In vivo stratum corneum over-hydration and water diffusion coefficient measurements using opto-thermal radiometry and TEWL Instruments

Skin over-hydration is a common problem that affects many people who wear incontinence pads or diapers. The aim of this study is to develop a new method for stratum corneum (SC) over-hydration and SC water diffusion coefficient measurements using opto-thermal transient emission radiometry (OTTER) and evaporimetry. With OTTER, we can measure the SC surface hydration and hydration gradient. With evaporimetry, we can measure the time-dependent evaporative drying curves of water vapour flux density (WVFD). The combination of hydration results and WVFD results can yield information on the SC water diffusion coefficient and how it depends on the SC surface hydration level. The results show that SC water diffusion coefficient is non-linearly proportional to the SC surface hydration level. The results also show strong correlations between evaporative drying flux measured using the Evaporimeter and surface hydration estimated from OTTER measurements.     

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.     

Studies on isolated corneocytes

The function of the stratum corneum is dependent on the functions of its constituent parts. This paper focusses on corneocyte dimensions and the implications of change in dimension for function. A technique in which it is possible to obtain a value for corneocyte thickness is described. The changes in corneocyte dimensions with age and site and after topical applications are documented.
Techniques which measure the rate of desquamation are also described. The rate of passive corneocyte loss has been estimated by counting the numbers of corneocytes shed into chambers fixed to the skin surface. In addition the corneocyte loss after a standardized rotational stimulus to the skin surface has been measured. Scanning and transmission electron microscope examination of corneocytes obtained by a scrub procedure is also described. A preliminary report on an immunological approach to the study of the biochemistry of corneocytes and stratum corneum is also included.
Etude de cornéocytes isolés     

Stratum corneum barrier integrity controls skin homeostasis

The stratum corneum water barrier controls structural and functional properties of both the epidermis and the dermis. Treatments which chronically disrupt the stratum corneum water barrier can induce changes similar to those seen with 'anti-aging' treatments such as (-Hydroxy acids (AHAs) and Retin Atrade mark. Barrier disruption via daily tape stripping increases epidermal and dermal thickness, superficial and integral skin firmness, and improves skin surface texture. Modest or transitory disruption did not produce such effects. Similar results were observed with topical application of AHAs, retinoids or mild irritants after about 4-6 weeks provided such treatments resulted in prolonged elevation in TEWL (trans-epidermal water loss). Treatments that did not chronically elevate TEWL could also produce positive cosmetic effects, but such effects were in general restricted to the skin surface or epidermis. Irritation, which was observed with some treatments, was not solely responsible for the positive effects observed.     

Seasonal influences on stratum corneum ceramide 1 fatty acids and the influence of topical essential fatty acids

Ceramide 1 is the main repository of stratum corneum linoleic acid, and changes in the levels of ceramide 1 linoleate are associated with cutaneous abnormalities: essential fatty acid deficiency, atopic dermatitis and acne. We have previously reported seasonal variation in stratum corneum lipids where lipids, particularly ceramides, are very much reduced in winter which probably influences the appearance of skin xerosis in this season. However, investigations into the seasonal variation in ceramide 1 fatty acid levels and the effects of topical essential fatty acids on the composition of these lipid species have not been conducted in healthy subjects. We determined the composition of stratum corneum ceramide 1 esterified fatty acids in leg skin from Caucasian women in winter and summer and also investigated the effects of topical triglycerides rich in linoleic acid on stratum corneum ceramide 1 fatty acid levels in winter. A dramatic seasonal variation was observed. Whereas decreased levels of linoleic acid and some saturated fatty acids (C-15 and C-24) were observed in the winter months, increased levels of oleic acid were found. Nevertheless, it was possible to normalize the levels of stratum corneum ceramide 1 linoleate following topical application of formulations containing linoleic acid-rich triglycerides for 4 weeks. The reduction in ceramide 1 linoleate may lead to impaired stratum corneum function and increased susceptibility to skin xerosis in winter. However, by using formulations containing linoleic acid esters it is possible to correct these changes and possibly improve barrier resilience.     

Skin efficacy of liposomes composed of internal wool lipids rich in ceramides

Ceramides from intercellular lipids of skin stratum corneum are known to play an essential role in maintaining and structuring the lipid barrier of the skin. Internal wool lipids (IWL), which are also rich in ceramides, have a composition similar to that of the stratum corneum lipids. IWL extracted with chloroform/methanol azeotrope at the laboratory scale have been shown to be capable of forming liposomes with a stable bilayer structure. Furthermore, topical application of these IWL liposomes on intact and compromised skin has been demonstrated to improve barrier skin properties. In this study we evaluated the effect on human skin repair of different IWL extract compositions obtained by two extraction methodologies. The formation and characteristics of the liposomes prepared were greatly influenced by the IWL composition, primarily the sterol sulfate content. The IWL liposomes improved skin barrier integrity and increased skin hydration when applied onto intact skin. These improvements were slightly enhanced in the case of IWL liposomes that were richer in polar lipids.     

In vivo transepidermal water loss and skin surface hydration in assessment of moisturization and soap effects

Transepidermal water loss (TEWL), skin surface water loss (SSWL) and water content of the stratum corneum are utilized to assess the hydration effects of moisturizers and soaps. The relationship among these parameters may help differentiating hydration obtained via occlusion or by water-holding in the stratum corneum. Furthermore, skin function (hydration, dehydration, barrier damage) can be studied comparing the data obtained with these techniques. In this study, the effects of glycerol, petrolatum, soaps and commercial moisturizers on the skin are investigated and discussed.     

In vivo confocal fluorescence imaging of skin surface cellular morphology: a pilot study of its potential as a clinical tool in skin research

The cellular morphology of the stratum corneum was studied in vivo using a novel imaging technique that uses confocal fluorescence microscopy in combination with topical application of a fluorescent contrast agent. Images obtained with this method show a strong variation in skin surface cellular morphology among healthy subjects. The results of several clinical studies suggest that cellular morphology is affected by the efficiency of the process of desquamation. As such, cellular morphology shows strong potential to serve as an indicator of skin health that yields mechanistic insight into the origins of skin ailments, such as xerosis, and the effectiveness of their treatments.     

Age‐related changes in skin barrier function – Quantitative evaluation of 150 female subjects

The protection against water loss and the prevention of substances and bacteria penetrating into the body rank as the most important functions of the skin. This so‐called ‘skin barrier function’ is the natural frontier between the inner organism and the environment, and is primarily formed by the epidermis. An impairment of the skin barrier function is often found in diseased and damaged skin. An influence of ageing on skin barrier function is widely accepted, but has not been conclusively evaluated yet. Therefore, the aim of this clinical study was to assess the potential influence of ageing on skin barrier function, including transepidermal water loss (TEWL), stratum corneum hydration, sebum content and pH value. One hundred and fifty healthy women aged 18–80, divided into five age groups with 30 subjects each, were evaluated in this study. TEWL, hydration level, sebum secretion and pH value of hydro‐lipid acid film were measured with worldwide acknowledged biophysical measuring methods at cheek, neck, décolleté, volar forearm and dorsum of hand. Whereas TEWL and stratum corneum hydration showed only very low correlation with subject's age, the sebum production decreased significantly with age, resulting in the lowest skin surface lipids levels measured in subjects older than 70 years. The highest skin surface pH was measured in subjects between 50 and 60 years, whereas the eldest age group had the lowest mean pH. The dorsum of the hand was the location with the highest TEWL and lowest stratum corneum hydration in all age groups. The results show that only some parameters related to skin barrier function are influenced by ageing. Whereas sebum production decreases significantly over lifetime and skin surface pH is significantly increased in menopausal woman, TEWL and stratum corneum hydration show only minor variations with ageing.     

A new technique for the evaluation of cosmetics effect on mechanical properties of stratum corneum and epidermis in vitro

The mechanical properties of stratum corneum and epidermis have been measured in vitro and a dispersity of approximately 30% to 50% between samples from the same donor has been found. To overcome such a limitation, a technique was developed in order to compare the mechanical properties of the same skin sample before and after treatment. The chosen parameter (initial slope of the stress-strain curve) appeared to be influenced by the topical application of products. A different time response was found with stratum corneum and epidermis reflecting their structural differences.
Une nouvelle technique d'evaluation de l'effet des produits cosmétique sur les propriétés mécaniques du stratum corneum et de l'épiderme humains (etude in vitro )     

Original semiologic standardized evaluation of stratum corneum hydration by Diagnoskin stripping sample

In a normal and healthy skin, the regular elimination of the superficial corneocytes, called desquamation, is a fundamental physiologic process intended to protect the barrier function of the skin. This invisible loss of corneocytes, individually or in small groups, is incessantly compensated by the divisions of the proliferative layer and the upward cellular maturation in order to maintain the harmonious renewal of the epidermis and the integrity of the stratum corneum. The harmony of this desquamation process is intimately conditioned by a sufficient hydration of the stratum corneum: (i) an abnormal desquamation leads to a disruption of the water barrier function and consequently to a dehydration tendency of the stratum corneum, and (ii) a cutaneous dryness (whatever the cause) is able to disturb the desquamation process. Protecting the water content of the stratum corneum has always been a major preoccupation of the cosmetic industry scientists. Consequently, the moisturizing properties of a cosmetic product are objectively measured by various explorations directly targeted on the hydration (corneometry) and on the level of the water barrier function (transepidermal water loss (TEWL) measurements), which depends directly on the skin hydration state. This intimate linkage of the desquamation process and the water content of the stratum corneum enable us to suggest an indirect assessment of the hydration from a direct study of the desquamation by examining a skin-stripping sample (D-Squames) by an optical microscope (linked to a computer). We will describe this already known technique and mainly its new and unpublished semiologic exploitation, named Diagnoskin, whose advantages are its simplicity and its reproducibility particularly interesting in the case of sequential appraisal of dermatologic or cosmetic treatments.     

J. Cosmet. Sci., 59, 217–224 (May/June 2008) In vivo spectrophotometric evaluation of skin barrier recovery after topical application of soybean phytosterols

The skin's uppermost thin layer, stratum corneum, plays a crucial role in protecting the body against unwanted influences from the environment. Disruption of the stratum corneum, by tape stripping or chemical injury, results in epidermal recovery of the skin barrier. Soy phytosterols are widely used in the cosmetic field as active ingredients in creams and lipsticks. Furthermore, they deserve an important place among nutracosmeceuticals; in fact, after their absorption from the diet they are transferred from the plasma to the skin, playing an important role in the constitution of skin surface lipids. The aim of the present work was to study the effect of the topical application of soybean phytosterols on skin barrier recovery in human volunteers using the extent of methyl nicotinate (MN)-induced erythema in damaged skin as a parameter to evaluate the rate of stratum corneum recovery. MN was chosen as an erythematogenous substance for its capability to cause an erythema whose intensity and duration are proportional to the quantity of the substance that has entered the living epidermis over time. MN-induced erythema was monitored using reflectance spectrophotometry as a non-invasive instrumental technique. The results show clearly that soy phytosterols exert positive results on skin repair; in fact, 3 days after tape stripping, the sites treated with a formulation containing phytosterols showed an appreciable recovery of barrier function compared to those treated with a vehicle control without soy phytosterols.     

Clinical and instrumental evaluation of a food supplement in improving skin hydration

Topically applied cosmetic products can be helpful in improving skin hydration. The study shows how oral supplementation could be helpful in improving and preventing the skin dehydration. A total of 32 healthy female volunteers entered the study. Of which, 16 were treated with a food supplement based on vegetable ceramides, amino acids, fish cartilage, antioxidants and essential fatty acids for 40 days and 16 with placebo. The results of the clinical and instrumental evaluations carried out in this study, have highlighted how the active treatment is effective in improving skin hydration and in reducing the cutaneous smoothness and roughness and the depth of furrows, in comparison to the placebo. In fact, concerning several important parameters, as stratum corneum hydration and skin roughness, the improvement measured exceeded 25%. We therefore suggest that a combination of treatments (oral and topical) can be more effective in improving skin hydration.     

Formulating for efficacy

Lip chapping is a serious cosmetics problem, though remedies other than moisturization have not been proposed. We investigated changes in the surface configurations of lip corneocytes and activities of desquamation-regulating proteinases associated with lip chapping. Using scanning electron microscopy, villus-like projections were observed on the inner surfaces of most corneocytes from normal lips, whereas those with flatter surface were predominantly found in chapped lips. Furthermore, cell surface area increased with the severity of lip chapping. Cathepsin D-like and chymo-trypsin-like proteinase, which are also present in skin as desquamation-regulating proteinases, were detected in lip corneocytes, though only cathepsin D activity was found to decrease in severely chapped lips. Hydration was also lower in areas of lip chapping. Sequential topical application of apricot extract essence increased cathepsin D activity and improved chapping severity. Our results suggest that lip chapping can be characterized as similar to senile xerosis rather than dry skin such as winter xerosis, as it shows a delayed transition of corneocytes through the stratum corneum, and the reduced cathepsin D activity may be one of the mechanisms that is further decreased by low hydration. We propose that an enhancement of both cathepsin D activity and lip moisture may be effective to improve lip chapping.     

Application of substrate curvature method to differentiate drying stresses in topical coatings and human stratum corneum

Despite the extensive use of topical coatings in cosmetics, their effect on the mechanical properties of human skin and the perception of skin tightness in the form of drying stresses is not well understood. We describe the application of a recently developed substrate curvature technique to characterize stresses in drying and non‐drying occlusive topical coatings. We then extend the technique to measure the combined effects of the coating applied to human stratum corneum (SC) where the overall drying stresses may have contributions from the coating, the SC and the interaction of the coating with the SC. We show how these separate contributions in the coating and SC layers can be differentiated.     

In vivo measuring of skin softness using the Gas Bearing Electrodynamometer

The bio-mechanical properties of the stratum corneum are of great importance for the subjective perception of skin conditions (dry vs. normal), as well as for its multiple functions. The recently developed Gas Bearing Electrodynamometer allows us to measure objectively the viscoelastic properties of the stratum corneum in vivo and to evaluate, in terms of skin softness, the changes of this parameter induced by the application of emollients.
The principle of the technique, as designed by Christensen et al. , is briefly described in context with our results.
During the probe movement, there was a modification of the skin surface relief: compression of the skin furrows ahead of the moving probe, and extension-flattening of the relief behind the probe.
The variations of the skin softness were studied in context with the reproducibility of the measurements: the coefficient of variation was found to be about 3%. Considerably higher variations were found as related to: the anisotropy of the skin surface relief, various areas of the body, and individual differences.
The effects of emollients and water on the skin softness were studied. Water had a clear effect but of very short duration (around 10 minutes). Water-containing emollients of the type o/w emulsions displayed a distinct long-lasting effect (followed up to 6 hours). Its magnitude was generally higher than that obtained with w/o emulsions. It was found that the emollient's non-volatile components (oil phase) play an important role in the product's softening effect.
In conclusion, results to date have established this technique as a promising tool for studying skin softness in general, and effects of topical applications, new formulae and specific ingredients in particular.
Mesure de la souplesse de la peau avec un Electrodynamomètre     

Skin surface profile technique and its applications

In this study the 'skin surface profile'(SSP) technique and its applications are described. With this technique the SSP is replicated by means of silicone impression materials. The SSP preparations were studied by a stereomicroscope or microscopic projector. Some applications of SSP technique are as follows: a. measurement of the stratum corneum hydration, b. study of the profile of primary crests and lines, c. measurement of the primary crests reservoir available for extension, and d. estimation of the true area of the skin surface in a body region.     

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.     

Moisturization processes in living human skin studied by magnetic resonance imaging microscopy

Magnetic resonance imaging is a non-invasive, non-destructive and chemically specific imaging method widely used in medicine to reveal information about both tissue structure and function. It can measure water in tissue, but it has been difficult to achieve the necessary sensitivity and resolution when applying it to studies of the dry, thin stratum corneum. In this paper the use of magnetic resonance imaging to image the outer layers of the skin with a resolution of 0.06 mm is reported. Configuring the magnetic resonance imaging method in this way has made it possible for the first time to actually 'see'directly the moisturization in the stratum corneum. It is no longer necessary to rely upon methods which can only show side-effects of moisturization, such as changes in the appearance of the skin cells. As magnetic resonance imaging is harmless, it can be used repeatedly on the same skin and so produce a series of stills, or a time-lapse video, clearly showing the actual process of moisturization and related phenomena. The behaviour of skin has been observed during both hydration and dehydration; the two processes follow different time courses. Two layers have been observed in the stratum corneum, which appear to be different when the skin is hydrated.
For the first time the actual surface of normal skin has been revealed on magnetic resonance images.     

A liquid crystal application in skin care cosmetics

Intercellular lipids of the stratum corneum contribute threefold to the maintenance of a healthy skin: by hydration, cell adhesion, and reduction of transepidermal water loss. All of these functions can be attributed to the self-organizing property of the amphiphilic molecules of the stratum corneum lipids. A new type of skin care product called Lamellar Gel was developed, which contains a (synthesized) pseudo-ceramide. Its structure is similar to that of ceramide found among the stratum corneum lipids, which allows it to control intramolecular interactions. Compared to regular emulsions the Lamellar Gel demonstrated better skin care characteristics regarding permeability, skin hydration, and skin occlusion. This was attributed to the fact that it formed the same self-organizing structure as natural stratum corneum lipids, hence showing a high affinity to the skin. A high moisturizing effect was observed as the Lamellar Gel combines the benefits of both O/W and W/O emulsions: it provides the same initial hydration as an O/W emulsion and at the same time the same occlusivity as a W/O emulsion. Transepidermal water loss increases under dry environmental conditions. This especially affects the skin around the eyes, where the skin is very thin, and wrinkles are very easily formed. Treatment with the Lamellar Gel recovered these wrinkles promptly and hydrated the stratum corneum for a long time.