Stratum corneum lipids: the effect of ageing and the seasons

Stratum corneum lipids play a predominant role in maintaining the water barrier of the skin. In order to understand the biological variation in the levels and composition of ceramides, ceramide 1 subtypes, cholesterol and fatty acids, stratum corneum lipids collected from tape strippings from three body sites (face, hand, leg) of female Caucasians of different age groups were analysed. In addition, we studied the influence of seasonal variation on the lipid composition of stratum corneum from the same body sites. The main lipid species were quantified using high-performance thin-layer chromatography and individual fatty acids using gas chromatography. Our findings demonstrated significantly decreased levels of all major lipid species, in particular ceramides, with increasing age. Similarly, the stratum corneum lipid levels of all the body sites examined were dramatically depleted in winter compared with spring and summer. The relative levels of ceramide 1 linoleate were also depleted in winter and in aged skin whereas ceramide 1 oleate levels increased. The other fatty acid levels remained fairly constant with both season and age, apart from lignoceric and heptadecanoic acid which showed a decrease in winter compared with summer. The decrease in the mass levels of intercellular lipids and the altered ratios of fatty acids esterified to ceramide 1, are likely to contribute to the increased susceptibility of aged skin to perturbation of barrier function and xerosis, particularly during the winter months. Received: 17 October 1995     

Ceramide profile in hypohidrotic ectodermal dysplasia

Background. Hypohidrotic ectodermal dysplasia (HED) is a rare genetic disease. The clinical presentation includes lack of sweating ability, and an often widely spread dermatitis resembling atopic dermatitis (AD). In AD, the skin‐barrier defect is partly ascribed to the altered lipid profile in the stratum corneum and partly to mutations of the filaggrin genes. To our knowledge, no data are available about the epidermal lipid profile of HED. Aim. To compare the ceramide profile for patients with HED and AD. Methods. The ceramide profile and ceramide/cholesterol ratio were compared between patients with HED (n = 7) and patients with AD (n = 21), using cyanoacrylate to take biopsy samples from the stratum corneum. Lipids were extracted from the biopsies and analysed using high‐performance thin‐layer chromatography. Results. The lipid profiles of HED and AD were similar in distribution, apart from ceramide 1, which was significantly higher in HED (P = 0.04). Conclusions. The increased ceramide 1 level found in HED compared with AD is known to play a role in the structure of the lipid bilayers. However, further studies are needed to identify the functional significance of these observations and thereby elucidate differences in the skin barrier between HED and AD.     

Existence of a lipid gradient in the upper stratum corneum and its possible biological significance

The internal stratum corneum lipid composition was investigated in relation to depth in vivo in healthy human volunteers by extraction following one, three or five strippings. Automated multiple development high-performance thin-layer chromatography (AMD-HPTLC) and gas chromatography (GC) followed by normalized principal component analysis showed a decrease in the amount of lipids extracted after one, three and five strippings. Between levels 0, 1, 3 and 5 the stratum corneum lipid composition showed an increase in phospholipids and cholesterol-3-sulphate at level 3, a decrease in ceramide, cholesterol and free fatty acids after level 1, and a slight decrease in sterol esters at level 3. Lipids extracted after three strippings displayed a characteristic composition with an increase in the proportion of phospholipids and cholesterol-3-sulphate. Free fatty acid analysis in relation to depth revealed a decrease in the amounts of C14:0, C16:0, C16:1, C18:0 and C18:1 between levels 1 and 5 and an increase in the C24:0. A decrease in the unsaturated/ saturated chain ratio with depth was also observed, reflecting a greater decrease in unsaturated than saturated free fatty acids. A decrease in the ratios of free fatty acids to cholesterol and free fatty acids to ceramides after three and five strippings, respectively, and previously reported results, confirm the importance of this level of stratum corneum lipids in skin barrier properties. Received: 12 March 1996     

Comparison of lipids from epidermal and palatal stratum corneum.

Stratum corneum has been isolated by tryptic digestion of porcine epidermis and palatal epithelium, and the lipid concentrations and compositions have been compared by thin-layer chromatography in conjunction with photodensitometry. Palatal stratum corneum contained 47 +/- 6 micrograms lipid/mg tissue or 115 +/- 16 micrograms lipid per cm2 of stratum corneum surface, whereas epidermal stratum corneum contained 105 +/- 17 micrograms lipid/mg tissue or 135 +/- 16 micrograms/cm2. The difference in total lipid content does not account for the tenfold higher permeability constant for the permeation of water through the former tissue compared to the latter; therefore, the difference in permeability must be based on differences in lipid composition. In this regard, palatal stratum corneum includes 12.1% phospholipids, although phospholipids were undetected in epidermal stratum corneum. Differences in the content and location of non-polar liquid-phase lipids may also be of significance for permeability. Other factors that may contribute to the greater permeability of the palatal horny layer relative to epidermal stratum corneum include generally lower proportions of cholesterol, fatty acids, and ceramides, a dramatically lower proportion of the linoleate-containing acylceramide, and a tenfold lower content of covalently bound lipids associated with the corneocyte envelope.     

Microanalytical screening of all major stratum corneum lipids by sequential high-performance thin-layer chromatography

For rapid and sensitive screening of lipid biochemical abnormalities of scaling skin disorders a sequential, one-dimensional high-performance thin-layer chromatographic method (HPTLC) has been developed. All major human stratum corneum lipid classes, i.e., cholesterol sulfate, glucosylceramides, six major ceramide fractions, free sterols, free fatty acids, triglycerides, sterol esters, squalene, and n-alkanes, are separated and quantitated after a stepwise development of a single silica gel 60 HPTLC-plate using three consecutive solvent systems. Reproducible results have been obtained by degradative charring as well as fluorescence detection. By fluorescence detection the method is particularly suitable for the determination of minor amounts of cholesterol sulfate and other sterols.     

Barrier characteristics of different human skin types investigated with X-ray diffraction, lipid analysis, and electron microscopy imaging

The stratum corneum requires ceramides, cholesterol, and fatty acids to provide the cutaneous permeability barrier. The lipids are organized in intercellular membranes exhibiting short- and long-periodicity lamellar phases. In recent years, the phase behavior of barrier lipid mixtures has been studied in vitro. The relationship of human stratum corneum lipid composition to membrane organization in vivo, however, has not been clearly established. Furthermore, the special function of the different ceramide species in the stratum corneum is largely unknown. We examined lipid organization and composition of stratum corneum sheets from different subtypes of healthy human skin (normal, dry, and aged skin). Lipid organization was investigated using X-ray diffraction and demonstrated that the 4.4 nm peak attributed to the long periodicity phase was frequently missing for skin with a low Cer(EOS)/Cer(total) ratio, indicating an important part for Cer(EOS), which contains omega-hydroxy fatty acid (O) ester-linked to linoleic acid (E) and amide-linked to sphingosine (S). A deficiency in the 4. 4 nm peak was predominantly observed in young dry skin. In one case of aged skin, however, and less often in young normal skin this peak was also missing. Furthermore, the ceramide composition of samples without the 4.4 nm peak showed a deficiency of Cer(EOH), which contains 6-hydroxy-4-sphingenine (H), and an increase in Cer(NS) and Cer(AS), which contain nonhydroxy (N) or alpha-hydroxy fatty acids (A). In addition, a 3.4 nm peak attributed to crystalline cholesterol occurred in most cases of aged and dry skin, but was not observed in young normal skin. Our results do not indicate a definite pattern of correlation between lipid organization and types of human skin. They demonstrate, however, that Cer(EOS) and Cer(EOH) are key elements for the molecular organization of the long periodicity lamellar phase in the human stratum corneum.     

Effect of cyproterone acetate-ethinyl estradiol treatment on the proportions of linoleic and sebaleic acids in various skin surface lipid classes

Summary The effect of high-dose cyproterone acetateethinyl estradiol treatment on rates of sebum secretion and on the proportions of linoleic acid (1829,12) and sebaleic acid (1825,8) in the skin surface lipids of three female acne patients was examined. Changes in rates of sebum secretion were evaluated indirectly by measuring the ratio of wax esters/(cholesterol +cholesterol esters) in the subjects' skin surface lipid. In two of the subjects, this ratio indicated a reduction of sebum secretion rates to the childhood range. Concomitantly, there was an increase in linoleic acid and a decrease in sebaleic acid in all lipid classes. In the third subject, in whom there was only a small reduction in sebum secretion rate, the proportion of linoleic acid in the cholesterol esters more than doubled, but the changes in the other lipid classes were small or nonexistent. The results indicate that the proportions of linoleic acid and sebaleic acid in sebum are influenced by sebum secretion rates.     

Adverse effects of gasoline on the skin of exposed workers

Gasoline is widely used as a solvent in industry. To study its adverse effects on the skin and to understand their mechanisms, a matched epidemiological study (1:1, 52 exposed workers and 52 control subjects) was developed. Information about general conditions, history of dermatosis, changes in skin after exposure to gasoline, etc., was obtained. Ceramide, fatty acid and cholesterol collected from the backs of the hands were analyzed by high‐performance thin‐layer chromatography (HPTLC), because stratum corneum lipids play a predominant rôle in maintaining the physiological function of skin. The results showed that prevalences of hyperkeratosis, dryness, onychosis and dermatitis were clearly higher in exposed workers than in the control group, prevalence ratios being 3.33 (p<0.05), 3.00 (p<0.001), 11.25 (p<0.001), 5.00 (p<0.001), respectively. Fissures and onychorrhexis were the common symptoms in exposed workers. The stratum corneum lipid levels of ceramide, fatty acid and cholesterol were significantly lower in the exposed group than in the control group (p<0.05). Findings indicated that prolonged or repeated contact with gasoline could cause fissuring of the skin and nail disorders, and that the mechanism was perhaps depletion of stratum corneum lipids.     

New aspects of the skin barrier organization

In the superficial layer of the skin, the stratum corneum (SC), the lipids form two crystalline lamellar phases with periodicities of 6.4 and 13.4 nm (long-periodicity phase). The main lipid classes in SC are ceramides, free fatty acids and cholesterol. Studies with mixtures prepared with isolated ceramides revealed that cholesterol and ceramides are very important for the formation of the lamellar phases, and the presence of ceramide 1 is crucial for the formation of the long-periodicity phase. This observation and the broad-narrow-broad sequence of lipid layers in the 13.4-nm phase led us to propose a molecular model for this phase. This consists of one narrow central lipid layer with fluid domains on both sides of a broad layer with a crystalline structure. This model is referred to as 'the sandwich model'. While the presence of free fatty acids does not substantially affect the lipid lamellar organization, it is crucial for the formation of the orthorhombic sublattice, since the addition of free fatty acids to cholesterol/ceramide mixtures results in transition from a hexagonal to a crystalline lipid phase. Studies examining lipid organization in SC derived from dry or lamellar X-linked ichthyosis skin revealed that in native tissue the role of ceramide 1 and free fatty acids is similar to that observed with mixtures prepared with isolated SC lipids. From this we conclude that the results obtained with lipid mixtures can be used to predict the SC lipid organization in native tissue.     

Quantitative Analysis of Stratum Corneum Lipids in Xerosis and Asteatotic Eczema

Sphingolipids, a major constituent of intercellular lipids, are an important determinant for both water-holding and permeability barrier function in the stratum corneum. To assess the pathogenic role of sphingolipids in the stratum corneum of dry skin disorders such as xerosis and asteatotic eczema in leg skin, ceramides were quantified by thin layer chromatography after n-hexane/ethanol extraction of resin-stripped stratum corneum and evaluated as μg/mg stratum corneum. In healthy leg skin (n=49), there was age-related decline in the total ceramide, whereas xerosis (n=25) and asteatotic eczema (n=16) suffering significantly reduced water-holding properties, exhibited no definite decrease, rather slight increase in ceramide quantity with the same composition of each individual ceramide as compared to healthy age-matched controls. These data indicate that the seemingly elevated level of ceramide is an artificial effect due to inflammatory processes which result from susceptibility to dryness. Analysis of sebum-derived lipids present in the stratum corneum revealed that there was a significant decline in free fatty acids in xerosis and asteatotic eczema as compared to age-matched healthy controls, and a similar decline in triglycerides in the above three groups when compared to younger controls. Although the observed decrease in the stratum corneum lipids may well explain the high incidence of winter dry skin in older people, the progression toward asteatotic eczema can not be accompanied solely by a decrease in ceramide quantity, suggesting that the evolution of xerotic skin is associated with other moisturizing factors and/or environmental stimuli.     

特应性皮炎动物模型表皮脂合成的研究

目的:探讨特应性皮炎(AD)皮肤生理改变(包括表皮水分丢失量和角质层水分含量)是否与表皮脂的代谢有关。方法:在小鼠的背部和躯干外涂2,4-二硝基氟苯(DNFB),建立小鼠特应性皮炎模型,利用14[C]乙酸对AD模型表皮脂的代谢进行研究,并用电子显微镜对AD皮肤的超微结构进行观察。结果:AD表皮的胆固醇和脂肪酸的合成速度明显低于对照组,正常对照组的角质细胞间均为正常的复层板层膜结构。而皮炎组的深层角质细胞间虽可见正常的复层板层结构,但有许多没有加工完全的膜结构存在。结论:推测AD皮肤生理的异常改变可能是由于表皮脂的合成减少和角质细胞间膜异常所致。     

Validation of an in vivo extraction method for human stratum corneum ceramides

A topical acetone/diethylether (A/E) lipid extraction method was evaluated for its suitability for use in the study of stratum corneum lipids in various skin disorders. Its efficiency was compared in vitro with topical chloroform/methanol (C/M) extraction and with the classical integral C/M extraction (submerged tissue) of stratum corneum or whole epidermis. To estimate the depth of lipid removal by A/E extraction, light microscopic and freeze-fracture electron microscopic studies were carried out on A/E and C/M topically treated skin samples. The in vivo experiments consisted of topical A/E extraction and of classical C/M extraction of scrapings of the stratum corneum. Transepidermal water loss (TEWL) was measured before and after topical A/E extraction and after every scraping procedure, and correlated with TEWL values found after stripping of the stratum corneum. The total amount of lipid found with both topical extraction procedures was lower than that found with the integral extraction of the stratum corneum. Light microscopy showed that topical C/M extraction induced cell damage in the living epidermal cell layers. Great interindividual variation in overall lipid composition was shown in the in vitro experiments irrespective of the extraction protocol used. However, the ceramide (CER) profiles in a single skin sample from the same subject were similar irrespective of the protocol used, and a uniformity in the CER profiles was found in skin samples from different subjects. Similar results were obtained with in vivo topical A/E extractions: marked interindividual variation was seen in overall lipid composition, but not in the CER profile. Furthermore, the CER profiles found using the A/E extraction procedure both in vivo and in vitro were similar. The CER profiles were also found to be identical throughout the stratum corneum, as revealed by scraping experiments. Since the CER profiles are though to play a major role in the stratum corneum barrier function, the non-invasive A/E extraction of epidermal lipids seems to be suitable for clinical application.     

Distinct barrier integrity phenotypes in filaggrin-related atopic eczema following sequential tape stripping and lipid profiling

Background Filaggrin gene (FLG) loss‐of‐function mutations have been shown to represent the strongest so far known genetic risk factor for atopic dermatitis (AD). Whereas the barrier characteristics in FLG mutation carriers under baseline conditions have been investigated, there are only limited data on the permeability barrier function in filaggrin‐AD under compromised conditions. Aim We investigated: (i) stratum corneum (SC) integrity/cohesion; (ii) barrier recovery after controlled mechanical and irritant‐induced barrier abrogation; and (iii) the lipid composition of the non‐lesional and lesional skin of AD patients harbouring the European R501X, 2282del4, 3702delG, R2447X or S3247X FLG variants. Methods Thirty‐seven AD patients (14 FLG mutation carriers and 23 non‐carriers) and 20 healthy controls participated in the study. Stratum corneum integrity/cohesion was assessed by measurement of transepidermal water loss (TEWL) and amount of removed protein following sequential tape stripping. Barrier recovery was monitored by repeated measurements of TEWL and erythema up to 96 h after barrier abrogation. Samples for lipid analysis were obtained from non‐lesional and lesional skin using the cyanoacrylate method. Results Tape stripping revealed distinct genotype‐related impairment of the SC integrity/cohesion. No differences in the rate of barrier recovery among the groups were found. The SC lipid analysis revealed significant differences regarding the percentage amount of cholesterol, ceramide/cholesterol ratio and triglycerides in the uninvolved skin as well as the amounts of free fatty acids, CER[EOH] and triglycerides in the skin lesions of the AD FLG mutation carriers. Conclusions Our results provide evidence for discernible FLG‐related barrier integrity phenotypes in atopic eczema.     

New acylceramide in native and reconstructed epidermis

Culturing of normal human keratinocytes at the air-liquid interface results in the formation of fully differentiated epidermis under in vitro conditions. Although the reconstructed epidermis shows a close resemblance to native tissue, there are still some differences in the stratum corneum lipid profile and intercellular lipid organization. As ceramides belong to one of the major stratum corneum lipid classes, the aim of this study was to characterize this fraction in more detail. For this purpose, individual ceramide fractions were isolated by column chromatography and characterized by a combination of nuclear magnetic resonance spectroscopy, high-performance thin-layer chromatography, and gas chromatography. The results of this study show that in both the native and reconstructed human epidermis the extractable ceramide fraction contains, in addition to the well known acylceramides (EOS, EOH), a new acylceramide in which the omega-O-acylhydroxyacid is amide-linked to phytosphingosine (EOP). The same three sphingoid base moieties (S, P, H) are also found in ceramides with amide-linked nonhydroxy and alpha-hydroxyacids. Whereas the same types of ceramides were present in both tissues, some differences in their fatty acid profiles have been found. In reconstructed epidermis the content of linoleic acid in all three acylceramides fraction was significantly lower; the ceramide(NS) fraction was enriched in short fatty acids and the ceramide(AS) fraction was enriched in long chain alpha-hydroxyacids. These differences together with a lower content of free fatty acids may explain the differences between native and reconstructed tissue in stratum corneum lipid organization observed earlier by X-ray diffraction.     

Free fatty acids and sterols in human eccrine sweat

The lipid content of human sweat was determined in thermally induced sweat collected over a Vaseline or silicone barrier placed on the skin (clean sweat) and in sweat scraped from the skin surface without a barrier coating (scraped sweat). Lipids were extracted from concentrated sweat samples into chloroform:methanol and estimated by thin-layer chromatography in conjunction with photodensitometry. Scraped sweat contained 4 to 10 times more lipid than clean sweat and included cholesterol sulphate and ceramides resembling those found in the stratum corneum. In contrast, clean sweat contained only small amounts of free fatty acids and sterol. A marked individual and daily variation in sweat lipid content was also noted. The study indicates the importance of avoiding epidermal contamination when collecting sweat and the usefulness of our sweat collection method.     

Palmitoleic acid isomer (C16:1delta6) in human skin sebum is effective against gram-positive bacteria

The percent lipid composition of pooled human sebum analyzed by thin-layer chromatography was: ceramides (13%), fatty acid (47%), cholesterol (7%), cholesterol esters (2%), squalene (11%), triglycerides (3%), and wax esters (17%). Total sebum lipids (2- 4 mg/ml), sonicated into bacterial culture medium, caused 4- to 5-fold log reduction in growth of gram-positive bacteria, Staphylococcus aureus, Streptococcus salivarius and the anaerobe Fusobacterium nucleatum, but was ineffective against most gram-negative bacteria. Fractionation of the sebum lipids showed that both saturated and unsaturated fatty acids contained the bulk of the antimicrobial activity. Lauric acid (C12:0) was the most active saturated fatty acid. The unsaturated fatty acid, palmitoleic acid (C16:1delta6, cPA) was both the most predominant monoene and the most active antimicrobial fatty acid. Purified cPA (>99%) yielded typical minimal inhibitory concentration (MIC) values of 10-20 microg/ml against gram-positive bacteria. Organically synthesized cPA isomer gave MIC values comparable to the natural material. Both natural and synthetic cPA were found to be the most active sebum lipid fraction in blocking the adherence of a pathogenic strain of Candida albicans to porcine stratum corneum. Ethanol in combination with cPA exerts a synergistic bactericidal activity against gram-negative pathogenic bacteria, including Pseudomonas aeruginosa, Propionibacterium acnes, Escherichia coli, and several methacillin-resistant strains of S. aureus. Palmitoleic acid may be useful in topical formulations for treatment of secondary gram-positive bacterial infections, as a gram-positive bacteria antimicrobial in wound dressings, and as a natural gram-positive antimicrobial preservative in skin and hair care products.     

Human epidermal glucosylceramides are major precursors of stratum corneum ceramides

Ceramides are the major component of the stratum corneum, accounting for 30%-40% of stratum corneum lipids by weight, and are composed of at least seven molecular groups (designated ceramides 1-7). Stratum corneum ceramides, together with cholesterol and fatty acids, form extracellular lamellae that are responsible for the epidermal permeability barrier. Previous studies indicated that beta-glucocerebrosidase- and sphingomyelinase-dependent ceramide production from glucosylceramides and sphingomyelins, respectively, is important for epidermal permeability barrier homeostasis. A recent study indicated that sphingomyelins are precursors of two stratum corneum ceramide molecular groups (ceramides 2 and 5). In this study, we have examined the role of glucosylceramides in the generation of each of the seven stratum corneum ceramide molecular groups. First, the structures of various glucosylceramide species in human epidermis were determined by gas chromatography-mass spectrometry, fast atom bombardment-mass spectrometry, and nuclear magnetic resonance. The results indicate that total epidermal glucosylceramides are composed of six distinct molecular groups, glucosylceramides 1-6. Glucosylceramide 1 contains sphingenine and nonhydroxy fatty acids, glucosylceramide 2, phytosphingosine and nonhydroxy fatty acids, glucosylceramide 3, phytosphingosine with one double bond and nonhydroxy fatty acids, glucosylceramide 4, sphingenine and alpha-hydroxy fatty acids, glucosylceramide 5, phytosphingosine and alpha-hydroxy fatty acids, and glucosylceramide 6, phytosphingosine with one double bond and alpha-hydroxy fatty acids. The nonhydroxy fatty acids typically have 16-24-carbon-length chains, whereas alpha-hydroxy fatty acids are limited to 24-, 25-, and 26-carbon chains. The sphingosine bases are C18 or C20 chains. Next, acylglucosylceramides and glucosylceramides were treated with beta-glucocerebrosidase and the ceramides released were compared with stratum corneum ceramides. Ceramide moieties of acylglucosylceramides and glucosylceramides 1, 2, 4-6 correspond to stratum corneum ceramides 1-7. These results, together with those of our previous reports characterizing epidermal sphingomyelins, indicate that all ceramide species, including omega-hydroxy fatty-acid-containing ceramides, are derived from glucosylceramides, and fractions of ceramides 2 and 5 are from sphingomyelins. Furthermore, structural analysis of glucosylceramides revealed that human epidermal glycosphingolipids display a unique lipid profile that is rich in very long chain hydroxylated (alpha- and omega-hydroxy) fatty acids and phytosphingosine.     

Decreased level of ceramides in stratum corneum of atopic dermatitis: an etiologic factor in atopic dry skin?

Stratum corneum lipids are an important determinant for both water-retention function and permeability-barrier function in the stratum corneum. However, their major constituent, ceramides, have not been analyzed in detail in skin diseases such as atopic dermatitis that show defective water-retention and permeability-barrier function. In an attempt to assess the quantity of ceramides per unit mass of the stratum corneum in atopic dermatitis, stratum corneum sheet was removed from the forearm skin by stripping with cyanoacrylate resin and placed in hexane/ethanol extraction to yield stratum corneum lipids. The stratum corneum was dispersed by solubilization of cyanoacrylate resin with dimethylformamide, and after membrane filtration, the weight of the stratum corneum mass was measured. The ceramides were quantified by thin-layer chromatography and evaluated as microgram/mg stratum corneum. In the forearm skin of healthy individuals (n = 65), the total ceramide content significantly declined with increasing age. In atopic dermatitis (n = 32-35), there was a marked reduction in the amount of ceramides in the lesional forearm skin compared with those of healthy individuals of the same age. Interestingly, the non-lesional skin also exhibited a similar and significant decrease of ceramides. Among six ceramide fractions, ceramide 1 was most significantly reduced in both lesional and non-lesional skin. These findings suggest that an insufficiency of ceramides in the stratum corneum is an etiologic factor in atopic dry skin.     

Selective recovery of deranged water-holding properties by stratum corneum lipids

Selective removal of stratum corneum lipids following applications of acetone/ether to the human forearm for extremely prolonged periods of 5-20 min induced an enduring (more than 4 days), chapped and scaly appearance of the skin which was accompanied by a significant decrease in the water-holding properties of the stratum corneum. In order to further elucidate the significance of lipids in the water-holding properties, lipids, which were extracted as sebaceous-rich lipids (SL) for the first 10-min acetone/ether treatment and as stratum corneum lipids (SCL) for the additional 30-min treatment, were topically applied daily on lipid-depleted forearm skin which had been pretreated with acetone/ether for 40 min. Two daily applications of the SCL which were solubilized in squalane containing 1% alpha-monomethyl heptadecyl glyceryl ether (GE) caused a significant increase of conductance, accompanied by a marked improvement in the level of scaling as compared with nontreatment or GE/squalane base, whereas the SL in the GE/squalane base did not exhibit any significant recovery in either conductance value or scaling. To clarify which components of the SCL are primarily responsible for the observed recovery of the water-holding properties, chromatographically separated fractions of the SCL were also topically applied in the same manner for 2 successive days. Out of the following separated fractions: cholesterol, cholesterol ester, free fatty acid, glycolipids, and ceramide, 2 daily topical applications of ceramide fraction induced a significant and the highest increase in the conductance value as compared with GE/squalane base. Furthermore, glycolipids and cholesterol fractions also exhibited a significant recovery when compared with no application at all. In contrast, free fatty acid and cholesterol ester fractions did not indicate any significant increase in the conductance value. These findings strengthen the hypothesis that structural lipids present in the intercellular spaces of the stratum corneum, especially ceramide, play a critical role in the water-holding properties of the stratum corneum.     

Aberrant lipid organization in stratum corneum of patients with atopic dermatitis and lamellar ichthyosis

There are several skin diseases in which the lipid composition in the intercellular matrix of the stratum corneum is different from that of healthy human skin. It has been shown that patients suffering from atopic dermatitis have a reduced ceramide content in the stratum corneum, whereas in the stratum corneum of lamellar ichthyosis patients, the amount of free fatty acids is decreased and the ceramide profile is altered. Both patient groups also show elevated levels of transepidermal water loss indicative of an impaired barrier function. As ceramides and free fatty acids are essential for a proper barrier function, we hypothesized that changes in the composition of these lipids would be reflected in the lipid organization in stratum corneum of atopic dermatitis and lamellar ichthyosis patients. We investigated the lateral lipid packing using electron diffraction and the lamellar organization using freeze fracture electron microscopy. In atopic dermatitis stratum corneum, we found that, in comparison with healthy stratum corneum, the presence of the hexagonal lattice (gel phase) is increased with respect to the orthorhombic packing (crystalline phase). In lamellar ichthyosis stratum corneum, the hexagonal packing was predominantly present, whereas the orthorhombic packing was observed only occasionally. This is in good agreement with studies on stratum corneum lipid models that show that the presence of long-chain free fatty acids is involved in the formation of the orthorhombic packing. The results of this study also suggest that the ceramide composition is important for the lateral lipid packing. Finally, using freeze fracture electron microscopy, changes in the lamellar organization in stratum corneum of both patient groups could be observed.