The stratum corneum (SC), the top layer of skin, dictates the rate of both water loss through the skin and absorption of exogenous molecules into the body. The crystalline organization of the lipids in the SC is believed to be a key feature associated with the very limited permeability of the skin. In this work, we characterized the organization of SC lipid models that include, as in native SC, cholesterol, a series of FFAs (saturated with C16-C24 chains), as well as a ceramide bearing an oleate chain-linked to a very long saturated acyl chain [-melissoyl-oleoyloxy hexacosanoyl-D--sphingosine (Cer EOS)]. The latter is reported to be essential for the native SC lipid organization. Our H-NMR, infrared, and Raman spectroscopy data reveal that Cer EOS leads to the formation of highly disordered liquid domains in a solid/crystalline matrix. The lipid organization imposes steric constraint on Cer EOS oleate chains in such a way that these hydrocarbon nanodroplets remain in the liquid state down to -30°C. These findings modify the structural description of the SC substantially and propose a novel role of Cer EOS, as this lipid is a strong modulator of SC solid/liquid balance.

The skin, the largest organ of the human body, forms a flexible interface between our internal and external environment that protects our organism from exogenous compounds as well as excessive water loss. The stratum corneum (SC), the outermost layer of mammal epidermis, is mainly responsible for the skin impermeability. The SC is formed by corneocytes embedded in a lipid matrix, which is mostly constituted of ceramides (Cer), free fatty acids (FFA), and cholesterol (Chol), organized in two coexisting crystalline lamellar phases. This arrangement of lipids is crucial to skin barrier function. The aim of this paper is to determine the impact of FFA chain length on the phase behavior of SC model lipid membranes using solid-state deuterium NMR and IR spectroscopy. We studied ternary mixtures of N-lignoceroyl-d-erythro-sphingosine (Cer24), cholesterol, and palmitic (FFA16) or lignoceric (FFA24) acid in an equimolar ratio. This proportion replicates the lipid composition found in the SC lipid matrix. Our studies revealed that the phase behavior of Cer24/FFA/Chol ternary mixtures is strongly affected by the length of the FFA. We found the formation of phase-separated crystalline lipid domains when using palmitic acid whereas the use of lignoceric acid results in a more homogeneous mixture. In addition, it was observed that mixtures with lignoceric acid form a gel phase, a very unusual feature for SC model mixtures.

Little is known about the interaction of very long-chain saturated fatty acids (VLCFAs) with biological membranes. However, this could play an important role on interleaflet interactions and signal transduction mechanisms in cells. The aim of this work is to determine how VLCFA structurally adapts in fluid phospholipid bilayers, since both species must exhibit a significant hydrophobic mismatch. The membrane organization has been described by means of (2)H NMR and molecular dynamics simulations. Our results show that the protonation state affects the position and order of free fatty acids (FFAs) in phospholipid membranes. It was shown that the protonated FFA-C24 carboxyl group is located slightly under the POPC head group and therefore its acyl chain can interact with the lipids of the opposite leaflet. This interdigitation of the end of the acyl chain causes a second plateau observed in SC-D profiles, a very unusual feature in lipid systems.

No abstract

Stratum corneum (SC), the outermost layer of the epidermis, is the primary barrier to percutaneous absorption. The diffusion of substances through the skin occurs through the SC lipid fraction, which is essentially constituted of an equimolar mixture of ceramides, free fatty acids, and cholesterol. The lipid constituents of SC are mainly forming continuous multilamellar membranes in the solid/crystalline state. However, recent findings suggest the presence of a highly disordered (liquid) phase formed by the unsaturated C18 chain of ceramide EOS, surrounded by a highly ordered lipid environment. The aim of the present work was to study the lipid spatial distribution of model SC membranes composed of ceramide EOS, ceramide NS, a mixture of free fatty acids, and cholesterol, using Raman microspectroscopy and AFM-IR spectroscopy techniques. The enhanced spatial resolution at the tens of nanometers scale of the AFM-IR technique revealed that the lipid matrix is overall homogeneous, with the presence of small, slightly enriched, and depleted regions in a lipid component. No liquid domains of ceramide EOS were observed at this scale, a result that is consistent with the model proposing that the oleate nanodrops are concentrated in the central layer of the three-layer organization of the SC membranes forming the long periodicity phase. In addition, both Raman microspectroscopy and AFM-IR techniques confirmed the fluid nature of the unsaturated chain of ceramide EOS while the rest of the lipid matrix was found highly ordered.