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OBJECTIVE--We investigated the impact of two different injection strategies on the pharmacokinetics and pharmacodynamics of insulin aspart in vivo in an open-label, two-period crossover study and verified changes in the [...]

Objective: To present a new imaging method for the anterior segment of the eye. Methods: Transscleral optical coherence tomographic images were generated in healthy volunteers using a slitlamp-adapted prototype equipped with a superluminescence diode with an infrared wavelength of 1310 nm. Results: The optical coherence tomographic system used allowed penetration of human sclera in vivo and high-resolution, cross-sectional imaging of the anterior chamber angle and the ciliary body. Conclusion: The 1310-nm optical coherence tomographic image shows a significant potential as a noninvasive diagnostic tool for the anterior segment of the eye. Arch Ophthalmol. 2002;120:816-819

We demonstrate the highest resolution (1.5×1.5×1 µm) micrometer optical coherence tomography (µOCT) imaging of the morphologic micro-structure of excised swine and non-human primate corneas. Besides epithelial, stromal, and endothelial cell morphology, this report focuses on investigating the most peripheral corneal nerve fibers, the nerve fibers of the subbasal plexus (SBP). Alterations of SBP nerve density and composition are reportedly linked to major neurologic disorders, such as diabetic neuropathy, potentially indicating earliest onsets of denervation. Here, the fine, hyperreflective, epithelial nerve structures located just above Bowman’s membrane, are i) visualized using our µOCT prototype, ii) validated by comparison to fluorescence confocal microscopy (including selective immunohistochemical staining), and iii) segmented using state-of-the-art image processing. Here, we also introduce polarization sensitive (PS) µOCT imaging, demonstrating, to the best of our knowledge, the highest resolution corneal PS-OCT scans reported to date.

PURPOSE:To evaluate a new non-ablative and adjustable procedure for laser ablative refractive corneal surgery in hyperopia using the injection of a biocompatible liquid filler material into a stromal pocket.METHODS:A total of 120 stromal pockets were created using a clinical femtosecond laser system in 96 rabbit corneoscleral discs and 24 whole globes. Pockets were cut at a depth of 120 or 250 µm below the epithelial surface. Hyaluronic acid was injected manually into the pocket. To determine the refractive changes, three-dimensional optical coherence tomography images and a specifically developed picture recognition Matlab (The Mathworks) routine were used.RESULTS:After injection, a steepening of the anterior and flattening of the posterior corneal surface was observed, which led to hyperopic correction. The two main factors determining the amount of correction were the pocket depth and the injected volume. After the pocket was homogeneously filled, an initial refractive increase was observed, followed by a linear relation between the injected volume and the refraction increase.CONCLUSIONS:This possible clinical protocol for controlled refraction correction of hyperopia suggests a potential readjustable clinical application.[[J Refract Surg. 2020;36(6):406–414.]

Eicosanoids are lipid-mediator molecules with key roles in inflammatory skin diseases, such as psoriasis. Eicosanoids are released close to the source of inflammation, where they elicit local pleiotropic effects and dysregulations. Monitoring inflammatory mediators directly in skin lesions could provide new insights and therapeutic possibilities. Here, we analyzed dermal interstitial fluid samples obtained by dermal open-flow microperfusion in a rat model of skin inflammation. We developed a solid-phase extraction ultra-HPLC/MS/MS method to reliably and precisely analyze small-volume samples and quantified 11 eicosanoids [thromboxane B2, prostaglandin (PG) E2, PGD2, PGF2α, leukotriene B4, 15-HETE, 12-HETE, 5-HETE, 12-hydroxyeicosapentaenoic acid, 13-HODE, and 17-hydroxydocosahexaenoic acid]. Our method achieved a median intraday precision of approximately 5% and interday precision of approximately 8%. All calibration curves showed excellent linearity between 0.01 and 50 ng/ml (R2> 0.980). In the rat model, eicosanoids were significantly increased in imiquimod-treated inflamed skin sites compared with untreated control sites. Oral treatment with an anti-inflammatory glucocorticoid decreased eicosanoid concentrations. These results show that a combination of tissue-specific sampling with LC/MS analytics is well suited for analyzing small sample volumes from minimally invasive sampling methods such as open-flow microperfusion or microdialysis to study local inflammation and the effect of treatments in skin diseases.

Objective:Ischaemia is one of the biggest problems in wound healing. It causes chronic wounds and also prevents normal wound healing because the tissue is oxygen deprived. Most oxygen-supplying therapies are only feasible in a clinical setting, but topical haemoglobin applications, such as Granulox, can be used in a non-clinical setting. For home application, the haemoglobin solution is sprayed topically onto the wound using a pressurised ready-to-use device with a bag-on-valve system. Although this system does not mix product and propellant, the risk of product inhalation by the patient, user or bystanders has to be minimised. This safety study aimed to determine particle size and product concentration in the surroundings after application to determine if there is a risk that product particles enter the respiratory tract.Methods:Measurements were performed using a laser scattered light photometer and a Scanning Mobility Particle Sizer (SMPS)-Spectrometer at different distances from the measuring devices to determine the inhalation risk for a possible user, patient and bystander. At all measuring points the amount of particles, their size and the formation of dust were measured.Results:No nanoparticles or dust were created during the application of the haemoglobin spray. The concentrations of the measured particles are below the allowed limits defined by Austrian law.Conclusion:There is no risk of inhaling nanoparticles or being exposed to harmful concentrations of larger particles of the tested product. All the product's ingredients can be degraded and excreted by the human body through natural pathways.

Dupilumab is a monoclonal antibody approved for the treatment of atopic dermatitis (AD); however, its effects on molecular, cellular, and immunological levels remain to be elucidated. In this study, blood and dermal interstitial fluid (ISF) from nonlesional (NL) and lesional (L) skin were collected from eight patients with moderate to severe AD, before (visit 2-v2) and at the end of a 16-week treatment with dupilumab (visit 10-v10). Clinical treatment effect was demonstrated by significantly decreased AD severity scores at the end of treatment. At v10 versus v2, the percentages of CD4+ interleukin-producing cells showed a decreasing trend in ISF L and NL, unbound IL-4 levels in plasma were increased, IL-5 levels in ISF L reduced, and levels of factors involved in anti-inflammatory pathways and re-epithelization increased. At v2, ISF L showed that AD lesions might have altered amino acid pathways and lipid signaling compared to ISF NL. At v10, ISF L exhibited raised levels of long- and very-long-chain fatty acids and lipids compared to v2. Furthermore, dupilumab administration caused reduced expression of miR-155–5p and miR-378a-3p in ISF L. In conclusion, results from the present study provided novel knowledge by linking local immune and metabolic alterations to AD pathogenesis and treatment response.

Background:The GLP-1 receptor agonist liraglutide is marketed for obesity treatment where it induces body weight reduction possibly via the hypothalamus, which regulates energy homeostasis. In animal studies, acute liraglutide treatment triggers satiety, weight loss and activates thermogenesis in adipose tissue. However, the precise mechanisms how liraglutide affects in particular chronic weight loss are still under investigation.Objectives:We aimed to evaluate whether chronic hypothalamic or chronic subcutaneous administration of liraglutide induces sustained weight loss through altered adipose tissue function and to what extent hypothalamic neuronal appetite regulators are involved in the liraglutide-induced weight loss in healthy lean rats on a normal diet.Materials/Methods:We continuously administered liraglutide either intrahypothalamically (10 μg per day) or subcutaneously (200 μg kg−1per day) for 28 days to lean Sprague Dawley rats (n=8 each). We assessed changes in body weight, adipose tissue mass, adipocyte size and adipose tissue volume in the abdominal region by using micro-CT. We analyzed genetic expression patterns of browning, thermogenic and adipocyte differentiation regulators in adipose tissues as well as particular neuronal appetite regulators in the hypothalamus.Results:Intrahypothalamic liraglutide administration induced an 8% body weight reduction at day 9 compared with the control group (P<0.01) and a 7% body weight loss at day 9 compared with subcutaneous liraglutide treatment (P<0.01), supported by a significant reduction in adipose tissue mass and volume with intrahypothalamic liraglutide administration (P<0.05). Our data show that chronic intrahypothalamic liraglutide treatment triggered an 18-fold induction of the hypothalamic mc4r gene (P<0.01) accompanied by a significant increase in circulating thyroxine (T4) levels (P<0.05).Conclusions:Chronic intrahypothalamic liraglutide administration resulted in a profound reduction in body weight and fat mass loss most likely mediated by the hypothalamic melanocortin system rather than by adipose tissue browning or improved thermogenesis.

With a simple setup, mainly composed of a low coherence light source and a camera, full-field optical coherence tomography (FF-OCT) allows volumetric tissue imaging. However, fringe washout constrains its use in retinal imaging. Here, we present a novel motion-insensitive approach to FF-OCT, which introduces path-length differences between the reference and the sample light in neighboring pixels using an off-axis reference beam. The temporal carrier frequency in scanned time-domain OCT is replaced by a spatial carrier frequency. Volumetric in-vivo FF-OCT measurements of the human retina were acquired in only 1.3 s, comparable to the acquisition times of current clinically used OCT devices.

Frequency‐doubled diode laser technology (FDDL) enables direct modulation of 561 nm lasers with unprecedented speeds. It also reaches an ultimate high on/off contrast with zero photons in the off state, which is favorable for laser scanning microscopes and multicolor microscopy techniques.

The transfer of academic innovation to clinical practice and industry is taking on an increasingly important role in academic centers, but significant challenges remain. This work identifies some of the pivotal elements that are required to assemble, manage and accelerate the pace of successful translational research. These elements can be incorporated in new models of translational research facilitation that improve the process of converting academic innovations to commercial products where they can be used to benefit patient health on a large scale.

The selective inhibition of intracellular and nuclear molecules such as Ki-67 holds great promise for the treatment of cancer and other diseases. However, the choice of the target protein and the intracellular delivery of the functional agent remain crucial challenges. Main hurdles are (a) an effective delivery into cells, (b) endosomal escape of the delivered agents, and (c) an effective, externally triggered destruction of cells. Here we show a light-controlled two-step approach for selective cellular delivery and cell elimination of proliferating cells. Three different cell-penetrating nano constructs, including liposomes, conjugates with the nuclear localization sequence (NLS), and conjugates with the cell penetrating peptide Pep-1, delivered the light activatable antibody conjugate TuBB-9-FITC, which targets the proliferation associated protein Ki-67. HeLa cells were treated with the photosensitizer benzoporphyrin monoacid derivative (BPD) and the antibody constructs. In the first optically controlled step, activation of BPD at 690 nm triggered a controlled endosomal escape of the TuBB-9-FITC constructs. In more than 75% of Ki-67 positive, irradiated cells TuBB-9-FITC antibodies relocated within 24 h from cytoplasmic organelles to the cell nucleus and bound to Ki-67. After a second light irradiation at 490 nm, which activated FITC, cell viability decreased to approximately 13%. Our study shows an effective targeting strategy, which uses light-controlled endosomal escape and the light inactivation of Ki-67 for cell elimination. The fact that liposomal or peptide-assisted delivery give similar results leads to the additional conclusion that an effective mechanism for endosomal escape leaves greater variability for the choice of the delivery agent.

Oxygen plays an important role in wound healing, as it is essential to biological functions such as cell proliferation, immune responses and collagen synthesis. Poor oxygenation is directly associated with the development of chronic ischemic wounds, which affect more than 6 million people each year in the United States alone at an estimated cost of $25 billion. Knowledge of oxygenation status is also important in the management of burns and skin grafts, as well as in a wide range of skin conditions. Despite the importance of the clinical determination of tissue oxygenation, there is a lack of rapid, user-friendly and quantitative diagnostic tools that allow for non-disruptive, continuous monitoring of oxygen content across large areas of skin and wounds to guide care and therapeutic decisions. In this work, we describe a sensitive, colorimetric, oxygen-sensing paint-on bandage for two-dimensional mapping of tissue oxygenation in skin, burns, and skin grafts. By embedding both an oxygen-sensing porphyrin-dendrimer phosphor and a reference dye in a liquid bandage matrix, we have created a liquid bandage that can be painted onto the skin surface and dries into a thin film that adheres tightly to the skin or wound topology. When captured by a camera-based imaging device, the oxygen-dependent phosphorescence emission of the bandage can be used to quantify and map both the pO_2 and oxygen consumption of the underlying tissue. In this proof-of-principle study, we first demonstrate our system on a rat ischemic limb model to show its capabilities in sensing tissue ischemia. It is then tested on both ex vivo and in vivo porcine burn models to monitor the progression of burn injuries. Lastly, the bandage is applied to an in vivo porcine graft model for monitoring the integration of full- and partial-thickness skin grafts.

Visualizing retinal photocoagulation by real-time OCT measurements may considerably improve the understanding of thermally induced tissue changes and might enable a better reproducibility of the ocular laser treatment. High speed Doppler OCT with 860 frames per second imaged tissue changes in the fundus of enucleated porcine eyes during laser irradiation. Tissue motion, measured by Doppler OCT with nanometer resolution, was correlated with the temperature increase, which was measured non-invasively by optoacoustics. In enucleated eyes, the increase of the OCT signal near the retinal pigment epithelium (RPE) corresponded well to the macroscopically visible whitening of the tissue. At low irradiance, Doppler OCT revealed additionally a reversible thermal expansion of the retina. At higher irradiance additional movement due to irreversible tissue changes was observed. Measurements of the tissue expansion were also possible in vivo in a rabbit with submicrometer resolution when global tissue motion was compensated. Doppler OCT may be used for spatially resolved measurements of retinal temperature increases and thermally induced tissue changes. It can play an important role in understanding the mechanisms of photocoagulation and, eventually, lead to new strategies for retinal laser treatments.

Phase sensitive OCT enables the measurement of thermal expansion in laser irradiated material at high lateral and temporal resolution. In principle, a calculation of the 3D temperature distribution and its temporal evolution should be possible by evaluating the local expansion. This could be utilized for a non-invasive and very fast temperature measurement, e.g. to realize an online dosimetry for photocoagulation. The possibilities of quantitative investigations at high axial and lateral resolution are demonstrated by imaging the reversible thermal expansion in laser irradiated multilayer silicone phantoms.

Retinal photocoagulation is an established treatment for various retinal diseases. The temperature development during a treatment can be monitored by applying short laser pulses in addition to the treatment laser light. The laser pulses induce optoacoustic pressure waves that can be detected at the cornea. Aim of this work is the investigation of the accuracy of the determined temperatures during a treatment. To calibrate the temperature dependency of the measured pressure, whole enucleated porcine eyes were heated using an infrared laser beam, while probing the retina optoacoustically. The temperatures and the optoacoustic pressure waves were measured simultaneously using thermocouples and a piezoelectric element, respectively. From the deviation of the individual measurements an error of less than 15% in the calibration regime between 37 °C to 55 °C was found. Furthermore, the spatial and temporal temperature course was investigated. Calculations were performed to simulate the temporal and spatial temperature development during photocoagulation. A theoretical model to determine the peak temperature of the irradiated tissue from the mean temperature measured by optoacoustics was developed. The validity of the model was experimentally examined by heating the retina of porcine eyes with a laser beam diameter of 500 m while successively measuring the temperature optoacoustically with a probe beam diameter of 500 m and 100 m at the center of the heated area, respectively. The deviation of the theoretical model and the experimental results were found to be less than 7%.

Considerable improvement in the reproducibility of retinal photocoagulation is expected if degree and extend of the heat-induced tissue damage can be visualized on-line during the treatment. Experimental laser treatments of the retina with enucleated pig eyes were investigated by high speed phase-sensitive OCT. OCT could visualize the increase of tissue scattering during the photocoagulation in a time-resolved way. Immediate and late tissue changes were visualized with more than 15 µm resolution. Changes of the reflectance in the OCT images had a similar sensitivity in detecting tissue changes than macroscopic imaging. By using Doppler OCT slight movements of the tissue in the irradiated spot were detected. At low irradiance the thermal expansion of the tissue is observed. At higher irradiance irreversible tissue changes dominate the tissue expansion. OCT may play an important role in understanding the mechanisms of photocoagulation. This may lead to new treatment strategies. First experiments with rabbits demonstrate the feasibility of in-vivo measurements.

Retinal laser photocoagulation is an established treatment method for many retinal diseases like macula edema or diabetic retinopathy. The selection of the laser parameters is so far based on post treatment evaluation of the lesion size and strength. Due to local pigment variations in the fundus and individual transmission the same laser parameters often lead to an overtreatment. Optoacoustic allows a non invasive monitoring of the retinal temperature increase during retinal laser irradiation by measuring the temperature dependent pressure amplitudes, which are induced by short probe laser pulses. A 75 ns/ 523 nm Nd:YLF was used as a probe laser at a repetition rate of 1 kHz, and a cw / 532 nm treatment laser for heating. A contact lens was modified with a ring-shaped ultrasonic transducer to detect the pressure waves at the cornea. Temperatures were collected for irradiations leading to soft or invisible lesions. Based on this data the threshold for denaturation was found. By analyzing the initial temperature increase, the further temperature development during irradiation could be predicted. An algorithm was found to calculate the irradiation time, which is needed for a soft lesion formation, from the temperature curve. By this it was possible to provide a real-time dosimetry by automatically switching off the treatment laser after the calculated irradiation time. Automatically controlled coagulations appear softer and more uniformly.

Selective retina treatment (SRT) is a laser based method to treat retinal diseases associated with disorders of the retinal pigment epithelium (RPE) while preserving photoreceptors and choroid. Applying microsecond laser pulses to the 100- 200 strongly absorbing melanin granules inside the RPE cells induces transient micro bubbles which disrupt the cells. Aim of this work is to understand bubble dynamics in clusters with respect to the influence of the adjacent retina. Bubble dynamics were investigated in vitro on porcine RPE. An about 200 m thick layer of agarose gel was applied to the RPE layer in order to simulate the mechanical properties of retina. Different laser pulse durations from 1 ns (532 nm, Nd:YAG) to 1.7 s (527 nm, Nd:YLF) were used. The bubbles were investigated interferometrically (fiber interferometer @ 830 nm) and with fast flash photography (25 ns flash duration). Bubble lifetimes were measured. The results show that with retina phantoms the bubble formation threshold was reached at 2.5 times higher irradiation than without retina phantom for 1.7 s laser pulses. The microbubbles generated with 1 ns laser pulses were almost not influenced by the agarose layer. Irradiation twofold over bubble formation threshold resulted in 3.5 times longer bubble lifetimes for s and 2 times longer for ns pulse durations, respectively.

Retinal photocoagulation is a long time established treatment for a variety of retinal diseases, most commonly applied for diabetic macular edema and diabetic retinopathy. The damage extent of the induced thermal coagulations depend on the temperature increase and the time of irradiation. So far, the induced temperature rise is unknown due to intraocular variations in light transmission and scattering and RPE/choroidal pigmentation, which can vary inter- and intraindividually by more than a factor of four. Thus in clinical practice, often stronger and deeper coagulations are applied than therapeutically needed, which lead to extended retinal damage and strong pain perception. The final goal of this project focuses on a dosimetry control, which automatically generates a desired temperature profile and thus coagulation strength for every individual coagulation spot, ideally unburden the ophthalmologist from any laser settings. In this paper we present the first realtime temperature measurements achieved on patients during retinal photocoagulation by means of an optoacoustic method, making use of the temperature dependence of the thermal expansion coefficient of retinal tissue. Therefore, nanosecond probe laser pulses are repetitively and simultaneously applied with the treatment radiation in order to excite acoustic waves, which are detected at the cornea with an ultrasonic transducer embedded in the contact lens and then are processed by PC.

Abstract: Background: Central serous chorioretinopathy (CSC) is a disease with a localized breakdown of the outer blood–retinal barrier located within the retinal pigment epithelium (RPE) causing subretinal fluid accumulation. Selective retina therapy (SRT) is a new, minimally invasive laser technology that has been designed to selectively target the RPE. SRT spares retinal tissue.Methods: Twenty-seven eyes of 27 patients with active CSC were treated with SRT using a pulsed double-Q-switched Nd-YLF prototype laser (λ=527 nm, t=1.7 μs). At baseline, best-corrected visual acuity was determined and fluorescein angiography and optical coherence tomography were performed. The patients were followed for up to 3 months.Results: After 4 weeks 85.2% of patients showed complete resolution of subretinal fluid and in 96.3% there was no leakage visible on fluorescein angiography. After 3 months 100% of patients demonstrated no subretinal fluid and 100% of patients had no leakage activity on fluorescein angiography. Visual acuity, 20/40 at baseline, improved to 20/28 after 4 weeks and to 20/20 after 3 months.Conclusion: SRT is a safe and effective treatment for active CSC. Especially if the RPE leak is located close to the fovea, SRT is the favoured therapeutic option. We recommend earlier treatment of patients with acute CSC in order to prevent development of chronic changes due to CSC with irreversible anatomical and functional damage. SRT might be considered as a first-line treatment for active CSC.

New diagnostic tools such as the retinal thickness analyzer (RTA), optical coherence tomography (OCT), and topographic angiography (TAG) were introduced into clinical ophthalmology during the last years giving the examiner new insights into anatomical and functional aspects of macular disease. In this study, advantages and disadvantages of the new imaging methods have been evaluated in patients with serous (sPED) and fibrovascular pigment epithelial detachments (fPED) secondary to age-related macular degeneration (AMD).TAG, using fluorescein angiography (FA), provides a three-dimensional profile of the fluorescein pattern based on the analysis of a set of 32 confocal images over a depth of 4 mm. RTA and OCT provide cross-sectional images of the neurosensory retina and the retinal pigment epithelium–choriocapillary complex as well as retinal thickness data encoded in a false color map. We compared and evaluated these modalities in 15 patients with fPED and 15 patients with sPED secondary to AMD.In patients with classic fPED, TAG detected neovascular structures and delineated their configuration. In sPEDs, pooling of extravascular fluid was detected in a dome-shaped configuration. OCT provided detailed information on the neurosensory retina’s structures but failed to detect the neovascular membrane in fPED. Mapping the retinal thickness, RTA and OCT both failed to detect the PED and showed typical algorithm error-based patterns.TAG OCT and RTA are useful imaging modalities in the evaluation of AMD cases. TAG visualizes the vascular configuration, dynamic perfusion, and leakage changes. OCT and RTA are able to complementarily document intra-, subretinal, and sub-RPE fluid accumulation secondary to CNV. However, OCT seems to be more efficient in imaging AMD-related pathologies than RTA, as this modality is often compromised by intra- or subretinal structural abnormalities. Nevertheless, all modalities may provide further valuable insight into AMD pathogenesis, enhance diagnostic quality, and improve the assessment of therapeutic effects.

Current PDT treatment for age-related macular degeneration uses a standard radiant exposure of 50 J/cm2at an irradiance of 600 mW/cm2. However; there is a general problem with the unusually high irradiance; in fact, the rate of photochemical production of singlet oxygen may be limited by insufficiently oxygenized neovascular tissue. It was the aim of this study to evaluate the efficacy of verteporfin (Visudyne) photoactivation to induce thrombosis of choriocapillaries and in experimentally induced corneal neovascularizations in rabbits by varying irradiance and retinal radiant exposure.The light-dose threshold to induce micro-thrombosis in the choriocapillaries (seven eyes) and in corneal neovascularizations (eight eyes) of Chinchilla-Bastard rabbits using different retinal irradiances (100 and 600 mW/cm2) at different radiant exposures (20, 10, 5, 2.5, 1.25, 0.62, and 0.3 J/cm2) was evaluated. Induction of neovascularizations was performed 7 days prior to PDT treatment using intracorneal silk sutures. A dose of 2 mg/kg verteporfin was intravenously infused 10 min before standard PDT. The criterion for vascular thrombosis was vessel closure as determined by fluorescein angiography 1 h and 1 day post exposure.Experiments on the choroid revealed vessel closure 1 h after irradiation at ED50= 10.8 J/cm2(both 600 and 100 mW/cm2) and after 24 h at ED50= 2.4 J/cm2(600 mW/cm2) versus 1.8 J/cm2(100 mW/cm2). Vessel closure was enhanced at irradiation with 100 mW/cm2. Regarding corneal neovascularizations, vessel thrombosis was observable by dark appearance of irradiated clotted neovascular tissue and angiographically by a lack of leakage at ED50thresholds of 0.62 J/cm2(1 h) and 0.41 J/cm2(1 day) for 100 mW/cm2and of 0.99 J/cm2(1 h), and 0.67 J/cm2(1 day) for 600 mW/cm2. Thus in both experiments thresholds for vessel closure were reduced by a factor of 1.5 for the lower intensity. Histology revealed more selective vessel occlusion without RPE and photoreceptor damage for 100 mW/cm2rather than 600 mW/cm2intensity at threshold irradiation.Low-intensity PDT with verteporfin for neovascular tissue seems to be more effective than regular high-intensity PDT. Future preclinical trials should address the issue of proper dosimetry for effective PDT in age-related macular degeneration.

Purpose Topographic angiography (TAG) using confocal scanning laser angiography and optical coherence tomography (OCT) are new imaging modalities that have been introduced during recent years. OCT and TAG imaging were compared to specify the characteristics of each imaging modality.Methods TAG using fluorescein angiography (FA) provides a three-dimensional profile of the vascular structures based on the analysis of a set of 32 confocal images over a depth of 4 mm. OCT provides cross-sectional images of the neurosensory retina and the retinal pigment epithelium-choriocapillary complex (RPE-CC). The authors compared and evaluated both modalities in 10 patients with predominantly classic choroidal neovascularization (CNV), 10 patients with serous pigment epithelial detachment (PED), and 10 patients with geographic RPE atrophy, all secondary to age-related macular degeneration (ARMD).Results In patients with classic CNV, TAG detected neovascular structures and delineated their configuration. In PEDs pooling of extravascular fluid is demonstrated, and in geographic RPE atrophy TAG showed reduced choroidal perfusion. Classic CNV was demonstrated by OCT as a hyperreflective band at the level of the RPE-CC, and PED showed a dome-shaped RPE detachment. In geographic RPE atrophy, OCT imaged loss of the RPE band and had an increased depth resolution.Conclusions TAG and OCT are useful imaging modalities in the evaluation of ARMD cases. TAG visualizes the vascular configuration and dynamic perfusion and leakage changes. OCT is able to document intra-, subretinal, and sub-RPE fluid accumulation secondary to CNV. Both modalities may provide further valuable insight into ARMD pathogenesis, enhance diagnostic quality, and improve the assessment of therapeutic effects.

We investigate the relationship between the laser beam at the retina (spot size) and the extent of retinal injury from single ultrashort laser pulses. From previous studies it is believed that the retinal effect of single 3-ps laser pulses should vary in extent and location, depending on the occurrence of laser-induced breakdown (LIB) at the site of laser delivery. Single 3-ps pulses of 580-nm laser energy are delivered over a range of spot sizes to the retina of Macaca mulatta. The retinal response is captured sequentially with optical coherence tomography (OCT). The in vivoOCT images and the extent of pathology on final microscopic sections of the laser site are compared. With delivery of a laser pulse with peak irradiance greater than that required for LIB, OCT and light micrographs demonstrate inner retinal injury with many intraretinal and/or vitreous hemorrhages. In contrast, broad outer retinal injury with minimal to no choriocapillaris effect is seen after delivery of laser pulses to a larger retinal area (60 to 300 m diam) when peak irradiance is less than that required for LIB. The broader lesions extend into the inner retina when higher energy delivery produces intraretinal injury. Microscopic examination of stained fixed tissues provide better resolution of retinal morphology than OCT. OCT provides less resolution but could be guided over an in vivo, visible retinal lesion for repeated sampling over time during the evolution of the lesion formation. For 3-ps visible wavelength laser pulses, varying the spot size and laser energy directly affects the extent of retinal injury. This again is believed to be partly due to the onset of LIB, as seen in previous studies. Spot-size dependence should be considered when comparing studies of retinal effects or when pursuing a specific retinal effect from ultrashort laser pulses. © 2004 Society of Photo-Optical Instrumentation Engineers.

Background Fundus autofluorescence (AF) is derived from the lipofuscin contained by the retinal pigment epithelial cells. Using a scanning laser ophthalmoscope, two-dimensional AF measurements of the ocular fundus can be achieved. Directly after conventional photocoagulation and also after selective RPE laser treatment (SRT) with ophthalmoscopically non-visible laser lesions, irradiated areas reveal reduced AF, indicating RPE damage. Since the green treatment laser beam could also be used for AF excitation, the aim of this study was to evaluate whether absolute measurements of AF can be performed, and also possible changes in AF detected, online during SRT. Methods SRT was carried out by use of a frequency-doubled Nd:YLF laser (wavelength 527 nm, pulse duration 1.7 µs, repetition rate 500 and 100 Hz, number of pulses 100 and 30, single pulse energy 50–130 µJ) in vitro (porcine RPE; retinal spot size 160 µm) and during patient treatment (retinal spot size 176 µm). During irradiation, fluorescence light from the RPE was decoupled from the laser light inside the slit lamp and detected by a photomultiplier or photodiode at wavelengths above 550 nm. Additionally, temperature-dependent fluorescence intensity measurements of A2-E, the main fluorescent component of lipofuscin, were performed in a different in-vitro setup. Results The intensity of AF decreased over the number of applied pulses during laser irradiation, and this trend was more pronounced in porcine RPE samples than during human treatment. In vitro, the AF intensity decreased by about 22%; however, only a weak signal was detected. When treating patients, the AF intensity was strong and the rate of decay of fluorescence intensity with number of pulses was greater when irradiating at 500 Hz than at the 100 Hz repetition rate. However, for both repetition rates the AF decay was merely up to 6–8% over the number of pulses per laser spot. Fluorescence intensity of A2-E decreased linearly with increasing temperature at about 1% per 1°C and was completely reversible. Conclusions Online measurements of AF during selective RPE laser treatment are possible and reveal a decay in AF as a function of the number of laser pulses applied to the RPE. If A2-E results can be transferred to RPE fluorescence, the AF decay could be related to the temperature increase within the tissue during treatment. Further clinical studies—in SRT as well as in conventional laser photocoagulation—might be able to show online AF changes on different areas of the retina and on different pathologies. Due to the temperature dependence of the fluorescence, on-line AF measurements during laser treatments such as photocoagulation or TTT may be able to be used as a real-time method for temperature monitoring.

Abstract: Three-dimensional topography of perfused vascular structures is possible via confocal laser scanning of intravascular fluorescence. The lateral resolution is given by the spot size of the scanning laser beam (optimally 10 µm at the retina). The axial resolution, however, depends on the accuracy of detection of the surface of the fluorescent structure, which is typically one order of magnitude higher (30 µm at the retina) than the confocal resolution. The vascular structure is stained with an appropriate fluorescent dye prior to the investigation using standard systemic dye injection. Confocal scanning of the fluorescence in planes of different depths within the vascular structure under investigation leads to a three-dimensional data set. Signal processing includes passive eye tracking, lateral averaging and axial determination of the surface of the fluorescent structure. The potential of this new technique is demonstrated by showing the topography of physiological vessel structures as well as of selected vascular diseases such as cone dystrophy, RPE detachment, choroidal haemangioma and retinal laser coagulation. Confocal laser angioscopic fluorescence topography (CLAFT) measures the three- dimensional surface structure of functional (perfused) vasculature and surrounding leakage. CLAFT may help to diagnose and quantify status and time course of vascular diseases.

· Purpose: To evaluate the diagnostic potential of a slit-lamp-adapted optical coherence tomography (OCT) system as an in vivo imaging device for routine clinical examination of the anterior segment of the eye. · Patients and methods: In a pilot study, healthy volunteers and patients with different pathologies of the anterior segment were examined with a slit-lamp-adapted OCT system using 100-200 axial scans with 100-Hz line-scan frequency. The scan length is variable up to 7 mm, and the axial depth is 1.5 mm in tissue. · Results: The slit-lamp-adapted OCT system allowed direct biomicroscopic imaging of the measured area. Anatomic structures and morphological changes anterior to the attenuating iris pigment epithelium could be visualized with high accuracy. Biometric analyses of the cornea, the chamber angle, the iris and secondary cataract were possible. Complete demonstration of the chamber angle was difficult due to the backscattering properties of the anterior part of the sclera and the consequent shadowing of the most peripheral part of the iris. · Conclusions: Slit-lamp-adapted OCT is a useful diagnostic tool which allows in vivo microscopic cross-sectional imaging of the anterior segment and precise measurement of ocular structures.

Background Subthreshold (retinal pigment epithelium) photocoagulation is a new photocoagulation method, which treats the retinal pigment epithelium (RPE) and avoids damage to the neural retina. The initial results in this prospective pilot study on various macular diseases are presented. Methods 12 patients with diabetic maculopathy (group I), 10 with soft drusen (group II), and four with central serous retinopathy (CSR) (group III) were treated and followed up for 1 year. Treatment was achieved using a train of repetitive short laser pulses (1.7 μs) of a green Nd:YLF laser (parameters: 527 nm, 100 and 500 pulses, repetition rate: 500 Hz, spot size: 160 μm, energies: 70-100 μJ). Laser energy was based on the visibility of test lesions on fluorescein angiography (50-130 μJ). Patients were examined at various times by ophthalmoscopy, fluorescein and ICG angiography, and infrared imaging. Results After 6 months hard exudates disappeared in six out of nine patients in group I and leakage disappeared in six out of 12 diabetic patients. In group II drusen were less in seven out of 10 patients. In group III serous detachment disappeared in three out of four cases. Visual acuity was stable in all cases. None of the laser lesions was clinically visible immediately. After 1 day most lesions were visible as yellowish RPE depigmentation. After 3 months some of the lesions were visible as hyperpigmented areas but most were not. Fluorescein angiography showed leakage only in the first week. Infrared imaging showed that most lesions can be visualised in groups I and II after a period longer than 1 week as hyperreflective areas. Conclusion This study showed that subthreshold (RPE) photocoagulation is effective in some cases of diabetic maculopathy, drusens, and in CSR. Visibility of laser burns is not always necessary in the treatment of macular diseases presented here. Infrared imaging is an effective and non-invasive way of visualising subthreshold (RPE) laser burns.

Der Einsatz von gepulster Laserstrahlung erlaubt es, das Ausmaß des Gewebeschadens zu begrenzen und Strukturen, die natürlicherweise oder durch Anfärbung mit einem Kontrastmittel eine höhere Absorption als ihre Umgebung besitzen, selektiv zu schädigen. Dieses Verfahren, die selektive Photothermolyse, wird unter anderem mit Pulslängen von einigen hundert Mikrosekunden bis Millisekunden erfolgreich zur selektiven Zerstörung von Gefäßen angewandt. Ob es auch möglich ist, einzelne Zellen, Zellbereiche oder sogar Makromoleküle thermisch selektiv zu zerstören, ist zur Zeit nicht geklärt.