Effects of 6-Hydroxyceramides on the Thermotropic Phase Behavior and Permeability of Model Skin Lipid Membranes

Abstract

Ceramides (Cer) based on 6-hydroxysphingosine are important components of the human skin barrier, the stratum corneum. Although diminished concentrations of 6-hydroxyCer have been detected in skin diseases such as atopic dermatitis, our knowledge on these unusual sphingolipids, which have only been found in the skin, is limited. In this work, we investigate the biophysical behavior of N-lignoceroyl-6-hydroxysphingosine (Cer NH) in multilamellar lipid membranes composed of Cer/free fatty acids (FFAs) (C16-C24)/cholesterol/cholesteryl sulfate. To probe the Cer structure-activity relationships, we compared Cer NH membranes with membranes containing Cer with sphingosine (Cer NS), dihydrosphingosine, and phytosphingosine (Cer NP), all with the same acyl chain length (C24). Compared with Cer NS, 6-hydroxylation of Cer not only increased membrane water loss and permeability in a lipophilic model compound but also dramatically increased the membrane opposition to electrical current, which is proportional to the flux of ions. Infrared spectroscopy revealed that Cer hydroxylation (in either Cer NH or Cer NP) increased the main transition temperature of the membrane but prevented good Cer mixing with FFAs. X-ray powder diffraction showed not only lamellar phases with shorter periodicity upon Cer hydroxylation but also the formation of an unusually long periodicity phase (d = 10.6 nm) in Cer NH-containing membranes. Thus, 6-hydroxyCer behaves differently from sphingosine- and phytosphingosine-based Cer. In particular, the ability to form a long-periodicity lamellar phase and highly limited permeability to ions indicate the manner in which 6-hydroxylated Cer contribute to the skin barrier function.

Behavior of 1-Deoxy-, 3-Deoxy- and N-Methyl-Ceramides in Skin Barrier Lipid Models

“…The steady-state flux of TH for the control model sample, i.e., Cer/LIG/Chol/CholS, was 0.11 ± 0.02 µg/h/cm 2 . This is in good agreement with published data 9 . The permeability coefficient for TH through these solid lipid sample (1.22 × 10 −9 cm.s −1 ; calculated according to Mitragotri et al 51 from the TH flux and its concentration in the donor sample 52 ) is approximately 5 orders of magnitude lower compared to its permeability through a fluid bilayer (2.9 × 10 −4 cm.…”

“…This method is mostly used to evaluate the skin barrier condition in vivo 41,42 , but is also useful in in vitro studies on skin [43][44][45] and model systems 43,46 . The water loss through the control sample, i.e., Cer/ LIG/Chol/CholS, was 1.29 ± 0.11 g/h/m 2 , which is consistent with the value published previously (1.40 ± 0.10 g/h/ m 2 ) 9 . A direct comparison of the water loss through the membrane and skin is not possible because of the differences in the instrument used, lipid amount, absence/presence of corneocytes, sweat glands, etc, but these values are mostly within the same order of magnitude 34,47 .…”

“…2A and Supporting Table S1). The d of this short lamellar phase (SLP) is consistent with those found in analogous SC lipid models containing LIG 8 or a mixture of free fatty acids with C16-C24 chain lengths 9,22,23 . In addition, a separated Chol phase with d (001) = 3.41 nm was found.…”

Seasonal changes in epidermal ceramides are linked to impaired barrier function in acne patients

“…Recent findings from studies using model membranes provide mixed evidence on their roles in barrier function. Model membranes with CER[NH] instead of CER[NS] had reduced ion permeability, although this was accompanied by increased water permeability . Additionally, the presence of multiple hydroxy groups (3 in CER[NH], 4 in CER[AH]) alters lipid packing in a way similar to inclusion of phyto‐ceramides in model membranes .…”

“…[7] While CER[EOS] and CER[EOH] levels are crucial for SC organisation and barrier function, [15,16] this was accompanied by increased water permeability. [17] Additionally, the presence of multiple hydroxy groups (3 in CER[NH], 4 in CER[AH]) alters lipid packing in a way similar to inclusion of phyto-ceramides in model membranes. [18,19] The physiological relevance of these findings remains uncertain, and it seems likely that an effective barrier depends on the inclusion of correct proportions of different ceramide classes.…”