CXCL13 as a biomarker in the diagnostics of European lyme Neuroborreliosis - A prospective multicentre study in Austria.

Background: 'Definite Neuroborreliosis (NB)' is diagnosed with the presence of NB-specific symptoms, cerebrospinal fluid (CSF) pleocytosis and an elevated Borrelia Burgdorferi antibody index. However, some diagnostic uncertainties exist. The B-cell chemokine CXCL13 represents an emerging biomarker for the diagnosis and treatment of NB because its intrathecal concentration rises prior to the Borrelia antibody index and drops rapidly after antibiotic therapy. Nevertheless, due to lacking prospective data, a definite CXCL13 cut-off for the diagnosis of NB is still pending. Objective: Definition of a CSF CXCL13 cut-off for the diagnosis of acute and untreated NB in a prospective study setting. Design and methods: This multicentre prospective study involved 6 neurological departments treating patients in the Lower Austria district (1.7 million inhabitants). The controls were patients scheduled for a spinal tap but not clinically diagnosed with NB. Demographic data, clinical characteristics and blood counts, as well as inflammatory CSF values and CSF CXCL13-concentration were analysed. Results: We recruited 440 adult patients, of whom 42 have been diagnosed as having an acute and untreated 'definite NB'. Three hundred ninety-eight patients were assigned to the control group. The median intrathecal CXCL13 concentration was 2384 pg/ml for patients with NB and 0 pg/ml for controls. The difference was highly statistically significant (P ≤ .001). A CSF CXCL13 cut-off of 271 pg/ml resulted in a sensitivity of 95.2% and a specificity of 97.2% for the confirmation or exclusion of NB. Conclusion: Based on our results, we propose a CSF CXCL13 cut-off of 271 pg/ml with Euroimmun-Elisa for the diagnosis of acute and untreated NB. Due to its high sensitivity and specificity, CXCL13 is a strong candidate biomarker for routine NB assessment, especially in clinically unclear cases.

A novel function of STAT3ß in suppressing interferon response improves outcome in acute myeloid leukemia.

Signal transducer and activator of transcription 3 (STAT3) is frequently overexpressed in patients with acute myeloid leukemia (AML). STAT3 exists in two distinct alternatively spliced isoforms, the full-length isoform STAT3α and the C-terminally truncated isoform STAT3ß. While STAT3α is predominantly described as an oncogenic driver, STAT3ß has been suggested to act as a tumor suppressor. To elucidate the role of STAT3ß in AML, we established a mouse model of STAT3ß-deficient, MLL-AF9-driven AML. STAT3ß deficiency significantly shortened survival of leukemic mice confirming its role as a tumor suppressor. Furthermore, RNA sequencing revealed enhanced STAT1 expression and interferon (IFN) signaling upon loss of STAT3ß. Accordingly, STAT3ß-deficient leukemia cells displayed enhanced sensitivity to blockade of IFN signaling through both an IFNAR1 blocking antibody and the JAK1/2 inhibitor Ruxolitinib. Analysis of human AML patient samples confirmed that elevated expression of IFN-inducible genes correlated with poor overall survival and low STAT3ß expression. Together, our data corroborate the tumor suppressive role of STAT3ß in a mouse model in vivo. Moreover, they provide evidence that its tumor suppressive function is linked to repression of the STAT1-mediated IFN response. These findings suggest that the STAT3ß/α mRNA ratio is a significant prognostic marker in AML and holds crucial information for targeted treatment approaches. Patients displaying a low STAT3ß/α mRNA ratio and unfavorable prognosis could benefit from therapeutic interventions directed at STAT1/IFN signaling.

Favorable impact of PD1/PD-L1 antagonists on bone remodeling: an exploratory prospective clinical study and ex vivo validation.

BACKGROUND: Skeletal morbidity in patients with cancer has a major impact on the quality of life, and preserving bone health while improving outcomes is an important goal of modern antitumor treatment strategies. Despite their widespread use in early disease stages, the effects of immune checkpoint inhibitors (ICIs) on the skeleton are still poorly defined. Here, we initiated a comprehensive investigation of the impact of ICIs on bone health by longitudinal assessment of bone turnover markers in patients with cancer and by validation in a novel bioengineered 3D model of bone remodeling. METHODS: An exploratory longitudinal study was conducted to assess serum markers of bone resorption (C-terminal telopeptide, CTX) and formation (procollagen type I N-terminal propeptide, PINP, and osteocalcin, OCN) before each ICI application (programmed cell death 1 (PD1) inhibitor or programmed death-ligand 1 (PD-L1) inhibitor) for 6 months or until disease progression in patients with advanced cancer and no evidence of bone metastases. To validate the in vivo results, we evaluated osteoclast (OC) and osteoblast (OB) differentiation on treatment with ICIs. In addition, their effect on bone remodeling was assessed by immunohistochemistry, confocal microscopy, and proteomics analysis in a dynamic 3D bone model. RESULTS: During the first month of treatment, CTX levels decreased sharply but transiently. In contrast, we observed a delayed increase of serum levels of PINP and OCN after 4 months of therapy. In vitro, ICIs impaired the maturation of preosteoclasts by inhibiting STAT3/NFATc1 signaling but not JNK, ERK, and AKT while lacking any direct effect on osteogenesis. However, using our bioengineered 3D bone model, which enables the simultaneous differentiation of OB and OC precursor cells, we confirmed the uncoupling of the OC/OB activity on exposure to ICIs by demonstrating impaired OC maturation along with increased OB differentiation. CONCLUSION: Our study indicates that the inhibition of the PD1/PD-L1 signaling axis interferes with bone turnover and may exert a protective effect on bone by indirectly promoting osteogenesis.

Prospective Analysis of Radiation-Induced Contrast Enhancement and Health-Related Quality of Life After Proton Therapy for Central Nervous System and Skull Base Tumors.

PURPOSE: Intracerebral radiation-induced contrast enhancement (RICE) can occur after photon as well as proton beam therapy (PBT). This study evaluated the incidence, characteristics, and risk factors of RICE after PBT delivered to, or in direct proximity to, the brain and its effect on health-related quality of life (HRQoL). METHODS AND MATERIALS: Four hundred twenty-one patients treated with pencil beam scanning PBT between 2017 and 2021 were included. Follow-up included clinical evaluation and contrast-enhanced magnetic resonance imaging at 3, 6, and 12 months after treatment completion and annually thereafter. RICE was graded according to Common Terminology Criteria for Adverse Events version 4, and HRQoL parameters were assessed via European Organisation for Research and Treatment of Cancer Quality of Life Questionnaire (EORTC QLQ)-C30 questionnaires. RESULTS: The median follow-up was 24 months (range, 6-54), and median dose to 1% relative volume of noninvolved central nervous system (D1%CNS) was 54.3 Gy relative biologic effectiveness (RBE; range, 30-76 Gy RBE). The cumulative RICE incidence was 15% (n = 63), of which 10.5% (n = 44) were grade 1, 3.1% (n = 13) were grade 2, and 1.4% (n = 6) were grade 3. No grade 4 or 5 events were observed. Twenty-six of 63 RICE (41.3%) had resolved at the latest follow-up. The median onset after PBT and duration of RICE in patients in whom the lesions resolved were 11.8 and 9.0 months, respectively. On multivariable analysis, D1%CNS > 57.6 Gy RBE, previous in-field radiation, and diabetes mellitus were identified as significant risk factors for RICE development. Previous radiation was the only factor influencing the risk of symptomatic RICE. After PBT, general HRQoL parameters were not compromised. In a matched cohort analysis of 54/50 patients with and without RICE, no differences in global health score or functional and symptom scales were seen. CONCLUSIONS: The overall incidence of clinically relevant RICE after PBT is very low and has no significant negative effect on long-term patient QoL.

Three-Dimensional Light Field Fundus Imaging: Automatic Determination of Diagnostically Relevant Optic Nerve Head Parameters.

Purpose: Morphological changes to the optic nerve head (ONH) can be detected at the early stages of glaucoma. Three-dimensional imaging and analysis may aid in the diagnosis. Light field (LF) fundus cameras can generate three-dimensional (3D) images of optic disc topography from a single shot and are less susceptible to motion artifacts. Here, we introduce a processing method to determine diagnostically relevant ONH parameters automatically and present the results of a subject study performed to validate this method. Methods: The ONHs of 17 healthy subjects were examined and images were acquired with both an LF fundus camera and by optical coherence tomography (OCT). The LF data were analyzed with a novel algorithm and compared with the results of the OCT study. Depth information was reconstructed, and a model with radial basis functions was used for processing of the 3D point cloud and to provide a finite surface. The peripapillary rising and falling edges were evaluated to determine optic disc and cup contours and finally calculate the parameters. Results: Nine of the 17 subjects exhibited prominent optic cups. The contours and ONH parameters determined by an analysis of LF 3D imaging largely agreed with the data obtained from OCT. The median disc areas, cup areas, and cup depths differed by 0.17 mm², -0.04 mm², and -0.07 mm, respectively. Conclusions: The findings presented here suggest the possibility of using LF data to evaluate the ONH. Translational Relevance: LF data can be used to determine geometric parameters of the ONH and thus may be suitable for future use in glaucoma diagnostics.

Modulation of Human Intraocular Pressure Using a Pneumatic System.

Purpose: To technically validate a novel pneumatically based system and method for modulation of intraocular pressure (IOP) and to test its application in the human eye. Special attention was paid to the applicability of the pneumatically driven balloon, which realizes the modulation of the IOP through its contact with the conjunctiva. Methods: A force sensor as key component of a customized measurement setup was used to check the applied pressure through the balloon. The IOP of 10 healthy subjects (4 female, 6 male, aged 28.8 ± 6.64 years) was modulated and increased linearly to at least 40 mmHg. At this point, the pressure inside the balloon was kept constant for 2 minutes, with IOP measurements taken every 40 seconds using a rebound tonometer. Results: The technical setup led to an IOP decrease of 0.71 mmHg within 2 minutes at an operating point of 40 mmHg. For all subjects, the IOP could be increased up to 42.8 ± 3.6 mmHg, whereby a mean pressure decrease of 2.4 mmHg/min was determined, which seems to be caused mainly by physiological processes. Conclusions: With the new pneumatically based setup, a targeted modulation in terms of level and constancy of the IOP can be realized. Translational Relevance: Additional and, compared with the technique according to Löw, a more precise and more constant methodology for the modulation of the IOP, can significantly simplify the determination of retinal vessel pressures for clinical application. It is suitable for practical questions concerning an enhanced retinal venous pressure.

3D retinal imaging and measurement using light field technology.

SIGNIFICANCE: Light-field fundus photography has the potential to be a new milestone in ophthalmology. Up-to-date publications show only unsatisfactory image quality, preventing the use of depth measurements. We show that good image quality and, consequently, reliable depth measurements are possible, and we investigate the current challenges of this novel technology. AIM: We investigated whether light field (LF) imaging of the retina provides depth information, on which structures the depth is estimated, which illumination wavelength should be used, whether deeper layers are measurable, and what kinds of artifacts occur. APPROACH: The technical setup, a mydriatic fundus camera with an LF imager, and depth estimation were validated by an eye model and in vivo measurements of three healthy subjects and three subjects with suspected glaucoma. Comparisons between subjects and the corresponding optical coherence tomography (OCT) measurements were used for verification of the depth estimation. RESULTS: This LF setup allowed for three-dimensional one-shot imaging and depth estimation of the optic disc with green light. In addition, a linear relationship was found between the depth estimates of the OCT and those of the setup developed here. This result is supported by the eye model study. Deeper layers were not measurable. CONCLUSIONS: If image artifacts can be handled, LF technology has the potential to help diagnose and monitor glaucoma risk at an early stage through a rapid, cost-effective one-shot technology.

Ocular direct current stimulation affects retinal ganglion cells.

Ocular current stimulation (oCS) with weak current intensities (a few mA) has shown positive effects on retinal nerve cells, which indicates that neurodegenerative ocular diseases could be treated with current stimulation of the eye. During oCS, a significant polarity-independent reduction in the characteristic P50 amplitude of a pattern-reversal electroretinogram was found, while no current stimulation effect was found for a full field electroretinogram (ffERG). The ffERG data indicated a trend for a polarity-dependent influence during oCS on the photopic negative response (PhNR) wave, which represents the sum activity of the retinal ganglion cells. Therefore, an ffERG with adjusted parameters for the standardized measurement of the PhNR wave was combined with simultaneous oCS to study the potential effects of direct oCS on cumulative ganglion cell activity. Compared with that measured before oCS, the PhNR amplitude in the cathodal group increased significantly during current stimulation, while in the anodal and sham groups, no effect was visible (α = 0.05, pcathodal = 0.006*). Furthermore, repeated-measures ANOVA revealed a significant difference in PhNR amplitude between the anodal and cathodal groups as well as between the cathodal and sham groups (p* ≤ 0.0167, pcathodal - anodal = 0.002*, pcathodal - sham = 0.011*).

Automatic segmentation of skin cells in multiphoton data using multi-stage merging.

We propose a novel automatic segmentation algorithm that separates the components of human skin cells from the rest of the tissue in fluorescence data of three-dimensional scans using non-invasive multiphoton tomography. The algorithm encompasses a multi-stage merging on preprocessed superpixel images to ensure independence from a single empirical global threshold. This leads to a high robustness of the segmentation considering the depth-dependent data characteristics, which include variable contrasts and cell sizes. The subsequent classification of cell cytoplasm and nuclei are based on a cell model described by a set of four features. Two novel features, a relationship between outer cell and inner nucleus (OCIN) and a stability index, were derived. The OCIN feature describes the topology of the model, while the stability index indicates segment quality in the multi-stage merging process. These two new features, combined with the local gradient magnitude and compactness, are used for the model-based fuzzy evaluation of the cell segments. We exemplify our approach on an image stack with 200 × 200 × 100  µm3, including the skin layers of the stratum spinosum and the stratum basale of a healthy volunteer. Our image processing pipeline contributes to the fully automated classification of human skin cells in multiphoton data and provides a basis for the detection of skin cancer using non-invasive optical biopsy.

Effects of Ocular Direct Current Stimulation on Full Field Electroretinogram.

Studies on weak current stimulation (1-2 mA) examine effects on neuronal cells for the treatment of neurological diseases, like depression. Ocular current stimulation showed positive effects on retinal nerve cells which indicate that neurodegenerative ocular diseases, e.g., glaucoma, can be treated with current stimulation of the eye. However, up to now it remains unclear which exact retinal cells can be influenced. During an ocular direct current stimulation, a significant reduction of the characteristic P50 amplitude of a pattern-reversal electroretinogram (PERG) was found for an anodal and a cathodal stimulation. This current stimulation effect could originate from the modulation of pre-ganglion cell activity or by changes in local ON and OFF responses of ganglion cells. For clarification, we investigate acute direct current stimulation effects on a full field electroretinogram (ERG), which represents the activity of pre-ganglion cells (specifically cones and bipolar cells). The ERG from 15 subjects was evaluated before (ERG 1) and during (ERG 2) an ocular direct current stimulation with 800 µA over 5 min. The current was applied through a ring rubber electrode placed around the eye and a 25 cm2 rubber electrode placed at the ipsilateral temple. For ERG measurements, sintered Ag/AgCl skin-electrodes were positioned on the lower eyelid (active), the earlobe (reference), and the forehead (ground). The volunteers were stimulated in three independent sessions, each with a different current application (randomized order): cathodal polarity, anodal polarity (referred to the electrode around the eye), or sham stimulation. The changes between the two ERG measurements of the characteristic full field ERG amplitudes, a-wave, b-wave, and b'-wave (b-wave measured from zero line) were tested with the Wilcoxon signed-rank test (α = 0.05). Comparing before to during the current stimulation for all applications, the ERG waves showed no effects on amplitudes or latencies. Furthermore, no significant difference between the cathodal, anodal, and sham stimulation could be found by a Friedman test. These results indicate an unlikely contribution of pre-ganglion cells to the previously reported stimulation effect on PERG signals.

The Influence of the Stimulus Design on the Harmonic Components of the Steady-State Visual Evoked Potential.

Steady-state visual evoked potentials (ssVEPs) are commonly used for functional objective diagnostics. In general, the main response at the stimulation frequency is used. However, some studies reported the main response at the second harmonic of the stimulation frequency. The aim of our study was to analyze the influence of the stimulus design on the harmonic components of ssVEPs. We studied 22 subjects (8 males, mean age ± SD = 27 ± 4.8 years) using a circular layout (r 1 = 0-1.6°, r 2 = 1.6-3.5°, r 3 = 3.5-6.4°, r 4 = 6.4-10.9°, and r 5 = 10.9-18°). At a given eccentricity, the stimulus was presented according to a 7.5 Hz square wave with 50% duty cycle. To analyze the influence of the stimulus eccentricity, a background luminance of 30 cd/m2 was added to suppress foveal stray light effects; to analyze the influence of simultaneous foveal and peripheral stimulations, stimulations are performed without stray light suppression. For statistical analysis, medians M of the amplitude ratios for amplitudes at the second harmonic to the first harmonic and the probability of the occurrence of the main response at the second harmonic P(MCSH) are calculated. For stimulations with foveal stray light suppression, the medians were M 0 -1.6^ ∘ = 0.45, M 1.6 -3.5^ ∘ = 0.45, M 3.5 -6.4^ ∘ = 0.76, M 6.4 -10.9^ ∘ = 0.72, and M 10.9 -18^ ∘ = 0.48, and the probabilities were P 0-1.6^ ∘ (MCSH) = 0.05, P 1.6 -3.5^ ∘ (MCSH) = 0.05, P 3.5 -6.4^ ∘ (MCSH) = 0.32, P 6.4 -10.9^ ∘ (MCSH) = 0.29, and P 10.9 -18^ ∘ (MCSH) = 0.30. For stimulations without foveal stray light suppression, the medians M were M 0 -1.6^ ∘ = 0.29, M 1.6 -3.5^ ∘ = 0.37, M 3.5 -6.4^ ∘ = 0.98, M 6.4 -10.9 ^^ ∘ = 1.08, and M 10.9 -18^ ∘ = 1.24, and the probabilities were P 0-1.6^ ∘ (MCSH) = 0.09, P 1.6 -3.5^ ∘ (MCSH) = 0.05, P 3.5 -6.4^ ∘ (MCSH) = 0.50, P 6.4 -10.9^ ∘ (MCSH) = 0.55, and P 10.9 -18^ ∘ (MCSH) = 0.55. In conclusion, the stimulus design has an influence on the harmonic components of ssVEPs. An increase in stimulation eccentricity during extrafoveal stimulation leads to a transition of the main response to the second harmonic. The effect is enhanced by a simultaneous foveal stimulation.

The Effects of an Ocular Direct Electrical Stimulation on Pattern-Reversal Electroretinogram.

Studies on transcranial current stimulation have shown that a direct current stimulation of the occipital cortex can influence the amplitude size of a visual evoked potential (VEP). The current direction (cathodal or anodal) determines whether the VEP amplitudes increase or decrease. The aim of this study was to design a new experimental setup that will enable a simultaneous ocular direct current stimulation and electroretinogram (ERG) recording which will broaden our understanding of current stimulation effects on the visual system. Furthermore, we examined whether a direct current stimulation on the eye has a similar effect on an ERG as on a VEP. The pattern-reversal ERG was measured with sintered Ag/AgCl skin-electrodes, positioned on the lower eyelid (active), the earlobe (reference), and the forehead (ground). Direct current was applied through a ring rubber electrode placed around the eye and a 5 cm × 5 cm rubber electrode placed at the ipsilateral temple with a current strength of 500 µA and a duration time of 5 min. Fifty-seven healthy volunteers were divided into three groups depending on the current direction (cathodal, anodal, and sham stimulation, n = 19 each). One ERG measurement (ERG 1) was performed before and another (ERG 2) during the direct current stimulation. The difference between ERG 1 and ERG 2 measurements for the characteristic P50, N95 and N95' (N95 minimum measured from zero line) amplitudes were evaluated by both confidence interval analysis and t-test for related samples (α = 0.05, after Bonferroni correction p ∗ = 0.0055). The P50 amplitude was significantly decreased for ERG 2 measurement in the cathodal and anodal stimulation group (cathodal p = 0.001, anodal p = 0.000). No significant changes could be found in the N95 and N95' amplitudes as well as in the sham-stimulation group. Additionally, the latencies did not undergo any significant changes. In conclusion, the newly designed experimental setup enables simultaneous current stimulation and ERG recording. The current influenced P50 amplitude although not the N95 and N95' amplitudes. Furthermore, the amplitude size decreased for both current directions and did not lead to contrary effects as expected.

Corrigendum: Pulsed Electrical Stimulation of the Human Eye Enhances Retinal Vessel Reaction to Flickering Light.

[This corrects the article DOI: 10.3389/fnhum.2019.00371.].

Corrigendum: The Influence of the Stimulus Design on the Harmonic Components of the Steady-State Visual Evoked Potential.

[This corrects the article DOI: 10.3389/fnhum.2020.00343.].

Pulsed Electrical Stimulation of the Human Eye Enhances Retinal Vessel Reaction to Flickering Light.

Recent studies indicate therapeutic benefits of electrical stimulation in cases of specific ophthalmic diseases that are associated with dysfunctional ocular microcirculation. This suggests effects of electrical stimulation on vascular functions. In the present study, we investigated the effects of electrical stimulation on retinal vessel reactions using dynamic vessel analysis (DVA). Eighty healthy subjects were randomly assigned to one of three groups receiving electrical stimulation with different current intensities: 400 µA (n = 26); 800 µA (n = 27); 1200 µA (n = 27). The electrode montage for electrical stimulation consisted of a ring-shaped active electrode surrounding one eye and a square return electrode at the occiput. Rectangular, monophasic, positive current pulses were applied at 10 Hz for a duration of 60 s per stimulation period. DVA was used to observe the stimulation-induced reactions of retinal vessel diameters in response to different provocations. In three DVA measurements, three stimulus conditions were investigated: flicker light stimulation (FLS); electrical stimulation (ES); simultaneous electrical and flicker light stimulation (ES+FLS). Retinal vasodilation caused by these stimuli was compared using paired t-test. The subjects receiving electrical stimulation with 800 µA showed significantly increased retinal vasodilation for ES+FLS compared to FLS (p < 0.05). No significant differences in retinal vessel reactions were found between ES+FLS and FLS in the 400 and 1200 µA groups. No retinal vasodilation was observed for ES for all investigated current intensities. The results indicate that positive pulsed electrical stimulation of an adequate intensity enhances the flicker light-induced retinal vasodilation.

Objective measurement of forward-scattered light in the human eye: An electrophysiological approach.

PURPOSE: Psychophysical measurements are used to examine the perception of ocular stray light, for example, with C-Quant. These measurements are subjective due to their principles. This work aims to determine ocular stray light objectively; thus, a psychophysical method is transferred into an electrophysiological setup. METHODS: Stray light perception was measured using steady-state visual evoked potentials (VEPs) in 10 healthy subjects (7 males, 3 females, mean age ± SD: 29.6 ± 4.1 years). Stray light emulating filters (Tiffen Black Pro Mist 2) were used for simulating the effect of cataracts to validate the results for increased scattered light conditions. Based on the direct compensation method, the stimulus consisted of a central test field (radius = 2°) with a luminance adjustable compensation light and surrounding ring-shaped stray light source (radius = 5 to 10°). Both flickered in the counter phase at a frequency of 7.5 Hz. The stimuli were presented for 15 luminance levels of the compensation light. The recorded steady-state VEPs at Oz channel were transformed by means of Fourier analysis. The magnitudes at the evoked frequency were plotted against the measured brightness levels of the compensation light. By fitting two linear functions to the resulting data points, a robust minimum log(Leq) was determined, which was correlated with the amount of stray light perception. We measured the stray light parameter log(sc) using C-Quant. For comparison, our results were converted into the C-Quant equivalent parameter log(sepm) and paired t-tests were performed for normal distributed results. RESULTS: A significant difference is observed between log(sepm) (without filter) and log(sepm) (with BPM 2 Filter) (p>0.05). No significant difference is observed between log(sepm) (without filter) and log(sc) (without filter) (p > 0.05) and between log(sepm) (with BPM 2 filter) and log(sc) (with BPM 2 filter) (p > 0.05). CONCLUSION: The electrophysiological approach offers the ability to measure stray light perception in an objective manner.

Bleaching effects and fluorescence lifetime imaging ophthalmoscopy.

This study investigates the influence of photopigment bleaching on autofluorescence lifetimes in the fundus in 21 young healthy volunteers. Three measurements of 30° retinal fields in two spectral channels (SSC: 498-560 nm, LSC: 560-720 nm) were obtained for each volunteer using fluorescence lifetime imaging ophthalmoscopy (FLIO). After dark-adaptation by wearing a custom-made lightproof mask for 30 minutes, the first FLIO-measurement was recorded (dark-adapted state). Subsequently, the eye was bleached for 1 minute (luminance: 3200 cd/m2), followed by a second FLIO-measurement (bleached state). Following an additional 10 minute dark adaptation using the mask, a final FLIO-measurement was recorded (recovered state). Average values of the fluorescence lifetimes were calculated from within different areas of a standardized early treatment diabetic retinopathy study (ETDRS) grid (central area, inner and outer rings). The acquisition time in the bleached state was significantly shortened by approximately 20%. The SSC did not show any significant changes in fluorescence lifetimes with photopigment bleaching, only the LSC showed small but significant bleaching-related changes in the fluorescence lifetimes τ1 and τ2 from all regions, as well as the mean fluorescence lifetime in the central area. The fluorescence lifetime differences caused by bleaching were by far less significant than pathological changes caused by eye diseases. The magnitudes of fluorescence lifetime changes are <10% and do not interfere with healthy or disease related FLIO patterns. Thus, we conclude that bleaching is not a relevant confounder in current clinical applications of FLIO.

Scotoma Simulation in Healthy Subjects.

SIGNIFICANCE: This article shows a successful concept for simulating central scotoma, which is associated with age-related macular degeneration (AMD), in healthy subjects by an induced dark spot at the retina using occlusive contact lenses. The new concept includes a control mechanism to adjust the scotoma size through controlling pupil size without medication. Therefore, a miniaturized full-field adaptation device was used. PURPOSE: The aim of this study was to design a novel concept to simulate AMD scotoma in healthy subjects using occlusive contact lenses. METHODS: To define an optimal set of lens parameters, we constructed an optical model and considered both the anatomical pupil diameter and the opaque central zone diameter of the contact lens. To adjust the scotoma size, we built a miniaturized full-field adaptation device. We demonstrate the validity of this novel concept by functional measurements of visual fields using automated threshold perimetry. Finally, we conducted a perception study including two tasks, consisting of pictograms and letters. The stimuli were presented at different eccentricities and magnifications. RESULTS: The visual fields of all 10 volunteers exhibited absolute scotomas. The loss of contrast sensitivity ranged within 27 and 36 dB (P < .05), and the scotoma localizations were nearly centered to the macula (mean variation, 2.0 ± 4.8° horizontally; 3.5 ± 4.7° vertically). The eccentric perception of letters showed the largest numbers of correctly identified stimuli. The perception of pictograms showed significantly reduced numbers (P < .0001) and revealed a dependency on magnification. The results suggest that best perception is possible for magnified stimuli near the scotoma. CONCLUSIONS: We demonstrated that the creation of an absolute simulated AMD scotoma is possible using occlusive contact lenses combined with a miniaturized full-field adaptation device.