[Clinical approach for solid intraocular metastases]. / Klinisches Vorgehen bei soliden intraokulären Metastasen.

BACKGROUND: Intraocular metastases arising from solid tumors are found in approximately 2% of patients with metastatic tumor diseases and are therefore more frequent than originally assumed. They often affect the uvea and are associated with a poor prognosis. Due to the difficult diagnosis and an inconsistent treatment regimen, ophthalmologists have a special responsibility here. OBJECTIVE: This article gives a summary of the various types of intraocular metastases with respect to clinical features, diagnostics, treatment and prognosis as well as recommendations for follow-up care. METHODS: A selective literature search was carried out on the topic of intraocular metastases using PubMed and Google Scholar. RESULTS: Intraocular metastases most frequently affect the uvea, specifically the choroid. In most cases the underlying disease is breast or lung cancer, but other rarer primary tumors have also been reported in the literature. Metastatic lesions can show very different morphological manifestations but can be distinguished based on the corresponding structure of manifestation in the eye and with the aid of targeted staging, thus providing valid information on the type of primary tumor. The treatment is partly experimental and usually depends on the primary tumor and leading symptoms of the patient. A differentiation between a curative or palliative treatment situation must always be made. CONCLUSION: Intraocular metastases are the most frequent intraocular tumor and are usually associated with a poor prognosis. Accurate diagnostics for finding the treatment as well as interdisciplinary collaboration and the presentation of the patient on the tumor board are essential.

Detection sensitivity of fluorescence lifetime imaging ophthalmoscopy for laser-induced selective damage of retinal pigment epithelium.

PURPOSE: To investigate the sensitivity of fluorescence lifetime imaging ophthalmoscopy (FLIO) to detect retinal laser spots by comparative analysis with other imaging modalities. METHODS: A diode laser with a wavelength of 514 nm was applied with pulse durations of 5.2, 12, 20, and 50 µs. The laser pulse energy was increased so that the visibility of the laser spot by slit-lamp fundus examination (SL) under the irradiator's observation covers from the subvisible to visible range immediately after irradiation. The irradiated areas were then examined by fundus color photography (FC), optical coherence tomography (OCT), fundus autofluorescence (AF), FLIO, and fluorescein angiography (FA). The visibility of a total of over 2200 laser spots was evaluated by two independent researchers, and effective dose (ED) 50 laser pulse energy values were calculated for each imaging modality and compared. RESULTS: Among examined modalities, FA showed the lowest mean of ED50 energy value and SL the highest, that is, they had the highest and lowest sensitivity to detect retinal pigment epithalium (RPE)-selective laser spots, respectively. FLIO also detected spots significantly more sensitively than SL at most laser pulse durations and was not significantly inferior to FA. AF was also often more sensitive than SL, but the difference was slightly less significant than FLIO. CONCLUSION: Considering its high sensitivity in detecting laser spots and previously reported potential of indicating local wound healing and metabolic changes around laser spots, FLIO may be useful as a non-invasive monitoring tool during and after minimally invasive retinal laser treatment.

In vivo evaluation of a nanotechnology-based microshunt for filtering glaucoma surgery.

To carry out the preclinical and histological evaluation of a novel nanotechnology-based microshunt for drainage glaucoma surgery. Twelve New Zealand White rabbits were implanted with a novel microshunt and followed up for 6 weeks. The new material composite consists of the silicone polydimethylsiloxane (PDMS) and tetrapodal Zinc Oxide (ZnO-T) nano-/microparticles. The microshunts were inserted ab externo to connect the subconjunctival space with the anterior chamber. Animals were euthanized after 2 and 6 weeks for histological evaluation. Ocular health and implant position were assessed at postoperative days 1, 3, 7 and twice a week thereafter by slit lamp biomicroscopy. Intraocular pressure (IOP) was measured using rebound tonometry. A good tolerability was observed in both short- and medium-term follow-up. Intraocular pressure was reduced following surgery but increased to preoperative levels after 2 weeks. No clinical or histological signs of inflammatory or toxic reactions were seen; the fibrotic encapsulation was barely noticeable after two weeks and very mild after six weeks. The new material composite PDMS/ZnO-T is well tolerated and the associated foreign body fibrotic reaction quite mild. The new microshunt reduces the IOP for 2 weeks. Further research will elucidate a tube-like shape to improve and prolong outflow performance and longer follow-up to exclude medium-term adverse effects.

Artificial Intelligence in Fluorescence Lifetime Imaging Ophthalmoscopy (FLIO) Data Analysis-Toward Retinal Metabolic Diagnostics.

The purpose of this study was to investigate the possibility of implementing an artificial intelligence (AI) approach for the analysis of fluorescence lifetime imaging ophthalmoscopy (FLIO) data even with small data. FLIO data, including the fluorescence intensity and mean fluorescence lifetime (τm) of two spectral channels, as well as OCT-A data from 26 non-smokers and 28 smokers without systemic and ocular diseases were used. The analysis was performed with support vector machines (SVMs), a well-known AI method for small datasets, and compared with the results of convolutional neural networks (CNNs) and autoencoder networks. The SVM was the only tested AI method, which was able to distinguish τm between non-smokers and heavy smokers. The accuracy was about 80%. OCT-A data did not show significant differences. The feasibility and usefulness of the AI in analyzing FLIO and OCT-A data without any apparent retinal diseases were demonstrated. Although further studies with larger datasets are necessary to validate the results, the results greatly suggest that AI could be useful in analyzing FLIO-data even from healthy subjects without retinal disease and even with small datasets. AI-assisted FLIO is expected to greatly advance early retinal diagnosis.

Fluorescence Lifetime Imaging Ophthalmoscopy of Mouse Models of Age-related Macular Degeneration.

Purpose: To investigate fluorescence lifetime of mouse models of age-related macular degeneration (AMD) by fluorescence lifetime imaging ophthalmoscopy (FLIO). Methods: Two AMD mouse models, apolipoprotein E knockout (ApoE-/-) mice and NF-E2-related factor-2 knockout (Nrf2-/-) mice, and their wild-type mice underwent monthly ophthalmic examinations including FLIO from 3 months of age. After euthanasia at the age of 6 or 11 months, blood plasma was collected to determine total antioxidant capacity and eyes were enucleated for Oil red O (ORO) lipid staining of chorioretinal tissue. Results: In FLIO, the mean fluorescence lifetime (τm) of wild type shortened with age in both spectral channels. In short spectral channel, τm shortening was observed in both AMD models as well, but its rate was more pronounced in ApoE-/- mice and significantly different from the other strains as months of age progressed. In contrast, in long spectral channel, both model strains showed completely opposite trends, with τm becoming shorter in ApoE-/- and longer in Nrf2-/- mice than the others. Oil red O staining at Bruch's membrane was significantly stronger in ApoE-/- mice at 11 months than the other strains. Plasma total antioxidant capacity was highest in ApoE-/- mice at both 6 and 11 months. Conclusions: The two AMD mouse models exhibited largely different fundus fluorescence lifetime, which might be related to the different systemic metabolic state. FLIO might be able to indicate different metabolic states of eyes at risk for AMD. Translational Relevance: This animal study may provide new insights into the relationship between early AMD-associated metabolic changes and FLIO findings.

Detection of Circulating Tumor Cells in Patients with Small Choroidal Melanocytic Lesions.

PURPOSE: To determine the presence of circulating tumor cells (CTCs) in patients with indeterminate small choroidal melanocytic lesions (SCMLs). DESIGN: Retrospective case series. PARTICIPANTS: Forty-seven patients with choroidal melanocytic lesions 2.5 mm or less in tumor thickness and ≤ 10 mm in largest basal diameter (LBD). METHODS: Blood samples were analyzed for CTCs and the presence of monosomy-3 (M3) in CTCs. Tissue biopsy was performed in the patients who were CTC-positive (pCTC). MAIN OUTCOME MEASURES: Presence and M3 status of the CTCs with regard to the clinical characteristics and results from tissue biopsy. RESULTS: Median thickness of all (n = 47) lesions was 1.1 mm (range: 0.2-2.5 mm), and LBD was 5.6 mm (range: 2.0-10.0 mm). Circulating tumor cells were found in 25 patients (n = 25). This group was classified as pCTC and compared with the CTC-negative (nCTC) group consisting of 22 patients (n = 22). Median tumor dimensions in the pCTC versus the nCTC group were 1.6 mm (range: 0.6-2.5 mm) versus 0.5 mm (range: 0.2-2.5 mm) for thickness and 6.6 mm (range: 4.1-10.0 mm) versus 4.0 mm (range: 2.0-8.0 mm) for LBD, respectively. Both LBD and thickness were positively associated (P < 0.001) with the presence of CTC. Compared with the nCTC group, a higher percentage of the pCTC group exhibited LBD > 5 mm (36% vs. 88%), subretinal fluid (9.1% vs. 56%), orange pigment (4.5% vs. 60%), sonographic hollowness (9.1% vs. 60%), and the presence of multiple risk factors (0% vs. 68% for ≥3 factors) with P < 0.001 for all parameters. No significant difference was detected in the clinical parameters of the patients who had disomy-3 (D3) (n = 7) versus M3 (n = 17) in their CTC. The tissue biopsy confirmed the uveal melanoma (UM) in 22 of the 25 pCTC patients (88%), whereas no conclusive diagnosis could be determined in the remaining 3 cases because of insufficient or invalid material. CONCLUSIONS: We report compelling evidence for the potential of liquid biopsy as an additional tool to screen SCMLs for malignancy. These findings pave the way toward the implementation of liquid biopsy to detect small UM and monitor melanocytic lesions. FINANCIAL DISCLOSURE(S): Proprietary or commercial disclosure may be found in the Footnotes and Disclosures at the end of this article.

Impact of cigarette smoking on fluorescence lifetime of ocular fundus.

Cigarette smoking is known to adversely affect cellular metabolism and is a risk factor for various retinal diseases. Fluorescence lifetime imaging ophthalmoscopy (FLIO) has the potential to detect metabolic changes in the ocular fundus. Aim of this study was to analyze the influence of cigarette smoking on fluorescence lifetime (FLT) of healthy eyes using FLIO. Twenty-six non-smokers and 28 smokers aged between 20 and 37 years without systemic and ocular diseases were investigated by FLIO (excitation: 473 nm, emission: short spectral channel (SSC) 498-560 nm, long spectral channel (LSC) 560-720 nm). The FLT at the ETDRS grid regions were analyzed and compared. In SSC, the mean FLT (τm) of smokers was significantly longer in the ETDRS inner ring region, whereas the τm in LSC was significantly shorter in the outer ring. For the long component (τ2), smokers with pack year < 7.11 showed significantly shorter τ2 in SSC than non-smokers and the smokers with pack year ≥ 7.11. There were no significant differences in retinal thickness. The lack of obvious structural differences implies that the observed FLT changes are likely related to smoking-induced metabolic changes. These results suggest that FLIO may be useful in assessing retinal conditions related to lifestyle and systemic metabolic status.

Zinc Oxide Tetrapods Modulate Wound Healing and Cytokine Release In Vitro-A New Antiproliferative Substance in Glaucoma Filtering Surgery.

Glaucoma filtering surgery is applied to reduce intraocular pressure (IOP) in cases of uncontrolled glaucoma. However, postoperative fibrosis reduces the long-term success of both standard trabeculectomy and microstents. The aim of this study was to test the antiproliferative and anti-inflammatory potential of ZnO-tetrapods (ZnO-T) on human Tenon's fibroblasts (HTFs) for glaucoma surgery. The toxicity of ZnO-T on HTFs was determined using an MTT test. For analysis of fibroblast proliferation, migration, and transdifferentiation, cultures were stained for Ki67, alpha-smooth muscle actin (α-SMA), and p-SMAD. A fully quantitative multiplex ELISA was used to determine the concentrations of different cytokines, platelet-derived growth factor (PDGF), and hepatocyte growth factor (HGF) in culture supernatants with and without previous ZnO-T treatment. Treatment with higher concentrations (10 and 20 µg/mL) was associated with HTF toxicity, as shown in the wound healing assay. Furthermore, the number of Ki67, α-SMA-positive, and pSMAD-positive cells, as well as IL-6 and HGF in supernatants, were significantly reduced following incubation with ZnO-T. In conclusion, we were able to show the antiproliferative and anti-inflammatory potentials of ZnO-T. Therefore, the use of ZnO-T may provide a new approach to reducing postoperative fibrosis in glaucoma filtering surgery.

In Vitro Evaluation of Zinc Oxide Tetrapods as a New Material Component for Glaucoma Implants.

In our previous study we were able to show that zinc oxide (ZnO) tetrapods inhibit wound healing processes. Therefore, the aim of this study was to test the antiproliferative effect of two types of porous polydimethylsiloxane (PDMS)/ tetrapodal zinc oxide (ZnO-T) materials, as well as their usability for glaucoma implants. To find the best implant material, two different porous PDMS/ZnO-T materials were examined. One consisted of 3D interconnected PDMS coarse-pored foams with protruding ZnO-T particles; the other consisted of fine-pored 3D interconnected ZnO-T networks homogeneously coated by a thin PDMS film in the nanometer range. Fibroblast cell viability was investigated for both materials via MTT dye, and some implant material samples were further processed for electron microscopy. Both PDMS/ZnO-T materials showed reduced cell viability in the MTT staining. Furthermore, the electron microscopy revealed barely any fibroblasts growing on the implant materials. At the surface of the fine-pored implant material, however, fibroblasts could not be observed in the etched control samples without ZnO-T. It was found that post-processing of the material to the final stent diameter was highly challenging and that the fabrication method, therefore, had to be adapted. In conclusion, we were able to demonstrate the antiproliferative potential of the two different PDMS/ZnO-T materials. Furthermore, smaller pore size (in the range of tens of micrometers) in the implant material seems to be preferable.

Fluorescence Lifetime Changes Induced by Laser Irradiation: A Preclinical Study towards the Evaluation of Retinal Metabolic States.

Fluorescence Lifetime (FLT) of intrinsic fluorophores may alter under the change in metabolic state. In this study, the FLT of rabbit retina was investigated in vivo after laser irradiation using fluorescence lifetime imaging ophthalmoscopy (FLIO). The retina of the Chinchilla bastard rabbits was irradiated with a 514 nm diode laser. FLIO, fundus photography, and optical coherence tomography (OCT) were conducted 30 min and 1 to 3 weeks after treatment. After strong coagulation, the FLT at laser spots was significantly elongated immediately after irradiation, conversely shortened after more than a week. Histological examination showed eosinophilic substance and melanin clumping in subretinal space at the coagulation spots older than one week. The FLT was also elongated right around the coagulation spots, which corresponded to the discontinuous ellipsoid zone (EZ) on OCT. This EZ change was recovered after one week, and the FLT became the same level as the surroundings. In addition, there was a region around the laser spot where the FLT was temporarily shorter than the surrounding area. When weak pulse energy was applied to selectively destroy only the RPE, a shortening of the FLT was observed immediately around the laser spot within one week after irradiation. FLIO could serve as a tool to evaluate the structural and metabolic response of the retina to laser treatments.

Investigations on Retinal Pigment Epithelial Damage at Laser Irradiation in the Lower Microsecond Time Regime.

Purpose: New lasers with a continuous wave power exceeding 15 W are currently investigated for retinal therapies, promising highly localized effects at and close to the Retinal Pigment Epithelium (RPE). The goal of this work is to evaluate mechanisms and thresholds for RPE cell damage by means of pulse durations up to 50 µs. Methods: A diode laser with a wavelength of 514 nm, a power of 15 W, and adjustable pulse durations between 2 µs and 50 µs was used. Porcine RPE-choroidal explants (ex vivo) and chinchilla bastard rabbits (in vivo) were irradiated to determine threshold radiant exposures for RPE damage \({\bar H_{Cell}}\) by calcein vitality staining and fluorescence angiography, respectively. Thresholds for microbubble formation (MBF) \({\bar H_{MBF}}\) were evaluated by time-resolved optoacoustics. Exemplary histologies support the findings. Results: \({\bar H_{{{MBF}}}}\) is significantly higher than \({\bar H_{Cell}}\) at pulse durations ≥ 5 µs (P < 0.05) ex vivo, while at 2 µs, no statistically significant difference was found. The ratios between \({\bar H_{{{MBF}}}}\) and \({\bar H_{Cell}}\) increase with pulse duration from 1.07 to 1.48 ex vivo and 1.1 to 1.6 in vivo, for 5.2 and 50 µs. Conclusions: Cellular damage with and without MBF related disintegration are both present and very likely to play a role for pulse durations ≥ 5 µs. With the lower µs pulses, selective RPE disruption might be possible, while higher values allow achieving spatially limited thermal effects without MBF. However, both modi require a very accurate real-time dosing control in order to avoid extended retinal disintegration in this power range.

Fluorescence Lifetime Imaging Ophthalmoscopy of the Retinal Pigment Epithelium During Wound Healing After Laser Irradiation.

PURPOSE: To investigate the change in fluorescence lifetime of retinal pigment epithelium (RPE) after laser irradiation by using an organ culture model. METHODS: Porcine RPE-choroid-sclera explants were irradiated with selective retina treatment laser (wavelength: 527 nm, beam diameter: 200 µm, energy: 80-150 µJ). At 24 and 72 hours after irradiation, the mean fluorescence lifetime (τm ) was measured with fluorescence lifetime imaging ophthalmoscopy (FLIO) (excitation wavelength: 473 nm, emission: short spectral channel: 498-560 nm, long spectral channel: 560-720 nm). For every laser spot, central damaged zone (zone 1: 120 × 120 µm), area including wound rim (280 × 280 µm except zone 1), and environmental zone (440 × 440 µm except zone 1 and 2) were analyzed. Peripheral zone at a distance from laser spots longer than 2000 µm was examined for comparison. Cell viability was evaluated with calcein-acetoxymethyl ester and morphology with fluorescence microscopy for filamentous-actin. RESULTS: The RPE defect after selective retina treatment was mostly closed within 72 hours. FLIO clearly demarcated the irradiated region, with prolonged τm at the center of the defect decreasing with eccentricity. In short spectral channel, but not in long spectral channel, τm in the environmental zone after 72 hours was still significantly longer than in the peripheral zone. CONCLUSIONS: FLIO may clearly demarcate the RPE defect, demonstrate its closure, and, moreover, indicate the induced metabolic changes of surrounding cells during wound healing. TRANSLATIONAL RELEVANCE: This ex vivo study showed that FLIO may be used to evaluate the extent and quality of restoration of the damaged RPE and to detect its metabolic change in human fundus noninvasively.

Diuretic state affects ascending thin limb tight junctions.

The nephron segments in the inner medulla are part of the urine concentrating mechanism. Depending on the diuretic state, they are facing a large range of extracellular osmolality. We investigated whether water homeostasis affects tubular transport and permeability properties in inner medullary descending thin limb (IMdTL) and ascending thin limb (IMaTL). Three experimental groups of rats under different diuretic states were investigated on metabolic cages: waterload, furosemide-induced diuresis, and control (antidiuresis). Urine production and osmolalities reflected the 3-day treatment. To functionally investigate tubular epithelial properties, we performed experiments in freshly isolated inner medullary thin limbs from these animals. Tubular segments were acutely dissected and investigated for trans- and paracellular properties by in vitro perfusion and electrophysiological analysis. IMdTL and IMaTL were distinguished by morphological criteria. We confirmed absence of transepithelial electrogenic transport in thin limbs. Although diffusion potential measurements showed no differences between treatments in IMdTLs, we observed increased paracellular cation selectivity under waterload in IMaTLs. NaCl diffusion potential was -5.64 ± 1.93 mV under waterload, -1.99 ± 1.72 mV under furosemide-induced diuresis, and 0.27 ± 0.40 mV under control. The corresponding permeability ratio PNa/Cl was 1.53 ± 0.21 (waterload), 1.22 ± 0.18 (furosemide-induced diuresis), and 0.99 ± 0.02 (control), respectively. Claudins are main constituents of the tight junction responsible for paracellular selectivity; however, immunofluorescence did not show qualitative differences in claudin 4, 10, and 16 localization. Our results show that IMaTLs change tight junction properties in response to diuretic state to allow adaptation of NaCl reabsorption.