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Interpreting diffuse reflectance for in vivo skin reactions in terms of chromophores
Abstract
The measurement and quantification of skin reactions to insults involves certain assumptions about the relation between intensity of color appearance of the skin and the concentration of endogenous chromophores. The underlying assumption is that the Beer-Lambert law is obeyed, i.e., that a linear relation exists between the absorbance and the concentration of each chromophore and that the total absorbance is the linear superposition of the contributions of each chromophore. In this paper the authors compiled the results from a number of interventions on human skin that result in changes in its appearance and small deviations from the homeostatic state, where the results may be accounted for by a single or multiple chromophores. The validity of the assumptions is found to hold for a limited range of responses. The biological constraints need to be considered in certain cases because as we move away from the homeostatic state, complex biological processes are induced.
... Skin is an important organ in the human body that is highly vulnerable to diseases such as cancer. The skin is a complex structure with a unique tissue biochemical composition that has to be carefully monitored for biochemical changes to detect and diagnose various skin diseases 58,106 Recently, fiberoptic spectroscopy techniques (e.g., reflectance, fluorescence and Raman spectroscopy) capable of probing changes in tissue optical properties that can reflect physiological parameters and tissue composition have been recognized as promising candidates for in vivo disease diagnosis in various organs of the human body. 1,3,8,23,28,66 The spectroscopy diagnostic instruments employ fiberoptic probe as an integral part of system, enabling non-invasive or minimally-invasive access to remote internal organs. ...
... Fiber based Diffuse Reflectance (DR) spectroscopy is a rapid, non-invasive spectroscopy technique that has been used in various biomedical applications like characterization of tissue biochemical content, studying the optical properties of biological tissues as well as disease diagnosis in a number of organs 10,11,36,106,123,132,133 The diagnostic capability of DR spectroscopy is predominantly due to its excellent sensitivity to reflect changes in biochemical concentration in target tissues. Fiber-based DR spectroscopy makes use of a fiber-optic probe that is in contact with the target tissue to reduce interferences due to refractive index mismatch, specular reflection, increases light penetration and transmittance and avoids inconsistent illumination-detection geometry during spectral acquisition which results in the enhancement of the diagnostic capability of the system. ...
... 31 The rise in hemoglobin concentration with pressure is mainly due to the occlusion of blood flow under the probe tip, in agreement with the reported literatures. 106,136 The higher concentrations of water, lipids and other endogenous chromophores with the magnitude of exerted probe pressure is associated with the compression of the tissue. 31 developed to differentiate DR spectral distortions introduced by the variation in the applied probe-tissue contact pressure. ...
A NOVEL PROBE DESIGN IS PRESENTED FOR REAL-TIME MONITORING OF PROBE-TISSUE CONTACT PRESSURE EFFECTS DURING IN VIVO SKIN OPTICAL SPECTROSCOPY MEASUREMENTS. THE DEVELOPED PRESSURE SENSITIVE PROBE IS VITAL TO PRESERVE IMPORTANT DIAGNOSTIC INFORMATION THAT MAY BE LOST DUE TO EXCESSIVE PROBE PRESSURE. OPTICAL SPECTRA WERE ACQUIRED DURING IN VIVO AUTOFLUORESCENCE (AF), DIFFUSE REFLECTANCE (DR) AND RAMAN SPECTROSCOPIC MEASUREMENTS ON HUMAN SKIN SITES (I.E. INDEX FINGERTIP, PALM AND VOLAR FOREARM) AT DIFFERENT PROBE-TISSUE CONTACT PRESSURES TO EVALUATE THE PROBE PRESSURE INDUCED VARIATIONS IN THE ACQUIRED SPECTRAL DATA. MULTIVARIATE STATISTICAL ANALYSIS WAS PERFORMED ON THE ACQUIRED SPECTRA TO ESTIMATE THE DIAGNOSTIC IMPORTANCE OF PROBE PRESSURE INDUCED VARIATIONS ON THE SPECTRAL DATA. THE RESULTS SHOW THAT PROBE PRESSURE IS AN ESSENTIAL PARAMETER THAT AFFECTS THE OPTICAL SPECTRA SIGNIFICANTLY. PROBE PRESSURE INDUCED SPECTRAL VARIATIONS CAN BE MITIGATED BY THE UTILIZATION OF THE DEVELOPED PRESSURE SENSITIVE FIBEROPTIC PROBE
... Quantitative comparisons of measured diffuse reflectance spectra (DRS) with theoretical predictions were used, e.g., to determine various tissue optical properties. [1][2][3][4][5][6] In addition, several groups have applied DRS for analysis of spatially heterogeneous organs, such as human skin, [7][8][9] with specific aims to assess concentrations of epidermal melanin [10][11][12] or other skin chromophores, [13][14][15] enable diagnostic characterization of skin lesions, [16][17][18] or monitor time evolution of traumatic bruises. [19][20][21] A common setup for diffuse reflectance spectroscopy involves an integrating sphere (IS) with internal broadband light source. ...
... Setting the fractional area of all openings combined (undocumented) to F ¼ 0.07-a reasonable value given the IS structure-results in an excellent match of the function's shape and amplitude with our experimental data [ Fig. 4(b)]. As can be seen from Eq. (14), Δ SBS becomes positive when R sam exceeds R wh . ...
... By using Eq. (14), this pertains, in particular, to finding the optimal value of parameter F. This task can be easily performed by each end user by following the steps described in the present article (see Fig. 4) and using the parameter values (f, ρ, and R wh ) appropriate for their own equipment. ...
Diffuse reflectance spectra (DRS) of biological samples are commonly measured using an integrating sphere (IS), in which spectrally broad illumination light is multiply scattered and homogenized. The measurement begins by placing a highly reflective white standard against the IS sample opening and collecting the reflected light at the signal output port to account for illumination field. After replacing the white standard with test sample of interest, DRS of the latter is determined as the ratio of the two values at each involved wavelength. However, because test samples are invariably less reflective than the white standard, such a substitution modifies the illumination field inside the IS. This leads to underestimation of the sample's reflectivity and distortion of measured DRS, which is known as single-beam substitution error (SBSE). Barring the use of much more complex dual-beam experimental setups, involving dedicated IS, literature states that only approximate corrections of SBSE are possible, e.g., by using look-up tables generated with calibrated low-reflectivity standards. We present a practical way to eliminate the SBSE using IS equipped with an additional " reference" output port. Two additional measurements performed at this port (of the white standard and sample, respectively) namely enable an accurate compensation for above described alteration of the illumination field. In addition, we analyze the dependency of SBSE on sample reflectivity and illustrate its impact on measurements of DRS in human skin with a typical IS.
... Quantitative comparisons of measured diffuse reflectance spectra (DRS) with theoretical predictions were used, e.g., to determine various tissue optical properties. [1][2][3][4][5][6] In addition, several groups have applied DRS for analysis of spatially heterogeneous organs, such as human skin, [7][8][9] with specific aims to assess concentrations of epidermal melanin [10][11][12] or other skin chromophores, [13][14][15] enable diagnostic characterization of skin lesions, [16][17][18] or monitor time evolution of traumatic bruises. [19][20][21] A common setup for diffuse reflectance spectroscopy involves an integrating sphere (IS) with internal broadband light source. ...
... Setting the fractional area of all openings combined (undocumented) to F ¼ 0.07-a reasonable value given the IS structure-results in an excellent match of the function's shape and amplitude with our experimental data [ Fig. 4(b)]. As can be seen from Eq. (14), Δ SBS becomes positive when R sam exceeds R wh . ...
... By using Eq. (14), this pertains, in particular, to finding the optimal value of parameter F. This task can be easily performed by each end user by following the steps described in the present article (see Fig. 4) and using the parameter values (f, ρ, and R wh ) appropriate for their own equipment. ...
Diffuse reflectance spectra (DRS) of biological samples are commonly measured using an integrating sphere (IS). To account for the incident light spectrum, measurement begins by placing a highly reflective white standard against the IS sample opening and collecting the reflected light. After replacing the white standard with the test sample of interest, DRS of the latter is determined as the ratio of the two values at each involved wavelength. However, such a substitution may alter the fluence rate inside the IS. This leads to distortion of measured DRS, which is known as single-beam substitution error (SBSE). Barring the use of more complex experimental setups, the literature states that only approximate corrections of the SBSE are possible, e.g., by using look-up tables generated with calibrated low-reflectivity standards. We present a practical method for elimination of SBSE when using IS equipped with an additional reference port. Two additional measurements performed at this port enable a rigorous elimination of SBSE. Our experimental characterization of SBSE is replicated by theoretical derivation. This offers an alternative possibility of computational removal of SBSE based on advance characterization of a specific DRS setup. The influence of SBSE on quantitative analysis of DRS is illustrated in one application example.
... [12][13][14] Several clinical studies in laser treatment of skin disorders have been reported employing optical techniques using reflection spectroscopy as a safe and accurate method. [15][16][17][18][19] Babadi et al. conducted a study in 2019 to quantitatively assess the erythema in hair removal laser therapy. In this study, the diffused reflection spectra before and after sessions of hair removal laser were recorded from the treated areas of the patients and analyzed. ...
... In analyzing patients' information, the hemoglobin spectrum In this light-based intervention, they evaluated the skin homeostasis condition by examining changes in blood concentration and measuring it through optical spectroscopy and spectrum analysis by fitting different slope diagram equations. 19 The result of our preliminary study seems to be according to the results of this reference in which the responses of skin lesion as the form of hyperthermia changes analyzed with quantitative tools. In patients with darker skin, subjective biases might be more prevalent in selecting energy laser parameters. ...
Introduction:
The lack of objectivity options for a specific individualzed therapy might cause challenges in laser treatment. In other words, we need optimally determined laser parameters for less side effects. Genrally, laser treatment procedures seem to be subjective. Then, the final evaluation of the patient for need for opoimezed better response with less laser sections and less side effects.Therefore, employing a reliable objective technique seems to be essential for better respons with less laser treatment sessions and also less side effects.
Method:
In this research, UV-visible diffused reflection spectra from normal skin and a lesion were taken.We obtained the differences in absorption intensity at 575 nm, the wavelength corresponds to the absorption peak of blood oxyhemoglobin for normal skin and hemangioma.. To calibrate the measurements, after using pulsed dye laser (PDL at 585 nm), the PDL treatment response of the patients were graded as "good (>50%), moderate(25-50%), and poor (0-25%)", by a specialist. Finally, patients were categorized based on the energy of the laser for the best treatment response to propose the recommended laser parameters RESULTS: Based on the differences in the absorption peak hemangioma compare with normal skin, the energy density of PDL for a good treatment response of hemangioma was obtained at peak wavelength 575 nm.
Conclusion:
Analysis of optical reflection spectroscopy cas assess the correlation of absorption peak differences of vascular lesions and normal skin. According to this data it seems to be effective in optimizing lasers parametes for the hemangioma treatmemnt.
... Likewise, scattering is used to determine physiological processes such as hemodynamics, 14 microcirculation 15 and helps determine tissue morphological characteristics like scatterer size and density. 16,17 Skin DRS spectra reflecting the contents from chromophores such as melanin, bilirubin, and oxy-deoxyhemoglobin could reveal many diseased conditions like skin inflammation, [18][19][20] jaundice, 21 vitiligo, 22 melanoma 23 , etc. A typical DRS instrumentation consists of a light source, spectrometer, fibre optic probe for transmitting, and collection of light. ...
... A typical DRS instrumentation consists of a light source, spectrometer, fibre optic probe for transmitting, and collection of light. 24 Although the instrumentation is simple in design, the strenuous task to determine the optical properties from the collected light 20,25 restrains the application of this technique in quantitative spectroscopy. Light propagation in tissues is appropriately illustrated by radiative transport equation (RTE). ...
Spatially Resolved Diffuse Reflectance Spectroscopy (SRDRS) is a non-invasive optical technique that helps in clinical diagnosis of various tissue microcirculation and skin pigmentation disorders based on collected backscattered light from multi-layered tissue. The extraction of the optical properties from the reflectance spectrum using analytical solutions is laborious. Model-based light tissue interaction studies help in quantifying the optical properties. This work presents the use of finite element models of light tissue interaction for this purpose. A bilayer model mimicking human skin was considered and the diffused reflectance spectra at multiple detector points were generated using finite element modeling for varying melanin concentration, epidermal thickness, blood volume fraction, oxygen saturation and scattering components. The reflectance value based on varying optical parameters from multiple detection points lead to the generation of a look-up Table (LUT), which is further used for finding the tissue parameters that contribute to the spatially resolved reflectance values. The tissue parameters estimated after inverse modelling showed a high degree of agreement with the expected tissue parameters for a test dataset different from the training dataset.
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... Normalmente, el proceso de curación depende de factores tales como el tipo de lesión y mediadores sistémicos empleados. Sin embargo, en la mayoría de los casos el monitoreo y control de la curación han sido tratados con técnicas de evaluación visual [1] basadas en la formación y experiencia previa del médico. Ello por supuesto no permite una cuantificación sistemática durante el proceso mismo. ...
... , (1) donde , , y son la longitud de onda, velocidad de fase, y frecuencia del sonido respectivamente. Esto ocurre debido a la condición de selectividad de Bragg, que establece que la longitud de onda de la luz difractada por el cristal obedece a la ecuación: ...
This paper presents a novel non-invasive methodology to study the cutaneous wound healing process in rabbits. Spectral images were used to establish a quantitative evaluation of the healing process time-line dynamics. Measurements were conducted by image analysis of the wounds of an experimental population with two different systems: an acousto-optical acquiring/processing system permitted the experimental quantitative study of different stages naturally occurring during the healing process time-line. Honey and saline solution application were used in a subgroup of the population under study as examples of treatments that could be employed to purportedly accelerate cutaneous tissue restoration, and their efficacy was evaluated. The methodology described on this paper allows for a non-invasive and effective way to quantitatively study/monitor the wound healing processes in terms of spectral changes of the wound area. Finally, it was found that this methodology allows quantitative parametrization of measurements to be used as accurate testing grounds for new wound healing assisting clinical treatments.
... Their analysis was thus applied to diagnosis of various pathologies in bladder [1], colon [2], brain [3], breast [4,5], esophagus [6], and skin [7,8]. Specifically, DRS from human skin reflect the contents of epidermal melanin, oxy-and deoxy-hemoglobin, carotenoids, lipids, etc. [9][10][11][12], with potential applications ranging from monitoring of skin inflammation [10], changes induced by exposure to ultraviolet light [11,13], aging of traumatic bruises [14], and evaluation of scars [15]. ...
... Their analysis was thus applied to diagnosis of various pathologies in bladder [1], colon [2], brain [3], breast [4,5], esophagus [6], and skin [7,8]. Specifically, DRS from human skin reflect the contents of epidermal melanin, oxy-and deoxy-hemoglobin, carotenoids, lipids, etc. [9][10][11][12], with potential applications ranging from monitoring of skin inflammation [10], changes induced by exposure to ultraviolet light [11,13], aging of traumatic bruises [14], and evaluation of scars [15]. ...
Diffusion approximation (DA) of the radiative transport equation allows derivation of enclosed solutions for diffuse reflectance from multi-layer scattering structures, such as human skin. Although the DA is known to be inadequate near tissue boundaries and light sources, analytical tractability makes such solutions very attractive for use in noninvasive characterization of biological organs based on measured diffuse reflectance spectra (DRS). For the presented three-layer model of human skin, which enables a good match with DRS in visible spectral range measured with an integrating sphere, the DA solutions systematically overshoot numerically simulated DRS (using Monte Carlo approach) by 1–2 percentage points. However, using the former in inverse analysis of the latter can result in much larger artifacts, most notably overestimations of the melanin and blood contents by up to 15%, which must be considered when analyzing experimental DRS. Despite such systematic errors, the described approach allows simple and robust monitoring of physiological changes in human skin, as demonstrated in tests involving temporary obstruction of blood circulation and seasonal variations due to extensive sun exposure.
... The significant variance of tissue absorption and scattering coefficients stems from their complexity due to tissue composition and different content of endogenous chromophores [60]. This problem is typical for other label-free optically-based diagnostic instruments [61]. ...
One of the urgent tasks of modern medicine is to detect microcirculation disorder during surgery to avoid possible consequences like tissue hypoxia, ischemia, and necrosis. To address this issue, in this article, we propose a compact probe with sapphire tip and optical sensing based on the principle of spatially resolved diffuse reflectance analysis. It allows for intraoperative measurement of tissue effective attenuation coefficient and its alteration during the changes of tissue condition, caused by microcirculation disorder. The results of experimental studies using (1) a tissue-mimicking phantom based on lipid emulsion and hemoglobin and (2) a model of hindlimb ischemia performed in a rat demonstrated the ability to detect rapid changes of tissue attenuation confirming the feasibility of the probe to sense the stressful exposure. Due to a compact design of the probe, it could be useful for rather wide surgical operations and diagnostic purposes as an auxiliary instrument.
... Melanin pigments, a natural sunscreen in the skin, are the main determinants of skin color thanks to its ability to absorb, reflect and scatter the incident light, thus contributing to the perceived color [21]. Briefly, the enzyme tyrosinase is involved in the first two steps of melanogenesis, i.e., the hydroxidation of L-tyrosine and L-dihydroxyphenylalanine (L-DOPA) and the subsequent oxidation of L-DOPA to L-Dopaquinone. ...
Oral formulations with natural plant-based extracts represent a safe and promising strategy for skin lightening and anti-dark-spot effects, especially in Asia. This study evaluated the effect of an oral formulation including polyphenol-rich extracts and vitamin C (Belight3TM) on in vitro tyrosinase inhibitory activity and investigated its skin lightening and anti-dark-spot effects in vivo. Tyrosinase inhibitory activity of the formulation was measured with spectrophotometry. A randomized, double-blind, placebo-controlled clinical study was carried out on 58 healthy Asian males and females, aged 45–65. Skin color was measured at baseline, 6 weeks and 12 weeks with digital photographs. Color of dark spots was assessed with spectrophotometry. In vitro, the formulation showed a significant synergistic tyrosinase inhibitory activity of 85% compared to the control. In vivo, 12-week oral administration of the formulation significantly lightened the skin and was significantly better than the placebo. In addition, this formulation induced a slight and significant lightening effect of the dark spots after 6 and 12 weeks. Our findings suggest that the daily oral administration of Belight3TM during 12 weeks appears as an efficient and safe nutricosmetic to lighten the color of the facial skin and dark spots in Asian subjects.
... In the context of non-destructive optical characterization of biological tissues, Diffuse Reflectance Spectroscopy (DRS) has been widely used for several years [1][2][3][4] for cancer detection. It consists in back-reflected intensity spectra measurement. ...
The estimation of skin optical properties by means of inverse problem solving from spatially resolved diffuse reflectance (SR-DR) spectra is one way to exploit the acquired clinical signals. This method requires the comparison between the experimental spectra collected with a medical device, and spectra generated by the photons transport numerical simulations. This comparison is usually limited to spectral shape due to the absence of intensity standardization of the experimental DR spectra. This study proposes to theoretically (using photometric calculation) and experimentally (from experimental spectra acquired on optical phantom) establish a corrective factor to obtain common intensity unit for experimental and simulated signals.
... The present work is devoted to the study of the applicability of the diffuse reflection spectroscopy (DRS) method for determining the thickness of dentin. This technique is widely used in biophotonics to determine the composition and thickness of layers of biological tissues such as skin, organ surfaces, etc. [20][21][22][23][24]. Diffuse reflection spectroscopy and imaging were earlier applied for caries detection, dental calculus visualization and investigation of teeth tissues ex vivo [25][26][27]. ...
The modern approach to the treatment of caries requires maximum preservation of tooth tissues and pulp viability, for which it is necessary to know the residual thickness of dentin during its removal. Currently existing methods (Cone-beam Computed Tomography, electrical impedance device, and optical coherence tomography) are not widely used in clinical practice due to the laboriousness of their use or low accuracy. We evaluated the capabilities of the diffuse reflectance spectroscopy (DRS) method for determining dentine thickness in situ. Dentin tissues transmit light well in the visible and near-IR range, which makes it possible to detect the optical response of the dental pulp. The pulp contains hemoglobin and water, while dentin contains no hemoglobin, and its water content is less than 10%. Thus, the selection of the contributions of these components allows estimating the thickness of the dentin. Our results show a strong correlation (> 0.9) between dentin thickness and the amplitudes of the water and hemoglobin components. However, hemoglobin content is more susceptible to changes, caused by inflammation or the action of anesthesia. Thus, the most promising approach is use the water component as a proxy. The proposed method can be the basis for the development of a fiber-optic laser probe for clinical dentistry.
... Diffuse Reflectance Spectroscopy (DRS) applied to biological tissues is a widely studied [1][2][3][4], non-destructive optical characterization technique. It consists of measuring back-reflected intensity spectra carrying information from light-tissue interactions, from which the optical and structural properties of the probed medium can then be extracted and analyzed. ...
In the context of cutaneous carcinoma diagnosis based on in vivo optical biopsy, Diffuse Reflectance (DR) spectra, acquired using a Spatially Resolved (SR) sensor configuration, can be analyzed to distinguish healthy from pathological tissues. The present contribution aims at studying the depth distribution of SR-DR-detected photons in skin from the perspective of analyzing how these photons contribute to acquired spectra carrying local physiological and morphological information. Simulations based on modified Cuda Monte Carlo Modeling of Light transport were performed on a five-layer human skin optical model with epidermal thickness, phototype and dermal blood content as variable parameters using (i) wavelength-resolved scattering and absorption properties and (ii) the geometrical configuration of a multi-optical fiber probe implemented on an SR-DR spectroscopic device currently used in clinics. Through histograms of the maximum probed depth and their exploitation, we provide numerical evidence linking the characteristic penetration depth of the detected photons to their wavelengths and four source–sensor distances, which made it possible to propose a decomposition of the DR signals related to skin layer contributions.
... Together with skin structures, these chromophores absorb, reflect, and scatter the incident light, thus contributing to the perceived color. 2,3 Skindepigmenting agents target melanin and are based on different approaches, by modifying either its biosynthesis or its transfer from melanocytes into keratinocytes or its degradation rate. It must be mentioned that accelerating the epidermal turnover and desquamation by exfoliation is another way to achieve a skin-lightening effect by removing the outermost layers of keratinocytes with their melanin content. ...
Background
Glutathione has become a potential skin‐lightening ingredient after the discovery of its anti‐melanogenic properties. Various mechanisms of action have been considered to explain this property, one of them being the skewing of the melanin synthesis pathway towards the production of lighter pheomelanin instead of darker eumelanin, consequently producing a lightening effect.
Aims
To evaluate the skin lightening and anti‐dark spot effects of oral supplementation with L‐Cystine associated with L‐Glutathione as compared to placebo and benchmark.
Methods
Effects of this L‐Cystine‐L‐Glutathione oral combination were investigated in a 12‐week randomized, double‐blind, parallel‐group, benchmark‐ and placebo‐controlled trial involving 124 Asian female subjects. Women were randomly allocated into 4 equal groups (500 mg L‐Cystine and 250 mg L‐Glutathione, 250 mg reduced L‐Glutathione, 500 mg L‐Cystine, or a placebo, daily). Skin color was measured at baseline, after 6 and 12 weeks by spectrophotometry. Size and color of facial dark spots were determined from digital photographs.
Results
A significant skin lightening was observed after 12 weeks of oral supplementation with L‐Cystine associated with L‐Glutathione. This combination also induced a significant reduction in the size of facial dark spots after 6 and 12 weeks. It is noteworthy that the observed effects were not only significantly better than those obtained with placebo, but also with L‐Cystine alone or L‐Glutathione alone.
Conclusion
The daily oral administration of 500 mg L‐Cystine and 250 mg L‐Glutathione during 12 weeks was a safe treatment to effectively lighten the skin and reduce the size of facial dark spots of Asian women.
... Assessment of microcirculatory hemoglobin oxygen saturation and concentration of blood provides important information of local metabolism and its regulation in health and disease. In skin, epidermal melanin and carotenoids are present affecting foremost short wavelengths, 1 whereas for higher wavelengths water and lipid absorption is significant. 2 The calculation of tissue chromophores, which is most often the aim for DRS methods, is commonly done using inverse modeling based on diffusion theory [3][4][5] or Monte Carlo techniques, 6,7 where modeled DRS data are fitted to measured *Address all correspondence to Ingemar Fredriksson, ingemar.fredriksson@liu.se ...
Significance:
Diffuse reflectance spectroscopy (DRS) is frequently used to assess oxygen saturation and hemoglobin concentration in living tissue. Methods solving the inverse problem may include time-consuming nonlinear optimization or artificial neural networks (ANN) determining the absorption coefficient one wavelength at a time.
Aim:
To present an ANN-based method that directly outputs the oxygen saturation and the hemoglobin concentration using the shape of the measured spectra as input.
Approach:
A probe-based DRS setup with dual source-detector separations in the visible wavelength range was used. ANNs were trained on spectra generated from a three-layer tissue model with oxygen saturation and hemoglobin concentration as target.
Results:
Modeled evaluation data with realistic measurement noise showed an absolute root-mean-square (RMS) deviation of 5.1% units for oxygen saturation estimation. The relative RMS deviation for hemoglobin concentration was 13%. This accuracy is at least twice as good as our previous nonlinear optimization method. On blood-intralipid phantoms, the RMS deviation from the oxygen saturation derived from partial oxygen pressure measurements was 5.3% and 1.6% in two separate measurement series. Results during brachial occlusion showed expected patterns.
Conclusions:
The presented method, directly assessing oxygen saturation and hemoglobin concentration, is fast, accurate, and robust to noise.
... DRS has been used in investigating divergent skin signs associated with cutaneous diseases [18,[65][66][67]. DRS systems were implemented utilizing multiple optical configurations including the integrating sphere (IS)-based DRS [68][69][70][71] and the spatially resolved steady-state DRS [72][73][74]. ...
Abstract
Background
Skin erythema may present due to many causes. One of the common causes is prolonged exposure to sun rays. Other than sun exposure, skin erythema is an accompanying sign of dermatological diseases such as acne, psoriasis, melasma, post inflammatory hyperpigmentation, fever, as well as exposure to specific electromagnetic wave bands.
Methods
Quantifying skin erythema in patients enables the dermatologist to assess the patient’s skin health. Therefore, quantitative assessment of skin erythema was the target of several studies. The clinical standard for erythema evaluation is visual assessment. However, the former standard has some imperfections. For instance, it is subjective, and unqualified for precise color information exchange. To overcome these shortcomings, the past three decades witnessed various methodologies that aimed to achieve erythema objective assessment, such as diffuse reflectance spectroscopy (DRS), and both optical and non-optical systems.
Discussion
This review article revises the studies published in the past three decades where the performance, the mathematical tactics for computation, and the limited capabilities of erythema assessment techniques for cutaneous diseases are discussed. In particular, the current achievements and limitations of the current techniques in erythema assessment are presented.
Conclusion
The profits and development trends of optical and non-optical methods are displayed to provide the researcher with awareness into the present technological advances and its potential for dermatological diseases research.
Keywords
Skin pigments; Erythema assessment; Dermatological diseases; Skin inflammation; Optical diagnosis
... Diffuse reflectance spectroscopy (DRS) measurement is a popular optical technique. It is used for monitoring various apparent cutaneous symptoms (17)(18)(19). The apparent symptoms usually change the concentration of pigments as well as the skin color tone. ...
... Diffuse reflectance spectroscopy (DRS) measurements is an erythema assessment tool with better spectral resolution. Unlike colorimetry and digital photography, DRS discriminates between skin color changes based on the variation in the skin's chromophores spectral profile (15,(27)(28)(29)(30)(31)(32)(33)(34). The spectral profile provides quantitative information out of the skin's chromophores concentration. ...
Surveillance and assessment of radiation-induced erythema is an important aspect of managing skin toxicity in radiation therapy treated patients. Upon receiving the early fractions of radiation, an inflammatory response and vascular dilation takes place due to damage of basal cells in the skin’s epidermal layer. This process of skin reddening known as erythema. The gold standard used for assessing and grading erythema is visual assessment (VA) by an experienced clinician/ radiotherapist using toxicity scoring tools. This method is limited by the assessor’s experience, vision acuity, and the subjectivity of qualitative scores. An alternative optical technique to VA, is diffuse reflectance spectroscopy (DRS). A comparison between both techniques performance in detecting radiation therapy-induced erythema is demonstrated in this pilot study. The results evidenced that DRS is capable of detecting skin erythema before an expert eye could do so.
... DRS is an analytical tool for investigation of optical properties of absorbed molecules to measure the scattering and absorption properties of the skin in which a beam of light penetrates into the skin. The detail for method of analysis is described by Kollias et al [7][8][9]. Briefly, the system consisted of a HL-2000 tungsten halogen light source (Dunedin, Florida, USA), a bifurcated optical fiber probe and a BWTEK BTC112E spectrometer (Newark, DE, USA) with a spectral range from 300nm-850nm and a 2 nm resolution. ...
Objective
This paper presents in vivo an in vitro studies demonstrating the induction of pigmentation in human skin by visible light which can be blocked by using formulation containing the correct amount of yellow iron oxide (YIO).
Methods
An in vitro absorption method was developed to determine the protection provided by a test formulation containing 4.5% YIO using an IPD UVA‐VIS action spectrum. Following the development of the in vitro method and in vivo study with 10 normal healthy volunteers with Fitzpatrick skin phototypes IV to VI was conducted to verify if the predictive model.
Results
The in vitro model for visible light protection provided a protection factor of 2.5 using the in vitro absorption spectrum of 4.5% of YIO with a very similar result from the in vivo study with a protection factor of 3.0. Multiple daily exposures of visible light have shown increase in skin pigmentation and the application of YIO provide less development of pigmentation when compared to unprotected skin.
Conclusion
In vitro testing of the absorbance of the pigmented formulation using a proposed action spectrum for immediate pigment darkening (IPD) response in the visible light range supports the in vivo protection observations for persistent pigment darkening (PPD) and can be used as predictor for skin pigmentation induced by visible light.
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... DRS has been used in investigating divergent skin signs associated with cutaneous diseases [21], [73]- [75]. DRS systems were implemented utilizing multiple optical configurations including the integrating sphere (IS)-based DRS [76]- [79] and the spatially resolved steady-state DRS [80]- [82]. ...
... The investigations show a variety of advanced biomedical technologies, which are in direct need of knowledge of the optical parameters of tissues. As the most impressive and useful examples of tissue quantified data application, there are the dosimetry of radiation during PDT and PTT 27,28 and in laser surgery, 29 various imaging technologies, 1,2,19,[30][31][32][33][34][35][36][37][38][39][40] the development of batteryless solar-powered cardiac pacemakers, 41 standardization of tissue-mimicking phantoms, 32,42-46 assessment of tissue chromophore concentration and distribution, 47,48 and proposition of adequate optical models of tissues. [49][50][51][52] This paper gives an overview of recent results on the measurements and control of tissue optical properties that were presented on the VI International Symposium "Topical Problems of Biophotonics 2017," St.-Petersburg, Nizhny Novgorod, Russia, July 28 to August 03, 2017, as an invited talk. ...
Nowadays, dynamically developing optical (photonic) technologies play an ever-increasing role in medicine. Their adequate and effective implementation in diagnostics, surgery, and therapy needs reliable data on optical properties of human tissues, including skin. This paper presents an overview of recent results on the measurements and control of tissue optical properties. The issues reported comprise a brief review of optical properties of biological tissues and efficacy of optical clearing (OC) method in application to monitoring of diabetic complications and visualization of blood vessels and microcirculation using a number of optical imaging technologies, including spectroscopic, optical coherence tomography, and polarization- and speckle-based ones. Molecular modeling of immersion OC of skin and specific technique of OC of adipose tissue by its heating and photodynamic treatment are also discussed.
... In 1985, Kollias et al described an instrument and method to quantify non-invasively melanin in human skin by measuring the difference in the remittance spectra of normal and vitiligo-involved skin assuming that the only difference was the melanin filter 4 . Again, this represented pioneering work in the field of photobiology and set off a series of papers on the melanin absorption spectrum after application of UVB, UVA and visible light 5-8 as well as several studies that elucidated the skin response to various challenges using diffuse reflectance spectroscopy (DRS) [9][10][11] . Importantly, Nik and his collaborators used FExS to study non-melanoma skin cancers (NMSC) 12 which includes basal cell carcinoma and squamous cell carcinomas and thus represents the most common malignancy worldwide. ...
In a paper published at the J Invest Dermatol in 1998 Nik Kollias and coworkers described distinct changes in skin native fluorescence associated with skin aging and photoaging, using in vivo fluorescence excitation spectroscopy. The assignment of the 295 nm band to tryptophan fluorescence had a profound significance influencing many later studies from multiple groups. The reproducible changes in skin native fluorescence suggested that aging causes predictable alterations in both the epidermis and the dermis, whereas chronic UV exposure induces the appearance of new fluorophores. This seminal, but insufficiently widely appreciated work deserves re-examination as it points to important horizons in future experimental dermatology, such as cancer diagnostics, diabetes, wound healing, and understanding skin aging and photoaging mechanisms. This article is protected by copyright. All rights reserved.
... Deeper melanin can be analyzed once more superficial melanin and hemoglobin are removed [56]. Skin erythema from surfactant irritation, histamine release, ultraviolet exposure, and reactive hyperemia have been quantified [58]. p0255 ...
The aim of this chapter is to examine image processing for skin conditions versus normal skin and engage readers from the clinical, healthcare, educational, research, and industrial settings. Dermatology is broadly defined to encompass skin diseases, skin cancers, photodamage, wounds, scars, skin restoration, cosmetics, and environmental effects. Contributions from academic medical, bioengineering, and skin-care industry researchers provide an overview of important concepts, research, and development. “Image processing” depends upon the particular purpose and the specific imaging modality being applied. The number and types of image processing techniques have increased considerably. To put them in context, varied objectives of skin imaging, skin imaging techniques, and the most up-to-date technologies are presented in this chapter. Image processing is discussed as an overview and specific examples illustrate various techniques for purpose of diagnosis and quantifying with change in condition over time. The vantage point also considers emerging methods, new applications, opportunities, and unmet needs.
... DRS is a non-invasive assessing tool to determine changes of chromophores in skin [42,43] and we employed DRS for both in vivo and ex-vivo studies. The system consisted of a light source (Xe arc lamp Newport, Stratford, CT), a bifurcated optical fiber probe (Multimode Fiber Optics, Inc., NJ) and a spectrometer (PIXIS 256, Princeton Instruments, Trenton, NJ). ...
Visible light (400-700 nm) lies outside of the spectral range of what photobiologists define as deleterious radiation and as a result few studies have studied the effects of visible light range of wavelengths on skin. This oversight is important considering that during outdoors activities skin is exposed to the full solar spectrum, including visible light, and to multiple exposures at different times and doses. Although the contribution of the UV component of sunlight to skin damage has been established, few studies have examined the effects of non-UV solar radiation on skin physiology in terms of inflammation, and limited information is available regarding the role of visible light on pigmentation. The purpose of this study was to determine the effect of visible light on the pro-pigmentation pathways and melanin formation in skin. Exposure to visible light in ex-vivo and clinical studies demonstrated an induction of pigmentation in skin by visible light. Results showed that a single exposure to visible light induced very little pigmentation whereas multiple exposures with visible light resulted in darker and sustained pigmentation. These findings have potential implications on the management of photo-aggravated pigmentary disorders, the proper use of sunscreens, and the treatment of depigmented lesions.
... [16][17][18][19] Spectroscopy-based methods, such as reflectance spectroscopy and hyperspectral imaging, are the most complex of the methods used for measuring skin color. [20][21][22][23][24] Spectroscopy provides quantitative data across a range of wavelengths, allowing for different parameters to be extracted from its measurements, depending on the scope of the investigation and the apparatus used. User-friendly commercial models capable of monitoring relative erythema and tissue oxygen saturation are expensive and use single-use detection probes. ...
The measurement of changes in blood volume in tissue is important for monitoring the effects of a wide range of therapeutic interventions, from radiation therapy to skin-flap transplants. Many systems available for purchase are either expensive or difficult to use, limiting their utility in the clinical setting. A low-cost system, capable of measuring changes in tissue blood volume via diffuse reflectance spectroscopy is presented. The system consists of an integrating sphere coupled via optical fibers to a broadband light source and a spectrometer. Validation data are presented to illustrate the accuracy and reproducibility of the system. The validity and utility of this in vivo system were demonstrated in a skin blanching/reddening experiment using epinephrine and lidocaine, and in a study measuring the severity of radiation-induced erythema during radiation therapy. (C) 2014 Society of Photo-Optical Instrumentation Engineers (SPIE)
... Skin color is primarily dominated by the presence of chromophores such as melanin, hemoglobin, bilirubin and carotene. 1,2 Those chromophores can be altered in concentration either by external insults such as chemical irritants, UV radiation or by changes of physiological environment. Colorimetry has been used as an objective measure of perceived skin color by human eye to document and score physiological responses of the skin from the various external insults. ...
Colorimetry has been used as an objective measure of perceived skin
color by human eye to document and score physiological responses of the
skin from external insults. CIE color space values (L*, a* and b*) are
the most commonly used parameters to correlate visually perceived color
attributes such as L* for pigment, a* for erythema, and b* for
sallowness of the skin. In this study, we investigated the relation of
Lab color scale to the amount of major skin chromophores (oxy-,
deoxyhemoglobin and melanin) calculated from diffuse reflectance
spectroscopy. Thirty two healthy human subjects with ages from 20 to 70
years old, skin types I-VI, were recruited for the study. DRS and
colorimetry measurements were taken from the left and right cheeks, and
on the right upper inner arm. The melanin content calculated from
630-700 nm range of DRS measurements was shown to correlate with the
lightness of skin (L*) for most skin types. For subjects with
medium-to-light complexion, melanin measured at the blue part spectrum
and hemoglobin interfered on the relation of lightness of the skin color
to the melanin content. The sallowness of the skin that is quantified by
the melanin contribution at the blue part spectrum of DRS was found to
be related to b* scale. This study demonstrates the importance of
documenting skin color by assessing individual skin chromophores with
diffuse reflectance spectroscopy, in comparison to colorimetry
assessment.
... 74,75 Because of the presence of melanin, SIA has also been applied to quantify the effects of long-term UV exposure leading to hyperpigmented spots, and to evaluate the perception of age, health status, and common skin conditions. 73,76,77 Skin Elasticity and Laxity Exposure to UV radiation also results in dermal changes, as evidenced by reduced levels of types I and III collagen, increased elastin, and poor organization of the collagen fibrils. These effects are manifested in the loss of tissue elasticity and slower recovery from applied mechanical stress. ...
The purpose of this article is to review the strategies and methods for quantifying treatment outcomes, perhaps defined by the consumer/patient as a "decrease in perceived age." The demand for the rejuvenation of facial skin is expected to increase as the population ages and seeks optimal outcomes from the array of available treatment modalities. This information will be of value to the plastic surgeon in collaborating with patients on evaluation and treatment strategies.
... 4 An accurate assessment of cutaneous microcirculation is still challenging, because probing small volumes of skin with variable content of melanin precludes the use of models based on diffusion approximation to quantify oxy-and deoxyhemoglobin that are pulsating or steady in the same volume of interrogation. In addition, from two independent in-vivo studies for as-sessing UV-induced erythema, different dose-response relations for blood concentration measured from time-averaged diffuse reflectance spectroscopy (DRS) 5 and blood perfusion measured from laser Doppler velocimeters 6 have been found. In this work, we present a simple and cost-effective functional diffuse reflectance spectroscopy (fDRS) system that enables us to study hemodynamics in the papillary dermal vasculature, where the contribution to the signal at the "superficial" vascular plexus is dominant. ...
We present a simple and cost-effective optical technique for the simultaneous assessment of pulsating and total blood noninvasively in an inflammatory skin lesion. Acquisitions of diffuse reflectance spectra in the visible range at 6 Hz are used to trace the oscillating components of reflectance. Measurements on erythematous lesions from a UV insult show slow changing signal at about 0.1 Hz and heart-driven regular oscillations at about 1 Hz simultaneously. The results demonstrate the potential of the technique in monitoring both pulsating and steady components of the blood in inflammatory lesions of the skin.
... SIA has also been applied to the quantitation of the effects of long term ultraviolet exposure on facial skin, e.g., hyperpigmented spots and to evaluate the perception of age and health status. 83 The applicability of SIA for the evaluation of common skin conditions has been demonstrated by Kollias et al. 84 Changes in erythema could be used to quantify the irritant response to sodium lauryl sulfate, inflammation due to histamine release, changes in blood flow (by application of pressure), inflammation induced by ultraviolet exposure and for conditions of reactive hyperemia. Their findings suggest applicability of this technique for the evaluation of common skin conditions including irritant contact dermatitis, allergic dermatitis, and pressure induced injury. ...
Skin imaging modalities relevant to the range of skin conditions encountered in clinical settings are described with respect to the information provided, advantages and limitations, current status and indications for further development. The methods use the interaction of energy with the skin, penetrating to various depths in the stratum corneum, epidermis, dermis and subcutaneous layers. They include a detection system such as the retina, film or a digital array, and a processing system to deconstruct, analyze and interpret the information. Similarly, the areas of interest, or targets, have common features. The skin conditions deviate from the ideal or normal state with respect to skin integrity and function. The deviations include evidence of barrier disruption, inflammation, dispigmentation, and vascular change. The user of skin imaging is often interested in the extent and severity of disease. Part of the task in skin imaging is to establish the criteria for the normal condition. The review encompasses the past, present and future of visual assessment, photographic image collection, spectrophotometric techniques, noninvasive histology, and three dimensional scanning. The analytical techniques for processing and extracting specific parameters that inform about the underlying biological status are presented.
Skin erythema may present due to many causes. One of the common causes is prolonged exposure to sunrays. Other than sun exposure, skin erythema is an accompanying sign of dermatologic diseases, such as psoriasis and acne. Quantifying skin erythema in patients enables the dermatologist to assess the patient's skin health. Quantitative assessment of skin erythema has been the target of several studies. The clinical standard for erythema evaluation is visual assessment; however, the former standard has some imperfections. For instance, it is subjective, and unqualified for precise color information exchange. To overcome these shortcomings, the past three decades has witnessed various methodologies that aimed to achieve erythema objective assessment, such as diffuse reflectance spectroscopy (DRS), and both optical and non-optical systems. This review appraises the studies published in the past three decades, where the performance, the mathematical tactics for computation, and the limited capabilities of erythema assessment techniques for cutaneous diseases are discussed. The current achievements and limitations of the current techniques in erythema assessment are presented. The profits and development trends of optical and non-optical methods are displayed to provide the researcher with awareness into the present technological advances and its potential for dermatological diseases research.
Background
The main chromophores of human skin are melanins and hemoglobins along with carotenoids, bilirubin and other compounds. In an effort to study the spectral signatures of skin melanin we measured absorption spectra in a variety of situations, including a method to show early signs of re‐pigmentation in vitiligo.
Methods
To measure skin in vivo, the essential component was a “Bifurcated Optical Fiber” with one end connected to the light source and the second end connected to the spectrometer while the common end was placed on the skin.
Results
In a typical in situ “melanin in skin” spectrum, the Absorbance Values first rise gradually, from 750 nm to 600 nm, then rise moderately from 600 nm to 450 nm, and rise sharply from 450 nm to a broad peak at 335 nm, below which it gradually rolls down to much lower values.
Conclusion
We successfully studied melanin spectroscopically in subjects with vitiligo lesions, obtaining the differential spectra. Higher melanin levels can be shown by steeper negative slopes of a straight line fitted between 620 nm to 720 nm. Also, absorption peak at 335 nm showed the presence of melanin.
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The ability to monitor changes in the concentration of hemoglobin in the blood of the skin in real time is a key component to personalized patient care. Since hemoglobin has a unique absorption spectrum in the visible light range, diffuse reflectance spectroscopy is the most common approach. Although the collection of the diffuse reflectance spectrum with an integrating sphere (IS) has several calibration challenges, this collection method is sufficiently user-friendly that it may be worth overcoming the initial difficulty. Once the spectrum is obtained, it is commonly interpreted with a log-inverse-reflectance (LIR) or "absorbance" analysis that can only accurately monitor changes in the hemoglobin concentration when there are no changes to the nonhemoglobin chromophore concentrations which is not always the case. We address the difficulties associated with collection of the diffuse reflectance spectrum with an IS and propose a model capable of retrieving relative changes in hemoglobin concentration from the visible light spectrum. The model is capable of accounting for concentration changes in the nonhemoglobin chromophores and is first characterized with theoretical spectra and liquid phantoms. The model is then used in comparison with a common LIR analysis on temporal measurements from blanched and reddened human skin. (C) 2015 Society of Photo-Optical Instrumentation Engineers (SPIE)
Prior to the introduction of reflectance spectrophotometry into anthropological field research during the 1950s, human skin color was most commonly classified by visual skin color matching using the von Luschan tiles, a set of 36 standardized, opaque glass tiles arranged in a chromatic scale. Our goal was to establish a conversion formula between the tile-based color matching method and modern reflectance spectrophotometry to make historical and contemporary data comparable. Skin pigmentation measurements were taken on the forehead, inner upper arms, and backs of the hands using both the tiles and a spectrophotometer on 246 participants showing a broad range of skin pigmentation. From these data, a second-order polynomial conversion formula was derived by jackknife analysis to estimate melanin index (M-index) based on tile values. This conversion formula provides a means for comparing modern data to von Luschan tile measurements recorded in historical reports. This is particularly important for populations now extinct, extirpated, or admixed for which tile-based measures of skin pigmentation are the only data available. Am J Phys Anthropol, 2013. © 2013 Wiley Periodicals, Inc.
Multispectral imaging (MSI) is becoming a powerful tool for tissue abnormality detection. Conventional MSI systems, however, are not readily suitable for challenges of routine clinical uses due to the fact that they are expensive, bulky, and time consuming to acquire the data. In this letter we report a novel approach to instrument MSI technology into a handheld, low-cost, standing-alone, real-time operational device that is suitable for home-based health care. It covers techniques used to produce multiple images at discrete signature wavelengths of tissues with a single shot.
Coherent anti-Stokes Raman scattering (CARS) microscopy is a label-free imaging technique that is capable of real-time, nonperturbative examination of living cells and organisms based on molecular vibrational spectroscopy. Recent advances in detection schemes, understanding of contrast mechanisms, and developments of laser sources have enabled superb sensitivity and high time resolution. Emerging applications, such as metabolite and drug imaging and tumor identification, raise many exciting new possibilities for biology and medicine.
The non-invasive determination of deep tissue three dimensional structure and biochemistry is the ultimate goal of optical biopsy. Two-photon microscopy has been shown to be a particularly promising approach. The use of infrared radiation in two-photon microscopy is critical for deep tissue imaging since tissue absorption and scattering coefficients for infrared light are much lower than for shorter wavelengths. Equally important, tissue photodamage is localized to the focal region where fluorescence excitation occurs. This report demonstrates that, by means of high resolution two-photon microscopy, skin and subcutaneous tissue structures can be imaged utilizing their endogenous fluorescence. From a freshly prepared tissue punch of a mouse ear, we were able to resolve in 3D both the living and cornified keratinocytes in the epidermis, the collagen/elastin fibers in the dermal layer and the cartilage in the subcutaneous layer. The ability to non-invasively acquire 3D structures of these tissue components may find application in areas such as non-invasive diagnosis of skin cancer and the study of wound healing processes.
A noninvasive tool for skin tumor diagnosis would be a useful clinical adjunct. The purpose of this study was to determine whether near-infrared spectroscopy can be used to noninvasively characterize skin lesions. In vivo visible- and near-infrared spectra (400--2500 nm) of skin neoplasms (actinic keratoses, basal cell carcinomas, banal common acquired melanocytic nevi, dysplastic melanocytic nevi, actinic lentigines, and seborrheic keratoses) were collected by placing a fiberoptic probe on the skin. Paired t tests, repeated measures analysis of variance and linear discriminant analysis were used to determine whether significant spectral differences existed and whether spectra could be classified according to lesion type. Paired t tests showed significant differences (p < 0.05) between normal skin and skin lesions in several areas of the near-infrared spectrum. In addition, significant differences were found between the lesion groups by analysis of variance. Linear discriminant analysis classified spectra from benign lesions compared with premalignant or malignant lesions with high accuracy. Near-infrared spectroscopy is a promising noninvasive technique for the screening of skin lesions.
The chromatic characteristics of skin color arise from the interactions of light (primarily absorption and scattering) with the epidermis and the dermis. The primary light absorbers in skin are hemoglobin and melanin. Most of scattering is attributed to collagen fibers and in pigmented skin to melanosomes. Traditionally skin redness is considered to arise due to locally elevated concentrations of hemoglobin, whereas skin pigmentation is attributed to melanin. In this study we attempt to understand better the contributions of these chromophores to the perceived skin color using spectral analysis of skin color reactions induced by ultraviolet (UV) irradiation or pressure. In the first experiment 12 individuals with skin phototypes III-IV were irradiated on the back using a solar simulator with doses ranging from 0.7 to 3 MED. The skin reactions were evaluated on days 1, 7, 14, and 21 after irradiation. Evaluations included diffuse reflectance spectroscopy (DRS) and clinical assessment of the erythema and the pigment reaction. Apparent concentrations of melanin, oxy-, and deoxy-hemoglobin were calculated from the absorption spectra. In the second experiment the levels of deoxy-hemoglobin of the volar forearm of ten volunteers were selectively altered by either application of a pressure cuff or by topical application of 3% H(2)O(2). Changes in skin color appearance were documented by photography, colorimetry, and DRS. In the UV exposure experiment all reactions were dose dependent. Oxy-hemoglobin values increased to a maximum on day 1, correlating well with the clinical evaluation of erythema, and then decreased exponentially to base line. Melanin showed a significant increase on day 7 and remained relatively constant for the next 3 weeks, correlating well with the clinical evaluation of pigmentation (tanning). Deoxy-hemoglobin increased slightly on day 1 and remained elevated for the next 2 weeks. Thus, deoxy-hemoglobin correlated moderately with the clinical erythema scoring on day 1 only, while it contributes significantly to what is clinically perceived as skin tanning on days 7 and 14. Application of pressure below the diastolic level increased deoxy-hemoglobin concentration as measured by DRS. This increase corresponded to a decrease of a "pigmentation" parameter (based on the L(*)a(*)b(*) scale) in a similar fashion that has been documented for increases in melanin concentration. Topical H(2)O(2) application reduced deoxy-hemoglobin levels as measured by DRS. This reduction coincided kinetically with a visible skin blanching. Application of pressure or H(2)O(2) did not significantly alter the levels of oxy-hemoglobin or melanin. In this report we present compelling evidence that deoxy-hemoglobin significantly contributes to the skin color appearance. Blood pooling, expressed as increased deoxy-hemoglobin, can contribute to what is visually perceived as pigmentation. Furthermore, we present that measurement of its contribution to the skin color appearance can only be accomplished with DRS.
We present the results of diffuse reflectance measurements made on the surface of a tissue-simulating phantom containing intact human erythrocytes. These measurements indicate that the absorption spectrum of hemoglobin in its natural environment is significantly different from that measured in homogeneous fluid solution, especially in the spectral regions of highest absorption. We show that this difference can be explained by the pigment packaging theory developed by Duysens [Biochim. Biophys. Acta 19, 1 (1956)] and that the adoption of basis spectra that take this effect into account improves the accuracy of fitting diffuse reflectance spectra.
Imaging living organisms with molecular selectivity typically requires the introduction of specific labels. Many applications in biology and medicine, however, would significantly benefit from a noninvasive imaging technique that circumvents such exogenous probes. In vivo microscopy based on vibrational spectroscopic contrast offers a unique approach for visualizing tissue architecture with molecular specificity. We have developed a sensitive technique for vibrational imaging of tissues by combining coherent anti-Stokes Raman scattering (CARS) with video-rate microscopy. Backscattering of the intense forward-propagating CARS radiation in tissue gives rise to a strong epi-CARS signal that makes in vivo imaging possible. This substantially large signal allows for real-time monitoring of dynamic processes, such as the diffusion of chemical compounds, in tissues. By tuning into the CH2 stretching vibrational band, we demonstrate CARS imaging and spectroscopy of lipid-rich tissue structures in the skin of a live mouse, including sebaceous glands, corneocytes, and adipocytes, with unprecedented contrast at subcellular resolution.
• nonlinear microscopy
• vibrational imaging
• back scattering
Tissue inflammation is often accompanied by local interstitial fluid accumulation expressed as edema. Edema can be the manifestation of infection, lymphatic blockage, wound healing, or even cancer, and is typically graded visually. Here we demonstrate that the edema reaction can be objectively quantitated in vivo by the use of spectral imaging. To this end we applied the method on a histamine-induced cutaneous edema model. Apparent concentrations of oxy-hemoglobin, deoxy-hemoglobin, and water were calculated for each pixel of a spectral image stack. These values were used to construct concentration maps for each of these molecules as well as an intensity map of an optical tissue-scattering parameter. The oxy-hemoglobin and the tissue water maps are two-dimensional quantitative representations of the skin areas involved in erythema and edema, respectively. These maps demonstrated characteristics of the wheal-and-flare reaction and their gray-level intensities were dependent on the applied histamine dose. We conclude that spectral imaging can be a valuable noninvasive tool in the study of edema pathology and can be used to monitor the edema reaction in vivo or follow the efficacy of treatments in a clinical setting.
The deep tissue penetration and submicron spatial resolution of multiphoton microscopy and the high detection efficiency and nanometer spectral resolution of a spectrograph were utilized to record spectral images of the intrinsic emission of mouse skin tissues. Autofluorescence from both cellular and extracellular structures, second-harmonic signal from collagen, and a narrowband emission related to Raman scattering of collagen were detected. Visualization of the spectral images by wavelength-to-RGB color image conversion allowed us to identify and discriminate tissue structures such as epidermal keratinocytes, lipid-rich corneocytes, intercellular structures, hair follicles, collagen, elastin, and dermal cells. Our results also showed morphological and spectral differences between excised tissue section, thick excised tissue, and in vivo tissue samples of mouse skin. Results on collagen excitation at different wavelengths suggested that the origin of the narrowband emission was collagen Raman peaks. Moreover, the oscillating spectral dependency of the collagen second-harmonic intensity was experimentally studied. Overall, spectral imaging provided a wealth of information not easily obtainable with present conventional multiphoton imaging systems.
Objective in situ measurements of skin pigmentation are needed for accurate documentation of pigmentation disorders, in studies of constitutive and induced skin pigmentation, for testing of the efficacy of pro-pigmentation or de-pigmentation agents, etc. Non-invasive instrumental measurements of skin pigmentation have been used for many decades. All are based on the ability of melanin to attenuate light. However, hemoglobin in dermal capillaries also attenuates light and needs to be accounted for when pigmentation is assessed. The methods under consideration include: (a) single point measurements, in which light reflected from a defined skin area is collected and a pigment index is calculated representing the average pigmentation over the examined area, and (b) imaging methods that attempt to generate a concentration distribution map of melanin pigment for the skin area being imaged. In this article, we describe the potentials and the limitations of the different approaches to both single point and imaging methods.
A combination of near-infrared spectroscopy and discrete wavelength near-infrared imaging is used to noninvasively monitor the forearm during periods of restricted blood outflow (venous outflow restriction) and interrupted blood inflow (ischemia), Multivariate analysis of image and spectral data time courses was used to identify highly correlated spectral and regional domains, while fuzzy C-means clustering of image time courses was used to reveal finer regional heterogeneities in the response of stressed tissues, Localized near-infrared spectroscopy was used to investigate the magnitude of the bulk changes in the tissue optical properties and the variation in tissue oxygenation saturation during venous outflow restriction and complete forearm ischemia, The imaging and spectroscopic analyses revealed highly localized regional variations in tissue oxygen saturation during forearm ischemia as compared to the more diffuse and global response of the forearm during venous outflow restriction.
The dose-response relationship in patch testing with sodium lauryl sulphate (SLS) was studied. The irritant skin response was quantified by visual scoring as well as by the following noninvasive methods: measurement of transepidermal water loss (TEWL) by an evaporimeter, measurement of skin color by a colorimeter, measurement of superficial blood flow by laser Doppler flowmetry, and measurement of edema in the skin by ultrasound A-scan. Twelve volunteers were patch tested with 0.12, 0.25, 0.50, and 1.00% SLS, and the skin response was evaluated after 24 and 48 h, respectively. We found a statistically significant linear dose-response relationship between dose of SLS and skin response evaluated by measurement of TEWL, skin color, superficial blood flow, and edema. Statistical evaluation by regression analysis proved measurement of TEWL to be the method best suited overall for quantification in relation to patch testing with SLS, whereas colorimetry was found to be the least sensitive of the applied methods. Ultrasound A-scan was found to be a promising method for quantification of the inflammatory response, being consistently more sensitive than measurement of skin color.
In this paper we present methods that we have developed to measure pigmentation in human skin. This involves the measurement of diffuse reflection spectra from human skin in vivo and referencing them to either totally depigmented skin, or the skin of the same individual if we are measuring variations in pigmentation. Changes in the pigmentary system brought about by UV radiation can be measured for each individual. Absolute measurements lead to estimates of the melanin concentration in the skin, while differential measurements lead to estimates of the quantity of additional or reduced pigment content of some skin lesions. The same instrumentation has been successfully used to assess UV-induced erythema as well as other vascular changes. The determination of the minimum detectable erythema dose can be performed even in the darkest-skinned subjects without loss of sensitivity as in the case of laser Doppler instruments. It has been shown that what is perceived by the eye as erythema is a very complex phenomenon, encompassing a large number of vascular reactions that can be studied in detail through diffuse reflection spectroscopy. Some of the possible responses are presented, as well as the contributing chromophores that have been identified so far.
In this paper we present the absorption characteristics of human melanin in the visible range of wavelengths and specifically in the range 620-720 nm. The spectroscopy of melanin is studied by measuring remittance spectra of normal skin and vitiligo-involved skin of volunteers-patients. It is assumed that the spectral differences between adjacent areas of normally pigmented skin, and to some degree amelanotic skin, can only be due to the variations of the melanin filter. The ratio of the remittance spectrum of the vitiligo-involved skin with the spectrum of the normal skin in the range 620-720 nm can be fitted with a straight line for all the volunteers. A very strong correlation is obtained between the intercept and the slope for all the volunteers, which leads us to conclude that it is indeed melanin that we are measuring in all the volunteers and that it is the same substance spectroscopically for all the volunteers.
Reflectance spectrophotometry was used to observe the degree of vasoconstriction induced by topically applied corticosteroids. The average blanching effectiveness of the corticosteroid preparations as determined by this technique was the same as that found by other workers using subjective assessments. Reflectance spectrophotometry can be utilized to quantify the degree of vasoconstriction produced by local corticosteroids applied to the skin and is particularly suitable for measuring the response of individual subjects.
An integrated review of the transfer of optical radiation into human skin is presented, aimed at developing useful models for photomedicine. The component chromophores of epidermis and stratum corneum in general determine the attenuation of radiation in these layers, moreso than does optical scattering. Epidermal thickness and melanization are important factors for UV wavelengths less than 300 nm, whereas the attenuation of UVA (320-400 nm) and visible radiation is primarily via melanin. The selective penetration of all optical wavelengths into psoriatic skin can be maximized by application of clear lipophilic liquids, which decrease regular reflectance by a refractive-index matching mechanism. Sensitivity to wavelengths less than 320 nm can be enhanced by prolonged aqueous bathing, which extracts urocanic acid and other diffusible epidermal chromophores. Optical properties of the dermis are modelled using the Kubelka-Munk approach, and calculations of scattering and absorption coefficients are presented. This simple approach allows estimates of the penetration of radiation in vivo using noninvasive measurements of cutaneous spectral remittance (diffuse reflectance). Although the blood chromophores Hb, HbO2, and bilirubin determine dermal absorption of wavelengths longer than 320 nm, scattering by collagen fibers largely determines the depths to which these wavelengths penetrate the dermis, and profoundly modifies skin colors. An optical "window" exists between 600 and 1300 nm, which offers the possibility of treating large tissue volumes with certain long-wavelength photosensitizers. Moreover, whenever photosensitized action spectra extend across the near UV and/or visible spectrum, judicious choice of wavelengths allows some selection of the tissue layers directly affected.
Analytical modeling that interrelates the optical properties of multilayered structures is applied to the skin. The mathematical approach is based on relations of diffuse reflectance and transmittance of a multilayered system and the diffuse reflectance and transmittance of each component layer. The formula can also be derived from the Kubelka-Munk theory of radiation transfer. Using both collimated and diffuse incident irradiance, the applicability of the model to human epidermis over the UV and visible region has been verified. The model has been applied to calculate to absorption and scattering coefficients of human epidermis in vitro, and to estimate the epidermal transmittance under simulated in vivo condition.
A theoretical treatment has been developed for the optical properties of a layered structure which absorbs and scatters light. This theory predicts that the logarithm of the inverse of reflectance (LIR) of the surface should be a useful parameter for the examination of that structure. This approach has been applied to a study of skin in vivo. An instrument was constructed for use in clinical situations to measure the LIR spectrum of skin over the visible region of the spectrum (450-760 nm). The contributions to the observed spectra made by pigments and the skin structure were deduced by reference to the theoretical model. Numerical indices were used to quantify the changes in skin haemoglobin content following the application of vasoconstricting preparations. The indices also provided a means of measuring erythema and melanin pigmentation induced in the skin by exposure to ultraviolet radiation. The assessments made using this instrument were more reproducible and sensitive than judgments made by eye.
Confocal scanning laser microscopy of live human skin was performed to investigate the correlation of in vivo cellular and morphologic features to histology, the effect of wavelength on imaging, and the role of melanin as a contrast agent. We built a video-rate confocal scanning laser microscope for in vivo imaging of human skin. Using a 100 x microscope objective, we imaged high-contrast optical "sections" of normal skin, vitiliginous skin, and a compound nevus. In vivo "confocal histology" correlated well with conventional histology. The maximum imaging depth increased with wavelength: the epidermis was imaged with visible 400-700-nm wavelengths; the superficial papillary dermis and blood cells (erythrocytes and leukocytes) in the deeper capillaries were imaged with the near infrared 800-900-nm wavelengths. For confocal reflectance imaging, melanin provided strong contrast by increased backscattering of light such that the cytoplasm in heavily pigmented cells imaged brightly. In vivo confocal microscopy potentially offers dermatologists a diagnostic tool that is instant and entirely non-invasive compared to conventional histopathology.
To quantify the dose-response relation of irritant-induced erythema, we examined inflammation in human skin after application of an irritant, using perpendicular polarized photography and diffuse reflectance spectroscopy as compared to clinical visual scoring. The ventral forearms of 11 healthy subjects were patch-tested for 24 h under occlusion in finn chambers with five concentrations of the irritant sodium lauryl sulfate. The tested sites and three control sites were evaluated clinically for erythema at 24, 48, and 72 h after occlusion, photographed using standard and perpendicular polarized photography, and measured by diffuse reflectance spectroscopy. All photographs were evaluated for erythema by three investigators. Diffuse reflectance spectra were analyzed, and changes in apparent oxyhemoglobin and deoxyhemoglobin concentrations were estimated. Clinical and photographic assessments of erythema yielded similar linear dose-response relations. A linear dose-response relation, with no minimum threshold, also was obtained for changes in the apparent oxyhemoglobin concentration with increasing irritant dose, whereas the apparent deoxyhemoglobin concentrations were unchanged with increasing dose. These results show that diffuse reflectance spectroscopy permits the characterization of irritant-induced inflammation in terms of a single parameter, the apparent concentration of oxyhemoglobin, and that irritant-induced inflammation primarily involves the capillaries and the superficial arterial plexus.
Although the ability of UV irradiation to induce pigmentation in vivo and in vitro is well documented, the intracellular signals that trigger this response are poorly understood. We have recently shown that increasing DNA repair after irradiation enhances UV-induced melanization. Moreover, addition of small DNA fragments, particularly thymine dinucleotides (pTpT), selected to mimic sequences excised during the repair of UV-induced DNA photoproducts, to unirradiated pigment cells in vitro or to guinea pig skin in vivo induces a pigment response indistinguishable from UV-induced tanning. Here we present further evidence that DNA damage and/or the repair of this damage increases melanization. (i) Treatment with the restriction enzyme Pvu II or the DNA-damaging chemical agents methyl methanesulfonate (MMS) or 4-nitroquinoline 1-oxide (4-NQO) produces a 4- to 10-fold increase in melanin content in Cloudman S91 murine melanoma cells and an up to 70% increase in normal human melanocytes, (ii) UV irradiation, MMS, and pTpT all upregulate the mRNA level for tyrosinase, the rate-limiting enzyme in melanin biosynthesis. (iii) Treatment with pTpT or MMS increases the response of S91 cells to melanocyte-stimulating hormone (MSH) and increases the binding of MSH to its cell surface receptor, as has been reported for UV irradiation. Together, these data suggest that UV-induced DNA damage and/or the repair of this damage is an important signal in the pigmentation response to UV irradiation. Because Pvu II acts exclusively on DNA and because MMS and 4-NQO, at the concentrations used, primarily interact with DNA, such a stimulus alone appears sufficient to induce melanogenesis. Of possible practical importance, the dinucleotide pTpT mimics most, if not all, of the effects of UV irradiation on pigmentation, tyrosinase mRNA regulation, and response to MSH without the requirement for antecedent DNA damage.
A technique employing diffuse reflectance spectroscopy (DRS) is described to assess and mirror dynamic changes of pancreatic tissue perfusion. An especially designed reflectance spectrophotometer was initially used to derive the quantitative relation between hemoglobin concentration ([Hb]) and reflectance measurements in vitro. Over a wide range of scattering related to the medium in which the measurements were made (scattering coefficient: 6.5-13 cm-1), a close, direct correlation existed with a slope of 0.376 +/- 0.012. In Sprague-Dawley rats under general anesthesia, the pancreas was isolated in situ and perfused with graded infusions of hemoglobin solutions. A correlation, comparable to the in vitro setting, was found between a [Hb] of 0 and 14 g/dl in the perfusate with slopes of 0.0037 and 0.0035. Changes in perfusion induced by adrenergic drugs produced changes in hemoglobin oxygen saturation and [Hb] that correspond with measured alterations of systemic arterial pressure and aortic blood flow. We conclude that diffuse reflectance spectroscopy reliably provides data on intrapancreatic hemoglobin oxygen saturation and [Hb] that can be a valuable tool for minimally invasive on-line evaluation of these aspects of pancreatic perfusion in the rat. This newly designed device is superior to previously used ones in that it analyzes the entire spectrum and therefore can account for changes in scattering that are very likely to occur with pathophysiological alterations such as edema formation.
The purpose of this study was to evaluate the blockage of polymorphonuclear neutrophil endothelial adhesion by using a monoclonal antibody to the intercellular adhesion molecule 1 (ICAM-1) ligand to prevent ischemia-reperfusion injury in rat skin flaps. A skin and subcutaneous tissue flap (3.0 cm x 4.5 cm) supplied by the superficial epigastric artery and vein including the femoral vessels was isolated unilaterally in 45 male Sprague-Dawley rats and clamped for 9 hours (groups II and III) or 12 hours (groups IV and V) of ischemia. Five animals in group I were sham-operated only with 5 minutes of ischemia. Animals in groups II (n = 10) and IV (n = 10) received 0.05 mg of monoclonal antibody to ICAM-1 (0.20 mg/kg) in 0.5 ml of 0.9% normal saline intravenously 15 minutes before reperfusion; those in groups III (n = 10) and V (n = 10) received 0.5 ml of normal saline. The flaps were assessed histologically, by measuring viable and nonviable areas, and by diffuse reflectance spectroscopy to determine the ratio of oxyhemoglobin to deoxyhemoglobin. Flap measurements revealed that the average area of flap survival was 90.6 +/- 12.8 percent in group II and 18.3 +/- 19.6 percent in the control group (III) (p < 0.002). In the animals subjected to 12 hours of ischemia, those treated with monoclonal antibody to ICAM-1 (group IV) were 57.1 +/- 23.1 percent viable, which was significantly greater than the control animals (group V), in which only 0.3 +/- 1.0 percent of the flap was viable. Analysis of the diffuse reflectance spectra showed a hyperemic response during the first 10 minutes after reperfusion in animals treated with monoclonal antibody to ICAM-1. In group III, however, the spectra demonstrated a decreased amount of oxyhemoglobin, indicating decreased reperfusion of the flap after ischemia when compared with group II. Histopathologically, few inflammatory changes could be observed in groups I, II, and the viable areas of group IV. Marked damage was observed in groups III and V. We concluded that treating ischemic skin flaps with monoclonal antibody to ICAM-1 was effective for alleviating reperfusion injury after 9 or 12 hours of warm ischemia. The reactive hyperemic response determined by diffuse reflectance spectroscopy in groups II and IV correlated with areas of flap survival. Antibodies to particular adhesion molecules, such as ICAM-1, have potential clinical utility in that they could be administered, individually or together, to patients immediately before reestablishing perfusion after free-tissue transfer or replantation to block the adverse effects attributed to reperfusion injury.
Infrared spectroscopy, by probing the molecular vibration of chemical bonds, directly indicates tissue biochemistry. An expanding body of literature suggests that infrared spectra distinguish diseased from normal tissue. The authors used infrared spectroscopy to examine basal cell carcinoma to explore distinctive characteristics of basal cell carcinoma versus normal skin samples and other skin neoplasms. Spectra of epidermis, tumor, follicle sheath, and dermis were acquired from unstained frozen sections, and analyzed qualitatively, by t-tests and by linear discriminant analyses. Dermal spectra were significantly different from the other skin components mainly due to absorptions from collagen in dermis. Spectra of normal epidermis and basal cell carcinoma were significantly different by virtue of subtle differences in protein structure and nucleic acid content. Linear discriminant analysis characterized spectra as arising from basal cell carcinoma, epidermis, or follicle sheath with 98.7% accuracy. Use of linear discriminant analysis accurately classified spectra as arising from epidermis overlying basal cell carcinoma versus epidermis overlying nontumor-bearing skin in 98.0% of cases. Spectra of basal cell carcinoma, squamous cell carcinoma, nevi, and malignant melanoma were qualitatively similar. Distinction of basal cell carcinoma, squamous cell carcinoma, and melanocytic lesions by linear discriminant analyses, however, was 93.5% accurate. Therefore, spectral separation of abnormal versus normal tissue was achieved with high sensitivity and specificity, which points to infrared spectroscopy as a potentially useful screening tool for cutaneous neoplasia.
In the present applications of optical coherence tomography (OCT), parameters besides pure morphology are evaluated in skin tissue under in vivo conditions. Spatially mapped refractive indices and scattering coefficients may support tissue characterization for research and diagnostic purposes in cosmetics/pharmacy and medicine, respectively. The sample arm of our OCT setup has been arranged to permit refractive index evaluation with little mechanical adjustment of a lens within the objective. A simple algorithm has been derived. Known from atmospheric work, the Klett algorithm [J. D. Klett, "Stable analytical inversion solution for processing LIDAR returns," Appl. Opt. 20(2), 211-220 (1981)] has been applied to the same data set for retrieval of scattering coefficients. Both parameters have been measured in layered structures in skin like stratum corneum, epidermis and dermis. Significant water content in a localized sweat gland duct has been observed by refractive index evaluation. Time studies over 1.5 h permitted a first understanding about physiological changes in skin which are not obtainable by intrusive methods.
Carotenoids are thought to play a significant part in the skin's anti-oxidant defense system, and may help prevent malignancy. Inability to measure skin carotenoid content readily has, however, made it difficult to establish the relationship between carotenoid concentration and the occurrence of cutaneous malignancy. We have measured in vivo carotenoid concentration using a noninvasive optical method, Raman spectroscopy. To validate our instrumentation, abdominoplasty skin was evaluated by both Raman spectroscopy and high-performance liquid chromatography determination for carotenoid content. Evaluation of the Raman signal in specific carotenoid solutions was also performed. Precision of Raman measurements within skin sites, within subjects, and between subjects was measured. Sensitivity of the method was evaluated as a function of anatomical region and the distribution of carotenoids within the stratum corneum. Lastly, we evaluated the Raman signal in actinic keratosis and basal cell carcinoma lesions and perilesional skin and compared this with region-matched sites in healthy subjects. Our results indicate that the Raman scattering method reflects the presence of carotenoids in human skin and is highly reproducible. Evaluation of five anatomical regions demonstrated significant differences in carotenoid concentration by body region with the highest carotenoid concentration noted in the palm. Comparison of carotenoid concentrations in basal cell carcinomas, actinic keratosis, and their perilesional skin demonstrate a significantly lower carotenoid concentration than in region-matched skin of healthy subjects. These results represent the first evidence that carotenoid concentration in the skin correlate with the presence or absence of skin cancer and precancerous lesions.
Confocal Raman spectroscopy is introduced as a noninvasive in vivo optical method to measure molecular concentration profiles in the skin. It is shown how it can be applied to determine the water concentration in the stratum corneum as a function of distance to the skin surface, with a depth resolution of 5 microm. The resulting in vivo concentration profiles are in qualitative and quantitative agreement with published data, obtained by in vitro X-ray microanalysis of skin samples. Semi-quantitative concentration profiles were determined for the major constituents of natural moisturizing factor (serine, glycine, pyrrolidone-5-carboxylic acid, arginine, ornithine, citrulline, alanine, histidine, urocanic acid) and for the sweat constituents lactate and urea. A detailed description is given of the signal analysis methodology that enables the extraction of this information from the skin Raman spectra. No other noninvasive in vivo method exists that enables an analysis of skin molecular composition as a function of distance to the skin surface with similar detail and spatial resolution. Therefore, it may be expected that in vivo confocal Raman spectroscopy will find many applications in basic and applied dermatologic research.
People who vacation in sunny places are exposed to the sun on multiple occasions at least on a daily basis. The clinical assessment of sun exposure is erythema in the first 48 h after exposure and pigmentation at times greater than 3-5 days. The purpose of this investigations was to determine the extent to which consecutive erythemogenic exposures result in additive erythema responses. Studies were conducted in which volunteers were first exposed to a graded series of fluences of UVB radiation and then on subsequent days (1-3 days) the same sites along with the surrounding unexposed skin were challenged with varying fluences of UVB radiation. The erythema reactions were assessed clinically and were objectively documented with diffuse reflectance spectroscopy. The sites that received two exposures always showed a reduced erythema response compared to a single erythemogenic exposure. The suppression of erythema was more pronounced when the second exposure was given 48 h after the first. The erythema suppression was maximal when the first exposure was at 1.3 minimum erythema dose (MED). The pigment response to the first exposure was completely suppressed for fluences less than 1.5 MED. We thus provide evidence for a decoupling of the classical sequence of erythema-pigmentation response. We also show that the erythema induced by a second exposure may be substantially suppressed by an earlier exposure, and that this cannot be due to melanin photoprotection or due to substantial thickening of the stratum corneum. We propose that the cause may be some diffusible element of yet unknown origin.
In vivo confocal Raman spectroscopy is a noninvasive optical method to obtain detailed information about the molecular composition of the skin with high spatial resolution. In vivo confocal scanning laser microscopy is an imaging modality that provides optical sections of the skin without physically dissecting the tissue. A combination of both techniques in a single instrument is described. This combination allows the skin morphology to be visualized and (subsurface) structures in the skin to be targeted for Raman measurements. Novel results are presented that show detailed in vivo concentration profiles of water and of natural moisturizing factor for the stratum corneum that are directly related to the skin architecture by in vivo cross-sectional images of the skin. Targeting of skin structures is demonstrated by recording in vivo Raman spectra of sweat ducts and sebaceous glands in situ. In vivo measurements on dermal capillaries yielded high-quality Raman spectra of blood in a completely noninvasive manner. From the results of this exploratory study we conclude that the technique presented has great potential for fundamental skin research, pharmacology (percutaneous transport), clinical dermatology, and cosmetic research, as well as for noninvasive analysis of blood analytes, including glucose.
Reflectance confocal microscopy (RCM) allows non-invasive visualization of human skin in vivo. It has been used to describe the histopathological features of acute contact dermatitis (CD). This work was designed to investigate the kinetics of both allergic and irritant CD (ACD and ICD) in vivo. Eighteen subjects with a prior diagnosis of ACD were patch tested with the specific allergen sodium lauryl sulfate as an irritant, and appropriate controls. RCM, transepidermal water loss (TEWL), and fluorescence excitation spectroscopy (FES) were performed at several time points within 2 wk after patch removal. After removal of the Finn chambers at 48 h, superficial epidermal changes, primarily involving the stratum corneum, and increased epidermal thickness were mainly present in ICD. ACD, on the other hand, showed microvesicle formation peaking at 96 h following patch removal. Both ACD and ICD showed exocytosis and similar degrees of spongiosis on RCM. TEWL and FES demonstrated a significant difference between ACD and ICD. RCM, TEWL, and FES are valuable non-invasive tools to quantitatively study the kinetics of the pathophysiology of acute CD reactions in vivo and monitor the changes at a cellular level.
We apply ultrasound-modulated optical tomography (UOT) to image ex-vivo methylene-blue-dyed sentinel lymph nodes embedded in 3.2-cm-thick chicken breast tissues. The UOT system is implemented for the first time using ring-shaped light illumination, intense acoustic bursts, and charge-coupled device (CCD) camera-based speckle contrast detection. Since the system is noninvasive, nonionizing, portable, relatively cost effective, and easy to combine with photoacoustic imaging and single element ultrasonic pulse-echo imaging, UOT can potentially be a good imaging modality for the detection of sentinel lymph nodes in breast cancer staging in vivo.
Various physical, chemical and biological insults, including exposure to ultraviolet (UV) radiation, cause erythema and change in pigmentation in human skin. These reactions provide an important measure of the cutaneous response to the insult.
To present a new implementation of a method for objective in vivo measurement of erythema and pigmentation.
The method is based on acquisition of reflectance spectra in the visible range using a commercially available spectrophotometer. The probe of this instrument incorporates an integrating sphere that captures the light remitted from the skin in a wide range of angles. We corrected the acquired reflectance spectra for the contribution of specular reflections by an amount given by the Fresnel equation and verified this correction experimentally. This correction is particularly important when measurements are performed on heavily pigmented skin. The corrected reflectance spectra are then transformed into absorbance spectra. To analyse these spectra, we developed an algorithm which can be used to calculate apparent concentrations of oxyhaemoglobin, deoxyhaemoglobin and melanin. This method was tested in clinical studies of skin reactions induced by exposure to UV radiation. These experiments involved three groups of subjects with progressively darker complexion (constitutive pigmentation). Each group consisted of 10 subjects. Erythema was measured 1 day after UV exposure, and pigmentation (melanin content) 1 week later. Results Distinct apparent absorbance spectra were obtained for dark, intermediate and fair skin. There was good agreement between reconstructed spectra and experimental data at relevant wavelengths. Difference absorption spectra were able to show the dose dependence of UV-induced responses, and erythema and pigmentation values obtained by the spectroscopic method showed good correlation with those derived by subjective visual grading.
The results demonstrate that the presented methodology provides an objective noninvasive way of measuring UV-induced reactions independently of the level of constitutive pigmentation.
Stress-induced changes in skin microcirculation allow staging of peripheral arterial vascular pathology using diffuse reflectance spectroscopy (DRS) of the skin.
The changes in relative concentration of oxyhemoglobin and deoxyhemoglobin in the cutaneous microvasculature were assessed at rest, during limb elevation, dependency, and cuff-mediated reactive hyperemia for the forearm of 25 normal subjects and 105 feet of patients with peripheral arterial occlusive disease (PAOD) (normal=28, claudication=34, limb threatening ischemia=44). Thirty-four patients who had revascularization procedures were again evaluated within the first week postoperatively.
Two measurements correlated with clinical staging: (1) the relative absorbance of oxyhemoglobin after 225 s of limb dependency and (2) the time to reach 50% of peak reactive hyperemia response (Spearman's rank: rs=0.625, P<0.001). Using these criteria alone, ischemic limbs were identified to a sensitivity of 69% and specificity of 95%. Significant post-revascularization improvement was identified in 14 of 34 patients' legs which had previously been classified as limb-threatening ischemia (n=14, W=105, P<0.001).
These simple bedside evaluations of the superficial skin microvasculature allow staging of large vessel vascular insufficiency and may suggest and differentiate focal areas of tissue at risk for ulceration or necrosis.
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