Kyung Ah Cheong

PDZK1 Upregulation in Estrogen-Related Hyperpigmentation in Melasma

The pathogenesis of melasma is unknown, although the potential role of estrogen has been considered. Microarray and real-time PCR analyses revealed that upregulation of PDZ domain protein kidney 1 (PDZK1) is clinically correlated with melasma. Although there has been no report that PDZK1 is involved in pigmentation and/or melanogenesis, PDZK1 expression can be induced by estrogen. In this study, the role of PDZK1 upregulation in melasma was examined, particularly in connection with estrogen, using biopsied skin specimens from 15 patients and monocultures and cocultures of melanocytes and keratinocytes with or without overexpression or knockdown of PDZK1. Estrogen upregulated PDZK1. Overexpression of PDZK1 increased tyrosinase expression and melanosome transfer to keratinocytes, whereas PDZK1 knockdown reduced estrogen-induced tyrosinase expression, through regulation of expression of estrogen receptors (ERs) ER-α and ER-β. The PDZK1-induced tyrosinase expression and melanosome transfer was regulated by ion transporters such as sodium-hydrogen exchanger (NHE), cystic fibrosis transmembrane conductance regulator (CFTR), and SLC26A3, which showed a specific association with each ER subtype. In the melanosome transfer, PDZK1 also increased phosphorylation of ezrin/radixin/moesin (ERM) and ras-related C3 botulinum toxin substrate 1, but not the expression of proteinase-activated receptor-2. Collectively, upregulation of PDZK1 could have an important role in the development of melasma in connection with estrogen through NHE, CFTR, and SLC26A3.

Glucosamine Improved Atopic Dermatitis‐like Skin Lesions in NC/Nga Mice by Inhibition of Th2 Cell Development

Dysregulated Th subset responses, characterized by Th2‐dominant allergic inflammation, are thought to be central to the pathogenesis of atopic dermatitis (AD). Glucosamine has been shown to have immunosuppressive properties, but its effect on AD has not been examined. In this study, the immunoregulatory effects of glucosamine, using dermatophagoides farinae (Df)‐induced AD‐like skin lesions in NC/Nga mice, were investigated. The clinical scores were reduced significantly by the treatment with glucosamine at 10 and 20 mg/day. Histological analysis of the skin also revealed that treatment of glucosamine at 10 and 20 mg/day significantly reduced the inflammatory cellular infiltrate, including mast cells and eosinophils. The levels of serum IgE and Th2 cytokines in spleen cells were reduced, whereas no significant change was detected in IFN‐γ, a Th1 cytokine. To determine the mechanism associated with inhibition of the Th2 immune response, the effects of glucosamine on the selective differentiation pathway of the Th subset in vitro was examined in NC/Nga mice. The results showed that glucosamine suppressed the differentiation of naïve CD4+ T cells to Th2 cells in vitro. On the basis of in vivo and in vitro results of the NC/Nga mice, the immunobiological effects of glucosamine on peripheral blood mononuclear cells from patients with AD were examined. The production of Th2 cytokines, such as IL‐4 and IL‐5, was significantly decreased after in vitro administration of glucosamine, which suggest that glucosamine might be a useful immunomodulatory agent for the treatment of human AD.

Mechanism underlying the effect of combined therapy using glucosamine and low-dose cyclosporine A on the development of atopic dermatitis-like skin lesions in NC/Nga mice.

Combination therapy is often used in the treatment of atopic dermatitis (AD) to improve clinical efficacy or to spare the dose of each drug. Cyclosporine A (CsA) is a calcineurin inhibitor that was developed for the treatment of AD. Glucosamine (Glu) is a potent immunosuppressant that inhibits Th2-mediated immunity. We previously reported that Glu has an ameliorative effect on the development of the pathology in NC/Nga mice. The aims of our study were to investigate the therapeutic efficacy of combination of Glu and low-dose CsA in dermatophagoides farina (Df)-induced AD-like skin lesions in NC/Nga mice and to determine the underlying therapeutic mechanisms. The Df-induced NC/Nga mice with a clinical score of 7 were used for treatment with Glu (500mg/kg) alone, low-dose CsA (2, 5, and 10mg/kg) or in combination. The clinical scores were reduced significantly by the combination treatment with Glu and low-dose CsA. The suppression of dermatitis by combined therapy was accompanied by decrease in the plasma level of IgE and in the splenic level of IL-4, IL-5, IL-13, TARC and eotaxin. Histological analysis of the skin also revealed that combination treatment significantly reduced the inflammatory cellular infiltrate, including mast cells and eosinophils. Particularly, immunological evaluation reveals an increase of CD4(+)CD25(+) Treg cells in the combined treatment. The induction of TSLP, which leads to systemic Th2 response, was reduced in the skin on combination treatment. The protein expression of filaggrin and involucrin was recovered by combination treatment in the skin lesions, whereas the protein expression of keratin-10 and keratin-14 decreased in the combination treatment. Collectively, our findings suggest that combination treatment of Glu and low-dose CsA leads to the therapeutic effects in Df-induced AD-like skin lesion in NC/Nga mice through inhibition of IgE, inflammatory cellular infiltrate, and recovery of skin barrier function via a mechanism that may inhibition of Th2-mediated immune responses, in part, increment of CD4(+)CD25(+) Treg cells. These results suggest that this combined immunosuppressive treatment may provide important implications for the design of therapeutic strategies aimed at AD treatment.

Therapeutic Effects of Combination Using Glucosamine Plus Tacrolimus (FK-506) on The Development of Atopic Dermatitis-Like Skin Lesions in NC/Nga Mice

Tacrolimus (FK‐506) has been found to exhibit potent inhibitory effects on spontaneously developed dermatitis. We previously showed that glucosamine prevents the development of Atopic dermatitis (AD)‐like skin lesions in NC/Nga mice. The aims of our study were to investigate the synergistic therapeutic efficacy of combination of glucosamine plus FK‐506 in dermatophagoides farina (Df)‐induced AD‐like skin lesions in NC/Nga mice and to determine the underlying therapeutic mechanisms. The Df‐induced NC/Nga mice with a clinical score of 8 were used for treatment with glucosamine (500 mg/kg) alone, FK‐506 (1.0 mg/kg) or in combination. The synergistic effects of combination therapy were evaluated by dermatitis scores, skin histology and immunological parameters such as IgE, Th2‐mediated cytokines and chemokines, CD3+ T cells and CLA+ T cells. Combined therapy using glucosamine plus FK‐506 improved the development of AD‐like skin lesions as exemplified by a significant decrease in total skin symptom severity scores. The suppression of dermatitis by combined therapy was accompanied by a decrease in the plasma level of IgE and in the splenic level of IL‐5, IL‐13, TARC and eotaxin. Histological finding indicated that the dermal infiltration of inflammatory cells including mast cells and eosinophils was greatly reduced. Particularly, immunohistological evaluation reveals a reduction in CD3+ T cells and CLA+ cells in the combined therapy. Our findings suggest that combination therapy of glucosamine plus FK‐506 was more synergistic efficacy than single‐modality treatment with either alone to improve the development of established dermatitis in NC/Nga mice model. This combined immunosuppressive therapy may provide an effective therapeutic strategy for the treatment of AD.

Combination of glucosamine improved therapeutic effect of low-dose cyclosporin A in patients with atopic dermatitis: A pilot study

Both glucosamine and cyclosporin have been reported to show immunomodulatory effect with inhibition of each different key transcription factor for cytokine gene expression and T‐cell function. The overall purpose of this pilot study was to assess the feasibility of the combination of cyclosporin with glucosamine for the treatment of patients with atopic dermatitis. Twelve patients more than 12 years old who required systemic cyclosporin were included in the study. Two of them dropped out due to violation of medication schedule. The single (S) and combination (C) regimens were crossed over every 2 weeks without a washout period between the cross‐over for 6 months. Five patients were randomly assigned to the S regimen first (SC sequence), whereas the other five were given the C first (CS sequence). The change of SCORAD index was analyzed as the primary efficacy end‐point by general linear model and piecewise linear mixed model. The SCORAD index was reduced with both SC and CS sequence regimens. In particular, index reduction with the C was more than that associated with S regimen; this difference increased as time lapsed. The glucosamine combination was predicted to cause an additive decrease in the mean percent change of the SCORAD index (~6%), with decreasing interleukin (IL)‐4 and IL‐5 cytokine levels but without increasing treatment‐related adverse events. This study suggests that the C would produce better clinical outcomes than the S regimen in patients with atopic dermatitis, although confirmatory clinical trials are warranted to determine the effect of combination.

Three New Single Nucleotide Polymorphisms Identified by a Genome-Wide Association Study in Korean Patients with Vitiligo

Genetic susceptibility is involved in the pathogenesis of vitiligo. Association studies with a whole genome-based approach instead of a single or a few candidate genes may be useful for discovering new susceptible genes. Although the etiology of non-segmental and segmental types is different, the association between gene polymorphisms and vitiligo has been reported, without defining types or in non-segmental type. Whole genome-based single nucleotide polymorphisms (SNPs) were examined in patients with non-segmental and segmental types of vitiligo using the Affymetrix GeneChip 500K mapping array, and 10 functional classes of significant SNPs were selected. Genotyping and data analysis of selected 10 SNPs was performed using real-time PCR. Genotype and allele frequencies were significantly different between both types of vitiligo and three of the target SNPs, DNAH5 (rs2277046), STRN3 (rs2273171), and KIAA1005 (rs3213758). A stronger association was suggested between the mutation in KIAA1005 (rs3213758) and the segmental type compared to the non-segmental type of vitiligo. DNAH5 (rs2277046), STRN3 (rs2273171), and KIAA1005 (rs3213758) may be new vitiligo-related SNPs in Korean patients, either non-segmental or segmental type.

Association of thymic stromal lymphopoietin gene −847C>T polymorphism in generalized vitiligo

Abstracts:  Thymic stromal lymphopoietin (TSLP) induces naïve CD4+ T cells to produce Th2 cytokines. In addition, to low production of Th2 cytokines, strong Th1 response, which plays an important role in vitiligo development, has been induced by blockade of TSLP or TSLP receptor. This study examined whether a functional TSLP polymorphism was associated with vitiligo. One hundred and sixty Korean patients with vitiligo and 568 healthy Korean individuals were examined for the four SNPs of TSLP gene. Luciferase activity was measured for promoter assay. The genotype and allele frequencies of −847C>T polymorphism were lower in vitiligo patients compared with the controls, whereas those of wild type were higher (P = 0.004, P = 0.017 respectively). None the less, the promoter activity of −847C decreased significantly (P = 0.013) compared with −847T, expecting lower TSLP mRNA levels in the polymorphism. Collectively, C allele at the TSLP −847C>T polymorphism may increase susceptibility to generalized vitiligo through decreasing TSLP mRNA expression levels.

850 nm light-emitting-diode phototherapy plus low-dose tacrolimus (FK-506) as combination therapy in the treatment of dermatophagoides farinae-induced atopic dermatitis-like skin lesions in NC/Nga mice

BACKGROUND Light emitting diode (LED) phototherapy is an effective alternative for the treatment of inflammatory skin disorders. Tacrolimus (FK-506) is a potent immunomodulating agent, which has been used to treat AD. Combination therapy is often used in the treatment of AD to improve therapeutic efficacy or to reduce the dose of each drug. OBJECTIVE To investigate the therapeutic efficacy of monotherapy with either 850nm LED phototherapy or low-dose FK-506, and combination therapy in Dermatophagoides farina (Df)-induced AD-like skin lesions in NC/Nga mice. METHODS The Df-induced NC/Nga mice with a clinical score of 7 were used for treatment with LED (10 and 25J/cm(2)) alone, low-dose FK-506 (1mg/kg) or in combination. The synergistic effects of combined therapy were evaluated by dermatitis scores, skin histology, skin barrier function, and immunological parameters, such as IgE, NO, Th2-mediated cytokines and chemokines. RESULTS Combination therapy with 850nm (25J/cm(2)) LED and low-dose FK-506 showed a significant reduction in the severity of skin lesions. Combined therapy decreased in the serum level of IgE, NO, and in the splenic level of Th2-mediated cytokines and chemokines. Combination therapy significantly also reduced the inflammatory cellular infiltrate into the skin lesions. Moreover, combination therapy led to recovery of skin barrier function in the skin lesions. CONCLUSIONS The use of combination of LED phototherapy and low-dose immunosuppressant improved Df-induced AD-like skin lesions in an NC/Nga mouse model by dominantly reducing IgE, NO, suppressing Th2-mediated immune responses, and inhibiting inflammatory cells, as well as improving skin barrier function.

S100B as a potential biomarker for the detection of cytotoxicity of melanocytes

Skin irritation is one of the most common adverse reactions in hydroquinone (HQ) and retinoic acid (RA). Although melanocytes have rarely been considered to be involved in skin irritation, RA and particularly HQ could induce melanocyte toxicity, resulting in depigmentation. We chose S100B as a candidate gene for melanocytotoxicity from a genome‐wide transcriptional profiling analysis after applying irritant doses of HQ, RA and sodium lauryl sulphate (SLS) to cultures of keratinocytes and/or melanocytes. In this study, the role of S100B on melanocyte viability and cytotoxicity was examined. S100B was detected in melanocytes, but not in keratinocytes or fibroblasts. Melanocytes after treatment with increasing concentrations of HQ, RA, SLS and urushiol showed significant increases in intracellular and extracellular S100B expression with reduced viable cell number and increased release of lactate dehydrogenase. No RAGE expression and no significant function of CD166/ALCAM in melanocyte survival and cytotoxicity favoured the role of intracellular S100B in chemically irritated melanocytes. S100B knock‐down increased apoptosis through inhibition of PI3K/AKT, NF‐κB and ERK activation, suggesting the increased intracellular S100B expression by chemical irritation as a compensatory reaction to reduce cytotoxicity. The numerical decrease in S100B/c‐kit‐double‐positive melanocytes was also examined in human skin epidermis irritated by HQ or RA with stronger staining intensities of S100B. Collectively, the decrease in viable cell number by reduced intracellular S100B levels in vitro and by chemical irritation in vivo suggests that S100B could be a potential biomarker for melanocytes cytotoxicity.

Combination of glucosamine and low-dose cyclosporine for atopic dermatitis treatment: A randomized, placebo-controlled, double-blind, parallel clinical trial

Our recent pilot study showed better outcomes using a combination of low‐dose cyclosporine and glucosamine than cyclosporine alone in the treatment of atopic dermatitis (AD). Here, a randomized, placebo‐controlled, double‐blind, parallel‐designed study was planned to compare the efficacy and safety of low‐dose cyclosporine and glucosamine combination to low‐dose cyclosporine alone for the treatment of patients with moderate to severe AD. AD patients with a Severity Scoring of Atopic Dermatitis (SCORAD) index ≥30 were randomly assigned in a 1:1 ratio to receive either cyclosporine 2 mg/kg and glucosamine 25 mg/kg (group A) or cyclosporine and placebo (group B) for 8 weeks. SCORAD indices, serum levels of chemokine ligand 17 and interleukin‐31, eosinophil counts, and blood cyclosporine levels were examined before and after treatment. The SCORAD indices for group A (n = 19) were significantly reduced after the treatment and a significant correlation between the changes in the SCORAD indices and changes in the serum levels of chemokine ligand 17, but not interleukin‐31, was detected. Glucosamine combined with cyclosporine did not increase adverse events and serum cyclosporine levels compared with cyclosporine alone. Therefore, combination of low‐dose cyclosporine and glucosamine may be useful to allow the long‐term use of cyclosporine in the treatment of patients with moderate to severe AD.