The skin barrier is fundamental to terrestrial life and its evolution; it upholds homeostasis and protects against the environment. Skin barrier capacity is controlled by lipids that fill the extracellular space of the skin's surface layer--the stratum corneum. Here we report on the determination of the molecular organization of the skin's lipid matrix in situ, in its near-native state, using a methodological approach combining very high magnification cryo-electron microscopy (EM) of vitreous skin section defocus series, molecular modeling, and EM simulation. The lipids are organized in an arrangement not previously described in a biological system-stacked bilayers of fully extended ceramides (CERs) with cholesterol molecules associated with the CER sphingoid moiety. This arrangement rationalizes the skin's low permeability toward water and toward hydrophilic and lipophilic substances, as well as the skin barrier's robustness toward hydration and dehydration, environmental temperature and pressure changes, stretching, compression, bending, and shearing.

Although ultraviolet B (UVB) irradiation induces local immune or systemic immune suppression, depending on the dose, the immune suppression by ultraviolet A (UVA) has not been fully investigated. In this study, we investigated the effect of UVA on the immune response in vitro and in vivo. The effect of UVA on the antigen-presenting function of epidermal cells was measured in terms of antigen-specific T cell proliferation. A murine epidermal cell suspension was exposed to UVA in vitro, pulsed with trinitrobenzenesulfonic acid, and cultured with T cells prepared from syngeneic mice previously sensitized with trinitrochlorobenzene. UVA (5-20 J per cm2) suppressed the antigen-presenting function of epidermal cells in a dose-dependent manner, accompanied with suppression of the expression of costimulatory molecules on Langerhans cells. In order to investigate the effect of an antioxidant on the immune suppression, an epidermal cell suspension was irradiated with UVA in the presence or absence of glutathione. The suppressions of antigen-presenting function and ICAM-1 expression were significantly prevented by glutathione in a dose-dependent manner. Further, the effect of UVA on the immune response at the induction phase of contact hypersensitivity was evaluated in terms of lymph node cell proliferation ex vivo. UVA irradiation suppressed the endogenous proliferation of lymph node cells in trinitrochlorobenzene-painted mice, and this suppression was significantly reversed by the application of glutathione to the skin during irradiation. These results suggest that UVA-induced immune suppression may be mediated by reactive oxygen species, at least in part.

The stratum corneum (SC) is the interface between the body and the environment, and is continuously exposed to oxidative stress that results in carbonyl modification of proteins. We previously developed a simple and non-invasive method to assess the stratum corneum carbonyl protein (SCCP) levels. In this study, we used this method to examine the seasonal changes in the SCCP levels and the relationship between the SCCP level and the physiological condition of the SC. SC was collected from the face of healthy Japanese volunteers by adhesive tape stripping and its carbonyl groups were determined by reaction with fluorescein-5-thiosemicarbazide. The average fluorescence intensity of the SC was calculated as the SCCP level. The SCCP level in the cheek was higher in winter than summer. The SCCP level was negatively correlated with the water content in the SC measured by the conductance and capacitance, and also negatively correlated with the extensibility of the skin measured by a Cutometer, suggesting that the mechanical properties of the skin can be affected by oxidative modification of the SC. These data suggest the involvement of oxidative modification of SC proteins in the generation of rough skin during winter.

Background: Acrolein is a degradation product of lipid peroxide as well as a well-known environmental pollutant. As a hallmark of oxidatively damaged protein, protein carbonyl, including acrolein-protein adduct, has been observed in the skin. However, the influence of protein carbonylation on the stratum corneum (SC) has not yet been clarified. Objectives: We explored the influence of oxidative protein modification, focusing on the major function of the SC, the water-holding capacity, by introducing experimental protein carbonylation. Methods: The level of carbonyls in the SC was evaluated by reaction with a labeled hydrazide. The water-holding capacities of the SC or keratin gels were evaluated by measurement of the surface conductance. The bound water (nonfreezing water) content in the SC was measured by the heat of fusion of frozen SC. Results: Acrolein caused protein carbonylation, decreased the water-holding capacity and the bound water of the porcine SC in vitro. The water-holding capacity of the keratin gels prepared from human SC was also decreased by acrolein in vitro. Water content in the human SC in vivo was decreased by sodium hypochlorite and accompanied by the increase in carbonyls. Conclusion: Exposure of SC to the oxidative environment damages the water-holding capacity of the SC through the modification of protein-water interaction.

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