Ceramides are key components of the skin's permeability barrier. In atopic dermatitis, pathological hydrolysis of ceramide precursors - glucosylceramides and sphingomyelin - into lysosphingolipids, specifically glucosylsphingosine (GS) and sphingosine-phosphorylcholine (SPC), and free fatty acids (FFAs) has been proposed to contribute to impaired skin barrier function. This study investigated whether replacing ceramides with lysosphingolipids and FFAs in skin lipid barrier models would exacerbate barrier dysfunction. When applied topically to human stratum corneum sheets, SPC and GS increased water loss, decreased electrical impedance, and slightly disordered lipid chains. In lipid models containing isolated human stratum corneum ceramides, reducing ceramides by ≥ 30% significantly increased permeability to four markers, likely due to loss of long-periodicity phase (LPP) lamellae and phase separation within the lipid matrix, as revealed by X-ray diffraction and infrared spectroscopy. However, when the missing ceramides were replaced by lysosphingolipids and FFAs, no further increase in permeability was observed. Conversely, these molecules partially mitigated the negative effects of ceramide deficiency, particularly with 5%-10% SPC, which reduced permeability even compared to control with "healthy" lipid composition. These findings suggest that while ceramide deficiency is a key factor in skin barrier dysfunction, the presence of lysosphingolipids and FFAs does not aggravate lipid structural or functional damage, but may provide partial compensation, raising further questions about the behavior of lyso(sphingo)lipids in rigid multilamellar lipid environments, such as the stratum corneum, that warrant further investigation.

• Keywords
• ceramide, fatty acid, glucosylsphingosine, lipid model, lysolipid, permeability, skin barrier, sphingosine-phosphorylcholine,
• MeSH
• Ceramides * metabolism   deficiency   MeSH
• Phosphorylcholine analogs & derivatives   MeSH
• Skin * metabolism   drug effects   MeSH
• Fatty Acids, Nonesterified metabolism   MeSH
• Humans MeSH
• Lysophospholipids * metabolism   MeSH
• Permeability MeSH
• Sphingolipids * metabolism   MeSH
• Sphingosine analogs & derivatives   metabolism   pharmacology   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Ceramides * MeSH
• Phosphorylcholine MeSH
• Fatty Acids, Nonesterified MeSH
• Lysophospholipids * MeSH
• Sphingolipids * MeSH
• Sphingosine MeSH
• sphingosine phosphorylcholine MeSH Browser

Ceramides belong to sphingolipids, an important group of cellular and extracellular lipids. Their physiological functions range from cell signaling to participation in the formation of barriers against water evaporation. In the skin, they are essential for the permeability barrier, together with free fatty acids and cholesterol. We examined the periodical structure and permeability of lipid films composed of ceramides (Cer; namely, N-lignoceroyl 6-hydroxysphingosine, CerNH24, and N-lignoceroyl sphingosine, CerNS24), lignoceric acid (LIG; 24:0), and cholesterol (Chol). X-ray diffraction experiments showed that the CerNH24-based samples form either a short lamellar phase (SLP, d ∼ 5.4 nm) or a medium lamellar phase (MLP, d = 10.63-10.78 nm) depending on the annealing conditions. The proposed molecular arrangement of the MLP based on extended Cer molecules also agreed with the relative neutron scattering length density profiles obtained from the neutron diffraction data. The presence of MLP increased the lipid film permeability to the lipophilic model permeant (indomethacin) relative to the CerNS24-based control samples and the samples that had the same lipid composition but formed an SLP. Thus, the arrangement of lipids in various nanostructures is responsive to external conditions during sample preparation. This polymorphic behavior directly affects the barrier properties, which could also be (patho)physiologically relevant.

• Publication type
• Journal Article MeSH

Epidermal omega-O-acylceramides (ω-O-acylCers) are essential components of a competent skin barrier. These unusual sphingolipids with ultralong N-acyl chains contain linoleic acid esterified to the terminal hydroxyl of the N-acyl, the formation of which requires the transacylase activity of patatin-like phospholipase domain containing 1 (PNPLA1). In ichthyosis with dysfunctional PNPLA1, ω-O-acylCer levels are significantly decreased, and ω-hydroxylated Cers (ω-OHCers) accumulate. Here, we explore the role of the linoleate moiety in ω-O-acylCers in the assembly of the skin lipid barrier. Ultrastructural studies of skin samples from neonatal Pnpla1+/+ and Pnpla1-/- mice showed that the linoleate moiety in ω-O-acylCers is essential for lamellar pairing in lamellar bodies, as well as for stratum corneum lipid assembly into the long periodicity lamellar phase. To further study the molecular details of ω-O-acylCer deficiency on skin barrier lipid assembly, we built in vitro lipid models composed of major stratum corneum lipid subclasses containing either ω-O-acylCer (healthy skin model), ω-OHCer (Pnpla1-/- model), or combination of the two. X-ray diffraction, infrared spectroscopy, and permeability studies indicated that ω-OHCers could not substitute for ω-O-acylCers, although in favorable conditions, they form a medium lamellar phase with a 10.8 nm-repeat distance and permeability barrier properties similar to long periodicity lamellar phase. In the absence of ω-O-acylCers, skin lipids were prone to separation into two phases with diminished barrier properties. The models combining ω-OHCers with ω-O-acylCers indicated that accumulation of ω-OHCers does not prevent ω-O-acylCer-driven lamellar stacking. These data suggest that ω-O-acylCer supplementation may be a viable therapeutic option in patients with PNPLA1 deficiency.

• Keywords
• PNPLA1 deficiency, Skin, acylceramides, barrier function, ceramides, linoleic acid, lipids, model membranes, sphingolipids, stratum corneum,
• MeSH
• Acyltransferases MeSH
• Ceramides * chemistry   MeSH
• Epidermis MeSH
• Ichthyosis MeSH
• Skin * MeSH
• Linoleic Acid MeSH
• Lipase MeSH
• Mice MeSH
• Animals MeSH
• Check Tag
• Mice MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Acyltransferases MeSH
• Ceramides * MeSH
• Linoleic Acid MeSH
• Lipase MeSH
• PNPLA1 protein, mouse MeSH Browser

Ceramides (Cer) are essential components of the skin permeability barrier. To probe the role of Cer polar head groups involved in the interfacial hydrogen bonding, the N-lignoceroyl sphingosine polar head was modified by removing the hydroxyls in C-1 (1-deoxy-Cer) or C-3 positions (3-deoxy-Cer) and by N-methylation of amide group (N-Me-Cer). Multilamellar skin lipid models were prepared as equimolar mixtures of Cer, lignoceric acid and cholesterol, with 5 wt% cholesteryl sulfate. In the 1-deoxy-Cer-based models, the lipid species were separated into highly ordered domains (as found by X-ray diffraction and infrared spectroscopy) resulting in similar water loss but 4-5-fold higher permeability to model substances compared to control with natural Cer. In contrast, 3-deoxy-Cer did not change lipid chain order but promoted the formation of a well-organized structure with a 10.8 nm repeat period. Yet both lipid models comprising deoxy-Cer had similar permeabilities to all markers. N-Methylation of Cer decreased lipid chain order, led to phase separation, and improved cholesterol miscibility in the lipid membranes, resulting in 3-fold increased water loss and 10-fold increased permeability to model compounds compared to control. Thus, the C-1 and C-3 hydroxyls and amide group, which are common to all Cer subclasses, considerably affect lipid miscibility and chain order, formation of periodical nanostructures, and permeability of the skin barrier lipid models.

• MeSH
• Cell Membrane metabolism   MeSH
• Ceramides chemistry   metabolism   MeSH
• Skin metabolism   MeSH
• Membranes, Artificial * MeSH
• Permeability MeSH
• Water metabolism   MeSH
• Phase Transition MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Ceramides MeSH
• Membranes, Artificial * MeSH
• Water MeSH

Ceramides (Cers) with ultralong (∼32-carbon) chains and ω-esterified linoleic acid, composing a subclass called omega-O-acylceramides (acylCers), are indispensable components of the skin barrier. Normal barriers typically contain acylCer concentrations of ∼10 mol%; diminished concentrations, along with altered or missing long periodicity lamellar phase (LPP), and increased permeability accompany an array of skin disorders, including atopic dermatitis, psoriasis, and ichthyoses. We developed model membranes to investigate the effects of the acylCer structure and concentration on skin lipid organization and permeability. The model membrane systems contained six to nine Cer subclasses as well as fatty acids, cholesterol, and cholesterol sulfate; acylCer content-namely, acylCers containing sphingosine (Cer EOS), dihydrosphingosine (Cer EOdS), and phytosphingosine (Cer EOP) ranged from zero to 30 mol%. Systems with normal physiologic concentrations of acylCer mixture mimicked the permeability and nanostructure of human skin lipids (with regard to LPP, chain order, and lateral packing). The models also showed that the sphingoid base in acylCer significantly affects the membrane architecture and permeability and that Cer EOP, notably, is a weaker barrier component than Cer EOS and Cer EOdS. Membranes with diminished or missing acylCers displayed some of the hallmarks of diseased skin lipid barriers (i.e., lack of LPP, less ordered lipids, less orthorhombic chain packing, and increased permeability). These results could inform the rational design of new and improved strategies for the barrier-targeted treatment of skin diseases.

• Keywords
• acylceramide, ceramides, disease models, epidermis, extracellular matrix, membrane nanostructure, membranes/model, permeability, sphingolipids,
• MeSH
• Ceramides analysis   metabolism   MeSH
• Skin Diseases metabolism   MeSH
• Skin chemistry   metabolism   MeSH
• Humans MeSH
• Membrane Lipids chemistry   metabolism   MeSH
• Models, Molecular MeSH
• Molecular Structure MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Ceramides MeSH
• Membrane Lipids MeSH

Ceramide (Cer) release from glucosylceramides (GlcCer) is critical for the formation of the skin permeability barrier. Changes in β-glucocerebrosidase (GlcCer'ase) activity lead to diminished Cer, GlcCer accumulation and structural defects in SC lipid lamellae; however, the molecular basis for this impairment is not clear. We investigated impaired GlcCer-to-Cer processing in human Cer membranes to determine the physicochemical properties responsible for the barrier defects. Minor impairment (5-25%) of the Cer generation from GlcCer decreased the permeability of the model membrane to four markers and altered the membrane microstructure (studied by X-ray powder diffraction and infrared spectroscopy), in agreement with the effects of topical GlcCer in human skin. At these concentrations, the accumulation of GlcCer was a stronger contributor to this disturbance than the lack of human Cer. However, replacement of 50-100% human Cer by GlcCer led to the formation of a new lamellar phase and the maintenance of a rather good barrier to the four studied permeability markers. These findings suggest that the major cause of the impaired water permeability barrier in complete GlcCer'ase deficiency is not the accumulation of free GlcCer but other factors, possibly the retention of GlcCer bound in the corneocyte lipid envelope.

• MeSH
• Administration, Topical MeSH
• Ceramides metabolism   MeSH
• X-Ray Diffraction MeSH
• Electric Impedance MeSH
• Glucosylceramides administration & dosage   metabolism   pharmacology   MeSH
• Indomethacin pharmacokinetics   MeSH
• Skin chemistry   drug effects   metabolism   MeSH
• Humans MeSH
• Membrane Lipids chemistry   metabolism   MeSH
• Cell Membrane Permeability * drug effects   MeSH
• Permeability MeSH
• Spectroscopy, Fourier Transform Infrared MeSH
• Theophylline pharmacokinetics   MeSH
• Check Tag
• Humans MeSH
• Female MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Ceramides MeSH
• Glucosylceramides MeSH
• Indomethacin MeSH
• Membrane Lipids MeSH
• Theophylline MeSH