Stratum Corneum Proteins Research Articles

Carbonylated proteins in the stratum corneum generate reactive oxygen species under short wavelength visible light

Carbonylated proteins in the stratum corneum generate reactive oxygen species under short wavelength visible light

Carbonylated proteins exposed to UVA and to blue light generate reactive oxygen species through a type I photosensitizing reaction

BackgroundCarbonylated proteins (CPs) are generated by the reaction of basic amino acid residues in proteins with aldehyde compounds produced during lipid peroxidation. CPs in the stratum corneum (SC) impact skin conditions such as skin moisture functions including water content and transepidermal water loss (TEWL). In addition, CPs can be frequently seen in the SC from sun-exposed sites compared with sun-protected sites. ObjectiveThe aim of this study was to reveal whether CPs could be a generation source of reactive oxygen species (ROS) in the SC following exposure to ultraviolet (UV) radiation and to identify the type of ROS and its generation mechanism. MethodsROS generation was detected using a methyl cypridina luciferin analog (MCLA) chemiluminescence system and an ESR spin-trapping method. CPs in porcine SC, in a keratin film and in bovine serum albumin (BSA) were prepared by reaction with acrolein. Levels of protein carbonylation were quantified by detecting aldehyde residues. ResultsCP levels in the SC were increased in a UVA energy-dependent manner. That result suggested that a source of ROS generation existed in the SC initiated and produced the carbonylation of SC proteins. Carbonylated BSA and carbonylated porcine SC sheets exhibited fluorescence spectra at an excitation wavelength of 430nm and an emission wavelength of 520nm. Irradiation of the SC with UVA increased protein carbonylation and the amount of autofluorescence in the SC. ROS generation in the SC caused by UVA and by short-wavelength visible light (blue light, 400–470nm) was detected by the MCLA chemiluminescence system. Artificially carbonylated porcine SCs and keratin films had increases of chemiluminescence intensity after exposure to both light sources as well. The addition of superoxide dismutase to the MCLA system completely abolished the incremental chemiluminescence intensity after both UVA and blue light exposure of the SC. In addition, acrolein-treated BSA gave ESR signals like hydroxyl radicals (OH) converted from superoxide anion radicals (O2−) during irradiation with a xenon arc lamp containing UVA and visible light. From the sum of these results, we consider that CPs are produced from O2− initially generated from exposure to UVA and blue light. ConclusionCPs are excited by absorbing sunlight, particularly UVA and blue light, and result in the generation of O2− through a CPs progress new protein carbonylation in stratum corneum through ROS generation. photosensitizing reaction. Further, the results suggest that the O2− produces CPs in the SC through lipid peroxidation in the sebum, and finally affects skin conditions including color and moisture functions.

ITRAQ-based quantitative proteomics of stratum corneum of dandruff scalp reveals new insights into its aetiology and similarities with atopic dermatitis.

The study aimed at detecting differentially expressed proteins in the stratum corneum of dandruff versus non-dandruff scalps to better understand dandruff aetiology. iTRAQ-based quantitative proteomic analysis revealed a total of 68 differentially expressed biomarkers. A detailed analysis of their known physiological functions provided new insights into the affected metabolic pathways of a dandruff scalp. Dandruff scalp showed (1) profound changes in the expression and maturation of structural and epidermal differentiation related proteins, that are responsible for the integrity of the skin, (2) altered relevant factors that regulate skin hydration, and (3) an imbalanced physiological protease-protease inhibitor ratio. Stratum corneum proteins with antimicrobial activity, mainly those derived from sweat and sebaceous glands were also found modified. Comparing our data with those reported for atopic dermatitis revealed that about 50% of the differentially expressed proteins in the superficial layers of the stratum corneum from dandruff and atopic dermatitis are identical.

The effects of polar excipients transcutol and dexpanthenol on molecular mobility, permeability, and electrical impedance of the skin barrier

In the development of transdermal and topical products it is important to understand how formulation ingredients interact with the molecular components of the upper layer of the skin, the stratum corneum (SC), and thereby influence its macroscopic barrier properties. The aim here was to investigate the effect of two commonly used excipients, transcutol and dexpanthenol, on the molecular as well as the macroscopic properties of the skin membrane. Polarization transfer solid-state NMR methods were combined with steady-state flux and impedance spectroscopy measurements to investigate how these common excipients influence the molecular components of SC and its barrier function at strictly controlled hydration conditions in vitro with excised porcine skin. The NMR results provide completely new molecular insight into how transcutol and dexpanthenol affect specific molecular segments of both SC lipids and proteins. The presence of transcutol or dexpanthenol in the formulation at fixed water activity results in increased effective skin permeability of the model drug metronidazole. Finally, impedance spectroscopy data show clear changes of the effective skin capacitance after treatment with transcutol or dexpanthenol. Based on the complementary data, we are able to draw direct links between effects on the molecular properties and on the macroscopic barrier function of the skin barrier under treatment with formulations containing transcutol or dexpanthenol.

Advantage of Sodium Polyoxyethylene Lauryl Ether Carboxylate as a Mild Cleansing Agent

AbstractFive anionic surfactants widely used in commercial skin cleansers were studied: sodium polyoxyethylene lauryl ether carboxylate (EC), sodium polyoxyethylene alkyl ether sulfate (ES), sodium dodecyl sulfate, potassium laurate, and N‐cocoyl‐l‐glutamic acid monosodium salt. The amount of surfactant from aqueous solution adsorbed into the stratum corneum (SC), the degree of SC swelling, the change of the secondary structure of SC proteins (denaturation). The surface tension of surfactant–zein mixed solutions, and the solubilization behavior of zein were measured. Results showed that EC had the lowest adsorption into SC, the lowest SC swelling, and lowest denaturation of SC proteins. Low interactions between surfactants and SC proteins were also observed for EC/ES mixture solutions as well as. Mixing EC with ES good foaming performance. The EC/ES mixture, at about 1:1 ratio, is an excellent surfactant system for skin cleanser applications having cleansing characteristics and mildness to the skin.

Tape Stripping Technique for Stratum Corneum Protein Analysis.

The aim of this study was to investigate the amount of protein in stratum corneum in atopic dermatitis (AD) patients and healthy controls, using tape stripping technique. Furthermore, to compare two different methods for protein assessment. Tape stripping was performed in AD patients and healthy controls to collect stratum corneum samples and subsequently analysed with two different methods: Squame Scan, which gives an estimate of total protein (soluble and insoluble) and Micro BCA protein determination kit which measures soluble protein. Significant differences in cumulative protein content between AD lesional, AD non-lesional and healthy control skin was found using the Squame Scan as well as the Micro BCA protein determination kit. AD patients had significantly lower amount of protein, both total protein and soluble protein compared to healthy controls. Furthermore, soluble protein formed 82% of total protein in AD lesional skin, compared to 17–24% for AD non-lesional skin and healthy control. A decreasing amount of total protein with increasing stratum corneum depth was found for all skin types. Significant differences in stratum corneum protein content between AD lesional, AD non-lesional and healthy control skin were revealed, independent of method used.

Keratins K2 and K10 are essential for the epidermal integrity of plantar skin

BackgroundK1 and K2 are the main type II keratins in the suprabasal epidermis where each of them heterodimerizes with the type I keratin K10 to form intermediate filaments. In regions of the ears, tail, and soles of the mouse, only K2 is co-expressed with K10, suggesting that these keratins suffice to form a mechanically resilient cytoskeleton. ObjectiveTo determine the effects of the suppression of both main keratins, K2 and K10, in the suprabasal plantar epidermis of the mouse. MethodsKrt2−/− Krt10−/− mice were generated by crossing Krt2−/− and Krt10−/− mice. Epidermal morphology of soles of hind-paws was examined macroscopically and histologically. Immunofluorescence analysis and quantitative PCR analysis were performed to analyze the expression of keratins in sole skin of wildtype and Krt2−/− Krt10−/− mice. Highly abundant proteins of the sole stratum corneum were determined by electrophoretic and chromatographic separation and subsequent mass spectrometry. ResultsK2 and K10 are the most prominent suprabasal keratins in normal mouse soles with the exception of the footpads where K1, K9 and K10 predominate. Mice lacking both K2 and K10 were viable and developed epidermal acanthosis and hyperkeratosis in inter-footpad epidermis of the soles. The expression of keratins K1, K9 and K16 was massively increased at the RNA and protein levels in the soles of Krt2−/− Krt10−/− mice. ConclusionsThis study demonstrates that the loss of the main cytoskeletal components of plantar epidermis, i.e. K2 and K10, can be only partly compensated by the upregulation of other keratins. The thickening of the epidermis in the soles of Krt2−/− Krt10−/− mice may serve as a model for pathomechanistic aspects of palmoplantar keratoderma.

Variation in stratum corneum protein content as a function of anatomical site and ethnic group.

Quantification of stratum corneum (SC) protein levels from tape strippings is frequently used to investigate skin conditions, to correct for amounts of SC protein removed in SC biomarker studies and to determine distribution of topically applied ingredients. In recent years, a rapid and convenient method for SC protein quantification from tape strippings has become available using infrared densitometry (IRD). However, standard curves have only been generated for Caucasian forearm and shoulder SC and have been assumed to be correct not only for facial SC but also for SC samples of other ethnic groups. The aim of this study was to investigate whether the use of IRD for SC protein measurement is valid for other body sites such as the cheek and for measuring SC protein content of darkly pigmented skin types. Ten Caucasian and ten Black African female subjects with self-assessed normal skin participated in the study. Tape strippings were collected from two different body sites (forearm and cheek). First from the tape strippings, the SC optical absorption was determined densitometrically. This obtained absorption (%) was compared with absolute SC protein extracted from the same tapes using a colorimetric microbicinchoninic acid (μBCA) assay. Higher amounts of SC protein were removed from the forearm compared with the cheek (P < 0.01). The absolute SC protein concentration quantified by μBCA assay and the absorption of SC proteins by IRD followed a similar profile. There was no significant difference found between the two ethnicities in SC protein (P > 0.05). The overall coefficient of determination (R(2) ) shows a good fit to the regression line between the two methods in both sites (forearm = 0.82, cheek = 0.77). Also, both ethnicities showed good correlation (R(2) ≥ 0.69, P = 0.01). Facial SC is morphologically distinct from the forearm, as demonstrated by the differences in amounts of SC removed. Although the data distribution in different subject groups varied, the regression was always quite similar between the two body sites and both ethnic groups. Also, the correlations were similar to previously published data on other body sites. The resultant calibration curves can be used as a rapid indirect protein assessment of tape strippings from the cheek.

Assessing the In Vivo Epidermal Barrier in Mice: Dye Penetration Assays

INTRODUCTION The formation of a properly functioning epidermal barrier is a prerequisite for terrestrial life. The epidermis not only protects from external influences such as pathogens, chemicals, and UV light, but also prevents dehydration. To maintain proper barrier function, the epidermis undergoes a dynamic turnover driven by proliferating keratinocytes in the basal layer that, upon induction of differentiation, will move upward through the stratum spinosum and stratum granulosum while undergoing a terminal differentiation process to ultimately form the stratum corneum (SC). This layer is in direct contact with the outside environment and provides important structural and innate immune barrier properties. Barrier function is initiated when stratum granulosum keratinocytes start secreting and crosslinking specific proteins and lipids into the intercellular space and at the same time transform their membrane into a so-called cornified envelope (e.g., by linking different structural proteins such as loricrin to the inner side of the cell membrane). Consequently, keratinocytes shed their nucleus to become corneocytes forming the SC. In recent years it has become clear that the SC is not the sole structure providing barrier properties to the epidermis. It is now clear that specialized intercellular junctions, called tight junctions (TJs), are present between cells of the granular layer, where they ensure proper barrier function (Furuse et al., 2002; Tunggal et al., 2005). TJs have been well characterized in simple epithelia where they form a paracellular sizeand ion-specific barrier to separate tissue compartments. The size and ion selectivity of TJs is mainly based on which members of the claudin family of transmembrane proteins are located in the TJs of different tissues. The epidermis expresses several claudins, and loss of claudin-1 in mice results in perinatal lethality attributable to rapid dehydration (Furuse et al., 2002). In addition, TJs are crucial for immune surveillance by Langerhans cells (Kubo et al., 2009). The importance of the two barriers is further confirmed by observations that mutations in SC proteins such as filaggrin are linked to diseases characterized by an impaired skin barrier, such as atopic dermatitis and ichthyosis vulgaris (Palmer et al., 2006; Smith et al., 2006). In addition, human claudin-1 mutations are associated with neonatal sclerosing cholangitis associated with ichthyosis (Hadj-Rabia et al., 2004). However, the exact molecular mechanisms of how TJs and the SC contribute to the formation, maintenance, and restoration of the skin barrier are not well understood. In addition, it is not known whether and how TJs and the SC cooperate to form a fully functional skin barrier. Transgenic mouse models have been helpful tools to investigate how proteins implicated in epidermal barrier function contribute to the formation and maintenance of skin barrier function. Next to assessing whether such mice have a BENEFITS OF DYE PENETRATION ASSAYS

Facial skin pigmentation is not related to stratum corneum cohesion, basal transepidermal water loss, barrier integrity and barrier repair.

Hypotheses have been developed for the evolutionary selection of skin pigmentation one of which relates to improved skin barrier function. The aim of this study was to compare facial skin condition on photoexposed (cheek) and photoprotected (post-auricular) sites of naturally pigmented subjects of different ethnicities (Fitzpatrick skin phototypes II/III and V/VI) and Albino African subjects to understand better the relationship between facial stratum corneum (SC) barrier function, skin surface pH and skin pigmentation. Expert grading of skin conditions, capacitance, skin surface pH and skin barrier function measurements were performed. For the latter, transepidermal water loss (TEWL) measurements before (basal TEWL), after 3, 6 and 9 consecutive tape strippings (SC integrity) and 3.5 and 24 h post tape stripping (barrier recovery) were taken. Amounts of SC protein removed during stripping were estimated using infrared densitometry (SC cohesion). Firstly, correlation analysis of the biometric data of the Black African and Caucasian subjects showed there to be no relationship between skin surface pH and ITA° values nor pH and ITA° with basal TEWL. Neither skin surface pH nor ITA° correlated with SC integrity and barrier recovery measurements, but skin surface pH correlated with SC cohesion. ITA° values were correlated with skin hydration. Secondly, on comparing the three ethnic groups, severe skin photodamage was observed in the Albino African subjects and their SC was thicker. Whereas their basal TEWL was elevated, superior values for SC integrity and barrier recovery were measured. No differences in basal TEWL, SC integrity and barrier recovery were found between the other two subject groups. Equally, SC cohesion and skin surface pH values were similar among the three groups. There was no relationship between ITA° values and basal TEWL, SC integrity, SC cohesion and barrier recovery, but ITA° was correlated with skin hydration. Skin surface pH, irrespective of ITA° values, correlated with SC cohesion, indicating a greater intracorneal cohesion at lower pH values. Thus, pigmentation has no effect on SC barrier properties but was related to skin hydration. On comparing the three ethnic groups, Albino African SC was found to be superior to the Caucasian and Black African subjects in terms of SC integrity and barrier recovery but not basal TEWL. The Albino African subjects also have a thicker SC which contributes to their better SC integrity. No differences in skin barrier functionality or skin surface pH were observed for the other two groups. Skin hydration was, however, greatest in the Black African subjects. Our data support the evolutionary hypothesis that pigmentation protects the skin from UV irradiation and thereby the skin barrier but not the skin pigmentation-/pH-driven adaptive skin barrier hypothesis.

Proteome analysis of stratum corneum from atopic dermatitis patients by hybrid quadrupole-orbitrap mass spectrometer

Proteome analysis of stratum corneum from atopic dermatitis patients by hybrid quadrupole-orbitrap mass spectrometer

Late Cornified Envelope Group I, a Novel Target of p53, Regulates PRMT5 Activity

p53 is one of the most important tumor suppressor genes involved in human carcinogenesis. Although downstream targets of p53 and their biologic functions in cancer cells have been extensively investigated, it is still far from the full understanding. Here, we demonstrate that Late Cornified Envelope Group I (LCE1) genes, which are located in the LCE gene clusters encoding multiple well-conserved stratum-corneum proteins, are novel downstream targets of p53. Exogenous p53 overexpression using an adenoviral vector system significantly enhanced the expression of LCE1 cluster genes. We also observed induction of LCE1 expressions by DNA damage, which was caused by treatment with adriamycin or UV irradiation in a wild-type p53-dependent manner. Concordantly, the induction of LCE1 by DNA damage was significantly attenuated by the knockdown of p53. Among predicted p53-binding sites within the LCE1 gene cluster, we confirmed one site to be a p53-enhancer sequence by reporter assays. Furthermore, we identified LCE1 to interact with protein arginine methyltransferase 5 (PRMT5). Knockdown of LCE1 by specific small interfering RNAs significantly increased the symmetric dimethylation of histone H3 arginine 8, a substrate of PRMT5, and overexpression of LCE1F remarkably decreased its methylation level. Our data suggest that LCE1 is a novel p53 downstream target that can be directly transactivated by p53 and is likely to have tumor suppressor functions through modulation of the PRMT5 activity.

The Effect of Formulation Excipients on the Penetration and Lateral Diffusion of Ibuprofen on and within the Stratum Corneum Following Topical Application to Humans

Distribution profiles of topically applied drugs can be influenced by the presence of excipients. This study investigated the effect of common topical excipients on the simultaneous lateral diffusion and stratum corneum (SC) penetration of a model compound, ibuprofen (IBU) in humans. IBU solutions with and without propylene glycol (PG), polyethylene glycol 200 (PEG 200), and/or octisalate (OS) were dosed onto the forearm of participants. At various times, 10 "tape-strippings" were obtained with perforated concentric tapes and analyzed for IBU concentration and SC protein mass. Complimentary in vitro permeation studies assessed the effect of excipients on the percutaneous absorption of IBU across human skin. Following in vivo application, IBU displayed a greater tendency for lateral diffusion when applied with OS, whereas IBU resisted lateral diffusion when dosed with PG and PEG 200. After 24 h, 25.3 ± 8.0% and 55.5 ± 18.6% of IBU was recovered from the SC in vivo with and without excipients, respectively. There was a twofold-to threefold enhancement in IBU flux in vitro when applied with excipients. The lower IBU recovery from the SC when applied with excipients may be attributed to the permeation enhancement effects of these excipients. The ability of IBU to laterally diffuse appears to be dependent on formulation excipients.

Defective barrier function accompanied by structural changes of psoriatic stratum corneum

Although barrier function of psoriatic skin is shown to be decreased by measuring transepidermal water loss (TEWL), few reports exist examining other physical skin properties and components including stratum corneum hydration, natural moisturizing factor (NMF), free fatty acids (FFA), β-sheet and α-helix ratio of structural protein(s), and sebum content. We compared the skin properties and components of normal, involved and uninvolved skin of psoriasis. Using a corneometer and attenuated total reflection-infrared spectrometer, we measured TEWL, stratum corneum hydration, NMF, FFA, β/α ratio and sebum in psoriasis vulgaris patients and healthy controls. TEWL and β/α ratio of involved psoriatic skin were significantly increased compared with uninvolved skin and normal control skin. In contrast, stratum corneum hydration, NMF and FFA, but not sebum, are significantly decreased in the involved skin compared with uninvolved skin and normal skin. TEWL and stratum corneum hydration returned to the normal levels following clinical improvement of the lesion. Barrier function and hydration of psoriatic skin are defective and secondary structure in stratum corneum protein is altered in the involved psoriatic skin.

In vivo studies of aquaporins 3 and 10 in human stratum corneum

Aquaporins (AQPs) constitute one family of transmembrane proteins facilitating transport of water across cell membranes. Due to their specificity, AQPs have a broad spectrum of physiological functions, and for keratinocytes there are indications that these channel proteins are involved in cell migration and proliferation with consequences for the antimicrobial defense of the skin. AQP3 and AQP10 are aqua-glyceroporins, known to transport glycerol as well as water. AQP3 is the predominant AQP in human skin and has previously been demonstrated in the basal layer of epidermis in normal human skin, but not in stratum corneum (SC). AQP10 has not previously been identified in human skin. Previous studies have demonstrated the presence of AQP3 and AQP10 mRNA in keratinocytes. In this study, our aim was to investigate if these aquaporin proteins were actually present in human SC cells. This can be seen as a first step toward elucidating the possible functional role of AQP3 and AQP10 in SC hydration. Specifically we investigate the presence of AQP3 and AQP10 in vivo in human SC using "minimal-invasive" technique for obtaining SC samples. SC samples were obtained from six healthy volunteers. Western blotting and immunohistochemistry were used to demonstrate the presence of AQP3 as well as AQP10. The presence of AQP3 and AQP10 was verified by Western blotting, allowing for detection of proteins by specific antibodies. Applying immunohistochemistry, cell-like structures in the shape of corneocytes were identified in all samples by AQP3 and AQP10 antibodies. In conclusion, identification of AQP3 and AQP10 protein in SC in an in vivo model is new. Together with the new "minimal-invasive" method for SC collection presented, this opens for new possibilities to study the role of AQPs in relation to function of the skin barrier.

Contribution of Tight Junction Proteins to Ion, Macromolecule, and Water Barrier in Keratinocytes

Tight junctions (TJs) form a selective barrier for ions, water, and macromolecules in simple epithelia. In keratinocytes and epidermis, TJs were shown to be involved in individual barrier functions. The absence of the TJ protein claudin-1 (Cldn1) in mice results in a skin-barrier defect characterized by lethal water loss. However, detailed molecular analyses of the various TJ barriers in keratinocytes and the contribution of distinct TJ proteins are missing. Herein, we discriminate TJ-dependent paracellular resistance from transcellular resistance in cultured keratinocytes using the two-path impedance spectroscopy. We demonstrate that keratinocyte TJs form a barrier for Na(+), Cl(-), and Ca(2+), and contribute to barrier function for water and larger molecules of different size. In addition, knockdown of Cldn1, Cldn4, occludin, and zonula occludens-1 increased paracellular permeabilities for ions and larger molecules, demonstrating that all of these TJ proteins contribute to barrier formation. Remarkably, Cldn1 and Cldn4 are not critical for TJ barrier function for water in submerged keratinocyte cultures. However, Cldn1 influences stratum corneum (SC) proteins important for SC water barrier function, and is crucial for TJ barrier formation for allergen-sized macromolecules.

The pilosebaceous unit—a phthalate-induced pathway to skin sensitization

Allergic contact dermatitis (ACD) is caused by low-molecular weight compounds called haptens. It has been shown that the potency of haptens can depend on the formulation in which they are applied on the skin. Specifically the sensitization potency of isothiocyanates, a group of haptens which can be released from e.g. adhesive tapes and neoprene materials, increases with the presence of phthalates; however, the underlying mechanisms are not clear. A better understanding of the mechanisms governing the potency of haptens is important, e.g. to improve the risk assessment and the formulation of chemicals in consumer products. In this study we have explored phthalate-induced effects on the sensitization potency, skin distribution, and reactivity of fluorescent model isothiocyanate haptens using non-invasive two-photon microscopy to provide new insights regarding vehicle effects in ACD. The data presented in this paper indicate that the sensitization potency of isothiocyanates increases when applied in combination with dibutylphthalate due to a specific uptake via the pilosebaceous units. The results highlight the importance of shunt pathways when evaluating the bioavailability of skin sensitizers. The findings also indicate that vehicle-dependent hapten reactivity towards stratum corneum proteins regulates the bioavailability, and thus the potency, of skin sensitizers.

Th1 cytokines accentuate but Th2 cytokines attenuate ceramide production in the stratum corneum of human epidermal equivalents: An implication for the disrupted barrier mechanism in atopic dermatitis

Although the mechanism(s) involved in the ceramide deficiency in the stratum corneum (SC) of atopic dermatitis (AD) skin is unknown, Th2 type cytokines have been reported to down-regulate ceramide levels in the epidermis. However, almost all research to date has focused on ceramide levels in the whole epidermis, not just in the SC layers alone, which are predominantly responsible for the skin barrier function. We highlighted the effects of Th1/Th2 cytokines on ceramide levels in the SC. We developed a modified system of human epidermal equivalents in which epidermis without a SC is cultured for 1 week to generate complete SC layers after which ceramides are extracted from the separated SC layers and are then quantified as per SC protein. The addition of Th2 cytokines (IL-4/IL-6) at a concentration of 10nM resulted in a marked decrease in SC ceramide levels. The reduced ceramide content in the SC was accompanied by the down-regulated expression of the genes encoding serine-palmitoyl transferase-2, acid sphingomyelinase and β-glucocerebrosidase in the epidermis. In contrast, the addition of Th1 cytokines (GM-CSF/IFN-γ/TNF-α) at concentrations of 2.5 or 10nM resulted in a slight increase in SC ceramide levels, which were accompanied by no change or an increase in the expression of those genes encoding sphingolipid metabolic enzymes in the epidermis. These findings suggest that the Th2 type of inflammation evoked in AD skin is one of the essential factors involved in down-regulating the levels of ceramide in the SC.

Comparison of gravimetric and spectroscopic approaches to quantify stratum corneum removed by tape-stripping

Skin surface tape-stripping is an extensively used technique to examine the distribution profile, penetration and safety of various active compounds. It is also a widely accepted method to probe skin barrier properties and more specifically, those of the stratum corneum (SC). The amount of SC removed by tape-stripping is generally determined either gravimetrically or by extraction and measurement of SC proteins. A novel infra-red densitometry (IRD) technique has recently been introduced to measure SC protein content. In the present study, IRD was investigated as an alternative method to measure the mass of SC removed by tape-stripping. Tape-stripping experiments were conducted on human volunteers. The weight of the stratum corneum removed was assessed by the gravimetric approach and by IRD. Transepidermal water loss (TEWL) was also measured before and after each tape-strip. A linear correlation coefficient was obtained for the data from the gravimetric and IRD measurements (r2=0.65; n=240). IRD is therefore proposed as a rapid, non-destructive alternative to the gravimetric approach to estimate the amount of SC removed by tape-stripping in vivo.

Skin integrity testing and monitoring of in vitro tape stripping by capacitance‐based sensor imaging

Despite the frequent use of porcine ear skin for tape-stripping experiments, the peculiarities of this skin type have not been characterised in detail yet. Thus, different techniques were employed to investigate the skin surface structure of porcine ear skin and the changes in barrier function during in vitro tape stripping. To this end, the potential of capacitance-based skin hydration imaging as a means of skin quality control was investigated for the first time. The porcine ear model was characterised before and during tape stripping using transepidermal water loss (TEWL) measurements, capacitance-based sensor imaging, attenuated total reflection Fourier transform infrared spectroscopy (ATR-FTIR) and optical light microscopy. The capacitance-based sensor was found to deliver precise information about the quality of the employed skin sites before and during tape stripping. The removal of stratum corneum proteins was highly reproducible even for different porcine ear types. The mean greyscale values showed an excellent linear correlation to the corresponding TEWL values and the respective penetration depth. Optical light microscopy confirmed the presence of canyons on the surface of porcine ear skin. The results suggest that the capacitance-based sensor is a suitable tool for skin integrity testing of porcine ear skin in vitro and for monitoring changes in skin barrier function.