Susanne Jentsch

Retinal venous oxygen saturation increases by flicker light stimulation.

PURPOSE Luminance flicker stimulation of the photoreceptors is known to increase retinal blood flow. Elevated blood velocity was determined using laser Doppler velocimetry, and increased vascular diameters during flicker were observed by measurements with a retinal vessel analyzer. Oxygen supply may be the target of the regulation of retinal blood flow. Thus, the oxygen saturation (SO(2)) in retinal arterioles and venules was investigated along with their diameters. METHODS Dual-wavelength (548 nm and 610 nm) fundus images were taken in 19 healthy volunteers (mean age, 26 ± 2.5 years) before (baseline) and during luminance flicker stimulation (12.5 Hz; modulation depth, 1:25). Retinal vessel SO(2) (dual-wavelength optical oximetry) and diameters (central retinal arterial and venous equivalents [CRAE and CRVE]) were determined. RESULTS CRAEs and CRVEs of 193 ± 20 μm and 228 ± 20 μm at baseline increased statistically significant to a maximum of 202 ± 19 μm (P < 0.0005) and 242 ± 17 μm (P < 0.0005), respectively, under flicker stimulation. Although the arterial SO(2) remained unchanged at 98%-99%, an increase of the venous saturation from 60% ± 5.7% to 64% ± 5.9% (P < 0.0005) was found. CONCLUSIONS In agreement with earlier investigations, the vessel dilation found here indicates an elevation of retinal blood flow by luminance flicker stimulation. This increase of the flow should meet the enhanced metabolic need of the neural retina under a physiological stimulus. The augmentation of venous oxygenation may indicate a higher capillary oxygen concentration, necessary to provide a sufficient diffusion rate of oxygen from the capillaries to the inner retinal tissue.

Retinal fluorescence lifetime imaging ophthalmoscopy measures depend on the severity of Alzheimer's disease.

To determine alterations in the retina of patients with Alzheimers disease (AD) by the newly developed technique of fluorescence lifetime imaging ophthalmoscopy (FLIO) in a pilot study.

Retinal Vessel Oxygen Saturation under Flicker Light Stimulation in Patients with Nonproliferative Diabetic Retinopathy

PURPOSE We investigated the response of retinal vessel diameters and oxygen saturation to flicker light stimulation of neuronal activity in patients with diabetic retinopathy. METHODS We included 18 patients with nonproliferative diabetic retinopathy (mean age 62.2 ± 8.3 years, diabetes type 1 in 4 patients and type 2 in 14, hemoglobin A1c 7.7 ± 0.9%, duration of diabetes 24.1 ± 9.3 years) and 20 age-matched healthy controls (age 66.7 ± 10.3 years). Dual wavelength (548 and 610 nm) fundus images were taken before and during luminance flicker stimulation (12.5 Hz, modulation depth > 1:25) for 90 seconds. Diameters (central retinal arterial [CRAE] and venous [CRVE] equivalents) and oxygen saturation (SO(2)) were determined, and averaged for all arterioles and venules in an annular area centered at the optic disk. RESULTS Flicker light increased CRAE, CRVE, and venous SO(2) by 0.6 ± 6.6%, 2.7 ± 6.1%, and 2.0 ± 2.4% (P < 0.05), respectively, in the patients as well as 4.7 ± 8.4% (P < 0.05), 8.7 ± 5.2% (P < 0.05), and 4.2 ± 3.5% (P < 0.05), respectively, in the controls. The arterial SO(2) remained unchanged in both groups. The increase of the venous SO2 correlated significantly (P = 0.027) with that of the CRAE. There was a trend (P = 0.06) for lower increase of the venous SO(2) with higher body mass index. CONCLUSIONS Our results support the thesis of an impaired regulation of oxygen supply to the diabetic retina. Whereas in healthy subjects the stimulation of neuronal activity increases the vascular diameters and, subsequently, the oxygen supply, this increase is reduced in diabetic retinopathy. This may hint at the role of endothelial dysfunction in the etiology of the disease.

Fluorescence lifetime imaging ophthalmoscopy in type 2 diabetic patients who have no signs of diabetic retinopathy

Abstract. The time-resolved autofluorescence of the eye is used for the detection of metabolic alteration in diabetic patients who have no signs of diabetic retinopathy. One eye from 37 phakic and 11 pseudophakic patients with type 2 diabetes, and one eye from 25 phakic and 23 pseudophakic healthy subjects were included in the study. After a three-exponential fit of the decay of autofluorescence, histograms of lifetimes τi, amplitudes αi, and relative contributions Qi were statistically compared between corresponding groups in two spectral channels (490<ch1<560  nm, 560<ch2<700  nm). The change in single fluorophores was estimated by applying the Holm–Bonferroni method and by calculating differences in the sum histograms of lifetimes. Median and mean of the histograms of τ2, τ3, and α3 in ch1 show the greatest differences between phakic diabetic patients and age-matched controls (p<0.000004). The lack of pixels with a τ2 of ∼360  ps, the increased number of pixels with τ2>450  ps, and the shift of τ3 from ∼3000 to 3700 ps in ch1 of diabetic patients when compared with healthy subjects indicate an increased production of free flavin adenine dinucleotide, accumulation of advanced glycation end products (AGE), and, probably, a change from free to protein-bound reduced nicotinamide adenine dinucleotide at the fundus. AGE also accumulated in the crystalline lens.

Simple and objective method for routine detection of the macular pigment xanthophyll

A new simple method for two-dimensional determination of optical density of macular pigment xanthophyll (ODx) in clinical routine is based on a single blue-reflection fundus image. Individual different vignetting is corrected by a shading function. For its construction, nodes are automatically found in structureless image regions. The influence of stray light in elderly crystalline lenses is compensated by a correction function that depends on age. The reproducibility of parameters in a one-wavelength reflection method determined for three subjects (47, 61, and 78 years old) was: maxODx = 6.3%, meanODx = 4.6%, volume = 6%, and area = 6% already before stray-light correction. ODx was comparable in pseudophakic and in an eye with a crystalline lens of the same 11 subjects after stray-light correction. Significant correlation in ODx was found between the one-wavelength reflection method and the two-wavelength autofluorescence method for pseudophakic and cataract eyes of 19 patients suffering from dry age-related macular degeneration (AMD) (R(2) = 0.855). In pseudophakic eyes, maxODx was significantly lower for dry AMD (n = 45) (ODx = 0.491±0.102 ODU) than in eyes with healthy fundus (n = 22) (ODx = 0.615±0.103 ODU) (p = 0.000033). Also in eyes with crystalline lens, maxODx was lower in AMD (n = 125) (ODx = 0.610±0.093 ODU) than in healthy subjects (n = 45) (ODx = 0.674±0.098 ODU) (p = 0.00019). No dependence on age was found in the pseudophakic eyes both of healthy subjects and AMD patients.

Macular Xanthophylls and ω-3 Long-Chain Polyunsaturated Fatty Acids in Age-Related Macular Degeneration: A Randomized Trial

IMPORTANCE It has been shown that the functionality of the macula lutea depends on the nutritional uptake of lutein and zeaxanthin and that it is inversely associated with the risk of age-related macular degeneration (AMD). Additionally, ω-3 long-chain polyunsaturated fatty acids (LC-PUFAs) may also be protective. OBJECTIVE To investigate the effect of a 12-month intervention with macular xanthophylls and ω-3 LC-PUFAs on xanthophylls and fatty acids in plasma, antioxidant capacity, and optical density of the macular pigment of patients with nonexudative AMD. DESIGN The LUTEGA study was a randomized, double-blind, placebo-controlled, parallel clinical trial that was conducted for 12 months. SETTING University Eye Hospital and Institute of Nutrition, Friedrich Schiller University Jena, Germany. PARTICIPANTS A total of 172 individuals with nonexudative AMD. INTERVENTION Individuals were enrolled and randomly divided as follows: placebo group, group 1 (a capsule containing 10 mg of lutein, 1 mg of zeaxanthin, 100 mg of docosahexaenoic acid, and 30 mg of eicosapentaenoic acid administered each day), and group 2 (same substances but twice the dose used in group 1). One hundred forty-five participants completed the study successfully. MAIN OUTCOME MEASURES Plasma xanthophyll concentrations and fatty acid profiles, optical density of the macular pigment, and antioxidant capacity in plasma (6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid [Trolox] equivalent antioxidant capacity and photochemiluminescence). RESULTS The concentrations of the administered carotenoids in plasma as well as the optical density of the macular pigment increased significantly in the groups randomized to receive supplementary macular xanthophylls and ω-3 LC-PUFAs after 1 month of intervention and remained at this level through the end of the study. Use of the double dose resulted in a beneficial alteration of the fatty acid profile in the plasma of patients with AMD in comparison with the dose in group 1. The lipophilic antioxidant capacity in plasma was significantly elevated with the intervention. CONCLUSIONS AND RELEVANCE A supplement containing a fixed combination of lutein, zeaxanthin, and ω-3 LC-PUFAs during 12 months significantly improved plasma antioxidant capacity, circulating macular xanthophyll levels, and the optical density of the macular pigment. TRIAL REGISTRATION clinicaltrials.gov Identifier: NCT00763659.

Vergleich von Parametern der zeitaufgelösten Autofluoreszenz bei Gesunden und Patienten mit früher AMD

BACKGROUND A fluorescence lifetime mapper (FLM) was tested for quantitative estimation of early alterations in age-related macular degeneration (AMD) which are assumed to be in cellular metabolism. METHOD In FLM time-resolved autofluorescence of the fundus is excited by picosecond (ps) laser impulses at 448 nm and detected in 2 spectral ranges (K1=490-560 nm and K2=560-700 nm) by time-correlated single photon counting. The time-dependent decrease in fluorescence intensity was approximated using 3 decay rates. The calculated lifetimes allow a comparison with endogenous fluorophores of cellular metabolism. RESULTS Initially mean lifetimes were determined for 8 healthy subjects (K1: tau1=118 ps, tau2=584 ps, tau3=2826 ps, K2: tau1=104 ps, tau2=477 ps, tau3=1623 ps). In 15 AMD patients (AREDS categories I and II) the lifetimes were longer (K1: tau1=166 ps, tau2=986 ps, tau3=3309 ps, K2: tau1=137 ps, tau2=583 ps, tau3=1924 ps). The best separation between healthy subjects and patients with early AMD was possible by parameters 1 and 2 in the short-wave channel. Fluorophore-specific alterations in the macula could be demonstrated in isolated cases with advanced AMD. CONCLUSION Measurements in the 30 degrees fundus field demonstrated that specific alterations were already present even in early AMD and also outside the macula. These act in the neuronal retina, because parameter tau2 is related to this layer. Increases in the lifetime of parameter tau2 in the short wave channel could at least partially be determined by an increase of protein bound NADH, the content of which increases with reduced cellular respiration.

[Comparison of parameters of time-resolved autofluorescence between healthy subjects and patients suffering from early AMD].

BACKGROUND A fluorescence lifetime mapper (FLM) was tested for quantitative estimation of early alterations in age-related macular degeneration (AMD) which are assumed to be in cellular metabolism. METHOD In FLM time-resolved autofluorescence of the fundus is excited by picosecond (ps) laser impulses at 448 nm and detected in 2 spectral ranges (K1=490-560 nm and K2=560-700 nm) by time-correlated single photon counting. The time-dependent decrease in fluorescence intensity was approximated using 3 decay rates. The calculated lifetimes allow a comparison with endogenous fluorophores of cellular metabolism. RESULTS Initially mean lifetimes were determined for 8 healthy subjects (K1: tau1=118 ps, tau2=584 ps, tau3=2826 ps, K2: tau1=104 ps, tau2=477 ps, tau3=1623 ps). In 15 AMD patients (AREDS categories I and II) the lifetimes were longer (K1: tau1=166 ps, tau2=986 ps, tau3=3309 ps, K2: tau1=137 ps, tau2=583 ps, tau3=1924 ps). The best separation between healthy subjects and patients with early AMD was possible by parameters 1 and 2 in the short-wave channel. Fluorophore-specific alterations in the macula could be demonstrated in isolated cases with advanced AMD. CONCLUSION Measurements in the 30 degrees fundus field demonstrated that specific alterations were already present even in early AMD and also outside the macula. These act in the neuronal retina, because parameter tau2 is related to this layer. Increases in the lifetime of parameter tau2 in the short wave channel could at least partially be determined by an increase of protein bound NADH, the content of which increases with reduced cellular respiration.

Fluorescence lifetime imaging ophthalmoscopy in glaucoma

Dear Editor: With the aim to detect retinal changes at cellular level, fluorescence lifetime imaging ophthalmoscopy (FLIO) was conducted in primary open-angle glaucoma. In addition to the loss of retinal ganglion cells, metabolic alterations and tissue remodeling are conceivable [1, 2]. Using a modified laserscanning ophthalmoscope, the autofluorescence of different fundus regions was investigated in 43 glaucoma patients (64.9 ±11.4 years) and 54 healthy controls (65.3±11.8 years, p= 0.85). Twenty-five patients and 39 controls were phacic, and 18 patients and 15 controls were pseudophacic. Subjects without serious systemic diseases, diabetes mellitus, or ocular pathologies (except previous cataract surgery) were included. Patients under anti-coagulant therapy with fluorescent coumarin-derivatives were excluded. All investigations were approved by a local institutional review board, and written informed consent was obtained. As the anti-glaucomatous treatment was not interrupted, the intraocular pressure was lower than 22 mmHg in all cases. The FLIO method was described by Schweitzer et al. [3]. Exciting the fundus with light of short wavelengths leads to fluorescence of different substances. Therefore, the measured signal is composed of the sum of several fluorophores. Investigating the fluorescence decay time allows, at least partly, a separation of single fluorophores [3, 4]. Using the fluorescence lifetime imaging ophthalmoscope, the fluorescence decay can be allocated to three components, and thereby described through the lifetime parameters τ1–3 and amplitudes α1–3. For a global characterization of the fluorescence, the amplitude-weightedmean of all decay times, τm, is calculated. Fluorescence was excited with a wavelength of 448 nm. The emission was captured in two spectral channels (Ch1: 490– 560 nm, Ch2: 560–700 nm) and the autofluorescence properties in different fundus regions were analyzed (Fig. 1). Fur-

Method for simultaneous detection of functionality and tomography

During clinical application of the fluorescence lifetime laser scanner ophthalmoscope, a stepped slope of ocular autofluorescence was found. Fitting of fluorescence results in wrong lifetimes if the left border of the fitting interval is set at the time channel of first appearance of fluorescence. A better fit was reached at least for the fluorescence decay if the left border of the fitting interval is set near the maximum of detected fluorescence. Analysing the appearance of the stepped slope, its origination was found by the different appearance time of fluorescence from the crystalline lens and from the fundus fluorescence. The extension of the exponential model function by a parameter tci results in an optimal fit of both the slope and of the fluorescence decay. This new parameter describes differences in the appearance time of fluorescence originating from different layers. Taking into account the refractive index between layers, the geometrical distance between them can be determined. In this way, functional information (lifetimes) and geometrical information (distances) can be determined by the same measurement. To reach geometrical resolution comparable with OCT, pulses and time resolution are required in the order of 30 fs.