein aflibercept, we studied its transport through immortalized bovine REC (iBREC) in detail. For shuttling of IgG or Fc fusion proteins like aflibercept, endothelial cells use the highly conserved neonatal Fc receptor (FcRn) also expressed in iBREC where it is down regulated by serum depletion. Therefore, we focused on studying intracellular localization and transport of aflibercept under conditions affecting its interaction with the FcRn. Intracellular localization of aflibercept was assessed by Western-blot analyses of subcellular protein fractions or by immunofluorescence staining. After uptake in a temperature-dependent process, aflibercept co-localized with early endosomes, which harbor FcRn. Similar amounts of aflibercept were co-extracted with proteins from membranes/organelles irrespectively of the amount of FBS in the culture medium. Lowering the concentration of FBS resulted in a strong, but reversible association with cytoskeletal proteins suggesting a block in intracellular transport. In accordance with this finding, aflibercept's transport through an iBREC monolayer grown on porous membrane inserts was markedly delayed in the absence of FBS in the culture medium indicating that aflibercept is taken up but not exocytosed under these conditions. Transcytosis of aflibercept was also strongly delayed by inhibition of phosphatidylinositol 3-kinase with LY294002, which affects FcRn-mediated IgG transport. A similar inhibition of aflibercept's transport was observed with IgG-binding proteins (i.e. protein A or protein G) that block interaction between FcRn and aflibercept. Interfering with aflibercept's binding to the FcRn with protein A (or protein G) or the inhibitory FcRn-specific monoclonal antibody 1G3 resulted in a reduced amount of intracellular aflibercept. Taken together, our results strongly suggest that FcRn is involved in transport of aflibercept through REC in vitro.

[Accurate OCT-angiography Interpretation - Detection and Exclusion of Artifacts]. / Präzise Befundung mit der OCT-Angiografie ­ Artefakte erkennen und ausschließen.

Background Optical coherence tomography angiography (OCTA) provides, non-invasively, a three-dimensional visualization of the microvasculature of the retina and choroid. However, image artifacts may occur in OCTA and have an impact on clinical interpretation. The aim of this article is to describe image artifacts of OCTA and to present a nomenclature. Methods OCTA examinations were performed with the AngioPlex™ OCTA-technology in combination with the CIRRUS HD-OCT 5000 (Carl Zeiss Meditec, Inc., Dublin, USA) as well as with the PlexElite 9000 (Carl Zeiss Meditec, Inc., Dublin, USA). Typical artifacts identified in the OCTA images are described and their causes are explained. Results There are three main groups of artifacts that can be distinguished: (a) artifacts that are inherent in the OCTA technology and occur with all types of devices (projection artifacts, masking, unmasking, loss of signal); (b) artifacts caused by data and image processing algorithms and whose frequency or severity may depend on the device type used (segmentation artifacts, duplications of vessels); (c) motion artifacts that vary in frequency and severity depending on the type of device used, as different methods (e.g., eye tracker) are used to reduce them. The occurrence of artifacts is also dependent on patient cooperation, the clarity of the optical media, and the pathology of the retina. Conclusion As in any other imaging method, artifacts also occur in OCTA images. Nevertheless, qualitative assessment of OCTA images is almost always possible and provides indispensable findings on the morphology and perfusion status of the retina and choroid. A good knowledge of possible artifacts, and a critical analysis of the complete OCTA data set, allows a correct interpretation and is essential for making a precise clinical diagnosis.