E. Göran Salerud

Evaluation of laser Doppler flowmetry in the assessment of intestinal blood flow in cat.

Laser Doppler flowmetry with a differential detector system has been used in the assessment of blood flow in the feline small intestine. Simultaneous mucosal and serosal laser Doppler flowmeter recordings were compared with total blood flow of a bowel segment measured by an optical drop-recorder unit in 6 cats. Blood flow through the muscularis layer was estimated using the 85Kr washout technique. A correlation coefficient of r = 0.96 (mucosal recordings = 90, serosal recordings = 80, p less than 0.001) was obtained between laser Doppler flowmeter output signals and total blood flow at different levels of vascular tone, regardless of whether the flowmeter recordings were made from the mucosal or the serosal side of the bowel wall. At intense vasodilation, the flowmeters showed a tendency to underestimate blood flow. The flowmeter signals were at variance with the muscularis blood flow but were clearly correlated to the calculated mucosal-submucosal blood flow. The uneven blood flow distribution in the intestinal wall did not affect the ability of the flowmeters to reflect total blood flow from either side of the bowel wall. A calibration curve could be constructed for approximate interpretation of the laser Doppler signals in absolute flow units. However, further experiments in humans and further development of the technique must be performed to elucidate clinical applications of the method.

Single-fibre laser Doppler flowmetry

A laser Doppler flowmeter with one optical fibre guiding light to and from the tissue under study has been developed. The outer diameter of the probe equals the optical fibre diameter (0·5 mm). The small size makes it useful for studying the deep tissue perfusion in organs. Differential-channel operation was compared with the single-channel operation and the benefit of this technique was evaluated theoretically as well as in a fluid model resembling tissue perfusion. The signal-to-noise improvement ratio was calculated and found to be related to the number of coherence areas detected and to the broadband noise of the laser. In vivo experiments in the gastrocnemius muscle of the pig were performed to compare the results from the single-fibre technique with those of the electromagnetic flowmeter. Linear regression analysis of femoral blood flow data obtained with the electromagnetic flowmeter and local muscle blood flow measured with the single-fibre technique showed a correlation coefficient of 0·88 (n=36, p<0·001).

Imaging, image processing and pattern analysis of skin capillary ensembles

Background/aims: The capillary bed is recognized as the site where metabolic and nutrient processes occur for living tissues at all levels. The evaluation of this vital process is a major concern in microcirculation. Unlike traditional approaches that concentrated on the extreme local properties of this process, a more global analysis toward capillary ensembles is employed here, since capillaries work as a cooperative entirety. As a first step toward ensemble analysis, the static and planar geometric parameters are investigated. Parameters such as the capillary adjacency and size information are very important in predicting and analysing certain malfunctions in the microvascular bed.

Improved model for myocardial diffuse reflectance spectra by including mitochondrial cytochrome aa3, methemoglobin, and inhomogenously distributed RBC

The aim of this study was to compare a previously used light transport model (I) comprising the chromophores hemo- and myoglobin, fat, and water, with two extended models, where the chromophores of cytochrome aa3, methemo- and metmyoglobin are added (model II), and in addition, accounting for an inhomogenous hemoglobin distribution (model III). The models were evaluated using calibrated diffuse reflectance spectroscopy measurements on the human myocardium. Model II proved a significantly better spectral fitting, especially in the wavelength ranges corresponding to prominent absorption characteristics for the added chromophores. Model III was significantly better than model II and displayed a markedly higher tissue fraction and saturation of hemo- and myoglobin. The estimated tissue chromophore fractions, saturation and oxidation levels, were in agreement with other studies, demonstrating the potential of diffuse reflectance spectroscopy measurements for evaluating open heart surgery. However, the choice of chromophores and vessel packaging effects in the light transport model has a major effect on the results.

Visible hyperspectral imaging evaluating the cutaneous response to ultraviolet radiation

In vivo diagnostics of skin diseases as well as understanding of the skin biology constitute a field demanding characterization of physiological and anatomical parameters. Biomedical optics has been successfully used, to qualitatively and quantitatively estimate the microcirculatory conditions of superficial skin. Capillaroscopy, laser Doppler techniques and spectroscopy, all elucidate different aspects of microcirculation, e.g. capillary anatomy and distribution, tissue perfusion and hemoglobin oxygenation. We demonstrate the use of a diffuse reflectance hyperspectral imaging system for spatial and temporal characterization of tissue oxygenation, important to skin viability. The system comprises: light source, liquid crystal tunable filter, camera objective, CCD camera, and the decomposition of the spectral signature into relative amounts of oxy- and deoxygenized hemoglobin as well as melanin in every pixel resulting in tissue chromophore images. To validate the system, we used a phototesting model, creating a graded inflammatory response of a known geometry, in order to evaluate the ability to register spatially resolved reflectance spectra. The obtained results demonstrate the possibility to describe the UV inflammatory response by calculating the change in tissue oxygen level, intimately connected to a tissues metabolism. Preliminary results on the estimation of melanin content are also presented.

A diffuse reflectance spectroscopic study of UV-induced erythematous reaction across well-defined borders in human skin.

Introduction: The colour of tissue is often of clinical use in the diagnosis of tissue homeostasis and physiological responses to various stimuli. Determining tissue colour changes and borders, however, often poses an intricate problem and visual examination, constituting clinical praxis, does not allow them to be objectively characterized or quantified. Demands for increased inter‐ and intra‐observer reproducibility have been incentives for the introduction of objective methods and techniques for tissue colour (e.g. erythema) evaluation. The aim of the present paper was to study the border zone of a UVB‐provoked erythematous response of human skin in terms of blood volume and oxygenation measured by means of diffuse reflectance spectroscopy using a commercial probe.

Spatial and temporal skin blood volume and saturation estimation using a multispectral snapshot imaging camera

Hyperspectral imaging (HSI) can estimate the spatial distribution of skin blood oxygenation, using visible to near-infrared light. HSI oximeters often use a liquid-crystal tunable filter, an acousto-optic tunable filter or mechanically adjustable filter wheels, which has too long response/switching times to monitor tissue hemodynamics. This work aims to evaluate a multispectral snapshot imaging system to estimate skin blood volume and oxygen saturation with high temporal and spatial resolution. We use a snapshot imager, the xiSpec camera (MQ022HG-IM-SM4X4-VIS, XIMEA), having 16 wavelength-specific Fabry–Perot filters overlaid on the custom CMOS-chip. The spectral distribution of the bands is however substantially overlapping, which needs to be taken into account for an accurate analysis. An inverse Monte Carlo analysis is performed using a two-layered skin tissue model, defined by epidermal thickness, haemoglobin concentration and oxygen saturation, melanin concentration and spectrally dependent reduced-scattering coefficient, all parameters relevant for human skin. The analysis takes into account the spectral detector response of the xiSpec camera. At each spatial location in the field-of-view, we compare the simulated output to the detected diffusively backscattered spectra to find the best fit. The imager is evaluated for spatial and temporal variations during arterial and venous occlusion protocols applied to the forearm. Estimated blood volume changes and oxygenation maps at 512x272 pixels show values that are comparable to reference measurements performed in contact with the skin tissue. We conclude that the snapshot xiSpec camera, paired with an inverse Monte Carlo algorithm, permits us to use this sensor for spatial and temporal measurement of varying physiological parameters, such as skin tissue blood volume and oxygenation.

Estimating skin blood saturation by selecting a subset of hyperspectral imaging data

Skin blood haemoglobin saturation (𝑠b) can be estimated with hyperspectral imaging using the wavelength (λ) range of 450-700 nm where haemoglobin absorption displays distinct spectral characteristics. Depending on the image size and photon transport algorithm, computations may be demanding. Therefore, this work aims to evaluate subsets with a reduced number of wavelengths for 𝑠b estimation. White Monte Carlo simulations are performed using a two-layered tissue model with discrete values for epidermal thickness (𝑇epi) and the reduced scattering coefficient (μs ), mimicking an imaging setup. A detected intensity look-up table is calculated for a range of model parameter values relevant to human skin, adding absorption effects in the post-processing. Skin model parameters, including absorbers, are; μs (λ), 𝑇epi, haemoglobin saturation (𝑠b), tissue fraction blood (𝑐b) and tissue fraction melanin (𝑐mel). The skin model paired with the look-up table allow spectra to be calculated swiftly. Three inverse models with varying number of free parameters are evaluated: A(𝑠b, 𝑐b), B(𝑠b, 𝑐b, 𝑐mel) and C(all parameters free). Fourteen wavelength candidates are selected by analysing the maximal spectral sensitivity to 𝑠b and minimizing the sensitivity to 𝑐b. All possible combinations of these candidates with three, four and 14 wavelengths, as well as the full spectral range, are evaluated for estimating 𝑠b for 1000 randomly generated evaluation spectra. The results show that the simplified models A and B estimated 𝑠b accurately using four wavelengths (mean error 2.2% for model B). If the number of wavelengths increased, the model complexity needed to be increased to avoid poor estimations.

A bone tissue integrated single fibre laser Doppler flowmeter probe.

Laser Doppler Flowmetry was developed and evaluated mainly for measuring blood flow in exposed or superficial tissue layers like skin. Results showed large variation coefficients between juxtaposed skin sites with apparently homogeneous blood perfusion. Spatial differences in blood flow as well as anatomical differences probably explain the obtained results. Vasomotion, hereto only discovered with a microscopic technique, was recorded and evaluated for the first time. Similar results had been obtained in previous bone perfusion studies. The LDF technique has also been used extensively for skin irritancy tests and microvascular effects of vasoactive substances and drugs. An in vivo model in calibrating LDF quantitatively showed a linear relationship between LDF output signal and total blood flow in the range 0-300 ml min-1 100 g-1. This quantitative calibration is however tissue specific. Most tissues of interest are not superficial. The single fibre LDF provides additional power in studying deep tissue physiology and trauma-induced changes in regulation of microvascular perfusion. Furthermore long-term studies require implantable and biocompatible probes and this could be fulfilled using the SFLDF technique in combination with a tissue-integrated titanium probe. Implantation of the titanium probes initially creates some trauma to the vascular bed in the bone marrow but almost none in the opposite cortical bone layer. The healing process enhances the blood perfusion during the first weeks. The increased perfusion can be due to different parameters such as angiogenesis. The histological findings support this hypothesis. This gives preference to the tissue-integrated probe where interference with the tissue should be avoided. The integrated probe technique has only been evaluated in bone tissue. Modification of the probe shape and dimension, however, would make it suitable for other tissues as well.

Noninvasive assessments of oxygen delivery from the microcirculation to skin in hypothermia-treated asphyxiated newborn infants.

Background:Therapeutic hypothermia (TH) has become standard treatment for severe and moderate hypoxic–ischemic neonatal encephalopathy (HIE). Our group has developed an optically based, noninvasive concept of assessing the capacity for oxygen delivery from the microcirculation to the cells of a tissue under investigation. The hypothesis was that mechanisms of reduced oxygen delivery due to reduced metabolism in cooled asphyxiated neonates could be characterized with this concept.Methods:The skin of 28 asphyxiated newborn infants was studied on days 1 and 3 during TH and on day 4 following rewarming with laser Doppler perfusion measurements (LDPM), computer-assisted video microscopy (CAVM), and diffuse reflectance spectroscopy (DRS). Twenty-five healthy neonates served as a control group.Results:The LDPM decreased during cooling (P < 0.01). Functional capillary density was higher both during and following TH compared with control infants (P < 0.01). Capillary flow velocities were reduced during TH (P < 0.05). The heterogeneity of the flow velocities was larger in the HIE infants than in the control infants. Tissue oxygen extraction was higher during TH (P < 0.01).Conclusion:This study indicates that assessments of skin microvascular density, capillary flow velocity, and oxygen extraction can be used to characterize reduced oxygen delivery to cells during TH.