Non-invasive methods are very useful for quantifying the atrophogenicity of topical corticosteroids.

The processes of aging and photoaging are associated with an increase in cellular oxidation. This may be in part due to a decline in the levels of the endogenous cellular antioxidant coenzyme Q10 (ubiquinone, CoQ10). Therefore, we have investigated whether topical application of CoQ10 has the beneficial effect of preventing photoaging. We were able to demonstrate that CoQ10 penetrated into the viable layers of the epidermis and reduce the level of oxidation measured by weak photon emission. Furthermore, a reduction in wrinkle depth following CoQ10 application was also shown. CoQ10 was determined to be effective against UVA mediated oxidative stress in human keratinocytes in terms of thiol depletion, activation of specific phosphotyrosine kinases and prevention of oxidative DNA damage. CoQ10 was also able to significantly suppress the expression of collagenase in human dermal fibroblasts following UVA irradiation. These results indicate that CoQ10 has the efficacy to prevent many of the detrimental effects of photoaging.

The non-involved skin of atopic eczema (NEAE) is characterized by severe dryness and an impaired barrier function of the stratum corneum as indicated by an increased transepidermal water loss. Previous studies have demonstrated that this barrier impairment coincides with marked alterations in the amount and composition of stratum corneum ceramides. The aim of this study was to identify specific alterations in NEAE that may be used in the diagnosis of the atopic eczema. Using a classical procedure for high performance thin layer chromatography we could confirm earlier results: apart from Cer(EOH), which contains omega-hydroxy fatty acid (O) ester-linked to linoleic acid (E) and amide-linked to 6-hydroxy-4-sphingenine (H), the quantities of all ceramide fractions were significantly decreased. Furthermore, Cer(EOH)/Certotal was significantly increased, whereas the percentage of Cer(EOS), which contains sphingosine (S), and Cer(NP), which contains non-hydroxy fatty acid (N) amide-linked to phytosphingosine (P), were significantly decreased. Using a modified procedure for high performance thin layer chromatography we could demonstrate the formation of a double peak in the position of Cer(AS), which contains alpha-hydroxy fatty acid (A), in lipids of NEAE. The subfractions of the double peak comprised 15% and 12% of Certotal. MALDITOF mass spectrometry suggested that the double peak was formed by a homologous series of mono-hydroxylated and mono-unsaturated ceramides of different chain length, e.g., Cer(AS) subfractions containing either (C16,18) or (C22,24,26) alpha-hydroxy fatty acids. In contrast, in normal skin a single peak in Cer(AS) position, which comprised 22% of Certotal, was mainly formed by the long chain subfraction. In some cases this single peak displayed a small shoulder at its right flank, but never showed a clear peak separation when developed with NEAE samples. Furthermore, even in senile xerosis, or in either non-involved skin of psoriasis or seborrhoic eczema, only a single peak occurred in Cer(AS) position. Accordingly, the double peak might be specific for NEAE and turn out to be a marker for atopic eczema.

The stratum corneum requires ceramides, cholesterol, and fatty acids to provide the cutaneous permeability barrier. The lipids are organized in intercellular membranes exhibiting short- and long-periodicity lamellar phases. In recent years, the phase behavior of barrier lipid mixtures has been studied in vitro. The relationship of human stratum corneum lipid composition to membrane organization in vivo, however, has not been clearly established. Furthermore, the special function of the different ceramide species in the stratum corneum is largely unknown. We examined lipid organization and composition of stratum corneum sheets from different subtypes of healthy human skin (normal, dry, and aged skin). Lipid organization was investigated using X-ray diffraction and demonstrated that the 4.4 nm peak attributed to the long periodicity phase was frequently missing for skin with a low Cer(EOS)/Cer(total) ratio, indicating an important part for Cer(EOS), which contains omega-hydroxy fatty acid (O) ester-linked to linoleic acid (E) and amide-linked to sphingosine (S). A deficiency in the 4. 4 nm peak was predominantly observed in young dry skin. In one case of aged skin, however, and less often in young normal skin this peak was also missing. Furthermore, the ceramide composition of samples without the 4.4 nm peak showed a deficiency of Cer(EOH), which contains 6-hydroxy-4-sphingenine (H), and an increase in Cer(NS) and Cer(AS), which contain nonhydroxy (N) or alpha-hydroxy fatty acids (A). In addition, a 3.4 nm peak attributed to crystalline cholesterol occurred in most cases of aged and dry skin, but was not observed in young normal skin. Our results do not indicate a definite pattern of correlation between lipid organization and types of human skin. They demonstrate, however, that Cer(EOS) and Cer(EOH) are key elements for the molecular organization of the long periodicity lamellar phase in the human stratum corneum.

The composition pattern of ceramides mirrors that of mid-gestational fetal epidermis. Vernix thus represents a 'homologous' substitute for the immature epidermal barrier in fetal skin. The differential role of individual ceramides in this process remains to be established.

The acidic pH of the horny layer, measurable on the skin surface, has long been regarded as a result of exocrine secretion of the skin glands. The 'acid mantle' was thought to regulate the bacterial skin flora and to be sensitive primarily to skin cleansing procedures. In recent years, an increasing number of investigations have been published on the changes in, and constituents and functions of, the pH of the deeper layers of the stratum corneum, as well as on the influence of physiological and pathological factors. A central role for the acidic milieu as a regulating factor in stratum corneum homeostasis is now emerging. This has relevance to the integrity of the barrier function, from normal maturation of the stratum corneum lipids through to desquamation. Changes in the pH and the organic factors influencing it appear to play a role, not only in the pathogenesis, prevention and treatment of irritant contact dermatitis, but also of atopic dermatitis and ichthyosis and in wound healing. On the basis of these findings, a broader concept, exceeding the superficial 'acid mantle' theory, has been formulated.

The stratum corneum (SC) requires ceramides, cholesterol, and fatty acids to provide the cutaneous permeability barrier. SC lipids can be analyzed by normal-phase high-performance thin-layer chromatography (HPTLC). However, without further analysis, some uncertainty remains about the molecular composition of lipids represented by every TLC band of an unknown sample. We therefore analyzed each ceramide band further by subjecting the isolated lipids to a direct coupling of reversed-phase high-performance liquid chromatography and electrospray ionization-mass spectrometry (HPLC/ESI-MS, or LC/MS). LC/MS analysis and ESI-MS/MS negative ion and collision-induced dissociation experiments revealed that ceramide band 4 contained not only N-(omega-OH-acyl)acyl-6-OH-sphingosine, Cer(EOH), but also N-(alpha-OH-acyl)-sphingosine. Band 5 exclusively contained N-acyl-6-OH-sphingosine. Our results demonstrate the benefit of LC/MS analysis for selective identification of human SC ceramides. Moreover, the combination of HPTLC for pre-separation and LC/MS for identification of lipids is an even more powerful tool for detailed ceramide analysis.

Recent studies have provided new insights into the occurrence, causes, and pathogenetic consequences of changes in the skin pH in atopic dermatitis, particularly with respect to skin barrier function and colonization with Staphylococcus aureus. Growing evidence suggests an impaired release of proton donors, such as amino acids, urocanic acid, and lactic acid, to the stratum corneum in atopic dermatitis, as a result of reductions in filaggrin proteolysis and sweat secretion. In addition, an impaired formation of free fatty acids from sebaceous lipids and epidermal phospholipids seems to be involved. Because both lipid organization and lipid metabolism in the stratum corneum requires an acidic pH, these alterations might contribute to the disturbance of skin barrier function observed in atopic dermatitis. Furthermore, bacterial growth and virulence of S. aureus, as well as defensive host mechanisms, have increasingly been delineated as pH dependent, giving rise to a new understanding of the pathophysiology underlying increased skin colonization seen in atopic dermatitis.