Bo Forslind

A new HPLC-based method for the quantitative analysis of inner stratum corneum lipids with special reference to the free fatty acid fraction.

Abstract The inner stratum corneum is likely to represent the location of the intact skin barrier, unperturbed by degradation processes. In our studies of the physical skin barrier a new high-performance liquid chromatography (HPLC)-based method was developed for the quantitative analysis of lipids of the inner stratum corneum. All main lipid classes were separated and quantitated by HPLC/light scattering detection (LSD) and the free fatty acid fraction was further analysed by gas-liquid chromatography (GLC). Mass spectrometry (MS) was used for peak identification and flame ionization detection (FID) for quantitation. Special attention was paid to the free fatty acid fraction since unsaturated free fatty acids may exert a key function in the regulation of the skin barrier properties by shifting the physical equilibrium of the multilamellar lipid bilayer system towards a noncrystalline state. Our results indicated that the endogenous free fatty acid fraction of the stratum corneum barrier lipids in essence exclusively consisted of saturated long-chain free fatty acids. This fraction was characterized as a very stable population (low interindividual peak variation) dominated by saturated lignoceric acid (C24:0, 39 molar%) and hexacosanoic acid (C26:0, 23 molar%). In addition, trace amounts of very long-chain (C32-C36) saturated and monounsaturated free fatty acids were detected in human forearm inner stratum corneum. Our analysis method gives highly accurate and precise quantitative information on the relative composition of all major lipid species present in the skin barrier. Such data will eventually permit skin barrier model systems to be created which will allow a more detailed analysis of the physical nature of the human skin barrier.

Stratum corneum swelling. Biophysical and computer assisted quantitative assessments.

Abstract The aim of this study was to characterize the swelling behaviour of the stratum corneum. Stratum corneum pieces isolated from the breast region of 20 different females were incubated in distilled water at two different temperatures (20° C and 45° C) for 90 min and 24 h, respectively. Half of the stratum corneum pieces were previously extracted with chloroformmethanol (2 : 1). The area-enlargement was photographically recorded. The thickness enlargement was determined using a confocal laser scanning microscope. The average swelling (99% confidence interval) in the area dimension at 20° C was 8.4% ± 1.4% ( n = 20), which corresponded to an average swelling in the length (lateral) dimension of approximately 4.1%. The swelling in the thickness dimension was 26.3% ± 16.3% ( n = 8). The swelling was most pronounced in the thickness dimension and was complete after 90 min of water immersion ( P < 0.01, n = 5). In addition, the removal of the intercellular lipids with chloroform/methanol (2 : 1) induced a decreased swelling in the samples ( P < 0.01, n = 20). An increase in temperature of the water from 20° C to 45° C resulted in an increase in swelling ( P < 0.01, n = 20). Taken together our results support the idea that the mechanism of stratum corneum swelling is linked to the intercellular lipid structure and hence to skin barrier function.

A novel approach to the understanding of human skin barrier function

The basis for externally caused skin disorders is penetration of the skin barrier. A recent model for the skin barrier, the domain mosaic model, based on current knowledge of the physics of lipid bilayer organization gave tentative explanations for several aspects of function. It is demonstrated here that a development of the model explains how the requirements are met for a water-tight structure that will still allow a controlled, minute loss of water, the perspiratio insensibilis, necessary for maintaining plasticity of the keratin. A major advantage of the extended model is that it allows an interpretation of the changes imposed on the structure when in contact with detergents and/or penetration enhancers.

Proton and Electron Microprobe Analysis of Human Skin

Abstract In order to demonstrate the feasibility of the proton microprobe in the analysis of dermatological material when a spatial resolution of a few micrometres is sufficient and to compare it with the electron microprobe technique, duplicate sections of human skin have been analysed with both methods. A skin sample was obtained from each of three healthy volunteers. After cryosectioning (12 μm) and freeze-drying adjacent sections of each sample were scanned by the electron microprobe and the proton microprobe, respectively.

Elemental Analysis Mirrors Epidermal Differentiation

Using a scanning nuclear microprobe, the distribution of elements and trace elements of skin cross sections of normal skin, non-lesional psoriatic skin and in dry atopic skin have been mapped. In non-lesional psoriatic skin and in dry atopic skin the epidermal Ca-gradient is higher than that of normal skin. In addition, abnormally high Fe and Zn levels were recorded in the stratum granulosum and corneum regions in the pathological skin. It is suggested that these findings correlate to an increased cell turnover in the basal cell layer of the psoriatic and atopic skins. The ratio of Ca/Zn in stratum corneum of paralesional psoriatic skin is approximately 8:1 compared to 12: 1 in normal skin and 15: 1 in atopic skin. This suggests that the differentiation process in paralesional psoriatic skin may actually be an example of disturbed programmed cell death.

On the Structure of the Normal Nail

SummaryA scanning electron microscopic (SEM) study of cut surfaces in normal human nails have confirmed the previous description of nail structure, i.e. the hard dorsal nail plate supported by the plastic intermediate nail plate.ZusammenfassungEine Raster-elektronenmikroskopische Untersuchung von Oberflächenschnitten von normalen menschlichen Nägeln konnte die frühere Beschreibung der Nagelstruktur bestätigen, d. h., daß die harte, dorsale Nagelplatte durch eine plastische intermediäre Nagelplatte abgestützt wird.

Applications in medicine using the new Lund microprobe

Abstract Using the Lund nuclear microprobe, application to medical research has been performed. It is known that people suffering from psoriasis have a skin composition that in some parts differs from normal skin. Using the microprobe, trace element profiles of the outer parts of the skin were obtained. From the results, e.g. the Fe distribution was found to be anomalous. In another study, calcification of arteries from patients affected by different degrees of atherosclerosis were compared with corresponding samples from normal individuals. Apart from Ca, which locally occurred in “granules”, the distributions of minor and trace elements were found to be different in the case of atherosclerosis.

The use of PIXE in experimental studies of the physiology of human skin epidermis.

In order to understand the normal and pathological physiologies of the epidermal cells, the simultaneous determination of several elements in the different cellular strata is of crucial importance. In recent years the electron microprobe (EMP) has become an established technique in this field. Its high spatial resolution, in principle, allows measurements of various cell organelles. However, the limited (intrinsic) sensitivity of the EMP represents a serious drawback to the technique. The introduction of the proton microprobe (PMP) has significantly improved the sensitivity, although the ultimate spatial resolution of the PMP is much less than that of the EMP.When studying the elemental profiles in skin epidermis, it is possible to use skin sections with a thickness of the order of 10 μm, then the spatial resolution of the PMP is equal to or better than that of the EMP since the electrons are scattered to a significant degree in the sample. The characteristics of the two methods have been compared by analysis of parallel duplicate freeze-dried sections of normal human skin. The distributions of the elements P, S, Cl, and K, obtained with the two techniques, were in good agreement. In addition, the PMP provided distributions of the important elements Ca, Fe, and Zn.In a recently started study, the useful features of the PMP will be used for studying how efficient a barrier the skin is to nickel and chromate ions. A preliminary experiment has been performed by exposing cadaverous skin, not older than 24-h postmortem, to solutions of the two ions. After an 18-h exposure, samples were prepared by shock-freezing and sectioning. The first results from PMP analysis of these samples demonstrate the presence of a nickel and chromium gradient in the outer strata in the epidermis (mainly stratum corneum).A third experiment deals with the physiology of psoriatic skin. Calcium is an important element in the differentiation. Hence, the higher sensitivity of the PMP has been used in analysis of sections from psoriatic skin epidermis. Preliminary results are presented.

ELECTROLYTES IN NAILS ANALYSED BY X-RAY MICROANALYSIS IN ELECTRON MICROSCOPY Considerations on a New Method for the Diagnosis of Cystic Fibrosis

ABSTRACT. Patients with cystic fibrosis (CF) have an increased concentration of sodium in their nails. Hitherto, only neutron activation analysis has been considered for the diagnosis of cystic fibrosis by analysis of electrolytes in nails. It has been thoroughly tested methodologically and clinically. However, the intrinsic advantages of X‐ray microanalysis and the results obtained in this study suggest that this method, after further testing, may be a useful diagnostic aid for cystic fibrosis. In comparison with neutron activation analysis, X‐ray micro‐analysis has the advantage of simultaneously giving the concentrations of several elements and may be accessible at any hospital with an electron microscope fitted with the necessary equipment. Nails of CF‐patients are here shown to have increased concentrations of Na, K and Cl, which will make the diagnosis of cystic fibrosis more reliable. The possibility of using sulphur as a reference element may eliminate the weighing procedure necessary in neutron activation analysis.

Electrical impedance and other physical parameters as related to lipid content of human stratum corneum.

Background/aims: In previous studies we have shown that variations in the properties of the stratum corneum are reflected by alterations in electrical impedance. The aim of this study was to explore the ability of the electrical impedance technique to detect changes in the lipid content of the stratum corneum, and to compare It with the other non‐invasive methods, measurement of transepidermal water loss and of skin moisture.