Lipofuscin-associated photo-oxidative stress during fundus autofluorescence imaging.

PURPOSE: Current standards and guidelines aimed at preventing retinal phototoxicity during intentional exposures do not specifically evaluate the contribution of endogenous photosensitizers. However, certain retinal diseases are characterized by abnormal accumulations of potential photosensitizers such as lipofuscin bisretinoids in the retinal pigment epithelium (RPE). We sought to determine these contributions by a numerical assessment of in-vivo photo-oxidative stress during irradiation of RPE lipofuscin. METHODS: Based on the literature, we calculated the retinal exposure levels, optical filtering of incident radiation by the ocular lens, media, photoreceptors, and RPE melanin, light absorption by lipofuscin, and photochemical effects in the RPE in two situations: exposure to short-wavelength (λ = 488 nm) fundus autofluorescence (SW-AF) excitation light and exposure to indirect (diffuse) sunlight. RESULTS: In healthy persons at age 20, 40, and 60, respectively, the rate of oxygen photoconsumption by lipofuscin increases by 1.3, 1.7, and 2.4 fold during SW-AF-imaging as compared to diffuse sunlight. In patients with STGD1 below the age of 30, this rate was 3.3-fold higher compared to age-matched controls during either sunlight or SW-AF imaging. CONCLUSIONS: Our results suggest that the RPE of patients with STGD1 is generally at increased risk of photo-oxidative stress, while exposure during SW-AF-imaging amplifies this risk. These theoretical results have not yet been verified with in-vivo data due to a lack of sufficiently sensitive in-vivo measurement techniques.

Self-mixing microprobe for monitoring microvascular perfusion in rat brain.

Measuring functional activity in brain in connection with neural stimulation faces technological challenges. Our goal is to evaluate, in relative terms, the real-time variations of local cerebral blood flow in rat brain, with a convenient spatial resolution. The use of laser Doppler flowmetry (LDF) probes is a promising approach but commercially available LDF probes are still too large (450 µm) to allow insertion in brain tissue without causing damage in an extension that may negatively impact local measurements. The self-mixing technique coupled to LDF is herein proposed to overcome limitations of the minimal diameter of the probe imposed by non-self-mixing probes (commercial available probes). Our Monte Carlo simulations show that laser photons have a mean penetration depth of 0.15 mm, on the rat brain with the 785 nm laser light microprobe. Moreover, three self-mixing signal processing methods are tested: counting method, autocorrelation method, power spectrum method. The perfusion signal computed shows a good linearity with the scatterers velocity, for the three methods (a determination coefficient close to one is obtained), for the in vitro measurements. Furthermore, we believe that these indicators can be used to monitor local blood flow changes in the rat brain.

Depth-kymography of vocal fold vibrations: part II. Simulations and direct comparisons with 3D profile measurements.

We report novel direct quantitative comparisons between 3D profiling measurements and simulations of human vocal fold vibrations. Until now, in human vocal folds research, only imaging in a horizontal plane was possible. However, for the investigation of several diseases, depth information is needed, especially when the two folds act differently, e.g. in the case of tumour growth. Recently, with our novel depth-kymographic laryngoscope, we obtained calibrated data about the horizontal and vertical positions of the visible surface of the vibrating vocal folds. In order to find relations with physical parameters such as elasticity and damping constants, we numerically simulated the horizontal and vertical positions and movements of the human vocal folds while vibrating and investigated the effect of varying several parameters on the characteristics of the phonation: the masses and their dimensions, the respective forces and pressures, and the details of the vocal tract compartments. Direct one-to-one comparison with measured 3D positions presents-for the first time-a direct means of validation of these calculations. This may start a new field in vocal folds research.

Time development models for perfusion provocations studied with laser-Doppler flowmetry, applied to iontophoresis and PORH.

OBJECTIVE: Clinical acceptance of laser-Doppler perfusion monitoring (LDPM) of microcirculation suffers from lack of quantitatively reliable signal data, due to varying tissue constitution, temperature, hydration, etc. In this article, we show that a novel approach using physiological models for response upon provocations provides quantitatively and clinically relevant time constants. METHODS: We investigated this for two provocation protocols: postocclusive reactive hyperemia (PORH) and iontophoresis shots, measured with LDPM on extremities. PORH experiments were performed on patients with peripheral arterial occlusive disease (PAOD) or diabetes mellitus (DM), and on healthy controls. Iontophoresis experiments were performed on pre-eclamptic patients and healthy controls. We developed two dynamical physical models, both based on two characteristic time constants: for PORH, an "arterial" and a "capillary" time constant and, for iontophoresis, a "diffusion" and a "decay" time constant. RESULTS: For the different subject groups, we could extract time constants that could probably be related to physiological differences. For iontophoresis, a shot saturation constant was determined, with very different values for different groups and administered drugs. CONCLUSIONS: With these models, the dynamics of the provocations can be investigated and quantitative comparisons between experiments and subject groups become available. The models offer a quantifiable standard that is independent of the type of LDPM instrumentation.

Depth-kymography: high-speed calibrated 3D imaging of human vocal fold vibration dynamics.

We designed and developed a laser line-triangulation endoscope compatible with any standard high-speed camera for a complete three-dimensional profiling of human vocal fold vibration dynamics. With this novel device we are able to measure absolute values of vertical and horizontal vibration amplitudes, length and width of vocal folds as well as the opening and closing velocities from a single in vivo measurement. We have studied, for the first time, the generation and propagation of mucosal waves by locating the position of its maximum vertical position and the propagation velocity. Precise knowledge about the absolute dimensions of human vocal folds and their vibration parameters has significant importance in clinical diagnosis and treatment as well as in fundamental research in voice. The new device can be used to investigate different kinds of pathological conditions including periodic or aperiodic vibrations. Consequently, the new device has significant importance in investigating vocal fold paralysis and in phonosurgical applications.

New laryngoscope for quantitative high-speed imaging of human vocal folds vibration in the horizontal and vertical direction.

We report the design of a novel laser line-triangulation laryngoscope for the quantitative visualization of the three-dimensional movements of human vocal folds during phonation. This is the first successful in vivo recording of the three-dimensional movements of human vocal folds in absolute values. Triangulation images of the vocal folds are recorded at the rate of 4000 fps with a resolution of 256x256 pixels. A special image-processing algorithm is developed to precisely follow the subpixel movements of the laser line image. Vibration profiles in both horizontal and vertical directions are calibrated and measured in absolute SI units with a resolution of +/-50 microm. We also present a movie showing the vocal folds dynamics in vertical cross section.

Normalization of vasomotion in laser Doppler perfusion monitoring.

Laser Doppler flux signals show temporal fluctuations caused by physiological phenomena like heartbeat, respiration, and local variation of vascular tonus, vasomotion. This study investigates the influence of fiber arrangement, equipment and two probe locations on the variations in laser Doppler flux signals in five frequency bands in the absence of provocations. Two probes with detecting optical fibers at several distances from the illuminating source were used, as well as instruments from two manufacturers. The results show that normalization of the filtered flux signals with the mean flux leads to an enormous decrease of the influence of fiber distance. The difference between instruments is small after normalization. Some influence of probe location remains after normalization. Development of a standard method for normalization of the variations in laser Doppler signals is recommended.

Diffusion model for iontophoresis measured by laser-Doppler perfusion flowmetry, applied to normal and preeclamptic pregnancies.

We present a physical model to describe iontophoresis time recordings. The model is a combination of monodimensional material diffusion and decay, probably due to transport by blood flow. It has four adjustable parameters, the diffusion coefficient, the decay constant, the height of the response, and the shot saturation constant, a parameter representing the relative importance of subsequent shots (in case of saturation). We test the model with measurements of blood perfusion in the capillary bed of the fingers of women who recently had preeclampsia and in women with a history of normal pregnancy. From the fits to the measurements, we conclude that the model provides a useful physical description of the iontophoresis process.

Integrated optoelectronic probe including a vertical cavity surface emitting laser for laser Doppler perfusion monitoring.

An integrated optoelectronic probe with small dimensions, for direct-contact laser Doppler blood flow monitoring has been realized. A vertical cavity surface emitting laser (VCSEL), and a chip with photodetectors and all necessary electronics are integrated in a miniature probe head connected to a laptop computer. The computer sound processor is utilized for acquisition and digital signal processing of the incoming Doppler signal. In this paper, the design of the laser Doppler perfusion monitor is described and its performance is evaluated. We demonstrate our perfusion monitor to be less sensitive to subject motion than a commercial fiber-optic device. For medium and high perfusion levels, the performance of our integrated probe is comparable to the fiberoptic flowmeter containing a normal edge-emitting laser diode. For very low perfusion levels, the signal-to-noise ratio of the fiber-optic device is higher. This difference can mainly be attributed to the shorter coherence length of the VCSEL compared with the edge-emitting laser diode.

A model for post-occlusive reactive hyperemia as measured with laser-Doppler perfusion monitoring.

To facilitate the quantitative analysis of post-occlusive reactive hyperaemia (PORH), measured with laser-Doppler perfusion monitoring (LDPM) on extremities, we present a flow model for the dynamics of the perfusion of the tissue during PORH, based on three parameters: two time constants (tau1 and tau2) and the ratio of the maximum flux and the resting flux. With these three constants quantitative comparisons between experiments will be possible and, therefore, we propose to adopt this approach as future standard. For this reason, we also developed a computer program to perform the fit of the model to measured data.

Imaging of small vessels using photoacoustics: an in vivo study.

BACKGROUND AND OBJECTIVES: The ability to correctly visualize the architectural arrangement of microvasculature is valuable to many diverse fields in medicine. In this study, we applied photoacoustics (PA) to obtain high-resolution images of submillimeter blood vessels. STUDY DESIGN/MATERIALS AND METHODS: Short laser pulses are used to generate ultrasound from superficial blood vessels in several animal models. From these ultrasound waves the interior of blood vessels can be reconstructed. RESULTS: We present results from a novel approach based on the PA principle that allows specific in vivo visualization of dermal blood vessels without the use of contrast agents or ionizing radiation. CONCLUSIONS: We show PA images of externalized blood vessels and demonstrate in vivo PA imaging of vasculature through layers of skin varying in thickness.

Photoacoustic imaging of blood vessels with a double-ring sensor featuring a narrow angular aperture.

A photoacoustic double-ring sensor, featuring a narrow angular aperture, is developed for laser-induced photoacoustic imaging of blood vessels. An integrated optical fiber enables reflection-mode detection of ultrasonic waves. By using the cross-correlation between the signals detected by the two rings, the angular aperture of the sensor is reduced by a factor of 1.9, from 1.5 to 0.8 deg. Consequently, photoacoustic images could be obtained in a manner analogous to the ultrasound B-scan mode. Next, the cross section of artificial blood vessels is visualized by reconstruction of the absorbed energy distribution. Finally, in vivo imaging and the subsequent reconstruction of the absorbed energy distribution is demonstrated for superficial blood vessels in the human wrist.

Photoacoustic determination of blood vessel diameter.

A double-ring sensor was applied in photoacoustic tomographic imaging of artificial blood vessels as well as blood vessels in a rabbit ear. The peak-to-peak time (tau(pp)) of the laser (1064 nm) induced pressure transient was used to estimate the axial vessel diameter. Comparison with the actual vessel diameter showed that the diameter could be approximated by 2ctau(pp), with c the speed of sound in blood. Using this relation, the lateral diameter could also precisely be determined. In vivo imaging and monitoring of changes in vessel diameters was feasible. Finally, acoustic time traces were recorded while flushing a vessel in the rabbit ear with saline, which proved that the main contribution to the laser-induced pressure transient is caused by blood inside the vessel and that the vessel wall gives only a minor contribution.

Suppression of dynamic laser speckle signals in multimode fibers of various lengths.

The effects of fiber coupling and fiber length on photocurrent fluctuations are studied when the light of a laser diode transmitted to and from a dynamic turbid medium by a step-index multimode fiber is studied. When the laser light is coupled asymmetrically, filling only the higher-order modes, the photocurrent fluctuations are suppressed significantly when fiber lengths of as much as 16 m are added between the laser and the medium. Addition of as much as 16 m of detection fiber, or any fiber in the case of symmetric light coupling, leads to much less or no suppression of the photocurrent fluctuations.

Temperature modulation of the visible and near infrared absorption and scattering coefficients of human skin.

We determine temperature effect on the absorption and reduced scattering coefficients (mu(a) and mu(s)(')) of human forearm skin. Optical and thermal simulation data suggest that mu( a) and mu(s)(') are determined within a temperature-controlled depth of approximately 2 mm. Cutaneous mu(s)(') change linearly with temperature. Change in mu(a) was complex and irreversible above body normal temperatures. Light penetration depth (delta) in skin increased on cooling, with considerable person-to-person variations. We attribute the effect of temperature on mu(s)(') to change in refractive index mismatch, and its effect on mu(a) to perfusion changes. The reversible temperature effect on mu (s)(' ) was maintained during more than 90 min. contact between skin and the measuring probe, where temperature was modulated between 38 and 22 degrees C for multiple cycles While temperature modulated mu(s)(' ) instantaneously and reversibly, mu(a) exhibited slower response time and consistent drift. There was a statistically significant upward drift in mu(a) and a mostly downward drift in mu( s)(') over the contact period. The drift in temperature-induced fractional change in mu(s)(') was less statistically significant than the drift in mu(s)('). Deltamu( s)(') values determined under temperature modulation conditions may have less nonspecific drift than mu(s)(') which may have significance for noninvasive determination of analytes in human tissue.

Poly(vinyl alcohol) gels for use as tissue phantoms in photoacoustic mammography.

Materials for solid photoacoustic breast phantoms, based on poly(vinyl alcohol) hydrogels, are presented. Phantoms intended for use in photoacoustics must possess both optical and acoustic properties of tissue. To realize the optical properties of tissue, one approach was to optimize the number of freezing and thawing cycles of aqueous poly(vinyl alcohol) solutions, a procedure which increases the turbidity of the gel while rigidifying it. The second approach concentrated on forming a clear matrix of the rigid poly(vinyl alcohol) gel without any scattering, so that appropriate amounts of optical scatterers could be added at the time of formation, to tune the optical properties as per requirement. The relevant optical and acoustic properties of such samples were measured to be close to the average properties of human breast tissue. Tumour simulating gel samples of suitable absorption coefficient were created by adding appropriate quantities of dye at the time of formation; the samples were then cut into spheres. A breast phantom embedded with such 'tumours' was developed for studying the applicability of photoacoustics in mammography.

Glass-fiber self-mixing intra-arterial laser Doppler velocimetry: signal stability and feedback analysis.

We have developed a blood velocimeter based on the principle of self-mixing in a semiconductor laser diode through an optical fiber. The intensity of the light is modulated by feedback from moving scattering particles that contain the Doppler-shift frequency. Upon feedback the characteristics of the laser diode change. The threshold current decreases, and an instable region may become present above the new threshold. The amplitude of the Doppler signal turns out to be related to the difference in intensity between situations with and without feedback. This amplitude is highest just above feedback. The suppression of reflection from the glass-fiber facets is of paramount importance in the obtaining of a higher signal-to-noise ratio. Using an optical stabilization of the feedback, we optimized the performance of the laser-fiber system and the Doppler modulation depth and clarified its behavior with a suitable physical model. We also investigated the effect of the finite coherence length of the laser. We tested the efficiency of the self-mixing velocimeter in vivo with the optical glass fiber inserted in the artery with endoscopic catheters, both in upstream and in downstream blood flow conditions. For the latter we used a special side-reflecting device solution for the fiber facet to allow downstream measurements.