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Excess amounts of skin surface oil can lead to adverse psychological consequences. Grease-spot photometry-based techniques measure sebum production rate. However, besides being tedious, these measurements are influenced by contact area, applied pressure, and time of application. Image analysis of polarized images has the potential to provide objective, quantitative information of skin oiliness. This study was designed to set up an imaging device for capturing and enhancing the changes in skin surface oiliness and to clinically and quantitatively, (via image analysis), evaluate varying levels of skin surface oiliness. Mineral oil was used to simulate skin surface oil. 40.5 µL of the mineral oil was applied within a two inch square area of interest on facial skin in twelve steps, from 1 to 40.5 µL, at 40% increments. The results indicate a strong correlation between the quantitative skin surface oiliness measurements and the clinical assessments. This sensitive technique has the potential to be utilized in future studies to evaluate product efficacies in reducing skin oiliness.

Solar radiation is a major contributor to the development of skin cancer. Recent studies have shown that visible light (VL), a major portion of solar spectrum, induces biologic effects on the skin. Ultraviolet filters in currently available broad-spectrum sunscreens do not offer protection against VL. This study was designed to identify the spectral characteristics of the skin responses induced by VL, which can be utilized for time efficient in vivo VL testing. Thirty-one subjects were irradiated with a light source emitting visible light with less than 0.5% long wavelength UVA1 (VL + UVA1, 370-700 nm), and 41 subjects were irradiated with pure visible light (pure VL, 400-700 nm). Assessments including clinical photography, investigator's global assessment of pigmentation and erythema, and diffuse reflectance spectroscopy (DRS) performed immediately and seven days after irradiation. Clinical and spectroscopic data showed that VL + UVA1 spectral output induced significantly darker and persistent skin responses as compared to those induced by pure VL. Spectroscopic signatures of skin responses induced by both radiation sources were identified. The signatures were found to be specific to the radiation source and time of collection. A method to evaluate VL protection factor, using quantitative information from the spectral signatures obtained, was proposed.

Excess amounts of skin surface oil can lead to adverse psychological consequences. Grease-spot photometry-based techniques measure sebum production rate. However, besides being tedious, these measurements are influenced by contact area, applied pressure, and time of application. Image analysis of polarized images has the potential to provide objective, quantitative information of skin oiliness. This study was designed to set up an imaging device for capturing and enhancing the changes in skin surface oiliness and to clinically and quantitatively, (via image analysis), evaluate varying levels of skin surface oiliness. Mineral oil was used to simulate skin surface oil. 40.5 µL of the mineral oil was applied within a two inch square area of interest on facial skin in twelve steps, from 1 to 40.5 µL, at 40% increments. The results indicate a strong correlation between the quantitative skin surface oiliness measurements and the clinical assessments. This sensitive technique has the potential to be utilized in future studies to evaluate product efficacies in reducing skin oiliness.

SummaryBackground Visible light (VL) induces multiple cutaneous effects. Sunscreen testing protocols recommended by regulatory bodies throughout the world require the use of solar simulators with spectral output in ultraviolet (UV) domain only. However, sunlight contains VL and infrared radiation also. Objective This study aimed to evaluate the contributions of VL and UVA on pigmentation and erythema, and optimize parameters for in-vivo testing. Methods Ten subjects with Fitzpatrick skin phototype IV-VI were enrolled. Subjects were irradiated on their back with VL using two light sources: one containing pure VL and one containing VL with less than 0.5% UVA1 (VL+UVA1). Four different irradiances were administered to investigate reciprocity behaviour. Assessments, including photography, investigator's global assessment, colorimetry, and spectroscopy, were performed immediately, 24 hours, 7 days and 14 days post-irradiation. Results Pigmentation was observed with both light sources; however, pigment intensity was greater with VL+UVA1 than pure VL. Reciprocity was observed in pure VL sites, but not VL+UVA1. Variation in spectral output had greater impact on pigment intensity than irradiance. Clinical erythema was observed on the VL+UVA1 side, but not on the pure VL side. Protocol for testing photoprotection product efficacy against VL induced effects has been proposed. Conclusions The findings suggest a synergistic relationship between VL and UVA1 and emphasize the need for developing means of photoprotection against VL. This article is protected by copyright. All rights reserved.

Human skin is exposed to visible light (VL; 400-700 nm) and long-wavelength ultraviolet A1 (UVA1) radiation (370-400 nm) after the application of organic broad-spectrum sunscreens. The biologic effects of these wavelengths have been demonstrated; however, a dose-response has not been investigated. Ten subjects with Fitzpatrick skin phototype IV-VI were enrolled. Subjects were irradiated with 2 light sources (80-480 J cm-2 ): one comprising VL with less than 0.5% UVA1 (VL+UVA1) and the other pure VL. Skin responses were evaluated for 2 weeks using clinical and spectroscopic assessments. 4-mm punch biopsies were obtained from nonirradiated skin and sites irradiated with 480 J cm-2 of VL+UVA1 and pure VL 24 h after irradiation. Clinical and spectroscopic assessments demonstrated a robust response at VL+UVA1 sites compared with pure VL. Histology findings demonstrated a statistically significant increase in the marker of inflammation (P < 0.05) and proliferation (P < 0.05) at the irradiated sites compared with nonirradiated control. Threshold doses of VL+UVA1 resulting in biologic responses were calculated. Results indicate that approximately 2 h of sun exposure, which equates to VL+UVA1 dose (~400 J cm-2 ), is capable of inducing inflammation, immediate erythema and delayed tanning. These findings reinforce the need of photoprotection beyond the UV range.

The pigment responses of human skin to broadband UVA radiation (320-400 nm) occur in three distinct phases. The first phase includes immediate pigment darkening (IPD), the pigment that appears immediately after irradiation. The second phase involves an intermediate step, termed persistent pigment darkening (PPD), which leads to the third phase of neomelanogenesis or delayed tanning (DT). Since DT results from synthesis of new melanin, it persists beyond 5-7 days. We conducted studies on human subjects to investigate the dynamic responses of the IPD and PPD reactions to broadband UVA radiation at threshold and superthreshold doses. The threshold doses for IPD, PPD, and DT were found to be approximately 1, 11, and 18 J/cm2 , respectively. The colorimetry ΔL* value corresponding to minimal clinically perceptible pigmentation was found to be 0.8 ± 0.1. IPD appeared immediately and had an associated decay constant of approximately 1.4 minutes. At doses greater than PPD threshold, IPD reaction decayed while PPD developed indicating toward IPD being used as a substrate in the formation of PPD.

Background/Objective Epidermal structure, function, and composition are different in white infants and adults. We investigated whether ethnicity and location contribute to differences in functional and clinical measurements of skin barrier function during the first years of life and in adults. Methods Children (n = 397, ages 3–49 mos) and women (n = 117, mean age 31 yrs) were enrolled at independent centers in Beijing, China (ethnic Chinese), Skillman, New Jersey (white, African American), and Mumbai, India (ethnic South Asian). Water barrier properties of the stratum corneum were assessed using high-frequency conductance and transepidermal water loss (TEWL) on the dorsal forearm and upper inner arm. Digital imaging was used to evaluate facial erythema and scaling. Results Despite differences in local climate, TEWL was similar in adults. In children, conductance and TEWL decreased monotonically from age 3 months to 4 years. In children from Beijing, TEWL values were higher in both arm locations than in children in Mumbai and Skillman. No significant differences were observed in TEWL or conductance between the white and African American groups. Conclusion In general, TEWL and conductance were greater on the upper inner arm than the dorsal forearm. Erythema and scaling were observed most often in subjects from Beijing and most infrequently in subjects from Mumbai. Stratum corneum water barrier properties were different in children and adults. Although there may be differences in these properties between ethnic groups in childhood, TEWL values were similar in adults across the three geographic locations and four ethnicities.

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 to 600 nm, then rise moderately from 600 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 and 720 nm. Also, absorption peak at 335 nm showed the presence of melanin.

Solar radiation has been accepted as a major contributor to the development of skin cancer. Recent studies have shown that visible light (VL), a major portion of solar spectrum, induces biologic effects. UV filters in currently available sunscreens do not offer protection against VL. Diffuse reflectance spectroscopy (DRS) is a noninvasive objective assessment technique. Previous studies have focused on physiological parameters including melanin concentration, hemoglobin concentration, scattering and oxygen saturation. However, the differential absorbance spectra, difference between absorbance spectra of irradiated site and adjacent un-irradiated skin, can aid in the identification of various phases of skin response after VL irradiation. We irradiated the back of 62 subjects with a VL source, which consisted of a halogen lamp with filters resulting in a spectral output with greater than 98% VL. Reflectance spectra was collected for VL irradiated sites (160 J/cm2 to 480 J/cm2) immediately post-irradiation, at 24 hours, and at 7 days post-irradiation. DRS aided in classifying skin responses in terms of clinical and sub-clinical erythema accompanied by pigmentation phases based on the corresponding spectral signatures. The response in terms of skin darkness was assessed by integrating the differential absorbance spectra between 400 and 700 nm (AUC, area under the curve) since it includes contribution from both melanin and hemoglobin. The findings demonstrate the potential of DRS in assessing photoprotection offered by a product against VL induced pigmentation and erythema, VL protection factor, which can be assessed by comparing the AUC of the protected skin to that of the unprotected skin.

Background There is a rationale for adding systemic photoprotective agents to the current photoprotection regimen. Objective This study was designed to objectively evaluate the molecular and photobiologic effects of oral administration of Polypodium leucotomos extract (PLE). Methods In all, 22 subjects with Fitzpatrick skin phototype I to III were enrolled. On day 1, subjects were irradiated with visible light, ultraviolet (UV) A1, and UVB (using 308-nm excimer laser). Evaluation was done immediately and 24 hours after irradiation. On days 3 and 4, irradiation and evaluation process was repeated after ingestion of PLE. Results Clinical assessments and colorimetry data showed a decrease in UVB-induced changes in 17 of 22 subjects post-PLE administration; histology findings demonstrated such a decrease in all 22 subjects. Limitations Only 2 doses of PLE were given. Furthermore, subjects with skin phototypes I to III only were studied. Conclusion The results suggest that PLE can potentially be used as an adjunctive agent to lessen the negative photobiologic effects of UVB.

Assessment of the optical properties of the skin is very helpful for the quantification of the content and spatial distribution of the various biological components in skin. It is also useful for the diagnosis of skin diseases, investigation of the impact of different environmental factors (chemical agents, drugs, UV-radiation, temperature, etc), and evaluation of the effectiveness of skin treatments. Currently, reflectance and fluorescence spectroscopy are probably the most developed among the available optical methods for investigation of skin in vivo. Reflectance and fluorescence from skin carry information about the structures of the epidermis and dermis, the quantity and density of blood vessels, the concentration and spatial distribution of chromophores and fluorophores in skin, and the nature of skin metabolic processes. The latest improvements in fiber optics, electronics, and computer technologies have made reflectance spectroscopy a common and popular technique for skin analysis. Typical applications include in vivo quantitative analysis of skin erythema and pigmentation, determination of cutaneous color variation, monitoring of dermatological treatment effects, and study of skin biophysics.

Erythema and pigment responses of human skin following an acute exposure to ultraviolet radiation (UVR) are frequently used to determine the photosensitivity of the skin. In this study we investigated the responses of the skin to a micro-scale area of UVR exposure (MiR) and compared the responses to a macro-scale area of exposure (MaR). Ten human volunteers were tested with solar-simulated radiation on their upper arm or back using a beam size of 8 mm and 0.2 mm in diameter. The fluence required to produce a minimally perceptible erythema (MED) using the MiR was found to be higher than that for the MaR. The erythema response extended beyond the exposed area and this became pronounced when the beam size was microscopic. Reflectance confocal microscopy in vivo revealed that MiR induced cellular alterations within a confined area of smaller dimensions than the area of exposure. Pigment responses were confined within the areas of cellular damage. The erythema expression of exposed skin recovered faster for the sites receiving MiR even when the applied fluence was higher than the MED for the MaR. Through the use of MiR we were able to visualize spatially dissimilar skin responses of erythema and pigmentation suggesting different cellular mechanisms.

SUMMARY Background/Purpose In the past56years, many differentin vitromethodologies have been developed and published to assess the sun protection factor (SPF) of products, but there is no method that has1:1correlation within vivomeasurements. Spectroscopic techniques have been used to noninvasively assess the UVA protection factor with good correlation toin vivoUVA-PF methodologies. To assess the SPF of sunscreen product by diffuse reflectance spectroscopy (DRS) technique, it is necessary to also determine the absorbance spectrum of the test material in the UVB portion of the spectrum (290‐320nm). However, because of the high absorbance characteristics of the stratum corneum and epidermis, the human skin does not remit enough UVB radiation to be used to measure the absorption spectrum of the applied product on skin. In this work, we present a new method combining the evaluation of the absolute UVA absorption spectrum, as measured by DRS with the spectral absorbance ‘shape’ of the UVB absorbance of the test material as determined with current in vitrothin film spectroscopy. Methods

Fluorescence signals depend on the intensity of the exciting light, the absorption properties of the constituent molecules, and the efficiency with which the absorbed photons are converted to fluorescence emission. The optical features and appearance of vitiligo have been explained primarily on the basis of reduced epidermal pigmentation, which results in abnormal white patches on the skin. The objective of this study is to explore the fluorescence properties of vitiligo and its adjacent normal skin using fluorescence excitation-emission matrix (EEM) spectroscopy. Thirty five (35) volunteers with vitiligo were acquired using a double-grating spectrofluorometer with excitation and emission wavelengths of 260-450 nm and 300-700 nm respectively. As expected, the most pronounced difference between the spectra obtained from vitiligo lesions compared to normally pigmented skin was that the overall fluorescence was much higher in vitiligo; these differences increased at shorter wavelengths, thus matching the characteristic spectral absorption of epidermal melanin. When comparing the fluorescence spectra from vitiligo to normal skin we detected three distinct spectral bands centered at 280nm, 310nm, and 335nm. The 280nm band may possibly be related to inflammation, whereas the 335 nm band may arise from collagen or keratin cross links. The source of the 310 nm band is uncertain; it is interesting to note its proximity to the 311 nm UV lamps used for vitiligo phototherapy. These differences are accounted for not only by changes in epidermal pigment content, but also by other optically active cutaneous biomolecules.

Synopsis An ageing study was conducted to capture skin colour parameters in the CIELab system from Caucasians of both genders and all available adult ages. This study produced a linear correlation between L* and age for a Caucasian population between 20 and 59 years of age as follows: (L* value) = −0.13 × (Age in years) + 63.01. Previous studies have addressed age-related changes in skin colour. This work presents a novel consumer correlated quantitative linear model of skin brightness by which to communicate age-related changes. Two product assessment studies are also presented here, demonstrating the ability of anti-ageing products to deliver on objective and subjective improvements in skin brightness. It was determined to be possible to use the fundamental Caucasian L*-age correlation to describe product benefits in a novel quantitative and consumer-relevant fashion, through the depiction of a ‘years back’ calculation. Resume Une etude de vieillissement a ete conduite en utilisant la capturer des parametres de couleur de peau dans le systeme Lab sur des sujets caucasiens des deux sexes et tous âges adultes disponibles. Cette etude a montre une correlation lineaire entre L* et l’âge comme suit (L* la valeur) = −0.13 × (l’âge en annees) +63.01. Des etudes precedentes ont mis en evidence des changements de la couleur de peau lies a l’âge. Ce travail presente un nouveau modele lineaire quantitatif correlea la luminosite de la peau par lequel il est possible de communiquer au grand public les changements lies a l’âge. Deux etudes d’evaluation de produits sont aussi presentees ici, demontrant la capacite de produits anti-âges a apporter des ameliorations objectives et subjectives dans la luminosite de la peau. Il a ete montre qu’il est possible d’utiliser la correlation âge, valeur fondamentale L* pour decrire chez les caucasiens des avantages produit d’une nouvelle facon quantitative et appropriee au grand public, par la description d’un calcul d’un “recul d’annees”.

Background: In vivo confocal scanning laser microscopy (CSLM) is a recently developed non-invasive technique for visualizing microscopic structures with the skin. CSLM has been used to characterize proliferative and inflammatory skin diseases, neoplastic skin lesions and pigmented lesions. Objective: Here, we assessed the ability of CSLM to evaluate the formation of neogenic hair follicles after a full-thickness wound in mice. Methods: Full-thickness wounds were made on the dorsal skin of 3-week-old mice. After scab detachment (SD), the number, width, length, space and volume of neogenic hair follicles were analyzed using CSLM. The results were compared with those from conventional methods, including staining for alkaline phosphatase (AP) and keratin 17 (K17) as well as histology. Results: Quantification of neogenic hair follicles using CSLM compared favorably with the results from direct measurements on isolated epidermal tissue after immunostaining for K17, a marker for the epithelial portion of new hair follicles. CSLM detected 89% of K17-stained follicles. CSLM more accurately quantified the number of new follicles compared with AP staining, which detects the dermal portion of the new follicle. The width and length measurement from CSLM and histology were very close and correlated with each other. The minimum length of a neogenic hair follicle that could be detected by CSLM was 21 μm. The space between neogenic hair follicles was decreased in histological sections compared with CSLM. Conclusion: CSLM is an accurate and valuable method for counting and measuring neogenic hair follicles non-invasively. CSLM produces images similar to histology in mice. Measurements of microstructures using CSLM more accurately reflect actual sizes as this technique avoids fixation artifacts. In vivo visualization of developing follicles with CSLM allows the detection of serial changes in hair follicle formation, thus conserving the numbers of mice required for studies and improving the detection of temporal changes in developing hair follicles.

Infant skin is often presented as the cosmetic ideal for adults. However, compared to adult skin it seems to be more prone to develop certain pathological conditions, such as atopic dermatitis and irritant contact dermatitis. Therefore, understanding the physiology of healthy infant skin as a point of reference is of interest both from the cosmetic as well as from the clinical point of view. Clinical research on healthy infants is, however, limited because of ethical considerations of using invasive methods and therefore until recently data has been scarce. Technical innovations and the availability of non-invasive in vivo techniques, such as evaporimetry, electrical impedance measurement, in vivo video and confocal microscopy, and in vivo fibre-optic based spectroscopy, opened up the field of in vivo infant skin physiology research. Studies incorporating such methods have demonstrated that compared to adult, infant skin continues to develop during the first years of life. Specifically, infant skin appears to have thinner epidermis and stratum corneum (SC) as well as smaller corneocytes at least until the second year of life. The water-handling properties are not fully developed before the end of the first year and infant SC contains more water and less amounts of natural moisturizing factors. Such findings re-evaluate the old notions that skin is fully matured at birth. Armed with this knowledge, we are in a position not only to better understand infant dermatological conditions but also to design better skin care products respecting the distinct qualities of infant skin.

Functional differences between infant and adult skin may be attributed to putative differences in skin microstructure. The purpose of this study was to examine infant skin microstructure in vivo and to compare it with that of adult skin. The lower thigh area of 20 healthy mothers (ages 25-43) and their biological children (ages 3-24 months) was examined using in vivo noninvasive methods including fluorescence spectroscopy, video microscopy, and confocal laser scanning microscopy. Stratum corneum and supra-papillary epidermal thickness as well as cell size in the granular layer were assessed from the confocal images. Adhesive tapes were used to remove corneocytes from the outer-most layer of stratum corneum and their size was computed using image analysis. Surface features showed differences in glyph density and surface area. Infant stratum corneum was found to be 30% and infant epidermis 20% thinner than in adults. Infant corneocytes were found to be 20% and granular cells 10% smaller than adult corneocytes indicating a more rapid cell turnover in infants. This observation was confirmed by fluorescence spectroscopy. Dermal papillae density and size distribution also differed. Surprisingly, a distinct direct structural relationship between the stratum corneum morphology and the dermal papillae was observed exclusively in infant skin. A change in reflected signal intensity at approximately 100 mum indicating the transition between papillary and reticular dermis was evident only in adult skin. We demonstrate in vivo qualitative and quantitative differences in morphology between infant and adult skin. These differences in skin microstructure may help explain some of the reported functional differences.

Summary Background Determination of skin surface texture is of particular importance in the field of dermatology as such measurements can be used for skin diagnostics and evaluation of therapeutic or cosmetic treatments. Profilometry of skin replicas, three-dimensional imaging and computer vision have been successfully used to measure and document skin texture. Nevertheless, the development of a simpler and faster technique may prove to be advantageous in a clinical setting. Objectives We propose the use of polarization imaging with high angles of incidence as a simple alternative to measure/document skin texture/roughness. Methods A system based on digital photography and polarization optics was developed to acquire and compute texture images. Optimization of the system configuration was conducted to enhance the contrast for measuring skin roughness. The method was validated against roughness standards and tested in clinical studies. Measurements were made in subjects aged from 9 to 70 years and image analysis was used to evaluate texture. Results The developed texture scale was shown to correlate closely to the results from clinical assessment and from roughness standards. Frequency domain analysis showed a significantly different power spectrum for the texture images of young subjects when compared with older subjects. The evaluation of texture as a function of age showed that facial skin roughness increased linearly from teenage to 40 years followed by a plateau thereafter. Conclusions The system proved to be a useful clinical tool for assessing skin texture. The age-related results may indicate that some skin texture features are formed before the age of 40 years.

Background: When the skin is exposed to solar irradiation, UVA photons interact with skin tissues and induce excessive reactive oxygen species, resulting in oxidative stress. We have shown in a previous study that in vivo chemiluminescence's measurement can be used to evaluate the overall level of UVA-induced oxidative stress in human skin. However, the origin of the observed chemiluminescence signals remains unclear. Methods: UVA-induced chemiluminescence measurements were conducted: (a) in vitro on collagen solutions and solid collagen sheet preparations, (b) ex vivo on human and mouse skin biopsies, and (c) in vivo on human skin of various constitutive pigmentation levels. Fluorescence was measured on collagen in vitro as well as on skin for the in vivo experiments. Results: We found in the in vitro experiments that UVA-induced chemiluminescence increases with the presence of collagen cross-links. When dermal sides were exposed to UVA irradiation, both mouse and human skin biopsies demonstrated significantly higher chemiluminescence levels than when epidermal sides were exposed to UVA. The amount of collagen cross-links decreases slightly following UVA exposure, as shown both by in vivo fluorescence and by UVA-induced chemiluminescence. Finally, there was less measurable UVA-induced chemiluminescence in dark skin compared with light pigmented skin in vivo. Conclusions: The dermis is very sensitive to UVA photons. Dermal cross-links are potential UVA sensitizers. The oxidative stress induced by UVA and measured by chemiluminescence may largely be attributed to the breakdown of dermal collagen cross-links.

Skin water barrier development begins in utero and is believed to be complete by week 34 of gestational age. The goal of this investigation was to assess the dynamic transport and distribution of water of the stratum corneum of infants and compare it to those of adults. The interaction of water with the stratum corneum was assessed by measuring capacitance, transepidermal water loss (TEWL), rates of absorption-desorption as well as Raman spectra as a function of depth (a total of 124 infants (3-12 months) and 104 adults (14-73 years)). The results show that capacitance, TEWL, and absorption-desorption rates had larger values consistently for infant stratum corneum throughout the first year of life and showed greater variation than those of adults. The Raman spectra analyzed for water and for the components of natural moisturizing factor (NMF) showed the distribution of water to be higher and have a steeper gradient in infants than in adults; the concentration of NMF was significantly lower in infants. The results suggest that although the stratum corneum of infants may appear intact shortly after birth (

Limited depth of penetration significantly limits photodynamic therapy of nodular basal cell carcinoma (BCC) using topical delta (5)-aminolevulinic acid (ALA). To demonstrate safety and efficacy of orally administered ALA in inducing endogenous protoporphyrin IX (PpIX) production in BCC, 13 patients with BCC ingested ALA in a dose-escalation protocol. All dose ranges (10, 20 or 40 mg/kg single doses) resulted in formation of PpIX in human skin and BCC, measurable by in vivo fluorescence spectrophotometry. The PpIX fluorescence peaked in tumors before normal adjacent skin from 1 to 3 h after ALA ingestion. Gross fluorescence imaging of ex vivo specimens revealed greater PpIX fluorescence in tumor than normal skin only at the 40 mg/kg dose. Fluorescence microscopy confirmed this finding by showing distinct, full-thickness PpIX fluorescence in all subtypes of BCC only after ALA given at 40 mg/kg. Side effects were dose dependent and self limited. Photosensitivity lasting less than 24 h and nausea coinciding with peak skin PpIX fluorescence occurred at 20 and 40 mg/kg doses. After 40 mg/kg ALA, serum hepatic enzyme levels rose to a maximum within 24 h, then resolved over 1-3 weeks. Transient bilirubinuria occurred in two patients.

Fluorescence excitation spectroscopy was used to assess cellular turnover in human skin by monitoring changes of endogenous fluorescence. Epidermal proliferation was induced with alpha-hydroxy acids. Commercially available glycolic acid creams (8 and 4% wt/wt concentration) and a vehicle cream (placebo) were applied in a randomized double blinded fashion on subjects' forearms, twice daily for 21 days. Excitation spectra were recorded (excitation 250-360 nm, emission 380 nm) at days 0, 1, 3, 7, 10, 11, 14, 17 and 21. The 295 nm excitation band (assigned to tryptophan moieties) was used in this study as a marker for cellular proliferation. To further reduce the day-to-day variability of the skin fluorescence the intensity of the 295 nm band was normalized to the 334 nm band (assigned to collagen crosslinks). The fluorescence emission intensity from placebo-treated skin remained practically unchanged over the period of the measurements while the fluorescence intensity measured from the glycolic acid-treated skin increased monotonically with treatment. The rate of increase of the excitation intensity with treatment was found to be dose dependent. The epidermal 295 nm band may be used as a quantitative marker to monitor the rate of proliferation of epidermal keratinocytes noninvasively.

Sheets of polysulphone film have been extensively used as detectors to monitor solar UVB radiation. The advantages of polysulphone detectors are that they are small in size, they have good thermal stability and they are sensitive to UVB radiation. The principal disadvantage of polysulphone detectors is that their spectral sensitivity includes part of the short-wavelength UVA. In this study, we investigate the spectral sensitivity of the polysulphone detector with a series of monochromatic (±2 nm) excitations. We then compare the polysulphone-effective solar radiation with the erythemally effective solar radiation by comparing solar UVB data obtained with polysulphone films with those obtained with a spectroradiometer. From polysulphone data on the seasonal variation of solar UVB radiation, we estimate the corresponding fluctuations of the absorption of the ozone layer. We show that the spectral sensitivity of the polysulphone film is closer to the erythema action spectrum than that indicated by earlier data and that polysulphone detectors can be used to predict the erythema risk of solar UVB. Measurements on solar UVB with polysulphone films and with a spectroradiometer were found to be strongly correlated (R2 > 0.95). Finally, polysulphone-based measurements provide a good measure of the fluctuations of the stratospheric ozone layer.

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.

Summary Background Throughout life facial skin is exposed to a variety of adverse environmental conditions and is constantly required to repair itself. The rate of epidermal cell proliferation is indicative of the skin's repair rate and can be monitored noninvasively in vivo using skin intrinsic fluorescence markers. Objectives The goal of the present study was to assess the effects of ageing, geographical region, ethnic origin and season on the ability of facial skin to repair itself in the presence of chronic environmental insults using in vivo fluorescence spectroscopy. Methods Skin fluorescence emission was measured on the cheeks of 522 individuals in winter and repeated in summer in five different geographical locations in the Asia-Pacific region. Fluorescence emission was also measured from 80 caucasians of fair complexion in the United States (New Jersey area) on the face and on a relatively protected area (upper inner arm). The age range of the participants was 14–75 years. Results We found that epidermal proliferation rates decrease monotonically with age, while the fluorescence of collagen and elastin cross-links increases with age indicating accumulation of advanced glycation end-products. These trends were independent of geographical region, ethnic origin and season of measurement. Epidermal proliferation rates of facial skin were higher than those of unexposed sites; they may be 10 times higher in younger (second decade) than in older (seventh decade) individuals, and they decrease with age at rates 10 times faster compared with those of unexposed sites. Conclusions This is the first time that epidermal proliferation and its dependence on ageing have been measured noninvasively on the human face. The higher tryptophan fluorescence values on the face vs. the protected site are indicative of accelerated rates of epidermal proliferation in the presence of chronic environmental insults. The repair potential of facial skin, i.e. its ability to maintain high proliferation rates, is maximal in younger populations and gradually decreases with age.

Summary Autofluorescent lipofuscin and advanced glycation end-products (age pigments) accumulate with age across phyla, yet little is understood about their formation under physiological conditions and their specific contributions to the aging process. We used in vivo spectrofluorimetry to quantitate autofluorescence in wild-type Caenorhabditis elegans and longevity mutants disrupted for distinct aspects of the aging process. In wild-type animals, age pigments increase into adulthood, accumulating slowly during the reproductive phase and more rapidly during the post-reproductive period. As in humans, insulin signaling influences age pigment accumulation – mutations that lower efficacy of insulin signaling and extend lifespan [daf-2(e1370) insulin receptor and age-1(hx546) PI3-kinase] dramatically lower age pigment accumulation; conversely, elimination of the insulin-inhibited DAF-16/FOXO transcription factor causes a huge increase in age pigment accumulation, supporting that the short-lived daf-16 null mutant is truly progeric. By contrast, mutations that increase mitochondrial reactive oxygen species production do not affect age pigment accumulation, challenging assumptions about the role of oxidative stress in generating these species in vivo. Dietary restriction reduces age pigment levels significantly and is associated with a unique spectral shift that might serve as a rapidly scored reporter of the dietary restricted state. Unexpectedly, genetically identical siblings that age poorly (as judged by decrepit locomotory capacity) have dramatically higher levels of age pigments than their same-aged siblings that appear to have aged more gracefully and move youthfully. Thus, high age pigment levels indicate a physiologically aged state rather than simply marking chronological time, and age pigments are valid reporters of nematode healthspan.

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.

Oxidative stress is defined as an imbalance between pro-oxidants and antioxidants in favor of pro-oxidants. Photon emission (also called chemiluminescence) has been widely used to study oxidative stress in biological systems in vitro. In vivo chemiluminescence has been proposed as a non-invasive method to assess oxidative stress in the skin. UVA (320-400 nm part of the ultraviolet radiation) exposure is generally accepted as a source of oxidative stress in the skin. In this study, UVA-induced oxidative stress was studied by using an in vivo chemiluminescence detection method. First, the dose response and the fluence rate response of the UVA-induced oxidative stress in human skin were investigated by examining the decay kinetics of the chemiluminescence signal following UVA exposure. A kinetic model was proposed to help differentiate these two responses. We found that the initial burst of the chemiluminescence signal depended on the UVA fluence rate, whereas the decay of the signal following exposure can be related to the UVA dose involved. Second, a significant reduction of UVA-induced chemiluminescence signal was observed after tape-stripping, indicating that stratum corneum is a major source of UVA-induced oxidative stress in the skin. Furthermore, the oxygen dependence of UVA-induced chemiluminescence signal was also confirmed by application of a pressure cuff, implying that some of the oxidative stress occurs in the deeper layers of the skin. Finally, topical application of vitamin C before exposure significantly reduced the UVA-induced chemiluminescence signal. We thus conclude that chemiluminescence is an effective method to assess the oxidative stress induced by UVA in human skin in vivo.

The purpose of this investigation was to study the ultraviolet A-induced effects on melanin pigmentation both in an in vitro model system and in vivo. Ultraviolet-Vis absorbance spectra of L-3,4-dihydroxyphenylalanine-melanin solutions at different concentrations were measured before and after ultraviolet A exposure (10–120 J per cm2). The difference spectra reveal that following ultraviolet A exposure the absorbance increases exponentially from 800 nm to 450 nm accompanied by a prominent decrease of absorbance in the ultraviolet A range. This change of spectral features depends on both ultraviolet A doses and melanin concentrations. The photo-bleaching effect observed in the ultraviolet A range also depends on oxygen. Human subjects were irradiated with ultraviolet A (40–80 J per cm2) on their back and diffuse reflectance spectra were collected at both irradiated and untreated sites. The absorption spectra of ultraviolet A-induced pigment were calculated as the difference of the two. The ultraviolet A-induced pigment in vivo has similar spectral characteristics and dose dependency as the in vitro system. Photo-oxidation of pheomelanin solutions presents distinctively different spectral and dose–response characteristics from eumelanin. After ultraviolet A irradiation pheomelanin absorbance decreases both in the visible and the ultraviolet A range. We conclude that irradiation with ultraviolet A induces significant photochemical alterations in the skin witnessed by increased photoprotection in the visible spectral range and reduced protection in the ultraviolet A range. We suggest that soluble melanin plays an important part in ultraviolet A-induced pigment in skin and two distinct absorption mechanisms of melanin may be involved in ultraviolet A photo-oxidation. We also propose that eumelanin and pheomelanin could be differentiated according to their spectral responses to ultraviolet A irradiation.

Background Biologically effective solar ultraviolet radiation is defined as the product of the intensity of the solar spectrum and the erythema action spectrum at each wavelength. In this way we may arrive at the weighted effectiveness of each wavelength of solar radiation to produce a sunburn reaction. There have been many measurements of the variation of the solar spectrum with the time of the day and the time of the year, but questions remain as to the variation of the quality of the spectrum and the contribution of the shortest wavelengths of solar terrestrial radiation. The purpose of the present study was to determine the variation of the biologically effective solar spectrum with the time of the day and the time of the year and to determine the variation of the shortest wavelength that contributes to the sunburn reaction with the time of the day and the time of the year. Methods Spectroradiometric measurements were made at ground level over the period of one year (1988–1989) and at different times of the day at latitude 29.5° north. The measured spectral irradiance was multiplied wavelength by wavelength by the erythema action spectra. Results We determined that the biologically effective solar spectrum remains essentially the same over the times of the day that sunburn may be experienced. The maximally effective wavelength of biologically effective solar radiation was determined to be 308 nm. The cut-off wavelength for biologically effective solar radiation (defined as the wavelength at which the biologically effective solar radiation is at 1% of its maximum) varied from 291 to 295 nm over the time of the year and from 292 to 296 nm over the day. Conclusion For all practical purposes the biologically effective spectrum of solar ultraviolet radiation may be considered to remain constant over the period when sunburn may occur and the minimal wavelength of sunlight that contributes to sunburn is in the range of 291–296 nm.

A number of noninvasive approaches have been developed over the years to provide objective evaluation of the skin both in health and in disease. The advent of computers, as well as of lasers and photonics, has made it possible to develop additional techniques that were impossible a few years ago. These approaches provide the dermatologist with sensitive tools to measure the skin's condition in terms of physiologic parameters (e.g., color, erythema and pigmentation, induration, sebaceous and stratum corneum lipids, barrier function, etc.). Yet, a typical dermatologic diagnosis relies primarily on the trained eyes of the physician and to a lesser extent on information from other senses, such as touch and smell. The trained senses of the dermatologist backed by his/her brain form a powerful set of tools for evaluating the skin. The golden rule in diagnosis remains the histologic examination of a skin biopsy, a rather invasive method. These tools have served the profession well. The advent of ever faster and cheaper computers and of sensitive, inexpensive optical instrumentation of minimal dimensions provides the professional with the possibility of making objective measures of a number of skin parameters.

The fluorescence properties of skin both human and of an animal model are reviewed in this article. Fluorescence spectroscopy is a valuable tool for the investigation of the optical properties of the skin in the ultraviolet range of the spectrum because the absorption bands are confluent. Epidermal and dermal fluorescence signals are each produced in distinct spectral regions. Biological processes modulate the fluorescence signals in predictable ways. Such cases include aging, epidermal proliferation, and photoaging as well as diseases such as psoriasis, acne and non-melanoma skin cancer. Fluorescence excitation spectroscopy has proved a valuable approach in the study of the structure and function of human skin. (C) 2002 Elsevier Science B.V. All rights reserved. Vibrational Spectroscopy

Topical application of active compounds on skin is common to both pharmaceutical and cosmetic industries. Quantification of the concentration of a compound deposited on the skin is important in determining the optimum formulation to deliver the pharmaceutical or cosmetic benefit. The most commonly used techniques to date are either invasive or not easily reproducible. In this study, we have developed a noninvasive alternative to these techniques based on spectrofluorimetry. A mathematical model based on diffusion approximation theory is utilized to correct fluorescence measurements for the attenuation caused by endogenous skin chromophore absorption. The limitation is that the compound of interest has to be either fluorescent itself or fluorescently labeled. We used the method to detect topically applied salicylic acid. Based on the mathematical model a calibration curve was constructed that is independent of endogenous chromophore concentration. We utilized the method to localize salicylic acid in epidermis and to follow its dynamics over a period of 3 d.

The cascade of events following an injury that significantly alters epidermal structures and leads to epidermal necrosis depends on the type of injury. We have studied epidermal necrosis caused by solar-simulated radiation, laser therapy, and liquid nitrogen (N2) with reflectance laser scanning confocal microscopy in vivo. In the present study we investigated injuries produced with liquid nitrogen (N2), or cryotherapy, a common therapeutic modality for solar lentigines, warts, and others. A 3 mm diameter skin site on the forearm of 7 healthy human volunteers was treated with liquid nitrogen for approximately 20 s, following written informed consent. The site was monitored by confocal microscopy and digital imaging until the epidermis appeared clinically fully repaired (>4 weeks). During the first 3 days following exposure to liquid nitrogen (N2), the epidermal cellular organization appears slightly perturbed and the epidermal cellular architecture becomes lost. The pigment distribution in the keratinocytes that line the papillae maintains its appearance and does not spread as the cell boundaries become imperceptible. The dead epidermis remains intact for approximately 2 weeks and protects the new restructuring epidermis under it. The wound repairs from the periphery. Approximately 2 weeks after injury, the new epidermis is infiltrated by large dendritic cells localized in the area where papillae form. The papillae appear to grow and produce the typical invaginations and surrounding structures of the dermal-epidermal junction, and the enclosed capillaries are fully engorged with flowing erythrocytes. These dendritic cells begin to disappear as the epidermis returns to normal appearance and pigment. The dendritic cells appear to play a role in tissue restructuring post-injury, as they are also present for a limited time following UV injury starting at 14 days as in the present case.

Acne is a pleomorphic skin disease. The evolution of an acne lesion involves several stages (a) preclinical manifestations (noninflammatory lesions), (b) emergence of inflammatory lesions, (c) clearance, and (d) leftover marks such as post-inflammatory hyperpigmentation (PIH) and scars (affected dermal structures).

Noninvasive and real-time analysis of skin properties is useful in a wide variety of applications. In particular, the quantitative assessment of skin in terms of hemoglobin and melanin content, as well as in terms of its light scattering properties, is a challenging problem in dermatology. We present here a technique for examining human skin, based on the in vivo measurement of diffuse reflectance spectra in the visible and near-infrared ranges of the electromagnetic spectrum. Spectra were measured by means of a fiber optic probe, and they were analyzed using an analytical model of light diffusion in the skin. The results of the analysis indicate that it is possible to obtain quantitative information about hemoglobin and melanin content, as well as basic information regarding the scattering properties of the skin.

We investigated the use of attenuated total reflection-Fourier transform infrared spectroscopy as a method to study differences in the molecular components of human stratum corneum in vivo. These variations as a function of the anatomic site and of the depth into its layered structure are important to understand the biology and physiology of the tissue. In this preliminary study we have investigated spectroscopic changes in 18 healthy individuals. Total reflection-Fourier transform infrared spectroscopy represents a potentially powerful tool to study biophysical properties of surfaces. We observed that, in vivo, biophysical parameters of the stratum corneum (such as hydration, lipid composition, and conformation of the aliphatic chains) are indeed dependent on the anatomic site. As in total reflection-Fourier transform infrared spectroscopy experiments the penetration depth of the evanescent field into the stratum corneum is comparable with the thickness of a layer of corneocytes, this technique can be used to follow the distribution of lipids, water, and proteins as a function of depth into the tissue. We found that, in vivo, these molecular components are non-uniformly distributed, in agreement with the presence of water and lipid reservoirs as observed with ex vivo ultrastructural investigations. Composition and conformational order of lipids are also a function of depth into the stratum corneum. Finally we compared the in vivo superficial hydration measured using the infrared absorption of the OH stretch of water, with the hydration measured using the Skicon hygrometer. Our results indicate that total reflection-Fourier transform infrared spectroscopy might be useful to measure important chemical and biophysical parameters of stratum corneum in vivo.