The stratum corneum (SC) lipid matrix, composed primarily of ceramides (CERs), cholesterol and free fatty acids (FFA), has an important role for the skin barrier function. The presence of the long... Show moreThe stratum corneum (SC) lipid matrix, composed primarily of ceramides (CERs), cholesterol and free fatty acids (FFA), has an important role for the skin barrier function. The presence of the long periodicity phase (LPP), a unique lamellar phase, is characteristic for the SC. Insight into the lipid molecular arrangement within the LPP unit cell is imperative for understanding the relationship between the lipid subclasses and the skin barrier function. In this study, the impact of the CER head group structure on the lipid arrangement and barrier functionality was investigated using lipid models forming the LPP. The results demonstrate that the positions of CER N-(tetracosanoyl)-sphingosine (CER NS) and CER N-(tetracosanoyl)-phytosphingosine (CER NP), two essentials CER subclasses, are not influenced by the addition of another CER subclass (N-(tetracosanoyl)-dihydrosphingosine (CER NdS), N-(2R-hydroxy-tetracosanoyl)-sphingosine (CER AS) or D-(2R-hydroxy-tetracosanoyl)-phytosphingosine (CER AP)). However, differences are observed in the lipid organization and the hydrogen bonding network of the three different models. A similar localization of CER NP and CER NS is also observed in a more complex lipid model, with the CER subclass composition mimicking that of human SC. These studies show the adaptability and insensitivity of the LPP unit cell structure to changes in the lipid head group structures of the CER subclasses. Show less
Beddoes, C.M.; Gooris, G.S.; Barlow, D.J.; Lawrence, M.J.; Dalgliesh, R.M.; Malfois, M.; ... ; Bouwstra, J.A. 2022
The stratum corneum's lipid matrix is a critical for the skin's barrier function and is primarily composed of ceramides (CERs), cholesterol (CHOL) and free fatty acids (FFAs). The lipids form a... Show moreThe stratum corneum's lipid matrix is a critical for the skin's barrier function and is primarily composed of ceramides (CERs), cholesterol (CHOL) and free fatty acids (FFAs). The lipids form a long periodicity phase (LPP), a unique trilayer unit cell structure. An enzyme driven pathway is implemented to synthesize these key lipids. If these enzymes are down- or upregulated as in inflammatory diseases, the final lipid composition is affected often altering the barrier function. In this study, we mimicked down regulation of enzymes involved in the synthesis of the sphingosine and CER amide bond. In a LPP lipid model, we substituted CER N-(tetracosanoyl)-sphingosine (CER NS) with either i) FFA C24 and free sphingosine, to simulate the loss of the CER amide bond, or ii) with FFA C24 and C18 to simulate the loss of the sphingosine headgroup. Our study shows the lipids in the LPP would not phase separate until at least 25% of the CER NS is substituted keeping the lateral packing and conformational ordering unaltered. Neutron diffraction studies showed that free sphingosine chains localized at the outer layers of the unit cell, while the remaining CER NS head group was concentrated in the inner headgroup layers. However, when FFA C18 was inserted, CER NS was dispersed throughout the LPP, resulting in an even distribution between the inner and outer water layers. The presented results highlight the importance of the CER NS headgroup structure and its interaction in combination with the carbon chain invariability for optimal lipid arrangement. Show less
The extracellular lipid matrix in the skin's outermost layer, the stratum corneum, is crucial for the skin barrier. The matrix is composed of ceramides (CERs), cholesterol (CHOL) and free fatty... Show moreThe extracellular lipid matrix in the skin's outermost layer, the stratum corneum, is crucial for the skin barrier. The matrix is composed of ceramides (CERs), cholesterol (CHOL) and free fatty acids (FFAs) and involves two lamellar phases: the short periodicity phase (SPP) and the long periodicity phase (LPP). To understand the skin barrier thoroughly, information about the molecular arrangement in the unit cell of these lamellar phases is paramount. Previously we examined the molecular arrangement in the unit cell of the SPP. Furthermore X-ray and neutron diffraction revealed a trilayer arrangement of lipids within the unit cell of the LPP [D. Groen et al., Biophysical Journal, 97, 2242-2249, 2009]. In the present study, we used neutron diffraction to obtain more details about the location of lipid (sub)classes in the unit cell of the LPP. The diffraction pattern revealed at least 8 diffraction orders of the LPP with a repeating unit of 129.6±0.5Å. To determine the location of lipid sub(classes) in the unit cell, samples were examined with either only protiated lipids or selectively deuterated lipids. The diffraction data obtained by means of D2O/H2O contrast variation together with a gradual replacement of one particular CER, the acyl CER, by its partly deuterated counterpart, were used to construct the scattering length density profiles. The acyl chain of the acyl CER subclass is located at a position of ~21.4±0.2Å from the unit cell centre of the LPP. The position and orientation of CHOL in the LPP unit cell were determined using tail and head-group deuterated forms of the sterol. CHOL is located with its head-group positioned ~26±0.2Å from the unit cell centre. This allows the formation of a hydrogen bond with the ester group of the acyl CER located in close proximity. Based on the positions of the deuterated moieties of the acyl CER, CHOL and the previously determined location of two other lipid subclasses [E.H. Mojumdar et al., Biophysical Journal, 108, 2670-2679, 2015], a molecular model is proposed for the unit cell of the LPP. In this model CHOL is located in the two outer layers of the LPP, while CER EOS is linking the two outer layers with the central lipid layers. Finally the two other lipid subclasses are predominantly located in the central layer of the LPP. Show less
