Hydration of the stratum corneum

Topically applied water, occlusion and topically applied glycerol were used to investigate and characterize some of the changes which occur in the hydrated stratum corneum. The effects of these treatments were monitored using non-invasive techniques under controlled conditions. The Servomed Evaporimeter was used to determine natural water flux from the skin surface before and after treatment. The performance of the Evaporimeter in this type of study had previously been improved by attaching a paper baffle to the detector. This eliminated the variance in output caused by atmospheric movement. Experiments were carried out at temperatures below the threshold of thermal sweating and emotional sweating was minimized. Skin surface topography was characterized by means of a new type of profilometer. The instrument's design allowed a diamond stylus to traverse the living skin surface without significantly altering its structure. Changes in skin surface roughness were further elucidated using scanning electron microscopy and macrophotography. In vivo penetration of glycerol was assessed by chemical analysis of stratum corneum layers of treated skin. Samples were obtained by sequential stripping of the stratum corneum using adhesive tape. Topically applied water produced only a transient benefit because of rapid evaporation. More prolonged hydration was achieved by suppressing transepidermal water loss with polyethylene film. This occlusive hyperhydration was characterized by a significant reduction in profile roughness and by a smoother macroscopic appearance. Glycerol achieved the same effects by reducing the magnitude of the natural water flux from the skin surface and by reducing the rate of evaporation of water from applied aqueous glycerol solution or cosmetic product. Both effects were seen as the result of lowered water activity in the proximity of glycerol. Smoothing effects of glycerol on the skin surface, and improved appearance, persisted for at least 24 h. This persistence was explained by evidence for diffusion of glycerol into the stratum corneum where it formed a reservoir. Hydration of the skin is known to affect its barrier function and thereby exert a profound effect on penetration of both lipophilic and hydrophilic molecules. Clinically, this effect may be achieved using liberal applications of occlusive petroleum jelly and ointments. The results presented in this paper suggest that the use of humectants could achieve useful hydration using cosmetically acceptable materials.     

Squamometry: a tool to move from exaggerated to more and more realistic application conditions for comparing the skin compatibility of surfactant-based products

Squamometry consists of sampling the most superficial layers of the stratum corneum by means of a sticky tape firmly pressed onto the skin. Harvested cells are stained and the intensity of staining is related to the level of skin surface alterations caused by surfactant-based products. This method is extremely sensitive and the current study demonstrates that, thanks to squamometry, product mildness can now be compared without causing skin irritation due to exaggerated application conditions. In a 15 minute patch test, squamometry assessments ranked a series of surfactants as expected from their known irritation potential, without causing clinical signs of irritation. Similarly, the counter-irritant effect of amphoteric and nonionic surfactants on the irritation potential of anionic surfactants was evidenced. In a 30 minute patch test, shampoos were ranked by squamometry as they were in a classical Soap Chamber Test (48 hours of application). Finally, four hand dishwashing liquids were tested for one week by consumers according to a procedure similar to normal usage of the products, and were classified by squamometry as they were in a previous Soap Chamber Test. In conclusion, this study suggests that squamometry is able to predict the skin compatibility of surfactant-based products even after very short and realistic application times to volunteers' skin. Providing further validation, this technique could allow a move to test conditions much closer to the normal usage procedure of the products, to design test protocols much more respectful of panelists' skin condition, and to get information on product tolerance very quickly.     

Infrared spectroscopic studies of sodium dodecyl sulphate permeation and interaction with stratum corneum lipids in skin

The barrier function of skin is primarily provided by the lamellar lipid matrix of the stratum corneum (SC), which has been shown in previous infrared (IR) and related studies to consist predominantly of ordered lipids packed in orthorhombic and hexagonal domains. In the current work, we investigate the effects of the anionic surfactant, sodium dodecyl sulphate (SDS), on SC lipid packing and phase behaviour, using FT-IR spectroscopy. The use of acyl chain perdeuterated SDS allows unequivocal spectroscopic detection of both endogenous lipid and exogenous material in intact tissue. IR spectra were acquired as a function of temperature from isolated human SC exposed to SDS for various incubation periods at 34°C. SDS is found to enter the SC and is observed to be in a more ordered state in the SC than in solution, indicating that the SDS interacts with the ordered SC lipids. The results reveal that SDS reduces the amount of orthorhombic phase in the SC and increases the amount of hexagonally packed lipid at physiologically relevant temperatures. In addition, a small decrease in the lipid T(m) (acyl chain melting temperature) is observed. Furthermore, these SDS-induced changes were found to be strongly dependent on the time of exposure.     

Change in optical properties of stratum corneum induced by protein carbonylation in vitro

The skin is the frontier against the external environment and continuously exposed to the environmental oxidative stress such as ultraviolet (UV) irradiation. Protein carbonyls are the major oxidative products of protein and may be introduced by reaction with aldehydes derived from lipid peroxide. Acrolein is one of the most reactive aldehydes generated during degradation of lipid peroxides and protein-acrolein adducts have been found in the oxidatively damaged lesion including UV-damaged skin. Recent studies revealed that protein carbonyls are also detected in thin outermost layer of the skin, the stratum corneum (SC). However, the effect of protein carbonylation on the function of SC was still unclear. In this study, we treated the SC sheets of reconstructed human epidermis and porcine epidermis with acrolein in the experimental conditions to explore the influence of protein carbonylation on the SC. Human and porcine SC sheets treated with acrolein showed less transmission at visible light than untreated SC sheets. Attenuated total reflection-infrared spectroscopy with curve fitting analysis of amide I region showed that acrolein induced alterations in protein secondary structure of the porcine SC sheets, which were accompanied by diminished fibrous keratin structure observed by transmission electron microscopy. These results show the possibility that carbonylation of the SC caused by environmental factors is one of factors altering the fibrous structure of keratin and decreasing the light transmission of SC, which changes the quality of the skin appearance.     

Profiling of serine protease activities in human stratum corneum and detection of a stratum corneum tryptase-like enzyme

Seasonal variation in stratum corneum (SC) biophysical and biological characteristics has been described previously. In particular, the winter season has been shown to affect more severely the properties of facial skin compared with forearm skin. Moreover, when compromised, such as in dry skin conditions, facial SC has been shown to contain increased inflammatory cytokines and proteases. Nevertheless, there have been no comparative studies of the activities and depth activity of several proteases in the SC on different body sites and at different times of the year. In this study, we examined the distribution of key serine protease activities (kallikrein 5, kallikrein 7, urokinase, plasmin and a tryptase-like enzyme) in different layers of the SC on the cheek and the forearm by analysis of consecutive tape strippings of healthy Caucasian subjects during winter and summer. The protein content of the tape strippings was quantified by absorption measurements with a recently developed and novel infrared densitometer SquameScan 850A while the SC enzyme activities were determined using fluorogenic peptide substrates. Transepidermal water loss (TEWL), skin pH and skin hydration were higher on the cheek than on the forearm. In the same way, the activity of the inflammatory-related proteases plasmin, urokinase and tryptase was approximately five to eight times and the activity of the desquamatory-related proteases kallikrein 5 and kallikrein 7 approximately two to four times higher on the cheek than on the forearm. There were no gender-related differences in these enzyme activities except for the increased kallikrein 7 in the forearm skin of the female subjects in winter. Reduced kallikrein 5 was associated with increased SC cohesion, as judged by increased protein removal, in forearm skin in the winter months of the year although the skin was clinically normal. It can be concluded that (i) protected skin areas show lower TEWL, skin pH and skin hydration and less protease activities than skin areas that are exposed to the environment, possibly indicating subclinical inflammation on these body sites, (ii) in normal healthy forearm skin, the outer SC exhibits greater serine protease activity than its deeper layers, (iii) compared with the forearm, urokinase- and plasmin-like activities are elevated on SC strippings from the cheek, confirming activation of the plasminogen cascade, and (iv) tryptase-like activity in the SC is also elevated in samples from the cheek, possibly indicating involvement of mast cells in these barrier-compromised body sites or the synthesis of a novel tryptase-like enzyme by keratinocytes. Although elevation of the activities of urokinase, plasmin, kallikrein 5, kallikrein 7 and now a tryptase-like enzyme was observed on SC derived from skin of clinically normal cheeks, we anticipate even higher activities in skin conditions where the epidermal barrier is further impaired.     

Reduced barrier efficiency in axillary stratum corneum

The skin of the axilla is cosmetically important with millions of consumers daily applying antiperspirant/deodorant products. Despite this, we know virtually nothing about axillary skin or how antiperspirant (AP) use impacts upon it. To characterize the axillary stratum corneum and determine whether this is a unique skin type, we have looked at stratum corneum composition and function, particularly its barrier properties, and compared it with other body sites. Transepidermal water loss (TEWL) and corneosurfametry (CSM) revealed a reduced barrier function in the axilla. HPTLC analysis of the stratum corneum lipids demonstrated statistically elevated levels of fatty acids, ceramides, and particularly cholesterol in the axilla. Both ceramide and cholesterol did not appear to change with depth, indicating that they were predominantly of stratum corneum origin. On the other hand, at least some of the fatty acid had a sebaceous origin. We hypothesized that the reduced barrier function might be owing to the changes in the crucial ceramide : cholesterol ratio. To address this, we used a combination of attenuated total reflectance-Fourier-transformed infrared spectroscopy (ATR-FTIR) with cyanoacrylate sampling. These results demonstrated more ordered lipid-lamellae phase behaviour in the axilla, suggesting that the elevated cholesterol might form crystal microdomains within the lipid lamellae, allowing an increase in water flux. Since an exaggerated application of antiperspirant had no effect upon the axilla barrier properties, it is concluded that this region of skin physiologically has a reduced barrier function.     

A study of the interaction of sodium dodecyl sulphate with the proteins of human heel stratum corneum

The aim of this research is to study the in vitro interaction of the anionic surfactant sodium dodecyl sulphate with human heel callus with special regard to the callus proteins.
The sorption of SDS by callus is analysed at different pH values, demonstrating the expected maximum binding of the anionic surfactant at low pH. In both surfactant and blank solutions, swelling of the callus pieces occurs but to different extents. The sorption of SDS by callus is accompanied by a loss of free amino acids and proteins. The protein composition of the callus residues after chemical treatment and of the corresponding treatment baths is examined by amino acid analysis and shows differences in the respective molar amino acid ratios. Results obtained with more specific techniques, such as gel electrophoresis and immunoblotting, demonstrate identical as well as different protein components in the treatment baths depending on the experimental conditions (pH, blank or SDS). Although effects due to the surfactant treatment are in principle more distinct than with blank experiments, those of the latter cannot be neglected.     

Effect of shaving on axillary stratum corneum

Removal of underarm hair is an intrinsic part of the care regimen for the majority of female consumers, with most using a wet shave with a disposable razor. However, little is known of the impact of shaving on axillary skin, and it is a particularly neglected area of research. To investigate this, we have studied the acute and chronic effects of shaving ultrastructurally, biochemically and functionally. A forearm patch test protocol was devised for antiperspirant (AP) product screening, which involved a pre-shave of the test site with a dry razor just prior to patching. Comparison of the irritation caused by a series of AP products confirmed that shaving leads to increased irritation consistent with enhanced sensitivity. The effect of regular shaving in the axilla was assessed in a 4-week in-use study with shaving either once a week or once a day, both combined with the application of an AP. Expert visual assessment of skin condition showed that more frequent shaving promoted a higher level of visible irritation. However, indirect measurement using corneosurfametry indicated no significant changes to the lipid barrier over the study period irrespective of shaving frequency. Nevertheless, digital images of the axillary skin after dry shaving show distinct opaque lines because of uplifting skin flakes with a corresponding increase in scaliness parameter. Moreover, histamine iontophoresis to assess skin sensitivity demonstrated a significant enhancement of histamine-induced itch and neurogenic flare.     

Approche osmotique de l'hydratation du stratum corneum

Afin de reprendre l'hypothèse selon laquelle l'absorption de l'eau par le stratum corneum est un phénomène osmotique et afin de d'effectuer des études, in vitro , de la biophysique de la diffusion de l'eau par osmose, nous avons mis au point un Modèle Osmotique de Cornéocyte (MOC), qui est constitué par de la kératine isolée par une membrane semi-perméable.
Immeré dans de l'eau, le MOC voit sa masse augmenter. En effet, bien qu'insoluble, la kératine est hydrophile et en accord avec le Deuxième principe de la thermodynamique, elle fait diffuser par osmose, de l'eau vers l'intérieur du MOC.
A saturation, l'eau absorbée représente 13 fois la masse de kératine.
La diffusion s'effectue selon une cinétique classique: la masse de l'eau qui pénètre est proportionelle à la racine carrée du temps.
Les éléments constitutifs du Facteur Naturel d'Hydratation comme l'urée, l'acide lactique, l'acide pyrrolidone carboxylique, accroissent la vitesse de diffusion et la quantité totale d'eau absorbée. Un humectant comme la glycérine produit un effet semblable. L'urée est le composé qui donne les meilleures performances.
L'accroissement de la vitesse de diffusion par osmose dans le MOC en présence de composés hydratants peut s'interpréter comme un accroissement de l'hydrophilie de la kératine induite par ces composés.
Le MOC semble pouvoir permettre, outre une étude théorique sur le phénomène osmotique dans le stratum corneum , des études préliminaires in vitro des produits hydratants hydrosolubles utilisables en cosmétologie.     

Drying stress and damage processes in human stratum corneum

The drying stresses that develop in stratum corneum (SC) are crucial for its mechanical and biophysical function, its cosmetic feel and appearance, and play a central role in processes of dry skin damage. However, quantitative methods to characterize these stresses are lacking and little understanding exists regarding the effects of drying environment, chemical exposures and moisturizing treatments. We describe the application of a substrate curvature technique adapted for biological tissue to accurately characterize SC drying stresses as a function of time following environmental pre‐conditioning and chemical treatment in a range of drying environments. SC stresses were observed to increase to stress levels of up to ～ 3 MPa over periods of 8 h depending on pretreatment and drying environment. A unique relationship between the SC stress and water in the drying environment was established. The effect of glycerol on lowering SC stresses and damaging surfactants on elevating SC stresses were quantified. Extensions of the method to continuous monitoring of SC stresses in response to changes in environmental moisture content and temperature are reported. Finally, a biomechanics framework to account for the SC drying stress as a mechanical driving force for dry skin damage is presented.     

Les enzymes de l'espace extra-cellulaire du stratum corneum

Bien que le stratum corneum soit composé de cellules ‘mortes’, il est le siège d'une activité métabolique très importante. Mais, contrairement à la plupart d'autres tissus, cette activité a la particularité d'étre extra-cellulaire. Elle est due à des enzymes excrétées par les Corps d'Odland avec les bicouches céramidiques. Ces enzymes sont des hydrolases, elles sont identiques ou très proches de celles des lysosomes. Les principales activités observées correspondent à une (ou des) glycosidase(s), une phospholipase, une sphingomyélinase, une phosphatase, une (ou des) estérase(s), des sulfatases, des protéases. Comme les hydrolases des lysosomes, elles semblent peu spécifiques. Ce pool enzymatique pourrait jouer plusieurs rôles fondamentaux, notamment: 1. La transformation des bicouches gluco-céramidiques en bicouches ceramidiques plus lipophiles; 2. L'élimination de la membrane plasmique, dont certains produits du catabolisme, comme les acides gras et les céramides, peuvent être intégrés aux bicouches céramidiques; 3. La diminution de la cohésion entre les cornéocytes; 4. La protection contre l'intrusion de corps étrangers. Comme tout système enzymatique, les hydrolases extra-cellulaires, sont certainement soumises à des régulations. Plusieurs de ces régulations sont envisagées. A partir de considérations d'enzymologie, le pH de la base du stratum corneum peut être estiméà environ 5. En cosmétologie, les enzymes du stratum corneum sont mises à contribution pour rendre actifs des précurseurs. L'étude des conséquences des modifications de l'activité enzymatique sur l'état de la peau pourrait constituer une future voie de recherche extrêmement prometteuse.     

Non-invasive assessment of stratum corneum structure and function

Methods for the study of the stratum corneum in situ are described and their application to clinical problems and cosmetic science are discussed. In vivo measurements of the thickness of skin and the components of skin can be made using high frequency pulsed ultrasound. Improved resolution of the device should allow measurement of epidermal thickness, and even that of the stratum corneum, which may vary with hydration. The protective ability of the stratum corneum can be estimated in a number of ways, the most familiar technique being transepidermal water loss measurements. Mechanical properties of the stratum corneum have been widely investigated in vivo by numerous researchers. A recent technique devised by us to deform the stratum corneum using an extendable metal frame has been used on patients with ichthyotic disorders. The deformation or compliance of the skin surface has been measured using profilometric methods, and compared to normal individuals. The results appear to be in agreement with the loss of flexural ability experienced by ichthyotic patients. Other mechanical methods more usually involve the measurement of the forces generated by skin while undergoing extension. Three techniques are described which have been used to investigate hydrational change with emollient application. Finally, the phenomenon of desquamation is discussed and methods presented for its evaluation. Passive collection of corneocytes using chambers is useful for estimation of the rates of desquamation, but are laborious and inconvenient. Forced desquamation using the desquamator enables rapid controlled harvesting of corneocytes. In addition to this, the dansyl chloride fluorescence technique has been adopted for estimation of stratum corneum kinetics using visual comparison and fluorimetric techniques. These techniques require further refinement and validation, but the tests available now are nonetheless important in understanding the biology and pharmacology of the stratum corneum.     

Effect of 2-hydroxyacids on guinea-pig footpad stratum corneum: mechanical properties and binding studies

The effect of aqueous solutions of 2-hydroxyacids of chain length C₃ to C₁₀ on the extensibility of undamaged and solvent-damaged guinea-pig footpad stratum corneum has been studied. The increase in extensibility of solvent-damaged corneum, caused by treatment with hydroxyacid, reached a maximum with 2-hydroxycaprylic acid (C₈); on undamaged corneum, 2-hydroxycaprylic acid was the only hydroxyacid studied to give a significant effect. The increase in corneum extensibility produced by 2-hydroxyacids decreases when the pH is raised from 3 to 4. This loss of effect correlates with the ionization of the hydroxyacid (p K ˜ 3.85).
The binding of radiolabelled 2-hydroxycaproic (C₆) and 2-hydroxycaprylic acids to stratum corneum has been studied. 2-Hydroxycaprylic acid binds much more strongly than 2-hydroxycaproic acid, the difference in the binding being consistent with the hydrophobic binding energy of two methylene groups. Raising the pH above 3.5 results in a large decrease in the binding of 2-hydroxycaprylic acid in line with the corresponding reduction in extensibility.
Treatment with 2-hydroxyacids results in a small increase in the water-binding capacity of solvent-damaged stratum corneum, but in a decrease in the water-binding capacity of undamaged stratum corneum. These data are discussed in terms of a possible mechanism for the plasticizing of stratum corneum.
Effet des 2-hydroxyacides sur la couche cornée du coussin des pattes de cobaye     

Assessment of human stratum corneum thickness and its barrier properties by in-vivo confocal Raman spectroscopy

IFSCC Magazine , 12 (2009) (1) 9–15
Measurement of water concentration profiles across living human skin by confocal Raman spectroscopy has developed into a powerful tool for a better understanding of distribution and function of water in the epidermis. From the water profile across the epidermis the border between stratum corneum and stratum granulosum can be estimated. This is due to the steep drop in water concentration from the inner to the outer side of the stratum corneum. Water content drops from approximately 70% at the inner stratum corneum to only 30% at the skin surface. This slope of the curve becomes clearly flatter in the stratum granulosum. A second parameter is usually taken from confocal Raman spectroscopy to define the stratum corneum border. This is the content of natural moisturization factor (NMF), which should be present only in the stratum corneum. Located at the depth at which the NMF content levels off and the slope of the water profile curve changes is the stratum corneum border. The goal of this work was to develop stratum corneum thickness detection into a robust and semi-automated measurement relying only on the water profile. Further, the aim was to base the empirical findings of water distribution in the epidermis on a well established theory, Fick's law of diffusion. A mathematical model was developed to fit the water profile curve for a robust and automated detection of the stratum corneum border. In addition, the new model automatically resulted in an accurately determined slope of the water concentration curve in the stratum corneum. This slope, or more exactly the gradient, is one of two parameters directly related to transepidermal water transport across the stratum corneum.
Keywords:  Barrier function, Fick's law, Raman spectroscopy, stratum corneum, transepidermal water loss     

The use of transepidermal water loss to measure and predict the irritation response to surfactants

The ability of baseline transepidermal water loss (TEWL) measurements to predict the response of healthy subjects to a standard soap chamber irritation insult was evaluated. Chambers containing surfactant solutions were applied to the forearms of normal volunteers for five consecutive days. TEWL was measured using a Servo Med Evaporimeter. TEWL measurements were made prior to the first application of the chambers and following the removal of the second set of chambers (the afternoon of Day 1). The correlation between the TEWL values obtained on Day 1 and the subjective evaluation of irritation resulting from the application of two different toilet soaps was statistically significant. This result indicates that damage to the skin is reflected by an increase in TEWL. Interestingly, there was also a statistically significant correlation between TEWL values obtained on the untreated skin at Time 0 and the five-day average irritation score for these two cleansing products. The results of this study indicate that a high baseline TEWL value may be predictive of a high susceptibility to soap irritation as measured by the chamber test.
Le role des pertes en eau transepidermique pour mesurer et prevoir l'irritation due aux tensio-actifs     

Irritancy ranking of 31 cleansers in the Indian market in a 24-h patch test

Cleansing trends promise freshness, sensory and health benefits but may also be accompanied by an increase in soap-induced skin irritation. The aim of this study was to evaluate the irritant effect of 31 cleansers (28 bar soaps and 3 liquid cleansers) available in the Indian market. Eight percent w/v solutions of the soaps/cleansers were made and 30 microL of each of the solutions were applied to Finn chambers and occluded for 24 h along with distilled water (negative control) and 20% sodium dodecyl sulphate (SDS) as positive control. The sites were graded for erythema and scaling 30 min after removing the patches. The pH of each of the soap solutions was determined. Mean with SD and ANOVA (F-value) was computed separately for each soap/cleanser with respect to the two parameters, erythema and scaling. The total of the means for both the parameters, erythema and scaling was also computed. The cleansers were listed based on this total from the least irritant to the most irritant. The differences between soaps (F-value) was significant for erythema and scaling [erythema = 4.106 (P = 0.000); scaling = 6.006 (P = 0.000)]. Cetaphil cleansing lotion had the lowest erythema score of 0.25. Lowest scaling score of zero was recorded for Cetaphil cleansing lotion and Elovera moisturizing body wash. Aquasoft and Lifebuoy soaps had the highest erythema score of 2.13. Acnex had the highest scaling score of 1.75; Aquasoft, Hamam scrub bath soap and Naturepower sandal soaps were the next with a scaling score of 1.63. Cetaphil cleansing lotion, Aquaderm liquid soap, Dove bar soap and Elovera moisturizing body wash proved to be the least irritant cleansers with a total score of less than 1. The four most irritant soaps/cleansers had an average score of 3.65. The irritant potential of the majority of the cleansers fell between these extremes. The pH of all the soap/cleanser solutions was neutral to alkaline (pH 7-9) except that of Dove bar, Cetaphil cleansing lotion, Aquaderm liquid soap and Elovera moisturizing body wash which tested acidic (pH 5-6). The pH of the positive control--20% SDS, was acidic (pH 6). The difference in the irritancy potential between soaps/cleansers as determined by the 24-h patch test was significant. There were individual variations in the irritant potential of the soaps/cleansers in the volunteers, thus when the patient queries on what soap to use, it may be advisable to test each patient separately and educate him/her regarding the soaps/cleansers less likely to cause irritation. The limitations of the study was that it was single blind and non-randomized as all the 14 soap solutions were applied on 15 volunteers in the first panel and subsequently all the 17 soap solutions were applied on eight volunteers in the second panel. However, we could compare the irritant potential of 31 cleansers. The results of 24-h patch testing of 31 soaps/cleansers in the Indian market in two panels of 14 and 17 soaps/cleansers on 15 and eight volunteers, respectively, are presented.     

The ultraviolet microscopic transmission characteristics of human stratum corneum

This study has evaluated the ultraviolet light transmission characteristics of human stratum corneum at the single cell level using a low light level video microscope to measure the mean percentage transmission of light at different wavelengths and the variation in transmission across the stratum corneum. Stratum corneum was isolated by an enzymic technique and examined on a low light level UV video microscope. Quantitative evaluation of the transmitted monochromatic light for the underside of the layer was measured directly using a Kontron UNIPS image processor or indirectly with a Quantimet 920 image processor after video recording. Transmission distribution histograms were obtained from samples of stratum corneum taken from human breast, scalp, abdomen and leg. Mean transmission values were also derived and compared with diffuse transmission values obtained using the same tissue mounted on an integrating sphere. The UV microscopic transmission characteristics of enzyme separated stratum corneum clearly demonstrated that this structure was not an ideal diffuser. Uniform light intensity on the surface of the stratum corneum led to areas of transmitted intensity in close proximity that differed by factors ofthree to six fold, e.g. between regions of high (>70%) and low (<20%) brightness. However, the average transmission was found to be compatible with published data obtained by diffuse transmission spectrophotometry, taking into account the enhanced transmission arising from stratum corneum immersion in phosphate buffered saline. This was confirmed by the elevated values obtained by diffuse transmission spectrophotometry in this study for samples of stratum corneum prepared for UV microscopy being higher than these found in published data. It is obvious from these findings that viable cells in the epidermis are not exposed to a uniform incident light intensity even when this is true for the surface of skin. Studies of skin response to ultraviolet light at the single cell level must take account of the possibility of preferential exposure of specific sites in any subsequent explanation of cell sensitivity. This is in addition to the already well established cell cycle dependent ultraviolet sensitivity.     

Investigation of interactions between silicones and stratum corneum lipids

Ingredients of topically applied skin care formulations have not only positive effects on the appearance of human skin but can also disturb the Stratum corneum (SC) lipid barrier. In the present study, the influence of silicones (PDMS), as often used cosmetic ingredients, on the microstructure of SC lipids was investigated. For this purpose the interactions of four different PDMS with excised human SC were examined first using differential scanning calorimetry (DSC) and wide angle X-ray diffraction for physical characterization. Because the physical properties of human stratum corneum strongly depend on the lipid composition, showing inter-and intra-individual differences, the interactions with an in vitro model lipid system containing SC fatty acids were also studied, using polarized light microscopy, transmission electron microscopy, small angle X-ray diffraction and DSC. The results revealed that the investigated PDMS do not change either the microstructure of excised human SC or the biphasic lamellar/inverse hexagonal structure of the in vitro model. We concluded that PDMS will not cause any side-effects when topically applied and that our simplified in vitro model could be helpful for estimating interactions between cosmetic ingredients and other topically applied substances and the skin barrier at an early moment of formulation development.     

Influence of cleansing on stratum corneum tryptic enzyme in human skin

Desquamation in human skin is a well-balanced process of de novo production of corneocytes and their shedding from the skin surface. The proteolysis of corneodesmosomes is an important step in the final desquamation process. In the degradation of these adhesion molecules, the stratum corneum tryptic enzyme (SCTE) plays a key role. In initial studies with extracts of porcine epidermis, SCTE was shown to be inactivated by low concentrations of sodium lauryl ether sulphate (SLES). These in vitro findings were supported by in situ results obtained by measuring the release of fluorescent dyes coupled to trypsin-specific substrates incubated on human skin cross-sections. Moreover, in further studies, it could be demonstrated that the SCTE activity in the human horny layer decreases after in vivo application of cleansing products containing SLES. After repeated washing of human volunteers with tap water, a standard market cleansing product (SLES/betaine system) or a new improved cleansing product (SLES/betaine/disodium cocoyl glutamate system), the specific SCTE activity was determined in extracts from the uppermost layers of the stratum corneum. It could be shown that after application of the new formula the remaining SCTE activity was significantly higher than after use of the standard market formula. This ex vivo approach has proven to be very helpful for measuring surfactant effects on human skin enzymes. Using this assay, we developed an improved shower gel formula, which leads to a significantly higher skin enzyme activity after application, compared to a standard market formula.