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PubMed1993

Dioxygen diffusion in the stratum corneum: an EPR spin label study.

The stratum corneum, the outer 10 microns of the skin, serves as a permeability barrier regulating the transport of molecules between the body and the environment. The purpose of this study is to understand this permeability barrier function as it pertains to the diffusion of molecular oxygen. The stratum corneum was investigated with EPR spectroscopy following inoculation with a stearic acid spin probe. The presence of paramagnetic molecular oxygen results in the broadening of the EPR spectral lines of the spin probe. The rate of oxygen diffusion across the stratum corneum, and then the oxygen diffusion coefficient, D(O2), was determined by studying this line-broadening as a function of time. D(O2) in human stratum corneum was found to be 3 x 10(-7) cm2/s at 37 degrees C with an activation energy of approx. 44 kJ/mol. The application of the permeation-enhancing chemicals, DeMSO and DMSO, to the stratum corneum increased D(O2) two- to three-fold.

Animals↗
PubMed1995

Interaction of phosphatidylcholine liposomes with the human stratum corneum.

The interaction of dimyristoylphosphatidylcholine liposomes with the human stratum corneum was investigated by confocal laser scanning microscopy and differential scanning calorimetry. Human skin is characterized by a high autofluorescence. By introducing appropriate optical filters the autofluorescence of the skin was depressed and the penetration profile of fluorescence labelled vesicles was investigated. From optical sectioning it was obvious that neither the vesicles nor the fluorophore N-(lissamine rhodamine B sulfonyl)diacylphophatidylethanolamine (Rho-PE) penetrates in detectable amounts into the human skin. Differential scanning calorimetry of human stratum corneum revealed, that the peak positions of the human stratum corneum specific endothermic transitions at 10 degrees C, 35 degrees C, 50 degrees C, 62 degrees C, 73 degrees C and 81 degrees C did not change significantly after 18 h of non-occlusive vesicle application. However, the enthalpy of the transitions at 35 degrees C, 50 degrees C, 62 degrees C and 73 degrees C, estimated through peak heights increased, relative to the protein related peak at 81 degrees C. A novel transition at 10 degrees C was observed. From these data we conclude that DMPC liposomes do not penetrate intact into the human skin. We deduce, however, that the vesicles disintegrate at the surface of stratum corneum after non-occlusive application. The individual lipid molecules then interact with the lipid barrier of the stratum corneum and penetrate into the latter, which results in an increase of the enthalpy, related to the lipid components of the SC.

Calorimetry, Differential Scanning↗
PubMed1996

Phase transitions of rat stratum corneum lipids by an electron paramagnetic resonance study and relationship of phase states to drug penetration.

In order to relate barrier function to stratum corneum structure and the thermal transitions of corneum lipids, samples from hairless rat skin were investigated by using ESR and drug penetration techniques. The phase transition of stratum corneum lipids was estimated using a deeper probe (16-doxyl-stearic acid) inserted in the lipid bilayers and measuring the rotational correlation time, tau(c). Results of ESR study showed that stratum corneum lipids underwent thermal transitions at 39.3 +/- 1.6 degrees C and 63.6 +/- 2.6 degrees C roughly similar to the data obtained by differential scanning calorimetry measurements. Cholesterol oxidase treatment decreased the fluidity of the lipids at lower temperatures. The treatment of stratum corneum with laurocapram (1%) and isopropyl myristate (IPM, 2%) little changed both phase transition temperatures, although the treatment highly increased the molecular motion of the lipids. The flux (J(s)) of lipophilic drugs (beta-estradiol, indomethacin and betahistine) through the skin was enhanced with increasing temperatures, with an increase in the diffusion constant within skin and a decrease in the lag time. There was a good relationship between log J(s) or log permeability coefficient (K(p)) and 1/tau(c) in the temperature range of 45 to 64 degrees C. The calculated activation energy (delta E) for diffusion of these drugs across skin was 17-40 kcal/mol. Judging from our data, stratum corneum lipids of rat probably exist as the gel, crystalline state below 39 degrees C, the mesomorphic state between 39 and 64 degrees C and the fluid, liquid-crystalline state at temperatures of 64 degrees C or above. These results are in line with the permeability of these lipophilic drugs through the intercellular lipids disordered is highly increased.

Animals↗
PubMed1991

Effects of time of day, season and stratum on Haemonchus contortus and Haemonchus placei third-stage larvae on irrigated pasture.

The effect of time of day, season and stratum of herbage and soil on the availability of Haemonchus contortus and Haemonchus placei third-stage larvae (L3) on pasture was assessed. Feces from infected calves and lambs were placed on pasture plots and samples of upper herbage, lower herbage, mat and soil were collected at five intervals per day throughout the daylight hours on 18 sample days over 12 months. Using recovery rate factors derived from a preliminary investigation on the efficacy of larval recovery from each stratum, the data on larval recoveries were analyzed for the effect of season, time and stratum, and their interactions. Significant (P less than 0.05) differences were found for season, stratum and the season-with-stratum interaction for both parasites. No significant differences were detected for larval counts at different times of the day. Larval recoveries of H. contortus were larger throughout the study than those of H. placei. Most H. contortus L3 were recovered in the summer and autumn, and H. placei in the spring and summer. For both parasites, the recoveries of larvae from the upper and lower herbage were larger than those from the mat and soil. The implications of these findings are discussed in terms of control strategies.

Animals↗
PubMed1981

Postnatal ontogenesis of hippocampal CA1 area in rats. II. Development of ultrastructure in stratum lacunosum and moleculare.

Fine structure of stratum moleculare and lacunosum of the hippocampal CA1 area was studied in rats 5, 10, 15, 24, 48 and 90 days old. By means of an electron microscope the detailed description of both layers as well as quantitative data about the development of neuropile were obtained. The density of dendrites per 1 mm3 decreased during the development. Adult values were reached in both layers at the age of 24 days. Formation of axon terminals extended in the stratum lacunosum till the 24th day; in the stratum moleculare the density of terminals was highest on the 15th day, but decreased during the later period. The density of axo-dendritic synapses did not change in the whole postnatal period while that of axo-spine synapses considerably increased. Dendritic spines increased their density in both layers, having reached their maximum on the 24th day in the stratum moleculare and on the 48th day in the stratum lacunosum. The significance of fine structure development becomes apparent in light of the distribution of afferent fibers and the functional activity of pyramidal neurons. The gradual development of individual afferent systems probably results from the input information pattern conveyed by the afferents. The resulting integration of pyramidal neurons should thus be subjected to a substantial change during development.

Aging↗
PubMed2005

The influence of topical formulations on the permeation of 5-aminolevulinic acid and its n-butyl ester through excised human stratum corneum.

The limited permeation of 5-aminolevulinic acid (ALA) through excised human stratum corneum could be improved by using 5-aminolevulinic acid-n-butyl ester (ABE). Furthermore drug permeation could be increased by choice of a permeation enhancing formulation. In this study, permeation of ALA and ABE was investigated from various formulations. In addition, differential scanning calorimetry (DSC) and wide angle X-ray diffraction (WAXD) experiments were performed in order to reveal an interaction between the tested formulations and stratum corneum lipid structure. Drug incorporation into Dolgit Mikrogel showed the highest increase in permeability with both ALA and ABE. Especially, ABE together with Dolgit Mikrogel was the most promising combination. Further permeation studies with poloxamer based ABE formulations, partially enriched with ibuprofen acid and medium chained triglycerides showed that both compounds promote permeation. The permeation coefficients of either drug from Excipial Creme and Basiscreme DAC were found to be very similar. These results were in accordance with those of DSC and WAXD experiments. Interaction between formulation and stratum corneum lipid structure resulting in an increased drug permeation only occurred after pretreatment with formulations enriched with ibuprofen acid. After pretreatment with Excipial Creme, Basiscreme DAC or Excipial Fettcreme stratum corneum structure and subsequently permeability remained unchanged. Nevertheless permeation of ALA from Excipial Fettcreme is slower than from the tested hydrophilic formulations and therefore believed to be influenced by the affinity of ALA to the vehicle and stratum corneum.

Administration, Cutaneous↗
PubMed2006

Combined effects of ethanol and L-menthol on hairless rat stratum corneum investigated by synchrotron X-ray diffraction.

Synchrotron X-ray diffraction was employed to evaluate the effect of ethanol and l-menthol on lipid arrangements in the stratum corneum of hairless rats. Two sharp diffractions (S=2.40 and S=2.67, corresponding to spacing of 0.417 nm and 0.374 nm respectively) were observed on the broad hump peak derived from soft keratin. To assist in understanding the effects of treatment with ethanol and l-menthol, an abundance ration of lipid hydrocarbon chain packing index (R(H/O)) was defined as R(H/O)=(Peak area at S=2.40 nm(-1))/(Peak area at S=2.67 nm(-1)). When ethanol was applied to the stratum corneum the intensities of diffraction peaks declined slightly. The R(H/O) values observed were not affected by variations in ethanol concentrations in the range 0-40% (w/w). The R(H/O) values did not change even when treatment with ethanol (40%, w/w) was extended to 8 h. These results suggested that lipid arrangements in the stratum corneum were not affected by ethanol. On the other hand, exposure of the stratum corneum to 2% (w/w) L-menthol caused a significant decrease in R(H/O) value. It was shown that L-menthol was dispersed through the stratum corneum, intruded mainly into hexagonal hydrocarbon chain packing, and disrupted the regular organization of these structures.

Animals↗
PubMed2005

Effect of vehicles and sodium lauryl sulphate on xenobiotic permeability and stratum corneum partitioning in porcine skin.

Dermal contact with potentially toxic agricultural and industrial chemicals is a common hazard encountered in occupational, accidental spill and environmental contamination scenarios. Different solvents and chemical mixtures may influence dermal absorption. The effects of sodium lauryl sulphate (SLS) on the stratum corneum partitioning and permeability in porcine skin of 10 agricultural and industrial chemicals in water, ethanol and propylene glycol were investigated. The chemicals were phenol, p-nitrophenol, pentachlorophenol, methyl parathion, ethyl parathion, chlorpyrifos, fenthion, simazine, atrazine and propazine. SLS decreased partitioning into stratum corneum from water for lipophilic compounds, decreased partitioning from propylene glycol and did not alter partitioning from ethanol. SLS effects on permeability were less consistent, but generally decreased permeability from water, increased permeability from ethanol and had an inconsistent effect on permeability from propylene glycol. It was concluded that, for the compounds tested, partitioning into the stratum corneum was determined by the relative solubility of the solute in the donor solvent and the stratum corneum lipids. Permeability, however, reflected the result of successive, complex processes and was not predictable from stratum corneum partitioning alone. Addition of SLS to solvents altered partitioning and absorption characteristics across a range of compounds, which indicates that partition coefficients or skin permeability from neat chemical exposure should be used with caution in risk assessment procedures for chemical mixtures.

Agrochemicals↗
PubMed1998

Entropy-driven binding/partition of amino acids/dipeptides to stratum corneum lipid vesicles.

This study employed large unilamillar vesicles composed of purchased stratum corneum lipids to investigate the binding/partition of amino acids/dipeptides to stratum corneum lipid vesicles. The partition coefficients of amino acids/dipeptides between the stratum corneum lipid vesicles and the acetate buffer were determined by HPLC. In addition, the binding/partition enthalpy of amino acids/dipeptides with the stratum corneum lipid vesicles was derived by directly measuring the binding/partition heat with isothermal titration calorimetry. According to the binding/petition Gibbs free energy and the binding/partition enthalpy, all the binding/partition of amino acids/dipeptides with the stratum corneum lipid vesicles is endothermic, implying an entropy-driven binding/partition. Also, the equilibrium binding/partition results demonstrate that the partition coefficients of amino acids/dipeptides do not correlate with the transdermal permeability. This finding suggests that either the interaction between the penetrants and the lipid bilayer between corneocytes may not be a determining step or that the paracellular path is not a dominant route of transdermal penetration.

Amino Acids↗
PubMed1999

Effects of iontophoresis and electroporation on the stratum corneum. Review of the biophysical studies.

This review focuses on the effects induced by iontophoresis and electroporation on the stratum corneum of the skin. Hence, the aims were: (1) to contribute to the understanding of the mechanisms of drug transport by these methods; (2) to evaluate the safety issues associated with current application. Complementary biophysical methods were used to provide a complete picture of the stratum corneum. Even though the mechanism of drug transport is believed to be different, i.e., electrophoresis for iontophoresis and creation of new aqueous pathways for electroporation, the effects on the stratum corneum detected minutes after current application are very similar. For both methods, the major findings were: (1) a disorganisation of the lipid bilayers of the stratum corneum; (2) an increase in skin hydration; (3) a larger decrease in skin resistance induced by electroporation as compared to iontophoresis. These changes were partly reversible and depended on the amount of electrical charges transferred. The mechanisms of stratum corneum perturbations are discussed. These perturbations could explain partly the increase in drug transport. If iontophoresis is considered as a safe method of drug delivery, the data augurs for the safety of electroporation.

Journal Article↗
PubMed1997

Activation of interneurons at the stratum oriens/alveus border suppresses excitatory transmission to apical dendrites in the CA1 area of the mouse hippocampus.

The consequences of activation or inactivation of interneurons at the CA1 stratum oriens/ alveus border for signal transmission at the apical dendritic region of pyramidal cells were investigated in slices from mice submerged in a perfusion chamber. A characteristic subpopulation of interneurons with a horizontal dendritic tree in this region, which sends a GABAergic projection to the apical dendrites of CA1 pyramidal cells is strongly excited by metabotropic glutamate receptor activation and receives GABAergic input from vasoactive intestinal polypeptide-containing interneurons. Pressure ejection of glutamate or the metabotropic agonist 1s,3r-aminocyclopentane dicarboxylic acid from micropipettes onto the stratum oriens/alveus border caused a long lasting (more than 90 min) decrease of field-excitatory postsynaptic potentials in the strata radiatum and lacunosum-moleculare. The GABAB antagonist CGP 35348 (100 microM in the perfusion fluid) partially and reversibly blocked this effect. Vasoactive intestinal polypeptide- (0.1 microM in the bath) excited neurons with response and firing properties characteristic for interneurons at the oriens/alveus border. Local pressure application of vasoactive intestinal polypeptide (10 microM) to the alveus region led, after a brief (2 min) and small (10%) increase, to a longer lasting (30-50 min) decrease (by 20-30%) in the slope of the field-excitatory postsynaptic potential in strata radiatum and lacunosum-moleculare. This action was completely blocked by bath application of CGP 35348. Local application of tetrodotoxin in the stratum oriens/alveus region markedly increased the slope of evoked dendritic excitatory postsynaptic potentials, and caused multiple firing of pyramidal cells. Thus, stratum oriens/alveus interneurons have a profound inhibitory effect on signal transmission in the apical dendritic area of CA1, which is, at least in part, mediated by GABAB receptors. It appears that the GABAB receptor-mediated effect in stratum lacunosum-moleculare is produced by vasoactive intestinal polypeptide-sensitive interneurons.

Animals↗
PubMed1997

Individual variation of human plantar stratum corneum lipids, determined by automated multiple development of high-performance thin-layer chromatography plates.

The stratum corneum lipids are unique in composition and have been used frequently as a model system of the skin's lipid barrier. Automated multiple development (AMD) of high-performance thin-layer chromatography plates in combination with a 25-step gradient, based on methanol, diethyl ether and n-hexane separated the six major human plantar stratum corneum lipids. Post-chromatographic staining of these lipids with a solution of MnCl2-H2SO4 at 130 degrees C or a solution of CuSO4-H3PO4 at 140 degrees C allowed visualization of the lipids and quantification. The MnCl2-H2SO4 solution stained saturated fatty acids less intensely. Therefore, the CuSO4-H3PO4 solution was used for quantification and we found, on average, 2.06% (w/w) cholesterol 3-sulphate, 20.16% (w/w) free fatty acids, 20.25% (w/w) ceramides, 43.53% (w/w) non-esterified sterols, 4.56% (w/w) triacylglycerols and 9.4% (w/w) sterolesters in the human plantar stratum corneum extracts. The concentration of phospholipids was less than 1% (w/w). In addition, the lipid composition of twenty different human plantar stratum corneum extracts was determined. Statistics revealed a correlation between the ratio of free fatty acids and non-esterified sterols (r=0.832, p<0.01, n=20). Several control experiments proved that this correlation is not due to the extraction method, the post-chromatographic staining procedure or bacterial contamination of the stratum corneum.

Animals↗
PubMed1999

A new model system for lipid interactions in stratum corneum vesicles: effects of lipid composition, calcium, and pH.

We prepared large unilamellar vesicles (LUVs) with three different stratum corneum lipid compositions: constant amounts of ceramides (55 wt %) and fatty acids (15%) with varying amounts of cholesterol sulfate (0-15%) and cholesterol (15-30%). One of the compositions served as a model for normal stratum corneum, while the second one served as a model for recessive X-linked ichthyosis stratum corneum. The third composition consisted of no cholesterol sulfate. Intervesicle lipid interactions in these LUVs were monitored by fluorescence methods for content leakage, and contents mixing at pH 9, in the absence and presence of Ca2+, and at pH 6. Since the content leakage and contents mixing assays were originally developed for phospholipid vesicles, we characterized the probe binding and the probe quenching properties for stratum corneum LUV systems, and modified the assays slightly accordingly. The time-dependent fluorescence intensity changes in the probe-containing LUVs at pH 9 and 6 and in response to the addition of calcium were monitored. Our results demonstrated that all three types of LUVs were relatively stable at pH 9. Addition of Ca2+ or decreasing the pH to 6 activated intervesicle lipid mixing followed by vesicle fusion and lysis. We found that the LUVs with no cholesterol sulfate and 30% cholesterol exhibited a more extensive Ca2+- or low-pH-activated intervesicle lipid interaction than LUVs with either 5% cholesterol sulfate and 25% cholesterol or 15% cholesterol sulfate and 15% cholesterol. These results suggest that fusogenic agents such as Ca2+ and H+ act to neutralize the fatty acids in the lipid bilayer of stratum corneum vesicles. The inclusion of 5-15% cholesterol sulfate helps to prevent the collapse of fused vesicles into other structures.

Animals↗
PubMed1998

Experimental penetration of Trichophyton mentagrophytes into human stratum corneum.

We present confirmation of the experimental penetration of Trichophyton mentagrophytes into human stratum corneum under designated conditions of temperature and humidity. When stratum corneum, obtained from healthy human heel region, was incubated at 100% humidity, mycelium was observed in the corneum layer on day 2 at 35 degrees C and 27 degrees C, and on day 4 at 15 degrees C. At 90% humidity, the hyphae penetrated into the stratum corneum on day 4 at 35 degrees C, and on day 6 at 27 degrees C. Whereas, at 80% humidity, no fungal elements were observed in the stratum corneum at both 27 degrees C and 35 degrees C for up to 7 days. These data suggested that humidity was a more important environmental factor for penetration than temperature, and that at least 90% humidity is necessary for dermatophytes to penetrate into the stratum corneum within a few days. Mean humidity in the interdigital space between the fourth and fifth toes was found to be approximately 98%.

Adult↗
PubMed1994

Subzero thermal analysis of human stratum corneum.

The thermal behaviour of human stratum corneum was studied using differential thermal analysis within the temperature range of -130 degrees C to 120 degrees C. Aside from thermal transitions at around 40 degrees C, 70 degrees C, 85 degrees C and 100 degrees C, which have been reported before, a particular transition below 0 degree C (subzero), at approx. -9 degrees C (264 K), was noticed. This transition was present in the analysis curves of dehydrated as well as hydrated stratum corneum sheets and could be distinguished from the water peak found only in hydrated stratum corneum samples. To further characterize this transition, thermal analysis was performed on stratum corneum sheets: (i) after lipid extraction, (ii) after pretreatment of propylene glycol and (iii) after pretreatment of oleic acid/propylene glycol solution. From the results, it was concluded that the subzero transition (-9 degrees C) belongs to low melting lipid components of stratum corneum.

Cold Temperature↗
PubMed1993

A critical comparison of some Raman spectroscopic techniques for studies of human stratum corneum.

This study evaluates a variety of techniques and sampling conditions for Raman spectroscopic investigations of human stratum corneum. Using a Fourier-transform Raman system and samples of stratum corneum in vitro, we demonstrated minimal inter- and intracadaver variations in molecular vibrations. We have also shown Raman spectroscopy to be relatively insensitive to the hydration state of human stratum corneum, indicating that the technique should be valuable for monitoring the transdermal delivery of drugs from aqueous solutions. The stability of human stratum corneum to near-infrared laser excitation was verified by spectral collection for approximately 1 hr. We have also compared FT-Raman spectra from human stratum corneum in vitro and in vivo. Of the different types of Raman instruments used in this study (visible-light excitation microprobe, visible-light excitation macroscopic sampling, and Fourier-transform Raman), the FT-Raman system provided good-quality spectra with high sample throughput, but systems using visible-light excitation should provide unique information for use in specialist applications.

Epidermis↗
PubMed1994

Organization of stratum corneum lipids in relation to permeability: influence of sodium lauryl sulfate and preheating.

The role of the structural organization of intercorneocyte lipids in the barrier function of human stratum corneum was evaluated by treatment with heat and sodium lauryl sulfate. Measurement of transepidermal water loss in treated samples was used to quantify variations in stratum corneum permeability. Thermodynamic transition of lamellar lipids and their degree of organization were evaluated by differential scanning calorimetry and small-angle X-ray diffraction, respectively. Progressively preheating stratum corneum samples from 75 degrees C to 90 degrees C increased stratum corneum permeability to water vapor, while the fusion temperature of lamellar lipids and the intensity of the X-ray diffraction peaks of the polar lipids decreased. Sodium lauryl sulfate induced similar variations of these three parameters. These results support the hypothesis that, in addition to the chemical nature of intercorneocyte lipids, their structural arrangement and thermodynamic properties play an important role in the barrier function of the stratum corneum to water vapor.

Calorimetry, Differential Scanning↗
PubMed2000

A study on structural and diffusion properties of porcine stratum corneum based on very small angle neutron scattering data.

PURPOSE: Generation of valuable information about the biphasic geometrical configuration of porcine stratum corneum from Very Small Angle Neutron Scattering (VSANS) data and investigation of its effect on the corresponding effective diffusivity. METHODS: Spectra of porcine stratum corneum are mathematically transformed in order to obtain the corresponding auto-correlation function (ACF). Model stratum corneum structures, matching this experimentally determined ACF, are then produced based on the "brick-and-mortar" configuration. The effective diffusivity through these model domains is calculated using an appropriate numerical method. RESULTS: The most appropriate geometry of porcine stratum corneum's lipid and protein phases in a "brick-and-mortar" configuration is quantitatively determined and correlated with the barrier properties (diffusivity) of the stratum corneum model structures. CONCLUSIONS: The ACF analysis indicates the most appropriate values for the dimensions of the corneocyte thickness and the surrounding lipid gap, while the corneocyte length is estimated from the diffusion study.

Animals↗