Different expression of cyclooxygenase-2 (COX-2) in selected nonmelanocytic human cutaneous lesions

The aim of our study was to elucidate the possible involvement of COX-2 in the development and/or progression of nonmelanocytic skin lesions. To evaluate the usefulness of that enzyme as a potential molecular marker, we examined the intensity and spatial distribution of COX-2 expression in selected types of such tumors using the same immunohistochemical procedure as in our earlier studies of melanocytic cancers. We examined 20 benign epithelial lesions, 11 precancerous lesions, 21 basal cell carcinomas (BCC), 14 squamous cell carcinomas (SCC) and eight fibromas. The levels of COX-2 expression detected in benign lesions and in normal skin were comparable. Elevated expression of this protein may play a role in the development of SCC, as indicated by strong immunostaining both in SCCs and precancerous lesions. Significantly stronger staining in SCCs compared to BCCs may indicate a role of COX-2 in cancer malignancy and serve as an indicator useful for differential diagnostics of the two types of cancer. Strong staining in all skin layers of SCC may help in detecting cancer cells infiltrating surrounding skin layers.     

Differential expression of CD44s and CD4v10 proteins and syndecan in normal and irradiated mouse epidermis

 The role of the CD44s adhesion molecule, its epithelial isoforms and its relationship to epidermal proteoglycans such as syndecan was studied in normal and irradiated mouse skin. In normal mouse skin, only 10% of basal cells are strongly CD44s-immunopositive, with a cytoplasmic expression pattern. Double-label experiments with the basal cell marker keratin 14 confirmed the epithelial nature of the strongly CD44s-positive cell type in the basal layer. Some spinous keratinocytes and the majority of the remaining basal cells exhibited a weak membranous staining pattern. In contrast, the epithelial isoform, CD44v10, was strongly present in all basal and suprabasal epithelial cells of the epidermis, with a membranous staining pattern. Syndecan was found in the granular layer of the normal epidermis only. After 1 week of daily irradiation, the entire basal cell layer of the epidermis expressed CD44s in the membrane, but with a varying degree of staining intensity. This reactivity spread to the upper spinous layer after 3 weeks of treatment. In hyperproliferative epidermis, there was no difference in the staining patterns between CD44s and CD44v10. The expression of syndecan switched from the granular layer to the basal and lower spinous layers after 2 weeks of daily irradiation. Immunoreactivity for syndecan was also strongly enhanced in the dermis of irradiated samples. The results suggest an important role for syndecan and CD44 in proliferative processes during radiation-induced accelerated repopulation. Accepted: 30 September 1996     

Increased IKKα expression in the basal layer of the epidermis of transgenic mice enhances the malignant potential of skin tumors

Non-melanoma skin cancer is the most frequent type of cancer in humans. In this study we demonstrate that elevated IKKα expression in murine epidermis increases the malignancy potential of skin tumors. We describe the generation of transgenic mice overexpressing IKKα in the basal, proliferative layer of the epidermis and in the outer root sheath of hair follicles. The epidermis of K5-IKKα transgenic animals shows several alterations such as hyperproliferation, mislocalized expression of integrin-α6 and downregulation of the tumor suppressor maspin. Treatment of the back skin of mice with the mitogenic agent 12-O-tetradecanoylphorbol-13-acetate causes in transgenic mice the appearance of different preneoplastic changes such as epidermal atypia with loss of cell polarity and altered epidermal tissue architecture, while in wild type littermates this treatment only leads to the development of benign epidermal hyperplasia. Moreover, in skin carcinogenesis assays, transgenic mice carrying active Ha-ras (K5-IKKα-Tg.AC mice) develop invasive tumors, instead of the benign papillomas arising in wild type-Tg-AC mice also bearing an active Ha-ras. Therefore we provide evidence for a tumor promoter role of IKKα in skin cancer, similarly to what occurs in other neoplasias, including hepatocarcinomas and breast, prostate and colorectal cancer. The altered expression of cyclin D1, maspin and integrin-α6 in skin of transgenic mice provides, at least in part, the molecular bases for the increased malignant potential found in the K5-IKKα skin tumors.     

Human 8-oxoguanine-DNA glycosylase 1 protein and gene are expressed more abundantly in the superficial than basal layer of human epidermis

Human 8-oxoguanine-DNA glycosylase 1 (hOGG1) repairs 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxo-dG) which results from oxidation of guanine. Reactive oxygen species (ROS) formed in response to ultraviolet (UV) radiation cause this DNA damage, which is involved in pathological processes such as carcinogenesis and aging. The initiation of skin tumors probably requires penetration of UV to the actively dividing basal layer of the epidermis in order for acute damage to become fixed as mutations. Previously, the majority of UVB fingerprint mutations have been found in the upper layers of human skin tumors, while UVA mutations have been found mostly in the lower layer. Our aim was to determine whether this localization of UVA-induced DNA damage is related to stratification of the repair-enzyme hOGG1. Anti-hOGG1 immunohistochemical staining of frozen sections of human foreskin, adult buttock skin, and reconstructed human skin samples showed the highest expression of hOGG1 in the superficial epidermal layer (stratum granulosum). Study of the hOGG1 mRNA expression again showed the highest level in the upper region of the epidermis. This was not regulated by UV irradiation but by the differentiation state of keratinocytes as calcium-induced differentiation increased hOGG1 gene expression. UVA-induced 8-oxo-dG was repaired more rapidly in the upper layer of human skin compared to the lower layers. Our results indicate that weaker expression of the nuclear form of hOGG1 enzyme in the basal cells of the epidermis may lead to a lack of DNA repair in these cells and therefore accumulation of UVA-induced oxidative DNA mutations.     

Downregulation of CXCR-2 but not CXCR-1 expression by human keratinocytes by UVB

Interleukin-8 (IL-8) belongs to the CXC chemokine family. IL-8 exerts its biological activities by binding to specific cell surface receptors, CXCR-1 and CXCR-2. Both receptors bind IL-8 with high affinity but they have different affinities for MGSA/Groalpha and NAP-2. It has been shown that the expression of epidermal CXCR-2 is increased in psoriasis, suggesting that activation of KC mediated by CXCR-2 contributes to the characteristic epidermal changes observed in psoriasis. In order to examine the mechanism(s) by which UVB therapy is effective for several dermatoses including psoriasis, we sought to examine if UVB would modulate the expression of CXCR-1 and CXCR-2 in human keratinocytes (KC). Constitutive expression of CXCR-1 and CXCR-2 mRNA was detected by RT-PCR in normal cultured human KC. After 100 or 300 J/m(2) irradiation, a decrease in CXCR-2 mRNA was detectable from 12 h after irradiation; this downregulation was observed until 48 h after irradiation. In contrast, the CXCR-1 mRNA level was unchanged. Immunohistochemical studies and flow cytometry analysis confirmed the suppressive effect of UVB on the expression of CXCR-2 protein in cultured human keratinocytes. Immunohistochemical studies on two minimal erythema doses (2MED)-exposed and 2MED-unexposed skin from healthy volunteers revealed that CXCR-2 staining occurred over the whole layer of the epidermis but at 24 h after 2MED irradiation, the positive staining of CXCR-2 was decreased. A faint CXCR-1 staining was observed in the lower part of the epidermis both in unexposed and exposed skins. Our results indicate that UVB-induced growth inhibition of KC in hyperproliferative skin disorders may, in part, be related to downregulation of CXCR-2.     

Evaluation of expression of Toll-Like Receptors 7 and 9, proliferation,and cytoskeletal biomarkers in plaque and guttate psoriasis: A pilot morphological study

This pilot study was aimed at comparing TLR7/TLR9 expression, cytoskeletal arrangement, and cell proliferation by indirect immunofluorescence in parallel lesional and non lesional skin samples of guttate psoriasis (PG) and psoriasis vulgaris (PV) in five male patients for each group (n=10). TLR7 expression was detected throughout all the epidermal compartment in PV samples, while in PG skin was restricted to the granular layer. TLR9 was present in the granular layer of non lesional skin and in the suprabasal layers of PV/PG lesional skin. Cell proliferation was localized in all the epidermal layers in lesional PG and PV, consistently with the immunopositivity for the “psoriatic keratin” K16. In the suprabasal layers of lesional PG and PV skin, a similar K17 expression was detected and K10 exhibited a patchy distribution. The present results suggest that TLR7 expression can be considered an intrinsic and differential histomorphological feature of PV.Key words: Keratinocyte proliferation, adaptive immunity, keratins, immunofluorescence     

Increased matriptase zymogen activation in inflammatory skin disorders

Matriptase, a type 2 transmembrane serine protease, and its inhibitor hepatocyte growth factor activator inhibitor (HAI)-1 are required for normal epidermal barrier function, and matriptase activity is tightly regulated during this process. We therefore hypothesized that this protease system might be deregulated in skin disease. To test this, we examined the level and activation state of matriptase in examples of 23 human skin disorders. We first examined matriptase and HAI-1 protein distribution in normal epidermis. Matriptase was detected at high levels at cell-cell junctions in the basal layer and spinous layers but was present at minimal levels in the granular layer. HAI-1 was distributed in a similar pattern, except that high-level expression was retained in the granular layer. This pattern of expression was retained in most skin disorders. We next examined the distribution of activated matriptase. Although activated matriptase is not detected in normal epidermis, a dramatic increase is seen in keratinocytes at the site of inflammation in 16 different skin diseases. To gain further evidence that activation is associated with inflammatory stimuli, we challenged HaCaT cells with acidic pH or H(2)O(2) and observed matriptase activation. These findings suggest that inflammation-associated reactive oxygen species and tissue acidity may enhance matriptase activation in some skin diseases.     

Pep-1 as a novel probe for the in situ detection of hyaluronan.

Hyaluronan (HA) is expressed by most tissues, including skin. Localization of HA in the skin is assessed by histology with HA-binding protein (HABP) serving as the probe. Reports have suggested that HA expression in skin is altered in a number of diseases. However, interlaboratory variations in HABP staining profiles, even in normal skin, suggest a need to standardize methods and/or identify new probes. We report the staining patterns of a HA-binding peptide (termed "Pep-1") in human and mouse skin. After acetone fixation, Pep-1 stained HA in the intercellular spaces of the epidermis, whereas staining in the dermis was weak and diffuse in both human and mouse skin. HABP staining of the epidermis and dermis were comparable in human skin but failed to stain the vital epidermis of mouse skin. In human skin, Pep-1 stained the basal, spinous, and granular layers, whereas HABP failed to stain the basal layer. Precipitation of HA in situ resulted in dermal staining but weak staining in the epidermis for HABP and Pep-1. Our results may suggest that Pep-1 is sensitive to HA conformation. Furthermore, Pep-1 may represent a new probe to study HA expression in the skin.     

Fluorochrome-coupled lectins reveal distinct cellular domains in human epidermis

The distribution of saccharide moieties in human interfollicular epidermis was studied with fluorochrome-coupled lectins. In frozen sections Concanavalin A (Con A), Lens culinaris agglutinin (LCA), Ricinus communis agglutinin I (RCAI), and wheat germ agglutinin (WGA) stained intensively both dermis and viable epidermal cell layers, whereas peanut agglutinin (PNA) bound only to living epidermal cell layers. Ulex europaeus agglutinin I (UEAI) bound to dermal endothelial cells and upper cell layers of the epidermis but left the basal cell layer unstained. Dolichos biflorus agglutinin (DBA) bound only to basal epidermal cells, whereas both soybean agglutinin (SBA) and Helix pomatia agglutinin (HPA) showed strong binding to the spinous and granular cell layers. On routinely processed paraffin sections, a distinctly different staining pattern was seen with many lectins, and to reveal the binding of some lectins a pretreatment with protease was required. All keratin-positive cells in human epidermal cell suspensions, obtained with the suction blister method, bound PNA, whereas only a fraction of the keratinocytes bound either DBA or UEAI. Such a difference in lectin binding pattern was also seen in epidermal cell cultures both immediately after attachment and in organized cell colonies. This suggests that in addition to basal cells, more differentiated epidermal cells from the spinous cell layer are also able to adhere and spread in culture conditions. Gel electrophoretic analysis of the lectin-binding glycoproteins in detergent extracts of metabolically labeled primary keratinocyte cultures revealed that the lectins recognized both distinct and shared glycoproteins. A much different lectin binding pattern was seen in embryonic human skin: fetal epidermis did not show any binding of DBA, whereas UEAI showed diffuse binding to all cell layers but gave a bright staining of dermal endothelial cells. This was in contrast to staining results obtained with a monoclonal cytokeratin antibody, which showed the presence of a distinct basal cell layer in fetal epidermis also. The results indicate that expression of saccharide moieties in human epidermal keratinocytes is related to the stage of cellular differentiation, different cell layers expressing different terminal saccharide moieties. The results also suggest that the emergence of a mature cell surface glycoconjugate pattern in human epidermis is preceded by the acquisition of cell layer-specific, differential keratin expression.     

Expression and Functional Role of Sox9 in Human Epidermal Keratinocytes

In this study, we investigated the expression and putative role of Sox9 in epidermal keratinocyte. Immunohistochemical staining showed that Sox9 is predominantly expressed in the basal layer of normal human skin epidermis, and highly expressed in several skin diseases including psoriasis, basal cell carcinoma, keratoacanthoma and squamous cell carcinoma. In calcium-induced keratinocyte differentiation model, the expression of Sox9 was decreased in a time dependent manner. When Sox9 was overexpressed using a recombinant adenovirus, cell growth was enhanced, while the expression of differentiation-related genes such as loricrin and involucrin was markedly decreased. Similarly, when rat skin was intradermally injected with the adenovirus expressing Sox9, the epidermis was thickened with increase of PCNA positive cells, while the epidermal differentiation was decreased. Finally, UVB irradiation induced Sox9 expression in cultured human epidermal keratinocytes, and keratinocytes are protected from UVB-induced apoptosis by Sox9 overexpression. Together, these results suggest that Sox9 is an important regulator of epidermal keratinocytes with putative pro-proliferation and/or pro-survival functions, and may be related to several cutaneous diseases that are characterized by abnormal differentiation and hyperproliferation.     

Localization of prolactin receptor in the dorsal and ventral skin of the frog (Rana ridibunda)

The dorsal and ventral skin in amphibians plays an important role in osmoregulation. Prolactin hormone is involved in regulation of amphibian skin functions, such as water and electrolyte balance. Therefore, amphibians may be useful as a model for determining the sites of the prolactin receptor. In this study, prolactin receptor was detected in frog dorsal and ventral skin using immunohistochemical staining method. Prolactin receptor immunoreactivity was localized in all epidermal layers except stratum corneum of dorsal skin epidermis, stratum germinativum layer of ventral skin epidermis, myoepithelial cells, secretory epithelium and secretory channel cells of granular glands in both skin regions. The mucous glands and secretory granules of granular glands did not show immunoreactivity for the prolactin receptor. According to our immunohistochemical results, the more widespread detection of prolactin receptor in dorsal skin epidermis indicates that prolactin is more effective in dorsal skin. Presence of prolactin receptors in epidermis points out its possible osmoregulatory effect. Moreover, detection of receptor immunoreactivity in various elements of poison glands in the dermis of both dorsal and ventral skin regions suggests that prolactin has a regulatory effect in gland functions.     

Multiple gap junction genes are utilized during rat skin and hair development.

The expression of four different gap junction gene products (alpha 1, beta 1, beta 2, and beta 3) has been analysed during rat skin development and the hair growth cycle. Both alpha 1 (Cx43) and beta 2 (Cx26) connexins were coexpressed in the undifferentiated epidermis. A specific, developmentally regulated elimination of beta 2 expression was observed in the periderm at E16. Coinciding with the differentiation of the epidermis, differential expression of alpha 1 and beta 2 connexins was observed in the newly formed epidermal layers. alpha 1 connexin was expressed in the basal and spinous layers, while beta 2 was confined to the differentiated spinous and granular layers. Large gap junctions were present in the basal layer, while small gap junctions, associated with many desmosomes, were typical for the differentiated layers. Although the distribution pattern for alpha 1 and beta 2 expression remained the same in the neonatal and postnatal epidermis, the RNA and protein levels decreased markedly following birth. Hair follicle development was marked by expression of alpha 1 connexin in hair germs at E16. Following beta 2 detection at E20, the expression increased for both alpha 1 and beta 2 in developing follicles. A cell-type-specific expression was detected in the outer root sheath, in the matrix, in the matrix-derived cells (inner root sheath, cortex and medulla) and in the dermal papilla. In addition, alpha 1 was specifically expressed in the arrector pili muscle, while sebocytes expressed both alpha 1 and beta 3 (Cx31) connexin. beta 1 connexin (Cx32) was not detected at any stage analysed. The results indicate that multiple gap junction genes contribute to epidermal and follicular morphogenesis. Moreover, based on the utilization of gap junctions in all living cells of the surface epidermis, it appears that the epidermis may behave as a large communication compartment that may be coupled functionally to epidermal appendages (hair follicles and sebaceous glands) via gap junctional pathways.     

E6/E7 expression of human papillomavirus type 20 (HPV-20) and HPV-27 influences proliferation and differentiation of the skin in UV-irradiated SKH-hr1 transgenic mice

The functional role of UV irradiation, in combination with the E6 and E7 proteins of the cutaneous human papillomavirus (HPV) types in the malignant conversion of benign papillomatous lesions, has not been elucidated. Transgenic SKH-hr1 hairless mice expressing HPV-20 and HPV-27 E6 and E7 proteins in the suprabasal compartment were generated and exposed to chronic UV irradiation. Histological and immunohistochemical examination of skin samples revealed enhanced proliferation of the epidermal layers and papilloma formation in both transgenic strains in comparison to what was observed with nontransgenic mice. Squamous cell carcinoma developed in the HPV-20 E6/E7 transgenic line as well as in the HPV-27 E6/E7 transgenic line. Several weeks after cessation of UV-B exposure, enhanced proliferation, as measured by BrdU incorporation, was maintained only in HPV-20 transgenic skin. Keratin 6 expression was increased in the transgenic mice throughout all cell layers. Expression of the differentiation markers involucrin and loricrin was reduced and disturbed. p63alpha expression was differentially regulated with high levels of cytoplasmic expression in clusters of cells in the granular layer of the skin in the transgenic lines 20 weeks after cessation of UV-B exposure, in contrast to uninterrupted staining in the nontransgenic lines. p53 was expressed in clusters of cells in nontransgenic and HPV-27 transgenic mice, in contrast to an even distribution in a higher number of cells in HPV-20 transgenic animals.     

Accelerated proliferation and abnormal differentiation of epidermal keratinocytes in endo-beta-galactosidase C transgenic mice

Transgenic (TG) mice that have systemically expressed endo-beta-galactosidase C (EndoGalC) have rough and flaky skin. This skin phenotype is detectable around 5 days postnatal and becomes obscure by 2 weeks after birth. Their epidermis is thickened but the dermis and hair follicles are normal in structure. EndoGalC, which removes the terminal Galalpha1-3Gal disaccharide (alphaGal epitope), was expressed in the epidermis of TG mice. GS-IB4 lectin staining showed that the alphaGal epitope did not exist in the epidermis in TG but existed in wild-type (WT) mice. In TG mice, N-acetylglucosamines were exposed by EndoGalC, which is detected using GS-II lectin. To understand the cause of the epidermal thickening and skin phenotype, we examined the proliferation and differentiation of kerationocytes. BrdU-pulse-labeling revealed that proliferating keratinocytes increased approximately three-fold in TG epidermis compared to WT one. In TG epidermis, the expression domain of cytokeratin 14 increased from 1-2 layers to 4-5 layers and co-expressed with cytokeratin 6 and 10 in the upper layers. The layers expressing involucrin and loricrin also increased but those expressing filaggrin and transglutaminase looked normal. The localization of E-cadherin was similar in both TG and WT mice. Although TG mice showed delayed development of the barrier function around 8 days postnatal, they acquired the function by 12 days after birth. These results suggest that the absence of the alphaGal epitope or the exposed N-acetylglucosamine terminal could play a critical role in the proliferation of basal keratinocytes and differentiation of them into the spinous cells in newborn mice.     

Inhibition of p38 mitogen-activated protein kinase and phosphatidylinositol 3-kinase decreases UVB-induced activator protein-1 and cyclooxygenase-2 in a SKH-1 hairless mouse model

Activation of activator protein-1 (AP-1) and increased expression of cyclooxygenase-2 (COX-2) have been clearly shown to play a functional role in UVB-induced skin tumor promotion. In this study, we examined UVB-induced signal transduction pathways in SKH-1 mouse epidermis leading to increases in COX-2 expression and AP-1 activity. We observed rapid increases in p38 mitogen-activated protein kinase (MAPK) signaling through activation of p38 MAPK and its downstream target, MAPK activated protein kinase-2. UVB also increased phosphatidylinositol 3-kinase (PI3K) signaling as observed through increases in AKT and GSK-3beta phosphorylation. Activation of the p38 MAPK and PI3K pathways results in the phosphorylation of cyclic AMP-responsive element binding protein, which was also observed in UVB-irradiated SKH-1 mice. Topical treatment with SB202190 (a specific inhibitor of p38 MAPK) or LY294002 (a specific inhibitor of PI3K) significantly decreased UVB-induced AP-1 activation by 84% and 68%, respectively, as well as COX-2 expression. Our data show that in mouse epidermis, UVB activation of the p38 MAPK and PI3K pathways leads to AP-1 activation and COX-2 expression.     

Rice bran supplement prevents UVB-induced skin photoaging in vivo

Although rice bran consumption is reportedly has numerous beneficial effects on human health, the relationship between rice bran and the prevention of photoaging has not been investigated in detail. We sought to investigate whether consumption of rice bran supplement (RBS) can elicit preventive effects against UVB-induced photoaging in vivo. Dorsal skin sections of hairless mice were exposed to UVB over 16 weeks. RBS consumption suppressed UVB-induced wrinkle formation and inhibited the loss of water content and epidermal thickening in the mouse skin. Western blot and immunohistochemical analyses revealed that repeated exposure to UVB upregulated matrix metalloproteinase-13 (MMP-13) and cyclooxygenase-2 (COX-2) expression, while consumption of RBS suppressed MMP-13 and COX-2 expression, as well as mitogen-activated protein kinase (MAPK) signaling pathways. These findings suggest that RBS could be a potential bioactive ingredient in nutricosmetics to inhibit wrinkle formation and water content loss via the suppression of COX-2 and MMP-13 expression.     

The bovine desmocollin family: a new gene and expression patterns reflecting epithelial cell proliferation and differentiation

We have discovered a third bovine desmocollin gene, DSC3, and studied expression of all three desmocollin genes, DSC1, 2, and 3, by Northern blotting, RT-PCR and in situ hybridization. DSC1 is strongly expressed in epidermis and tongue papillae, showing a "skin"-type pattern resembling that previously described for keratins 1 and 10. Expression is absent from the epidermal basal layer but appears in the immediate suprabasal layers and continues uniformly to the lower granular layer. In tongue epithelium, expression is suprabasal and strictly localized to papillae, being absent from interpapillary regions. In other epithelial low level DSC1 expression is detectable only by RT-PCR. The distribution of Dsc1 glycoproteins, detected by an isoform-specific monoclonal antibody, closely reflects mRNA distribution in epidermis and tongue. DSC2 is ubiquitously expressed in epithelia and cardiac muscle. In stratified epithelia, expression appears immediately suprabasal, continuing weakly to the lower granular layer in epidermis and to just above half epithelial thickness in interpapillary tongue, oesophageal, and rumenal epithelia. DSC3 expression is restricted to the basal and immediately suprabasal layers in stratified epithelia. In deep rete ridges DSC expression strikingly resembles the distribution of stem, transit-amplifying, and terminally differentiating cells described by others. DSC3 expression is strongly basal, DSC2 is strong in 5-10 suprabasal layers, and then weakens to be superseded by strong DSC1. These results suggest that desmocollin isoform expression has important functional consequences in epithelial proliferation, stratification, and differentiation. The data also provide a standard for nomenclature of the desmocollins.     

Sodium-dependent vitamin C transporter isoforms in skin: Distribution, kinetics, and effect of UVB-induced oxidative stress

Two sodium-dependent vitamin C transporter isoforms (SVCT1 and SVCT2) were identified as ascorbic acid transporters, but their roles in skin have, as yet, not been elucidated. Here we analyze the expression and function of SVCTs in healthy human skin cells and skin tissues, and in UVB-induced cutaneous tissue injury. SVCT1 was primarily found in the epidermis expressed by keratinocytes, whereas SVCT2 expression was in the epidermis and dermis in keratinocytes, fibroblasts, and endothelial cells. Uptake experiments revealed that ascorbic acid affinity of SVCT1 was lower than SVCT2 (K(m)=75 muM and K(m)=44 muM, respectively), but maximal velocity was 9-times higher (36 nmol/min/well). In keratinocytes, SVCT1 was found to be responsible for vitamin C transport, although SVCT2 gene expression was higher. On UVB irradiation, SVCT1 mRNA expression in murine skin declined significantly in a time- and dose-dependent manner, whereas SVCT2 mRNA levels were unchanged. Furthermore, UVB irradiation of keratinocytes in vitro was accompanied by reduced ascorbic acid transport. In summary, these data indicate that the two vitamin C transporter isoforms fulfill specific functions in skin: SVCT1 is responsible for epidermal ascorbic acid supply, whereas SVCT2 mainly facilitates ascorbic acid transport in the dermal compartment. UVB-induced oxidative stress in mice resulted in depletion of SVCT1 mRNA levels and led to significantly decreased ascorbic acid uptake in keratinocytes, providing evidence on why ascorbic acid levels are decreased on UVB irradiation in vivo.