Melanin accumulation in acanthotic seborrheic keratosis: Reduced proliferation and early differentiation of keratinocytes and increased number of melanocytes.

Seborrheic keratosis (SK) is a common benign tumour, often associated with hyperpigmentation. To investigate the mechanism of melanin accumulation in SK, we have conducted comprehensive gene expression and histological analyses. We obtained five pairs of skin samples, including non-lesional and SK samples, from the backs of three male Japanese participants aged 40-59 years. To examine melanocytes and keratinocytes in SK, three pairs of skin samples were separated by laser capture microdissection into the basal layer and the other layer in the epidermis. We performed a comprehensive gene expression analysis to identify differentially expressed genes between non-lesional and SK skin, followed by gene ontology and pathway analysis. We found abnormal morphogenesis and cell proliferation in the basal layer, along with increased immune response and impaired cell differentiation and metabolism in the other layer of SK. We focused on cell proliferation and differentiation, as these are directly associated with melanin accumulation. Immunohistochemical analyses of Ki67, keratin 10, and keratin 14 demonstrated the decreases in the proliferation and early differentiation of the epidermis. Contrarily, no significant changes were observed in terminal differentiation markers, filaggrin and loricrin. Although the number of melanocytes was higher in SK than in non-lesional skin, melanogenic activity showed no difference. These results indicated that melanin accumulation in SK is caused by delayed melanin excretion due to reduced turnover around the basal and spinous layers of the epidermis and melanin production due to an increased number of melanocytes. Our findings provide new insights for therapeutic approaches in SK.

Evaluation of subclinical chronic sun damage in the skin via the detection of long-lasting ultraweak photon emission.

BACKGROUND: It is well known that solar radiation accelerates skin photoaging. To evaluate subclinical photodamage in the skin especially from the early phase of ultraviolet (UV)-induced damage, we have focused on ultraweak photon emission (UPE), also called biophotons. Our previous study reported that the amount of long-lasting UPE induced by UV, predominantly from lipid peroxidation, is a valuable indicator to assess cutaneous photodamage even at a suberythemal dose, although it was only applied to evaluate acute UV damage. The aim of this study was to further investigate whether long-lasting UPE could also be a useful marker to assess subclinical chronic sun damage in the course of skin photoaging. MATERIALS AND METHODS: Forty-three Japanese females in their 20s were recruited and were divided into two groups according to their history of sun exposure based on a questionnaire (high- and low-sun-exposure groups). Several skin properties on the cheek and outer forearm were measured in addition to UV-induced UPE. RESULTS: Among the skin properties measured, water content, average skin roughness, and the lateral packing of lipids in the stratum corneum were significantly deteriorated in the high-sun-exposure group as were changes in some skin photoaging scores such as pigmented spots and wrinkles. In addition, those skin properties were correlated with the UPE signals, suggesting the possible impact of oxidative stress on chronic skin damage. CONCLUSION: Subtle oxidative stress detected by long-lasting UPE may contribute to subclinical cutaneous damage at the beginning phase of chronic sun exposure, which potentially enhances skin photoaging over a lifetime.

Impaired Tight Junctions in Atopic Dermatitis Skin and in a Skin-Equivalent Model Treated with Interleukin-17.

Tight junction (TJ) dysfunction in the stratum granulosum leads to aberrant barrier function of the stratum corneum (SC) in the epidermis. However, it is unclear whether TJs are perturbed in atopic dermatitis (AD), a representative aberrant SC-related skin disease, and whether some factors related to AD pathogenesis induce TJ dysfunction. To address these issues, we investigated the alterations of TJs in AD skin and the effects of Th2 and Th17 cytokines on TJs in a skin-equivalent model. The levels of TJ proteins were determined in the epidermis of nonlesional and lesional skin sites of AD. Western blot and immunohistochemical analyses revealed that the levels of zonula occludens 1 were decreased in the nonlesional sites of AD, and the levels of zonula occludens 1 and claudin-1 were decreased in the lesional sites relative to the levels in skin from healthy subjects. Next, we examined the effects of interleukin (IL)-4, tumor necrosis factor-α, IL-17, and IL-22 on the TJ barrier in a skin-equivalent model. Only IL-17 impaired the TJ barrier. Furthermore, we observed a defect in filaggrin monomer degradation in the IL-17-treated skin model. Thus, TJs are dysfunctional in AD, at least partly, due to the effect of IL-17, which may result in an aberrant SC barrier.

N-Terminal signal sequence is required for cellular trafficking and hyaluronan-depolymerization of KIAA1199.

Recently, we disclosed that KIAA1199-mediated hyaluronan (HA) depolymerization requires an acidic cellular microenvironment (e.g. clathrin-coated vesicles or early endosomes), but no information about the structural basis underlying the cellular targeting and functional modification of KIAA1199 was available. Here, we show that the cleavage of N-terminal 30 amino acids occurs in functionally matured KIAA1199, and the deletion of the N-terminal portion results in altered intracellular trafficking of the molecule and loss of cellular HA depolymerization. These results suggest that the N-terminal portion of KIAA1199 functions as a cleavable signal sequence required for proper KIAA1199 translocation and KIAA1199-mediated HA depolymerization.

KIAA1199, a deafness gene of unknown function, is a new hyaluronan binding protein involved in hyaluronan depolymerization.

Hyaluronan (HA) has an extraordinarily high turnover in physiological tissues, and HA degradation is accelerated in inflammatory and neoplastic diseases. CD44 (a cell surface receptor) and two hyaluronidases (HYAL1 and HYAL2) are thought to be responsible for HA binding and degradation; however, the role of these molecules in HA catabolism remains controversial. Here we show that KIAA1199, a deafness gene of unknown function, plays a central role in HA binding and depolymerization that is independent of CD44 and HYAL enzymes. The specific binding of KIAA1199 to HA was demonstrated in glycosaminoglycan-binding assays. We found that knockdown of KIAA1199 abolished HA degradation by human skin fibroblasts and that transfection of KIAA1199 cDNA into cells conferred the ability to catabolize HA in an endo-ß-N-acetylglucosaminidase-dependent manner via the clathrin-coated pit pathway. Enhanced degradation of HA in synovial fibroblasts from patients with osteoarthritis or rheumatoid arthritis was correlated with increased levels of KIAA1199 expression and was abrogated by knockdown of KIAA1199. The level of KIAA1199 expression in uninflamed synovium was less than in osteoarthritic or rheumatoid synovium. These data suggest that KIAA1199 is a unique hyaladherin with a key role in HA catabolism in the dermis of the skin and arthritic synovium.

Changes in epidermal hyaluronan metabolism following UVB irradiation.

BACKGROUND: Hyaluronan (HA) plays a role in keratinocyte proliferation and differentiation, and have shown different biological activities depending on its molecular mass. While many studies have shown changes in the amount of HA after UVB irradiation, molecular mass change remains to be elucidated. OBJECTIVE: To investigate the change in the molecular mass of HA after UVB irradiation in mouse epidermis. METHODS: The mice were irradiated with a single dose of UVB (0.15J/cm(2)). The amount of HA was examined using HABP sandwich assay. The molecular mass distribution was estimated by Sephacryl S-1000 chromatography. Has and Hyal mRNA expressions were detected by real-time PCR. RESULTS: On day 2 after UVB irradiation, both the amount of HA and the up-regulation of Has3 mRNA expression reached their maximum. The average HA molecular mass was about 1000 kDa, a level similar to that of the non-irradiated epidermis. On day 3, the average HA molecular mass drastically decreased to 100 kDa, while Hyal1, Hyal2, and Hyal3 mRNA expressions slightly increased. The amount of HA, however, remained high. On days 4 and 5, the amount of HA gradually decreased, but the molecular mass of HA remained low. A drastic reduction of the HA molecular mass after UVB irradiation was confirmed. CONCLUSION: UVB irradiation elicits remarkable changes in the molecular mass of HA, as well as amount. These qualitative and quantitative changes of HA might play an important role in UVB-induced cell proliferation and differentiation. Further study will be required to resolve the mechanism of HA degradation in the epidermis.

Novel effects of diosgenin on skin aging.

Extracts of Dioscorea coomposita or Dioscorea villosa are consumed as supplemental health foods at the time of climacteric. The extracts contain large amounts of the plant steroid, diosgenin. Here, we studied the safety and efficacy of diosgenin against skin aging at the time of climacteric. In vitro, diosgenin enhanced DNA synthesis in a human 3D skin equivalent model, and increased bromodeoxyuridine uptake and intracellular cAMP level in adult human keratinocytes. The increase of bromodeoxyuridine uptake by diosgenin was blocked by an adenylate cyclase inhibitor, but not by antisense oligonucleotides against estrogen receptor alpha, estrogen receptor beta or an orphan G-protein-coupled receptor, GPR30, indicating the involvement of cAMP but not estrogen receptor alpha, estrogen receptor beta or GPR30. In vivo, administration of diosgenin improved the epidermal thickness in the ovariectomized mice, a climacteric model, without altering the degree of fat accumulation. In order to examine the safety of diosgenin, diosgenin and 17beta-estradiol were administered to breast cancer-burdened mice. The results revealed that while 17beta-estradiol accelerated the tumor growth, diosgenin did not show this effect. Our finding, a restoration of keratinocyte proliferation in aged skin, suggests that diosgenin may have potential as a safe health food for climacteric.

Changes in responses of UVB irradiated skin of brownish guinea pigs with aging.

It is known that skin often shows irregular pigmentation during aging, which is frequently associated with hyperpigmentation. Many studies have utilized brownish A1 guinea pigs to investigate the pathogenesis of ultraviolet B (UVB)-induced skin pigmentation, however, responses associated with aging following UVB irradiation have not been elucidated. To characterize those responses, dorsal skin of A1 guinea pigs from 14-weeks to 5-yr old were investigated. The minimal erythema dose was found to increase with aging. Further, in pigmentation induced by UVB radiation, skin brightness (DeltaL*-value) decreased equally in both the 14-week old (young) group and in the 3-yr old (old) group of guinea pigs. The DeltaL*-value recovered in the young group from 21 d after UVB irradiation, whereas no such recovery was seen in the old group. In addition, the amount of melanin and the number of melanocytes returned near pre-irradiation levels in the young group, while they remained high in the old group. Our results therefore demonstrate for the first time that skin responses following UVB irradiation change with aging in A1 guinea pigs.

Pigmentation in intrinsically aged skin of A1 guinea pigs.

It is known that skin often shows irregular pigmentation during aging which is frequently associated with hyperpigmentation. Many studies have utilized brownish A1 guinea pigs to investigate the pathogenesis of ultraviolet (UV)-induced skin pigmentation, however, changes associated with intrinsic aging in A1 guinea pig skin have not been documented. To characterize such changes, skin from the dorsal and neck areas of 20-week, 1-, 2-, 3- and 5-yr-old guinea pigs was examined. Skin color was measured using a colorimeter, and biopsy specimens were stained with Masson-Fontana, L-3,4-dihydroxyphenylalanine (DOPA), and antibodies against KIT (ACK-45), gp100 (HMB-45) and S-100 proteins. The L* value of skin color decreased with aging and melanin deposits increased in the epidermis. Further, DOPA+, gp100+ and S-100+ melanocytes increased, indicating that the number of melanocytes had increased with age, whereas KIT+ melanocytes did not increase in dorsal skin and actually decreased in neck skin with aging. Further, rippled pigmented areas appeared in the neck skin of the 3-yr-old animals, and in the dorsal and neck skin of 5-yr-old guinea pigs in the absence of UV irradiation. Melanocytes were distributed uniformly in younger skin, whereas they were clustered in older skin. UV irradiation caused an increase in the number of melanocytes, although they were not clustered. These results are the first to provide evidence that pigmentation is induced in the skin of intrinsically aged A1 guinea pigs in the absence of UV irradiation, a process that differs from that elicited by UV irradiation.