Biomedical subjects

J Kempenaar

Publications and source records attributed to J Kempenaar.

At least 19 recordsLinked to original sources

PubMed2004

Human adipose tissue-derived cells delay re-epithelialization in comparison with skin fibroblasts in organotypic skin culture.

BACKGROUND: Wound healing of deep and extensive burns can induce hypertrophic scar formation. During the early steps of wound healing fibroblasts migrate into the wounded area. Fibroblastic cells present in tissues other than dermis may also migrate into the wounded area and participate in the wound healing process. OBJECTIVES: To examine the influence of human fibroblastic cells derived from subcutaneous fat or dermis on epidermal morphogenesis in vitro. METHODS: We prepared human skin equivalents (HSEs) made of a collagen type I matrix populated either with dermal fibroblasts or adipose tissue-derived cells (ADCs), on top of which keratinocytes were seeded and subsequently grown at the air-liquid interface. RESULTS: A fully differentiated epidermis was formed on matrices populated with ADCs. However, the HSE formed differed in a number of features from HSE generated with dermal fibroblasts. The major differences included: marked contraction of the dermal matrix, low lateral migration of keratinocytes, high keratin 17 expression indicating increased keratinocyte activation, delayed deposition of collagen IV at the epidermal/matrix junction, accumulation of alpha-smooth muscle actin-positive cells only underneath the epidermal compartment and positioning of these cells in a direction parallel to the epidermal compartment. The latter two phenomena have also been found in scar tissue. CONCLUSIONS: The possibility of generating HSEs with different cell types represents an attractive approach for in vitro studies focusing on the mechanism of wound healing.

Actins↗

PubMed2000

Skin equivalent: an attractive model to evaluate early melanoma metastasis.

The growth patterns and morphological phenotype of four human melanoma cell lines with different metastatic potentials were investigated in submerged and in air-exposed (skin equivalent) keratinocyte-melanoma cell co-cultures. In contrast to the submerged co-cultures, all four cell lines formed sharply demarcated tumour cell nests within the epidermal compartment of the skin equivalent model, with the morphology highly mimicking the in vivo situation. Differences among the melanoma cell lines tested were observed with respect to the number of clusters formed and the ability to exhibit invasive growth. Only the two metastatic cell lines were able to invade the dermal compartment. Screening of cellular adhesion molecules revealed that the expression patterns in different cell lines were heterogeneous and remained unchanged during the whole culture period, irrespective of whether the melanoma cells were located in the epidermal or dermal compartment. A correlation was found between expression of a lower number of different cellular adhesion molecules and the ability to acquire invasive growth capability. Our results indicate that melanoma cells exhibit a heterogeneous growth behaviour when co-cultured with human keratinocytes, and the air-exposed skin equivalent model was shown to be suitable for studying differences in growth patterns and potential invasive behaviour.

Air↗

PubMed1998

Temperature-sensitive regulation of epidermal morphogenesis and the expression of cornified envelope precursors by EGF and TGF alpha.

Epidermis reconstructed on de-epidermized dermis was used to investigate the effects of growth factors and culture temperature on epidermal morphogenesis and the expression of cornified envelope precursors. Cultures grown at 33 degreesC or 37 degreesC in the absence or presence of transforming growth factor alpha (TGFalpha), keratinocyte growth factor (KGF), basic fibroblast growth factor (bFGF), or insulin-like growth factor (IGF) show a similar morphology to that of native epidermis. Loricrin and SPRR2 are expressed in the stratum granulosum and SPRR3 is absent. Cultures grown in epidermal growth factor (EGF)-supplemented medium at 37 degrees C have a normal morphology, whereas cultures grown at 33 degrees C have a disorganized basal layer, no stratum granulosum, and nuclei are present in the stratum corneum. Loricrin is absent, and SPRR2 and SPRR3 expression extend into the spinous layers. Irrespective of the culture condition used, involucrin is aberrantly expressed in all suprabasal layers. EGF stimulated keratinocyte proliferation and migration to a greater degree than TGFalpha. Epidermis reconstructed on fibroblast-populated collagen gels at 33 degrees C led to the same disturbances in keratinocyte differentiation as seen in cultures grown on de-epidermized dermis at 33 degrees C in the presence of EGF, whereas parallel cultures grown at 37 degrees C have a similar morphology to that of native epidermis.

Cell Division↗

PubMed1997

Epidermal growth factor and temperature regulate keratinocyte differentiation.

The limited life-span and irregularities in epidermal differentiation and barrier function that have restricted the utility of presently available skin culture models for pharmacological and toxicological studies indicate that further modifications of culture conditions are required for optimization of these models. In the present study epidermis reconstructed on de-epidermized dermis was used to investigate the effects of temperature and epidermal growth factor (EGF) on epidermal differentiation and lipogenesis. When cultured at 37 degrees C, keratinocytes formed a well-differentiated epidermis whether EGF was present or not. However, the thickness of the epidermis, particularly of the stratum corneum, was higher in the presence of EGF. Both the differentiation-specific protein markers (keratins 1 and 10, involucrin and transglutaminase) and lipid markers (ceramides) were synthesized. EGF-induced increases in triglyceride content caused accumulation of lipid droplets within the stratum corneum which is indicative of a hyperproliferative effect of EGF. In the absence of EGF, a well-differentiated epidermis was generated at 33 degrees C with a morphology showing a higher resemblance to native epidermis than cultures grown at 37 degrees C. The stratum corneum was less compact and with practically no lipid droplets, irregularly shaped keratohyalin granules were abundant in the stratum granulosum, lamellar body extrusion was improved and the number of stratum corneum layers was reduced to normal levels. However, EGF supplementation had a deleterious effect on epidermal morphogenesis and differentiation of cultures grown at 33 degrees C. The epidermis lacked a stratum granulosum and the stratum corneum contained a high number of nuclear remnants. The synthesis of the early specific protein differentiation markers (keratins 1 and 10) was suppressed on both the protein and mRNA levels without significant interference with the synthesis of late differentiation lipid markers, such as ceramides. From this observation it can be concluded that the synthesis of keratins associated with terminal differentiation is profoundly affected by the presence of EGF and is sensitive to temperature and that of ceramides is not. The finding that TGF alpha did not modulate the morphogenesis and synthesis of keratins 1 and 10 in cultures grown at 33 degrees C indicates possible differences between the postreceptor binding processes of these EGF receptor ligands.

Biomarkers↗

PubMed1997

Culture of reconstructed epidermis in a defined medium at 33 degrees C shows a delayed epidermal maturation, prolonged lifespan and improved stratum corneum.

In this study we compared human keratinocyte cultures grown at the air-liquid interface on de-epidermized dermis at 33 degrees C or at 37 degrees C in two different culture media: medium I--a fully defined serum- and EGF-free medium; and medium II-a serum- and EGF-containing medium. Cultures grown in medium II were initially hyperproliferative followed rapidly by senescence, and had a high triglyceride content. The hyperproliferation was ascribed to the presence of EGF in the medium. In contrast, cultures grown in medium I at 33 degrees C showed a greatly improved balance between cell proliferation and differentiation. They had a prolonged lifespan of at least 32 days without a significant decrease in the number of living cell layers, a rate of proliferation similar to that of native epidermis and a low triglyceride content. Culturing at 37 degrees C increased the rate of differentiation without affecting the rate of proliferation. Furthermore, both at 33 degrees C and at 37 degrees C, keratin 6 was expressed only in the first suprabasal layer but was expressed in all suprabasal layers in cultures grown in medium II. High keratin 6 expression was not directly linked to hyperproliferation but to deregulated terminal differentiation. Involucrin, transglutaminase and SPRR1 were abnormally expressed irrespective of the culture conditions used, whereas SKALP expression was decreased in cultures grown in medium I. The epidermal lipid profile was better in cultures grown in medium I; the relative amounts of ceramides, free fatty acids and cholesterol being comparable to native epidermis. Small-angle X-ray diffraction showed a slightly improved structural organization of stratum corneum lipids as demonstrated by the appearance of second- and third-order peaks of the 12-nm long phase and a marked reduction in the polycrystalline cholesterol peak.

Cell Culture Techniques↗

PubMed1997

The formation of competent barrier lipids in reconstructed human epidermis requires the presence of vitamin C.

Our analysis of epidermal lipids revealed that (glucosyl)ceramide profiles in various human skin equivalents are different from those of native tissue. The main difference is the reduced content in skin equivalents of ceramides 4-7 and especially the very low content of the most polar ceramides 6 and 7, which contain hydroxylated sphingoid base and/or fatty acid. To facilitate hydroxylation, the culture medium was supplemented with vitamins C and E. Although in vitamin E-supplemented medium lipogenesis was not affected, in vitamin C-supplemented medium the content of glucosylceramides and of ceramides 6 and 7 was markedly increased, both in the presence and absence of serum and irrespective the substrate used (inert or natural, populated or not with fibroblasts). The improvement of the lipid profile was accompanied by a marked improvement of the barrier formation as judged from extensive production of lamellar bodies, their complete extrusion at the stratum granulosum/stratum corneum interface, and the formation of multiple broad lipid lamellar structures in the intercorneocyte space. The presence of well-ordered lipid lamellar phases was confirmed by small-angle x-ray diffraction. Some differences between native and reconstructed epidermis, however, were noticed. Although the long-range lipid lamellar phase was present in both the native and the reconstructed epidermis, the short lamellar phase was present only in native tissue. It remains to be established whether these differences can be ascribed to small differences in relative amounts of individual ceramides, to differences in fatty acid profiles, or to differences in cholesterol sulfate, pH, or calcium gradients. The results indicate the key role vitamin C plays in the formation of stratum corneum barrier lipids.

Ascorbic Acid↗

PubMed1995

Triglyceride metabolism in human keratinocytes cultured at the air-liquid interface.

Although epidermis reconstructed in vitro histologically demonstrates the presence of fully differentiated tissue with cornified strata, it does not synthesize or release epidermal barrier lipids in the same proportions as does native skin, causing the barrier function to be impaired. Lipids, the content of which deviates the most, include triglycerides that are present in high amounts and stored as lipid droplets. Our recent studies have revealed that a high triglyceride content may be a reflection of a high synthetic rate and a low turnover. Therefore, the present study was undertaken to examine whether the triglyceride accumulation in the air-exposed cultures may be a result of insufficient supplementation of cells with oxygen, an excessive supplementation of cells with glucose, dysregulation of lipogenesis, or an impaired catabolism of triglycerides caused either by insufficient activity of triglyceride lipase and/or accumulation of free fatty acids due to insufficient activity of beta-oxidase. When keratinocytes were cultured at the air-liquid interface in medium containing a standard glucose concentration, both the lactate and triglyceride production was high. Lowering glucose content in the medium resulted in a decrease in both lactate production and triglyceride synthesis. However, even when grown at a low glucose concentration the triglyceride content remained higher than found in vivo and synthesized triglycerides were stored in the cells as a stable pool, suggesting that the catabolism of triglycerides was impaired. Since both lipase and beta-oxidase were found to be active in cultured keratinocytes, another factor or other factors are probably implicated in the regulation of triglyceride metabolism.

Carnitine↗

PubMed1995

Use of human skin recombinants as an in vitro model for testing the irritation potential of cutaneous irritants.

Two human skin recombinants, the epidermis reconstructed on the deepidermized dermis (RE-DED) or on fibroblast-populated collagen matrix (Living Skin Equivalent, LSE), were used to study the irritating effect of sodium lauryl sulfate (SLS). The extent of cytotoxicity induced after a 24-hour exposure period to increasing concentrations of SLS (0-5%) was evaluated on the basis of (1) morphological perturbations, (2) changes in the expression of differentiation-specific protein markers (keratin 1, 10, 6, 16, involucrin and transglutaminase), (3) cell membrane integrity (LDH leakage) and (4) release of proinflammatory mediators (PGE2, IL-1, IL-6 and IL-8). SLS induced significant changes in epidermal morphology and changes in the expression and localization of differentiation-specific protein markers when applied topically in concentrations higher than 1% on RE-DED and higher than 0.1% on LSE. The exposure of both human skin recombinants to SLS resulted in a dose-dependent release of LDH, PGE2 and IL-1 alpha and in an increase in keratinocyte intracellular IL-1 levels. Upon application of 5% SLS on RE-DED the total (intra- and extracellular) IL-1 levels remained high but due to cell damage the intracellular IL-1 level was markedly decreased and the extracellular IL-1 level increased. Similar observations have been made with LSE after application of 0.5% SLS. However, with LSE the extracellular IL-1 alpha levels were found to be about 100 times lower than those measured with RE-DED. Exposure of LSE to SLS induced a marked increase of IL-6 production in fibroblasts incorporated in the collagen matrix. Contrary to LSE, both intra- and extracellular levels of IL-6 were low in unexposed controls and were only marginally modulated by the exposure of the RE-DED to SLS. In addition, a dose-dependent increase in IL-8 release was observed upon application of SLS on RE-DED. The results of the present study indicate that concentrations of SLS required to induce epidermal irritancy in vitro approximate those inducing irritation in human skin in vivo. All parameters used in the present study for evaluation of toxicity can serve as useful endpoints for screening of contact skin irritancy in vitro. Compared to RE-DED, the LSE seems to be more susceptible to SLS. The differences in sensitivity between LSE and RE-DEd can be ascribed to reported differences in their stratum corneum barrier function.

Adult↗

PubMed1995

Characterization of stratum corneum structure in reconstructed epidermis by X-ray diffraction.

The intercellular lipid regions in the stratum corneum (SC), the outermost layer of the skin, form the major barrier for diffusion of substances through the skin. The barrier function of in vitro reconstructed epidermis is still impaired. With respect to further optimization of the model, it is necessary to characterize its stratum corneum lipid structure. In this study, small and wide angle X-ray diffraction were used to characterize the lipid organization in stratum corneum isolated from 14-day-old reconstructed epidermis. The measurements were carried out at room temperature, and subsequently as a function of temperature between 25 degrees C and 109 degrees C, followed by measurements after cooling to room temperature. The results of the X-ray diffraction measurements revealed the following in reconstructed epidermis. 1) The lamellar ordering of stratum corneum lipids was much lower than that observed in native stratum corneum. 2) Crystalline anhydrous cholesterol was present. 3) Orthorhombic packing was present, but the corresponding reflections were very weak. The orthorhombic packing disappeared between 30 degrees C and 45 degrees C. 4) A hexagonal packing was present and disappeared between 60 degrees C and 75 degrees C. 5) Soft keratin is present. 6) A higher extent of lamellar ordering could be achieved by heating to 109 degrees C and cooling down to room temperature. Analysis of SC lipids revealed the presence of high amounts of triglycerides, the level of which could be decreased by lowering the glucose content. However, modulation of culture medium composition did not significantly affect lipid lamellae structures or hydrocarbon chain packing.

Cells, Cultured↗

PubMed1994

Role of extracellular calcium in the regulation of 1,25-dihydroxyvitamin D3 formation in cultured human keratinocytes.

Cultured normal human keratinocytes (NHK) provide a useful experimental model for studies of processes occurring during terminal differentiation, since the extent of keratinocyte maturation can be manipulated experimentally by modulation of extracellular calcium concentration. When NHK are maintained in low calcium (0.06 mM) medium they proliferate but do not stratify. Raising the level of calcium to 1-2 mM results within a few hours in induction of keratinocyte differentiation. Results of the present study show that formation of 1,25-(OH)2D3 is higher in NHK grown at 0.06 mM than in NHK grown at 1.6 mM calcium concentration. After 2 h exposure of low calcium cultures to 1.6 mM calcium the 1,25-(OH)2D3 production starts to decrease. On the other hand, exposure of cells cultured in 1.6 mM calcium medium to 0.06 mM calcium concentration induced already within 4 h an increase in 1,25-(OH)2D3 formation which was not accompanied by a decrease in cornified envelope formation. Thereby, the present study demonstrated that calcium can regulate 1,25-(OH)2D3 formation independently of changes in keratinocyte differentiation.

Calcitriol↗

PubMed1994

The involvement of protein kinase C in proliferation and differentiation of human keratinocytes--an investigation using inhibitors of protein kinase C.

Protein kinase C, the major cellular receptor for tumour-promoting phorbol esters, has been suggested as playing a key role in the regulation of proliferation and differentiation of epidermal cells. In the present study, we investigated the effects of various well-characterized inhibitors of protein kinase C on proliferation and differentiation of SV 40-transformed and normal human keratinocytes. The drugs were found to inhibit cell proliferation in a dose-dependent manner, displaying similar effects in both cell types and reflecting their potencies in inhibiting purified protein kinase C. In contrast, keratinocyte differentiation induced by treatment with a calcium ionophore or spontaneously, i.e. by exposure of cells grown in the presence of low calcium concentration (0.06 mM) to normal calcium concentration (1.6 mM), was not inhibited by the compounds tested. The potent protein kinase C inhibitor, staurosporine, was found even to enhance cell differentiation. Therefore, the present study provides evidence that the classical protein kinase C pathway plays a critical role in the regulation of keratinocyte proliferation rather than in calcium-induced differentiation.

Alkaloids↗

PubMed1994

Fumaric acid derivatives evoke a transient increase in intracellular free calcium concentration and inhibit the proliferation of human keratinocytes.

Systemic administration of fumaric acid (FA) derivatives was originally an empirical antipsoriatic treatment, which showed promising clinical results. In the present study, FURA-2-loaded suspensions of cultured normal keratinocytes and SV40-transformed keratinocytes (SVK-14 cells) were used to study the effects of FA derivatives on the intracellular free calcium concentration ([Ca2+]i). Monomethylfumarate (MMF), dimethylfumarate (DMF) and monoethylfumarate (MEF) induced a rapid, transient [Ca2+]i increase in both cell types. This immediate increase reached maximal values of 396 nmol/l 10s after addition of MMF, and fell to basal values within 90-120 s (173 nmol/l for normal keratinocytes and 68 nmol/l for transformed keratinocytes). This increase was not affected by the prior addition of EGTA, indicating that FA derivatives released Ca2+ mainly from intracellular stores into the cytoplasm. Subsequently, dose-dependent inhibitory effects of FA derivatives on keratinocyte proliferation were demonstrated. The results of these experiments revealed that DMF was the most potent, MMF and MEF intermediate, and FA and malonic acid the least potent growth inhibitors. These antiproliferative effects of FA derivatives might be linked to the observed, transient [Ca2+]i elevations.

Anticarcinogenic Agents↗

PubMed1993

Co-culture of human melanocytes and keratinocytes in a skin equivalent model: effect of ultraviolet radiation.

Melanocytes grown in pure monolayer culture lack the three-dimensional organization and many of the cellular interactions that exist in vivo. This can be partially overcome by growing melanocytes together with other epidermal cells in skin equivalent models. In this study skin equivalents were prepared by seeding mixtures of cultured human keratinocytes and melanocytes in various ratios onto de-epidermized dermis. They were cultured in DMEM/Ham's F12 (3:1) for 3 days and then lifted to the air-liquid interface and maintained for 11 days. Histological examination revealed a structure that closely resembled human interfollicular epidermis. Melanocytes, identified by their dendritic appearance, positive dopa reaction and positive staining with a melanocyte-specific antibody (MEL5), were located in the basal layer. Melanin was seen both in melanocytes and in neighbouring keratinocytes. Whilst the skin equivalent became more pigmented following UV irradiation (total UVB 4760 J/m2 over 3 days), the quantity and distribution of melanin at the light microscopic level appeared to be unchanged. However, the number and dendricity of melanocytes increased, as did their staining with dopa and MEL5. These results indicate that melanocytes are functional and capable of responding to UV irradiation.

Cell Count↗

PubMed1990

Modulation of the production of a parathyroid hormone-like protein in human squamous carcinoma cell lines by interaction with fibroblasts.

Normal human keratinocytes as well as human squamous cell carcinomas produce a parathyroid hormone-like protein (PLP). However, PLP production by these cells is not a constant phenomenon. Since nothing is known about factors which regulate the production of PLP, in vitro studies were performed with normal keratinocytes and squamous carcinoma cell lines in order to establish conditions under which PLP production may vary. PLP was measured as cyclic AMP production in parathyroid hormone target cells (osteoblasts) which could be inhibited by a parathyroid hormone antagonist. The presence of PLP was confirmed using a radioimmunoassay specific for PLP. Results from the bioassay correlated very well with the data obtained by radioimmunoassay for PLP. The results confirm that human squamous carcinoma cells and normal keratinocytes produce PLP. PLP production appeared to be very sensitive to modulation of coculture of squamous carcinoma cells with fibroblasts. The effect of fibroblasts was not mediated by an effect on squamous carcinoma cell viability. Murine transformed fibroblasts (3T3 cells) as well as human normal foreskin fibroblasts were equally effective in inducing PLP production in these cells. The fibroblastic factor was apparently present in a soluble form in the coculture system which prevented direct cell-cell contact but allowed communication through the medium. Nevertheless, conditioned medium from 3T3 cells failed to induce PLP production by squamous carcinoma cells. This suggests a more complicated interaction between the two cell types than a one way message from fibroblasts to keratinocytes. Production of PLP by a number of squamous carcinoma cell lines was variable and not evidently correlated with the ability of these carcinoma cells to differentiate. Production of parathyroid hormone-like protein not only is the expression of a disturbed metabolism of a specific cell type but also reflects the cell-cell interaction in tumor tissue.

Animals↗

PubMed1990

A new sensitive assay for measurement of cell-mediated cytotoxicity to intact layers of cultured human keratinocytes.

A cytotoxicity assay for sensitive measurement of cell-mediated lympholysis (CML) of human cultured keratinocytes (cK) is described. The usage of 51Cr-labeled keratinocytes in intact layers as target cells in this assay allows objective and accurate determination of lysis of keratinocytes which have not undergone trypsin- and suspension-induced membrane changes. Furthermore, the problem of high spontaneous 51Cr release values encountered with suspended keratinocytes is overcome. The assay was applied to study antigen-specific CML of cK by cloned cytotoxic T cells (CTL) and to determine the effect of IFN-gamma on the susceptibility of cK to lysis. The results showed that HLA-A2 specific CTLs could reproducibly lyse cK of HLA-A2 positive healthy skin donors both with and without incubation of cK with IFN-gamma. Applications of this keratinocyte cytotoxicity assay lie in determining the antigenic expression of human cK, in analysis of effector cell/keratinocyte interactions in CML and of the modulatory effects of cytokines on these mechanisms. The assay thus may provide a helpful tool in gaining insight into the role of CML of keratinocytes in the destruction of inflamed skin.

Cell Adhesion↗

PubMed1990

Use of human keratinocyte and fibroblast cultures for toxicity studies of topically applied compounds.

Penetration enhancers are often used as additives in pharmaceutical and dermatological preparations. It should be expected that in many cases penetration enhancers not only enter the stratum corneum but also reach the viable cells of the epidermis and exert a toxic effect. This study focused on a series of well-known compounds that are often used as skin penetration enhancers; namely, ethanol, propylene glycol, dimethylsulfoxide, dimethylformamide, and Brij 96. In order to obtain more insight in the potential skin toxicity of these agents, they were administrated to cultured human keratinocytes and fibroblasts and the following cytotoxicity assays were performed: inhibition of the proliferation of fibroblasts and keratinocytes; inhibition of collagen contraction by fibroblasts; and cell morphology changes in confluent cultures of fibroblasts and keratinocytes. In all assays performed, the same trend was observed: ethanol was the least toxic, propylene glycol, dimethylsulfoxide, and dimethylformamide were moderately potent, and Brij 96 was the most toxic agent. An obvious advantage of the in vitro model presented here is its immediate availability and reproducibility, which allows for the comparison of a large series of topical agents (e.g., penetration enhancers) with respect to their cell toxicity under standardized conditions. However, this single-cell model lacks some of the properties found in intact skin, such as the stratum corneum barrier, and interactions between keratinocytes and other cells, such as Langerhans cells. Hence, extrapolation of these data to in vivo should be done with caution.

Adjuvants, Pharmaceutic↗

PubMed1990

Freeze-fracture electron microscopy of in vitro reconstructed human epidermis.

Epidermis has been reconstructed in vitro by seeding human keratinocytes on a human dermal substrate in an air-exposed culture. The end product has been examined by freeze-fracture electron microscopy, transmission electron microscopy (TEM) of thin sections, light microscopy, and lipid analysis using thin-layer chromatography. Light microscopic observation of hematoxylin-eosin stained, paraffin embedded cross-sections of the cell culture revealed a strong resemblance to its intact human counterpart, especially with respect to the morphologic organization in basal, spinous, granular, and horny layers. Freeze-fracture electron microscopy and TEM of thin sections generally confirmed the observed resemblances and additionally suggested the presence of lamellar bodies in the stratum granulosum, and of lamellar (lipid) structures between the corneocytes. However, some imperfections were also observed, including some anomalous lipid structures in the intercellular space. Lipid analyses in conjunction with essential fatty acid enrichment studies suggested that the structural anomalies observed in the cultured system may be caused by a lack of linoleyl-ceramides resulting from "immobilization" of linoleyl moieties in the form of triglycerides and phospholipids. In its present form, the air-exposed cell culture already looks very promising as a model for studies of, e.g., skin differentiation disorders such as psoriasis or ichthyosis, studies of the percutaneous penetration and intra(epi)dermal biotransformation of drugs, and skin toxicity screenings. It is furthermore expected that the aforementioned imperfections in the air-exposed cell culture should be avoidable by changing culture conditions such as the relative humidity and the pH, the composition of the medium, or both.

Cells, Cultured↗

PubMed1990

Nitroglycerin and sucrose permeability as quality markers for reconstructed human epidermis.

In order to evaluate the epidermal permeability barrier of in vitro reconstructed epidermis, the penetration of nitroglycerin (NG) and sucrose were measured across human keratinocytes cultured at the air-liquid interface, using de-epidermized dermis (DED) as a substrate. In the presence of reconstructed epidermis on top of DED the penetration rate of sucrose is about 100 times and that of NG 2 times lower, as compared to DED only, indicating that the stratum corneum of the cultured epidermis exhibits considerable barrier capacity. The permeability of reconstructed epidermis was for both solutes higher (3- to 10-fold) than that of freshly excised human skin. Based on the impaired barrier function and distribution of various differentiation markers it can be concluded that the reconstructed human epidermis used in the present study shows a high extent of similarity with hyperproliferating epidermis.

Air↗