Influence of a Novel Dimeric Ceramide Molecule on the Nanostructure and Thermotropic Phase Behavior of a Stratum Corneum Model Mixture

Abstract

The stratum corneum (SC) is the outermost layer of the skin and is composed of a multilayered assembly of mostly ceramids (Cer), free fatty acids, cholesterol (Chol), and cholesterol sulfate (Chol-S). Because of the tight packing of these lipids, the SC features unique barrier properties defending the skin from environmental influences. Under pathological conditions, where the skin barrier function is compromised, topical application of molecules that rigidify the SC may lead to a restored barrier function. To this end, molecules are required that incorporate into the SC and bring back the original rigidity of the skin barrier. Here, we investigated the influence of a novel dimeric ceramide (dim-Cer) molecule designed to feature a long, rigid hydrocarbon chain ideally suited to forming an orthorhombic lipid phase. The influence of this molecules on the thermotropic phase behavior of a SC mixture consisting of Cer[AP18] (55 wt %), cholesterol (Chol, 25 wt %), steric acid (SA, 15 wt %), and cholesterol sulfate (Chol-S, 5 wt %) was studied using a combination of neutron diffraction and H NMR spectroscopy. These methods provide detailed insights into the packing properties of the lipids in the SC model mixture. Dim-Cer remains in an all-trans state of the membrane-spanning lipid chain at all investigated temperatures, but the influence on the phase behavior of the other lipids in the mixture is marginal. Biophysical experiments are complemented by permeability measurements in model membranes and human skin. The latter, however, indicates that dim-Cer only partially provides the desired effect on membrane permeability, necessitating further optimization of its structure for medical applications.

The Sphingosine and Acyl Chains of Ceramide [NS] Show Very Different Structure and Dynamics That Challenge Our Understanding of the Skin Barrier

“…In SC research, 2 H labeling is available for the Cer acyl chains, the fatty acids, and Chol, which were shown as highly rigid in many studies. [4,5,12] Methods that do not require isotopic enrichment ( 13 C MAS NMR [6] and FTIR [7,8] spectroscopy) are confronted with dynamic range problems as the highly intense NMR peaks or vibrational bands of the chain methylene groups render detection of low-intensity signals very difficult. These segments, however, could feature an alternative dynamic signature.…”

“…Such NMR spectra are obtained for rigid all-trans acyl chains. [5,12] Only the methyl groups undergo three-site hopping, reducing the quadrupolar splitting by 1/3. [21] In stark contrast, the Cer[NS]-sph-d 28 sphingosine moiety shows much different NMR spectra featuring two contributions.…”

Die unterschiedliche Struktur und Dynamik der Sphingosin‐ und Acylketten von Ceramid [NS] verändern unser Verständnis der Struktur der Hautbarriere

“…Die Lignocerylkette von Cer[NS]‐acyl‐ d 47 zeigte nahezu vollständig eine orthorhombische Phase bei 25 und 32 °C in beiden Mischungen. Solche NMR‐Spektren werden für rigide all‐ trans ‐Acylketten erhalten . Nur die Methylgruppen führen Sprünge mit dreizähliger Symmetrie aus, was die beobachtete Quadrupolaufspaltung auf 1/3 reduziert .…”

“…2 H-Markierung steht momentan nur für die Cer-Acylketten, die FFS und Chol-Segmente zur Verfügung, die sich in vielen Studien als sehr starr erwiesen. [4,5,12] Methoden, die keine Isotopenmarkierung bedingen wie 13 C-MAS-NMR- [6] oder IR-Spektroskopie, [7,8] sind oft unempfindlich für Signale geringer Intensität, deren Detektion aufgrund technischer Beschränkungen angesichts sehr intensiver NMR-Signale und IR-Banden erschwert ist. Diese Segmente kçnnten allerdings eine alternative dynamische Signatur aufweisen.…”

“…Solche NMR-Spektren werden für rigide all-trans-Acylketten erhalten. [5,12] Nur die Methylgruppen führen Sprünge mit dreizähliger Symmetrie aus, was die beobachtete Quadrupolaufspaltung auf 1/3 reduziert. [21] Im Gegensatz dazu lieferten die Sphingosinketten von Cer[NS]-sph-d 28 deutlich andere NMR-Spektren mit zwei Beiträgen: der erste zeigt eine spektrale Intensitätsverteilung, wie sie in Gelphasen von Phospholipidmembranen beobachtet wird.…”