Permeable Cornified Envelope Layer Regulates the Solute Transport in Human Stratum Corneum.

To unravel the diffusion mechanisms of percutaneous drug delivery, suitable numerical analysis of stratum corneum structure is essential. In this research paper, we accounted for the permeable envelope layer in the brick-and-mortar finite element models of human stratum corneum. Both penetration and desorption experiments for tritiated water were simulated by transient finite element analysis. Rivet-shaped corneodesmosomes were included in the brick and mortar model. Results showed that cornified lipid permeability (Penv) is a determinant in desorption of the solute, while lipid transverse diffusion coefficient (Dlip-trans) is prominent during penetration. These two major unknowns (Penv and Dlip-trans) were obtained by extensive fitting of the finite element model to the experimental water data. Penv and Dlip-trans were determined to be 1×10-2 cm/s and 5.7×10-10 cm2/s, respectively.

Stratum corneum ceramide profiles in vitro, ex vivo, and in vivo: characterization of the α-hydroxy double esterified ceramides.

The presence of a new ceramide subclass, the 1-O-acyl omega-linoleoyloxy ceramides [1-O-E (EO) Cer], has been previously highlighted in reconstructed human epidermis (RHE). These ceramides are double esterified on two positions. The first is the 1-O position of the sphingoid base moiety with a long to very long chain of acyl residues (1-O-E), and the second is the position of the ω-hydroxyl group of the fatty acid moiety with linoleic acid (EO). Considering its chemical structure and hydrophobicity, this subclass can contribute to the skin barrier. Thus, it is important to determine whether this subclass is also present in native human stratum corneum (SC). This work compares ceramide structures of this novel subclass between RHE (in vitro) and two sources of human SC (in vivo and ex vivo) using normal-phase high-performance liquid chromatography coupled to high-resolution mass spectrometry (NP-HPLC/HR-MSn). The results confirm the presence of this double esterified ceramide subclass [1-O-E (EO) Cer] in human SC. The molecular profile obtained from the RHE was very close to that found in the human SC (in vivo and ex vivo). In addition, thanks to the targeted MS2/MS3 analysis, a new ceramide subclass was discovered and characterized in the three studied samples. We propose to name it [A-1-O-E (EO) Cer] because in these ceramides species, the fatty acid-esterified with the sphingoid base on the 1-O position-is hydroxylated on the α position. These results highlight the potential of both the analytical method and the characterization approach employed in this study.

Ultrasound-Vortex-Assisted Dispersive Liquid-Liquid Microextraction Combined with High Performance Liquid Chromatography-Diode Array Detection for Determining UV Filters in Cosmetics and the Human Stratum Corneum.

This study explores the amounts of common chemical ultraviolet (UV) filters (i.e., avobenzone, bemotrizinol, ethylhexyl triazone, octocrylene, and octyl methoxycinnamate) in cosmetics and the human stratum corneum. An ultrasound-vortex-assisted dispersive liquid-liquid microextraction (US-VA-DLLME) method with a high-performance liquid chromatography-diode array detector was used to analyze UV filters. A bio-derived solvent (i.e., anisole) was used as the extractant in the US-VA-DLLME procedure, along with methanol as the dispersant, a vortexing time of 4 min, and ultrasonication for 3 min. The mass-transfer rate of the extraction process was enhanced due to vortex-ultrasound combination. Various C18 end-capped columns were used to investigate the separation characteristics of the UV filters, with XBridge BEH or CORTECS selected as the separation column. Calibration curves were constructed in the 0.05-5 µg/mL (all filters except octocrylene) and 0.1-10 µg/mL (octocrylene) ranges, and excellent analytical linearities with coefficients of determination (r2) above 0.998. The developed method was successfully used to analyze sunscreen. Moreover, experiments were designed to simulate the sunscreen-usage habits of consumers, and the cup method was used to extract UV filters from the human stratum corneum. The results suggest that a makeup remover should be employed to remove water-in-oil sunscreens from skin.

Sulforaphane-Loaded Ultradeformable Vesicles as A Potential Natural Nanomedicine for the Treatment of Skin Cancer Diseases.

Sulforaphane is a multi-action drug and its anticancer activity is the reason for the continuous growth of attention being paid to this drug. Sulforaphane shows an in vitro antiproliferative activity against melanoma and other skin cancer diseases. Unfortunately, this natural compound cannot be applied in free form on the skin due to its poor percutaneous permeation determined by its physico-chemical characteristics. The aim of this investigation was to evaluate ethosomes® and transfersomes® as ultradeformable vesicular carriers for the percutaneous delivery of sulforaphane to be used for the treatment of skin cancer diseases. The physico-chemical features of the ultradeformable vesicles were evaluated. Namely, ethosomes® and transfersomes® had mean sizes <400 nm and a polydispersity index close to 0. The stability studies demonstrated that the most suitable ultradeformable vesicles to be used as topical carriers of sulforaphane were ethosomes® made up of ethanol 40% (w/v) and phospholipon 90G 2% (w/v). In particular, in vitro studies of percutaneous permeation through human stratum corneum and epidermis membranes showed an increase of the percutaneous permeation of sulforaphane. The antiproliferative activity of sulforaphane-loaded ethosomes® was tested on SK-MEL 28 and improved anticancer activity was observed in comparison with the free drug.

Characterization of Water and a Model Lipophilic Compound in Human Stratum Corneum by NMR Spectroscopy and Equilibrium Sorption.

The stratum corneum (SC) is the outermost skin layer in humans and other mammals and the primary barrier to water loss and environmental exposure to chemicals and microorganisms. It consists of flattened, keratin-filled corneocytes surrounded by well-organized lipid layers. Human SC at varying degrees of hydration with and without addition of a model lipophilic compound, 2-(trifluoromethyl) benzonitrile (TFMB), was studied using proton (1H) and fluorine (19F) nuclear magnetic resonance techniques. Proton spectral analyses revealed that water mainly occupies the corneocytes in agreement with prior studies. Observations from 19F spectral and spin-lattice relaxation time (T1) analyses showed that TFMB is primarily present in the lipids with small amounts in water, which is located within the corneocytes. This is consistent with TFMB sorption, which was measured in SC with and without lipid extraction. The presence of TFMB within the corneocytes supports the hypothesis that transcellular diffusion of a lipophilic compound like TFMB may contribute to SC permeation.

Thermal phase transition behavior of lipid layers on a single human corneocyte cell.

We have improved the selected area electron diffraction method to analyze the dynamic structural change in a single corneocyte cell non-invasively stripped off from human skin surface. The improved method made it possible to obtain reliable diffraction images to trace the structural change in the intercellular lipid layers on a single corneocyte cell during heating from 24°C to 100°C. Comparison of the results with those of synchrotron X-ray diffraction experiments on human stratum corneum sheets revealed that the intercellular lipid layers on a corneocyte cell exhibit essentially the same thermal phase transitions as those in a stratum corneum sheet. These results suggest that the structural features of the lipid layers are well preserved after the mechanical stripping of the corneocyte cell. Moreover, electron diffraction analyses of the thermal phase transition behaviors of the corneocyte cells that had the lipid layers with different distributions of orthorhombic and hexagonal domains at 24°C suggested that small orthorhombic domains interconnected with surrounding hexagonal domains transforms in a continuous manner into new hexagonal domains.

Spectroscopic and thermal characterization of alternative model biomembranes from shed skins of Bothrops jararaca and Spilotis pullatus / Caracterização espectroscópica e térmica de biomembranas modelo alternativo de peles galpão de Bothrops jararaca e Spilotis pullatus

Recently, there has been an interest in the use of shed snake skin as alternative model biomembrane for human stratum corneum. This research work presented as objective the qualitative characterization of alternative model biomembranes from Bothrops jararaca and Spilotis pullatus by FT-Raman, PAS-FTIR and DSC. The employed biophysical techniques permitted the characterization of the biomembranes from shed snake skin of B. jararaca and S. pullatus by the identification of vibrational frequencies and endothermic transitions that are similar to those of the human stratum corneum.

Existe atualmente interesse no uso da muda de pele de cobra como modelos alternativos de biomembranas da pele humana. O presente trabalho apresentou como objetivo a caracterização qualitativa de modelos alternativos de biomembranas provenientes de mudas de pele de cobra da Bothrops jararaca e Spilotis pullatus por espectroscopia Raman (FT-Raman), espectroscopia fotoacústica no infravermelho (PAS-FTIR) e calorimetria exploratória diferencial (DSC). As técnicas biofísicas FT-Raman, PAS-FTIR e DSC permitiram caracterizar qualitativamente os modelos alternativos de biomembranas provenientes das mudas de pele de cobra da B. jararaca e S. pullatus e identificar freqüências vibracionais e transições endotérmicas similares ao estrato córneo humano.

Immunohistochemical study of crooke's hyaline change.

Crooke's hyaline change of the human pituitary gland appears as an intracytoplasmic accumulation of fine filaments under electron microscopy. This study was attempted to identify the fine filaments by immunohistochemical methods. Twenty-eight postmortem, formalin-fixed or chrome-alum-fixed, paraffin-embedded pituitary glands revealing unequivocal Crooke's hyaline change on hematoxylin and eosin stain were selected for this study. To demonstrate Crooke's cells and fine filaments simultaneously, mirror image sections were sliced and stained with the following monoclonal antibodies using an avidin-biotin-peroxidase complex method: an antibody against synthesized adrenocorticotropic hormone 1-24, human cytokeratins (55-57 kilodalton [kd] and 68 kd), porcine vimentin (57 kd), porcine desmin (53 kd), bovine neurofilaments (70, 160, and 210 kd), human glial fibrilfary acidic protein (GFAP) (56 kd), and chicken actin. Crooke's cells showed a variable intensity of cytoplasmic staining for 55- to 57-kd cytokeratins, from focal to more even and intense staining revealing a characteristic wide brown ring around the nucleus or beneath the cell membrane. The most severely affected cells were totally replaced by dark brown reaction products with no secretory granules detectable in the cytoplasm. However, 68-kd cytokeratin could not be unequivocally demonstrated. Crooke's cells were all negative for vimentin, desmin, neurofilaments, GFAP, and actin. Thus far, it could be concluded that Crooke's hyaline change was composed of intermediate-subunit molecular weight cytokeratins that are normal constituents of the ACTH cell.