This article is available online at http://www.jlr.org complex and specifi c architecture of the inter-corneocyte lipid matrix ( 2 ) composed of a mixture of ceramides, cholesterol, and long-chain fatty acids. Ultrastructural studies, using electron microscopy and diffraction techniques, have revealed that these lipids are organized in stacked bilayers that are predominantly parallel to the skin surface ( 3-5 ). Part of the lipids is covalently bound to the corneocytes whereas another part represents the "free lipids" removable by solvent extraction.X-ray diffraction is an excellent technique for studying the spatial organization of the intercellular matrix because the lipid molecules are highly ordered and oriented in a crystalline structure. This technique has provided a number of important results with regard to the SC structure. In particular, small-angle X-ray scattering (SAXS) gives information on the inter-bilayer distance in the multi-lamellar lipid structures, whereas the wide-angle X-ray scattering (WAXS) data contain information about the in-plane crystalline arrangement of lipids (lattice type).Most of the SAXS studies of human SC fi nd refl ections corresponding to ‫ف‬ 4.5 nm and ‫ف‬ 6.5 nm inter-bilayer spacings ( 4, 6, 7 ). Some studies have also detected a refl ection corresponding to a spacing of 11-13.5 nm ( 6, 8 ), referred to as the "long periodicity phase" ( 6 ). These data indicate that the SC lipid matrix has a complex organization into multiple sublayers or even phase-separated domains with different lamellar repeat distances.Similar complexity is found at the molecular level using WAXS, where two sets of peaks are found, corresponding to orthorhombic (peaks at 0.41 and 0.37 nm) and hexagonal (peak at 0.41 nm) packing of the lipid chains. In addition, a fraction of lipids might exist in a fl uid or amorphous state, which would then contribute to the broad band corresponding to an average distance of 0.46 nm. Abstract Lipid and protein components of the stratum corneum (SC) are organized in complex supramolecular arrangements. Exploring spatial relations between various possible substructures is important for understanding the barrier function of this uppermost layer of epidermis. Here, we report the fi rst study where micro-focus X-ray scattering was used for assessing fi ne structural variations of the human skin barrier with micrometer resolution. We found that the scattering profi les were unchanged when scanning in the direction parallel to the SC surface. Furthermore, small-angle scattering profi les did not change as a function of depth in the SC, confi rming that the lipid lamellar spacings remained the same throughout the SC. However, the wide-angle scattering data showed that the orthorhombic phase was more abundant in the middle layers of the SC, whereas the hexagonal phase dominated in the surface layers both at the external and the lowest part of the SC; i.e., the lipids were most tightly packed in the middle region of the SC. Taken together, our results demonstrate that micro...

Glycan spatial distribution and degradation is first observed on the surface of SC in native conditions and at high resolution. The method used can be extended to precisely localize the presence of other macromolecules on the surface of skin or other tissues where the maintenance of its native state is required.

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