When Two Eyes Don't Suffice-Learning Difficult Hyperfluorescence Segmentations in Retinal Fundus Autofluorescence Images via Ensemble Learning.

Hyperfluorescence (HF) and reduced autofluorescence (RA) are important biomarkers in fundus autofluorescence images (FAF) for the assessment of health of the retinal pigment epithelium (RPE), an important indicator of disease progression in geographic atrophy (GA) or central serous chorioretinopathy (CSCR). Autofluorescence images have been annotated by human raters, but distinguishing biomarkers (whether signals are increased or decreased) from the normal background proves challenging, with borders being particularly open to interpretation. Consequently, significant variations emerge among different graders, and even within the same grader during repeated annotations. Tests on in-house FAF data show that even highly skilled medical experts, despite previously discussing and settling on precise annotation guidelines, reach a pair-wise agreement measured in a Dice score of no more than 63-80% for HF segmentations and only 14-52% for RA. The data further show that the agreement of our primary annotation expert with herself is a 72% Dice score for HF and 51% for RA. Given these numbers, the task of automated HF and RA segmentation cannot simply be refined to the improvement in a segmentation score. Instead, we propose the use of a segmentation ensemble. Learning from images with a single annotation, the ensemble reaches expert-like performance with an agreement of a 64-81% Dice score for HF and 21-41% for RA with all our experts. In addition, utilizing the mean predictions of the ensemble networks and their variance, we devise ternary segmentations where FAF image areas are labeled either as confident background, confident HF, or potential HF, ensuring that predictions are reliable where they are confident (97% Precision), while detecting all instances of HF (99% Recall) annotated by all experts.

Real-Time Temperature-Controlled Retinal Laser Irradiation in Rabbits.

Purpose: Subdamaging thermal retinal laser therapy has the potential to induce regenerative stimuli in retinal diseases, but validated dosimetry is missing. Real-time optoacoustic temperature determination and control could close this gap. This study investigates a first in vivo application. Methods: Two iterations of a control module that were optically coupled in between a continuous-wave commercial laser source and a commercial slit lamp were evaluated on chinchilla rabbits. The module allows extraction of the temperature rise in real time and can control the power of the therapy laser such that a predefined temperature rise at the retina is quickly achieved and held constant. Irradiations with aim temperatures from 45°C to 69°C were performed on a diameter of 200 µm and a heating time of 100 ms. Results: We analyzed 424 temperature-guided irradiations in nine eyes of five rabbits. The mean difference between the measured and aim temperature was -0.04°C ± 0.98°C. The following ED50 values for visibility thresholds could be determined: 58.6°C for funduscopic visibility, 57.7°C for fluorescein angiography, and 57.0°C for OCT. In all measurements, the correlation of tissue effect was higher to the temperature than to the average heating laser power used. Conclusions: The system was able to reliably perform temperature-guided irradiations, which allowed for better tissue effect control than simple power control. This approach could enhance the accuracy, safety, and reproducibility of thermal stimulating laser therapy. Translational Relevance: This study is a bridge between preclinical ex vivo experiments and a pilot clinical study.

Optical coherence tomography angiography suggests choriocapillaris perfusion deficit as etiology of acute macular neuroretinopathy.

PURPOSE: Acute macular neuroretinopathy (AMN) can cause sudden-onset and permanent scotoma in healthy young patients. Analysis of optical coherence tomography (OCT) and OCT angiography (OCTA) of AMN patients may provide insights into disease mechanism. METHODS: We conducted a retrospective study of consecutive SARS-Cov-2-related AMN patients that presented in our clinic between Jan 1st, 2022, and April 30th, 2023, within 30 days of symptom onset. Retinal vessel area density (VAD) of AMN lesions in OCTA was quantified and compared to an adjacent tissue control (ATC). This quantification was performed for the superficial vascular plexus (SVP), the intermediate capillary plexus (ICP), the deep capillary plexus (DCP), the choriocapillaris (CC), and choroid. Furthermore, en face OCT images were analyzed. RESULTS: Nine AMN patients were identified, 6 of these (4 female, 2 male, average age 25 years) fulfilled the inclusion criteria and were included into this study. Average time from symptom onset to OCTA was 14.3 days. No VAD differences between AMN and adjacent tissue were found in either retinal layer (SVP, ICP, DCP). In contrast, VAD in CC was reduced by 27% against the ATC (p = 0.007) and choroidal VAD was reduced by 41% (p = 0.017). Further analysis of en face OCT could show that the pathognomonic infrared hyporeflectivity in AMN is caused by photoreceptor alterations rather than changes in the inner retinal layers. CONCLUSIONS: Our data suggests that a perfusion deficit in the choroidal layers is responsible for AMN rather than in the DCP, which is the predominant hypothesis in current literature.

Chronological Registration of OCT and Autofluorescence Findings in CSCR: Two Distinct Patterns in Disease Course.

Optical coherence tomography (OCT) and fundus autofluorescence (FAF) are important imaging modalities for the assessment and prognosis of central serous chorioretinopathy (CSCR). However, setting the findings from both into spatial and temporal contexts as desirable for disease analysis remains a challenge due to both modalities being captured in different perspectives: sparse three-dimensional (3D) cross sections for OCT and two-dimensional (2D) en face images for FAF. To bridge this gap, we propose a visualisation pipeline capable of projecting OCT labels to en face image modalities such as FAF. By mapping OCT B-scans onto the accompanying en face infrared (IR) image and then registering the IR image onto the FAF image by a neural network, we can directly compare OCT labels to other labels in the en face plane. We also present a U-Net inspired segmentation model to predict segmentations in unlabeled OCTs. Evaluations show that both our networks achieve high precision (0.853 Dice score and 0.913 Area under Curve). Furthermore, medical analysis performed on exemplary, chronologically arranged CSCR progressions of 12 patients visualized with our pipeline indicates that, on CSCR, two patterns emerge: subretinal fluid (SRF) in OCT preceding hyperfluorescence (HF) in FAF and vice versa.

Self-Examination Low-Cost Full-Field Optical Coherence Tomography (SELFF-OCT) for neovascular age-related macular degeneration: a cross-sectional diagnostic accuracy study.

OBJECTIVES: Self-Examination Low-Cost Full-Field Optical Coherence Tomography (SELFF-OCT) is a novel OCT technology that was specifically designed for home monitoring of neovascular age-related macular degeneration (AMD). First clinical findings have been reported before. This trial investigates an improved prototype for patients with AMD and focusses on device operability and diagnostic accuracy compared with established spectral-domain OCT (SD-OCT). DESIGN: Prospective single-arm diagnostic accuracy study. SETTING: Tertiary care centre (University Eye Clinic). PARTICIPANTS: 46 patients with age-related macular degeneration. INTERVENTIONS: Patients received short training in device handling and then performed multiple self-scans with the SELFF-OCT according to a predefined protocol. Additionally, all eyes were examined with standard SD-OCT, performed by medical personnel. All images were graded by at least 2 masked investigators in a reading centre. PRIMARY OUTCOME MEASURE: Rate of successful self-measurements. SECONDARY OUTCOME MEASURES: Sensitivity and specificity of SELFF-OCT versus SD-OCT for different biomarkers and necessity for antivascular endothelial growth factor (anti-VEGF) treatment. RESULTS: In 86% of all examined eyes, OCT self-acquisition resulted in interpretable retinal OCT volume scans. In these patients, the sensitivity for detection of anti-VEGF treatment necessity was 0.94 (95% CI 0.79 to 0.99) and specificity 0.95 (95% CI 0.82 to 0.99). CONCLUSIONS: SELFF-OCT was used successfully for retinal self-examination in most patients, and it could become a valuable tool for retinal home monitoring in the future. Improvements are in progress to reduce device size and to improve handling, image quality and success rates. TRIAL REGISTRATION NUMBER: DRKS00013755, CIV-17-12-022384.

[Ocular manifestations in patients with COVID-19]. / Okuläre Manifestationen bei COVID-19-Patienten.

BACKGROUND: There are only few reports on ocular symptoms and manifestations in association with coronavirus disease 2019 (COVID-19). OBJECTIVE: The aim of this study is to describe ocular manifestations in the anterior and posterior segments of the eye and to analyze viral prevalence in tears of patients with COVID-19. METHODS: Hospitalized COVID-19 patients treated from 16 April 2020 to 7 January 2021 at this hospital were screened for ocular manifestations in the anterior and posterior segments. Conjunctival swabs were analyzed for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA. RESULTS: A total of 37 patients were enrolled in this study. In the anterior segment we found chemosis of the conjunctiva (5), hyposphagma (2) and conjunctivitis (1). In 11 patients vascular alterations and potentially disease-specific manifestations of the fundus were found in one or both eyes: retinal hemorrhages (5), cotton wool spots (5) and tortuosity (5). One patient demonstrated branch artery occlusion, one had branch retinal vein occlusion and two patients had positive conjunctival swab results in one or both eyes. CONCLUSION: Our findings of the anterior segment are commonly known, although not specific for COVID-19. Various vascular fundus abnormalities were found in the study; however, it is unclear whether these were correlated to systemic comorbidities or whether they were caused or exacerbated by COVID-19. This study suggests that the risk of viral transmission via tears is low.

Selective retina therapy and thermal stimulation of the retina: different regenerative properties - implications for AMD therapy.

BACKGROUND: Selective Retina Therapy (SRT), a photodisruptive micropulsed laser modality that selectively destroys RPE cells followed by regeneration, and Thermal Stimulation of the Retina (TSR), a stimulative photothermal continuous wave laser modality that leads to an instant sublethal temperature increase in RPE cells, have shown therapeutic effects on Age-related Macular Degeneration (AMD) in mice. We investigate the differences between both laser modalities concerning RPE regeneration. METHODS: For PCR array, 6 eyes of murine AMD models, apolipoprotein E and nuclear factor erythroid-derived 2- like 2 knock out mice respectively, were treated by neuroretina-sparing TSR or SRT. Untreated litter mates were controls. Eyes were enucleated either 1 or 7 days after laser treatment. For morphological analysis, porcine RPE/choroid organ cultures underwent the same laser treatment and were examined by calcein vitality staining 1 h and 1, 3 or 5 days after irradiation. RESULTS: TSR did not induce the expression of cell-mediators connected to cell death. SRT induced necrosis associated cytokines as well as inflammation 1 but not 7 days after treatment. Morphologically, 1 h after TSR, there was no cell damage. One and 3 days after TSR, dense chromatin and cell destruction of single cells was seen. Five days after TSR, there were signs of migration and proliferation. In contrast, 1 h after SRT a defined necrotic area within the laser spot was seen. This lesion was closed over days by migration and proliferation of adjacent cells. CONCLUSIONS: SRT induces RPE cell death, followed by regeneration within a few days. It is accompanied by necrosis induced inflammation, RPE proliferation and migration. TSR does not induce immediate RPE cell death; however, migration and mitosis can be seen a few days after laser irradiation, not accompanied by necrosis-associated inflammation. Both might be a therapeutic option for the treatment of AMD.

Self-examination low-cost full-field OCT (SELFF-OCT) for patients with various macular diseases.

PURPOSE: The treatment guidelines for many macular diseases rely on frequent monitoring with optical coherence tomography (OCT). However, the burden of frequent disease control leads to low therapy adherence in real life. OCT home monitoring would address this issue but requires an inexpensive and self-operable device. With self-examination low-cost full-field OCT (SELFF-OCT), our group has introduced a novel technology that may fulfill both requirements. In this pilot study, we report the initial experiences with a clinical prototype. METHODS: Fifty-one patients with different macular diseases were recruited in a cross-sectional study. The most common diseases were age-related macular degeneration (AMD; 39/51), diabetic macular edema (DME; 6/51), and retinal vein occlusion (RVO; 3/51). Patients received a short training in device usage and then performed multiple self-scans with the SELFF-OCT device. For comparison, scans with a standard clinical spectral domain (SD-)OCT were taken. RESULTS: After a brief training, 77% of the patients were able to successfully acquire images that were clinically gradable. No significant influence on success could be found for age (p = 0.08) or BCVA (p = 0.97). Relevant disease biomarkers in the most common retinal diseases could be detected. CONCLUSIONS: SELFF-OCT was used successfully for retinal self-examination and in the future could be used for retinal home monitoring. Future improvements in technology are expected to improve success rates and image quality. TRIAL REGISTRATION: The Trial was registered in the German Trial Register under the number DRKS00013755 on 14.03.2018.

CRB1rd8 mutation influences the age-related macular degeneration phenotype of NRF2 knockout mice and favors choroidal neovascularization.

PURPOSE: We examined the influence of retinal degeneration 8 (rd8) mutation of crumbs homolog 1 (CRB1) gene on age-related macular degeneration (AMD) phenotype in nuclear factor E2-related factor 2 knock out (NRF2-/-) mouse model. METHODS: CRB1rd8 mutation genotype was determined by polymerase chain reaction from tail clips in 73 NRF2-/- mice originating from C57BL/6J background on mixed C57BL/6J and C57BL/6N ancestry. The clinical grade of AMD-like fundus alterations was determined by funduscopy, optical coherence tomography (OCT) and fluorescein angiography (FLA) at the age of 9 or 12 months. RESULTS: Twelve NRF2-/- mice were wildtype CRB1+/+, 61 NRF2-/- were homozygous CRB1rd8/rd8. NRF2-/-CRB1rd8/rd8 mice had a significantly higher probability to show an advanced grade (grade 4 and 5) of AMD-like fundus alterations known to appear in NRF2-/- mice. Choroidal neovascularization (CNV) was only detected in NRF2-/-CRB1rd8/rd8 homozygous mice. CONCLUSIONS: Homozygous CRB1rd8/rd8 mutation is common in commercial vendor mice strains of C57BL/6J origin if partly on C57BL/6N ancestry. The mutation has an influence on the extent of AMD-like retinal alterations in NRF2-/- mice and favors CNV formation.

Modulation of inflammatory processes by thermal stimulating and RPE regenerative laser therapies in age related macular degeneration mouse models.

PURPOSE: Inflammatory processes play a major role within the multifactorial pathogenesis of age-related macular degeneration (AMD). Neuroretina sparing laser therapies, thermal stimulation of the retina (TSR) and selective retina therapy (SRT), are known to reduce AMD-like pathology in vitro and in vivo. We investigated the effect of TSR and SRT on inflammatory processes in AMD mouse models. METHODS: One randomized eye of 8 months old apolipoprotein (Apo)E and 9 months old nuclear factor (erythroid-derived 2) -like 2 (NRF2) knock out mice were treated by TSR (10 ms, 532 nm, 50 µm2 spot size, mean 4.5 W, ~200 spots) or SRT (~1.4 µs pulses, 532 nm, 50 µm spot size, 100 Hz over 300 ms, mean 2.5 µJ per pulse, ~200 spots). Fellow eyes, untreated knock out mice and wild-type BL/6J mice acted as controls. All mice were examined funduscopically and by optical coherence tomography (OCT) at the day of laser treatment. Mice were euthanized and enucleated either 1 day or 7 days after laser treatment and examined by gene expression analysis of 84 inflammatory genes. RESULTS: The inflammatory gene expression profile of both knock out models compared to healthy BL/6J mice suggests a regulation of pro- and anti-inflammatory processes especially concerning T-cell activity and immune cell recruitment. TSR resulted in downregulation of several pro-inflammatory cell-mediators both in ApoE -/- and NRF2-/- mice compared to treatment naïve litter mates one day after treatment. In contrast, SRT induced pro-inflammatory cell-mediators connected with necrosis one day after treatment as expected following laser-induced selective RPE cell death. Seven days after laser treatment, both findings were reversed. CONCLUSIONS: Both TSR and SRT influence inflammatory processes in AMD mouse models. However, they act conversely. TSR leads to anti-inflammatory processes shortly after laser therapy and induces immune-cell recruitment one week after treatment. SRT leads to a quick inflammatory response to laser induced RPE necrotic processes. One week after SRT inflammation is inhibited. It remains unclear, if and to what extent this might play a role in a therapeutic or preventive approach of both laser modalities on AMD pathology.

[Effect of Selective Retina Therapy (SRT) on the Inflammatory Microenvironment of the Subretinal Space]. / Einfluss der Selektiven Retinatherapie (SRT) auf inflammatorische Zellmediatoren des subretinalen Raums.

PURPOSE: To investigate the effect of Selective Retina Therapy (SRT) on inflammatory key factors such as complement factor-C3 (CC3), tumor growth factor-beta2 (TGF-ß2), tumor necrosis factor-alpha (TNF-α) and interferon-gamma (IFN-γ). MATERIALS AND METHODS: Porcine RPE-Bruch's membrane-choroid explants were irradiated with two SRT laser systems, SRTYLF and SRTYAG (Nd : YLF laser, wave length 527 nm, pulse duration 1.7 µs and Nd : YAG laser, wave length 532 nm, pulse duration 2.4 - 3 µs). Laser irradiation was performed on a spot size of 200 × 200 µm, 30 pulses, with a repetition rate of 100 Hz, and a radiant exposure of 140 (threshold RPE death) and 180 mJ/cm2 per pulse (above threshold RPE death). Explants were cultivated in modified Ussing chambers and culture viability was assessed by calcein-AM cell staining. Secretion of inflammatory factors was analyzed by ELISA. Protein expression of tissue explants was assessed by Western blot. RESULTS: Regeneration of RPE was observed after 4 days. One day after SRT with 140 mJ/cm2 per pulse the secretion of basal CC3 decreased in ELISA. Following 180 mJ/cm2 radiant exposure, the level of IFN-γ decreased at day 4. CONCLUSION: SRT does not induce the release of the pro-inflammatory factors analyzed in this in-vitro study.

Selective Retina Therapy Reduces Bruch's Membrane Thickness and Retinal Pigment Epithelium Pathology in Age-Related Macular Degeneration Mouse Models.

PURPOSE: To investigate the effect of selective retina therapy (SRT) on age-related macular degeneration (AMD)-like alterations of retinal pigment epithelium (RPE) and Bruch's membrane (BrM) in AMD mouse models as therapeutic approach for the treatment of dry AMD. METHODS: In B6.129P2-Apoetm1Unc /J (ApoE-/-) and B6.129X1-Nfe2I2 tm1Ywk /J (NRF2-/-), one randomized eye of each mouse in groups of 15 mice was treated by SRT (532 nm, 300 ms, ∼1.4-µs pulse, 100 Hz, 50-µm spot), the fellow eye and healthy C57BL/6J mice served as controls. Clinical examinations were obtained at treatment day and 1 month later, followed by enucleation to analyze BrM thickness and ultrastructural RPE morphology. RESULTS: Nearly all ApoE-/- and NRF2-/- mice showed AMD-like retinal alterations. BrM thickness was increased in both mouse models, RPE had vacuoles within the cell body and shortened apical microvilli. SRT neither affected neuroretinal anatomy nor function. BrM thickness as well as AMD-like ultrastructural alterations of the RPE were significantly reduced in laser-treated eyes compared with fellow control and untreated control eyes. CONCLUSIONS: SRT reduces BrM thickness and AMD-like RPE alterations in AMD mouse models without damage to structural or functional properties of neuroretina. It may be a prophylactic or therapeutic option for dry AMD. TRANSLATIONAL RELEVANCE: SRT shows therapeutic effectivity in murine AMD models and might therefore become an option for the treatment of dry AMD.

Retinal volume change is a reliable OCT biomarker for disease activity in neovascular AMD.

PURPOSE: Current algorithms for automated computer interpretation of optical coherence tomography (OCT) imaging of patients suffering from neovascular age-related macular degeneration (AMD) mostly rely on fluid detection. However, fluid detection itself and correct interpretation of the fluid currently limits diagnostic accuracy. We therefore performed a detailed analysis of the requirements that would have to be met for fluid detection approaches. We further investigated if monitoring retinal volume would be a viable alternative to detect disease activity. METHODS: Retrospective analysis and manual grading of 764 OCT volume scans of 44 patients with exudative AMD treated with intravitreal anti-VEGF injections at a pro-re-nata (PRN) treatment regimen for at least 24 months. RESULTS: Detection of subretinal fluid (SRF) or intraretinal fluid (IRF) alone is not sufficient for disease detection. A combination of SRF and IRF can detect disease activity with a sensitivity of 98.6% and a specificity of 82%. With further characterization of IRF into exudative and degenerative cysts, specificity can be increased to 100%. However, correct characterization is currently not achieved by published fluid detection approaches. Change of macular retinal volume (MRV) can depict disease activity with sensitivity of 88.4% and specificity of 89.6%. Combination with the detection of SRF can further improve diagnostic accuracy to a specificity of 93.3% and sensitivity of 93.9% without relying on IRF or IRF characterization. CONCLUSION: Fluid detection without further characterization is not sufficient for AMD monitoring. Either further distinction between exudative and degenerative cysts is necessary, or other activity markers have to be taken into account. MRV offers good potential to fill this diagnostic gap and might become an important monitoring marker.

Thermal Stimulation of the Retina Reduces Bruch's Membrane Thickness in Age Related Macular Degeneration Mouse Models.

PURPOSE: To investigate the effect of thermal stimulation of the retina (TS-R) on Bruch's membrane (BrM) thickness in age-related macular degeneration (AMD) mouse models as a novel concept for the prophylaxis and treatment of dry AMD. METHODS: Two knockout AMD mouse models, B6.129P2-Apoetm1Unc/J (ApoE-/-) and B6.129X1-Nfe2I2tm1Ywk/J (NRF2-/-), were chosen. One randomized eye of each mouse in four different groups (two of different age, two of different genotype) of five mice was treated by TS-R (532 nm, 10-ms duration, 50-µm spot size), the fellow eye served as control. Laser power was titrated to barely visible laser burns, then reduced by 70% to guarantee for thermal elevation without damage to the neuroretina, then applied uniformly to the murine retina. Fundus, optical coherence tomography (OCT), and fluorescein angiography (FLA) images were obtained at the day of treatment and 1 month after treatment. Eyes were enucleated thereafter to analyze BrM thickness by transmission electron microscopy (TEM) in a standardized blinded manner. RESULTS: Fundus images revealed that all ApoE-/- and NRF2-/- mice had AMD associated retinal alterations. BrM thickness was increased in untreated controls of both mouse models. Subvisible TS-R laser spots were not detectable by fundus imaging, OCT, or FLA 2 hours or 1 month after laser treatment. TEM revealed a significant reduction of BrM thickness in laser-treated eyes of all four groups compared to their fellow control eyes. CONCLUSIONS: TS-R reduces BrM thickness in AMD mouse models ApoE-/- and NRF2-/- without damage to the neuroretina. It may become a prophylactic or even therapeutic treatment option for dry AMD. TRANSLATIONAL RELEVANCE: TS-R may become a prophylactic or even therapeutic treatment option for dry AMD.

Power-controlled temperature guided retinal laser therapy.

Laser photocoagulation has been a treatment method for retinal diseases for decades. Recently, studies have demonstrated therapeutic benefits for subvisible effects. A treatment mode based on an automatic feedback algorithm to reliably generate subvisible and visible irradiations within a constant irradiation time is introduced. The method uses a site-individual adaptation of the laser power by monitoring the retinal temperature rise during the treatment using optoacoustics. This provides feedback to adjust the therapy laser power during the irradiation. The technique was demonstrated on rabbits in vivo using a 532-nm continuous wave Nd:YAG laser. The temperature measurement was performed with 523-nm Q-switched Nd:YLF laser pulses with 75-ns pulse duration at 1-kHz repetition rate. The beam diameter on the fundus was 200 µm for both lasers, respectively. The aim temperatures ranged from 50°C to 75°C in 11 eyes of 7 rabbits. The results showed ophthalmoscopically invisible effects below 55°C with therapy laser powers over a wide range. The standard deviation for the measured temperatures ranged from 2.1°C for an aim temperature of 50°C to 4.7°C for 75°C. The ED50 temperature value for ophthalmoscopically visible lesions in rabbits was determined as 65.3°C. The introduced method can be used for retinal irradiations with adjustable temperature elevations.