Loss of IκBζ Drives Dentin Formation via Altered H3K4me3 Status.

Enforced enrichment of the active promoter marks trimethylation of histone H3 lysine 4 (H3K4me3) and acetylation of histone H3 lysine 27 (H3K27ac) by inhibiting histone demethylases and deacetylases is positively associated with hard tissue formation through the induction of osteo/odontogenic differentiation. However, the key endogenous epigenetic modulator of odontoblasts to regulate the expression of genes coding dentin extracellular matrix (ECM) proteins has not been identified. We focused on nuclear factor (NF)-κB inhibitor ζ (IκBζ), which was originally identified as the transcriptional regulator of NF-κB and recently regarded as the NF-κB-independent epigenetic modulator, and found that IκBζ null mice exhibit a thicker dentin width and narrower pulp chamber, with aged mice having more marked phenotypes. At 6 mo of age, dentin fluorescent labeling revealed significantly accelerated dentin synthesis in the incisors of IκBζ null mice. In the molars of IκBζ null mice, marked tertiary dentin formation adjacent to the pulp horn was observed. Mechanistically, the expression of COL1A2 and COL1A1 collagen genes increased more in the odontoblast-rich fraction of IκBζ null mice than in wild type in vivo, similar to human odontoblast-like cells transfected with small interfering RNA for IκBζ compared with cells transfected with control siRNA in vitro. Furthermore, the direct binding of IκBζ to the COL1A2 promoter suppressed COL1A2 expression and the local active chromatin status marked by H3K4me3. Based on whole-genome identification of H3K4me3 enrichment, ECM and ECM organization-related gene loci were selectively activated by the knockdown of IκBζ, which consistently resulted in the upregulation of these genes. Collectively, this study suggested that IκBζ is the key negative regulator of dentin formation in odontoblasts by inhibiting dentin ECM- and ECM organization-related gene expression through an altered local chromatin status marked by H3K4me3. Therefore, IκBζ is a potential target for epigenetically improving the clinical outcomes of dentin regeneration therapies such as pulp capping.

Three patients with advanced cutaneous angiosarcoma treated with eribulin: investigation of serum soluble CD163 and chemokine (C-X-C motif) ligand 10 as possible biomarkers predicting the biological behaviour of angiosarcoma.

Cutaneous angiosarcoma (CAS) is a highly aggressive vascular tumour that recurs locally and metastasizes early. Although chemoradiotherapy with taxanes shows a high response rate with prolonged survival, second-line therapy for advanced CAS remains contentious. This report describes three patients with advanced CAS treated with eribulin. In addition, we investigated serum soluble (s)CD163, chemokine (C-X-C motif) ligand 10 (CXCL10) and chemokine (C-C motif) ligand 2 levels at several time points of tumour progression in these patients, revealing serum levels of sCD163 and CXCL10 as potential biomarkers for progression of CAS.

Comparison of the measuring efficacy of transepidermal water loss of a reasonably priced, portable closed-chamber system device H4500 with that of rather expensive, conventional devices such as Tewameter® and Vapometer®.

BACKGROUND/PURPOSE: Although measuring transepidermal water loss (TEWL) is important to assess the barrier function of the stratum corneum (SC), the commercially available instruments are rather expensive. Recently launched Model H4500 employs a closed-chamber system to measure TEWL and is more reasonably priced compared to devices currently in general use. METHODS: To check the reproducibility of the obtained data with H4500, we conducted measurements on the volar forearms of healthy volunteers and compared these data with those measured with Vapometer® and Tewameter® . Then, we checked the correlations between the TEWL data obtained with these different devices on the same volar forearms of 15 healthy volunteers before and after the artificial production of barrier damage of the SC by tape stripping or by 0.5% aqueous solution of sodium lauryl sulfate. RESULTS: The obtained intra-class correlation coefficient (ICC, [1, 1]) with 95% CI of H4500 was 0.927 (0.835-0.978). Namely, an excellent correlation could be found in the values of TEWL measured with these three different instruments not only on healthy skin but also on the artificially barrier-damaged skin. CONCLUSIONS: H4500 is considered to be practical for daily use because of its performance as well as its reasonable price as compared with conventional devices.

Possible mechanisms of the crosstalk between Langerhans cells and regulatory T cells in extramammary Paget disease by receptor activator of nuclear factor kappa B (RANK) ligand/RANK pathways.

BACKGROUND: Extramammary Paget disease (EMPD) is a skin adenocarcinoma of apocrine gland origin, in which Paget cells express receptor activator of nuclear factor kappa B (RANK) ligand (RANKL) and matrix metalloproteinase (MMP)-7, and release soluble (s)RANKL into the tumour microenvironment. We previously reported that about 60% of the RANK+ cells among the stromal cells are M2 macrophages, but the identity of the remaining population of RANK+ cells is still unknown. OBJECTIVES: To investigate the unknown subpopulation of RANK-expressing cells in EMPD. METHODS: The main population of RANK-expressing cells in the epidermis was composed of epidermal Langerhans cells (LCs). To explore the effects of RANKL on LCs, we stimulated LCs generated from human CD34+ hematopoietic progenitor cells with graded concentrations of sRANKL. To further examine the correlation between LCs and regulatory T cells (Tregs) in EMPD, we employed immunohistochemical staining. RESULTS: sRANKL stimulation was shown to augment the production of C-C motif chemokine ligand 17 (CCL17) from LCs. We additionally demonstrated CCL17 expression by CD1a+ LCs in EMPD in an immunofluorescence study. Spearman's rank correlation test confirmed a correlation between the number of LCs and the number of Foxp3+ Tregs in the lesional skin of invasive EMPD. In addition, the numbers of Foxp3+ Tregs in the sentinel lymph nodes of metastatic EMPD were significantly higher than those of metastatic melanoma, which did not express RANKL. CONCLUSIONS: The findings suggest that the RANKL/RANK pathway in EMPD might contribute to the recruitment of Tregs and to maintenance of the tumour microenvironment.

Therapeutic drug monitoring of patients with psoriasis during tumour necrosis factor (TNF)-α antagonist treatment using a novel interleukin-8 reporter cell line.

BACKGROUND: Tumour necrosis factor (TNF)-α antagonist therapy is currently used for moderate and severe psoriasis. However, this treatment has several drawbacks, including interindividual variability in clinical response and secondary loss of effectiveness. OBJECTIVES: To evaluate quantitatively the TNF-α-neutralizing activity of the plasma of patients with psoriasis during TNF-α antagonist therapy and to determine poor responders objectively. METHODS: We used a human interleukin-8 reporter monocyte cell line, THP-G8, that harbours a stable luciferase orange (SLO) gene under the control of the interleukin-8 promoter. After confirming its dose-dependent response to exogenous TNF-α, we examined the suppressive activity of TNF-α antagonists and of the patients' plasma during TNF-α antagonist therapy on TNF-α-induced SLO luciferase activity (TNF-SLO-LA). RESULTS: Pretreatment of TNF-α with TNF-α antagonists or with the plasma of patients with psoriasis who achieved 75% improvement in Psoriasis Area and Severity Index (PASI 75) dose dependently suppressed TNF-SLO-LA. There was a significant correlation between change in PASI and percentage suppression (inhibitory rate of a 1 : 2 dilution of patient plasma on TNF-SLO-LA). A percentage suppression of 50·3% has a positive predictive value of 87% of achieving PASI 75, with a sensitivity of 93% and a specificity of 80%. CONCLUSIONS: Therapeutic monitoring of patients with psoriasis during TNF-α antagonist therapy using THP-G8 can provide a useful tool to determine objectively the efficacy of the administered TNF-α antagonists.

The spectrophotometrical analysis of rhododendrol-induced leucoderma using a novel multispectral camera.

BACKGROUND: Many users in Japan of skin brightening/lightening cosmetics containing rhododendrol (RD) have developed leucoderma. Leucoderma appears on skin areas repeatedly treated with RD-containing cosmetics. RD-induced leucoderma (RDIL) presents different degrees of well-defined hypopigmentation. It is crucial to determine the degree of hypopigmentation to differentiate RDIL from vitiligo vulgaris (VV). OBJECTIVES: To quantitatively evaluate hypopigmentation of RDIL lesions and the recovery of pigmentation, and to compare the hypopigmentation with VV and normal skin. MATERIALS AND METHODS: Sixteen cases of RDIL, nine cases of VV and 15 healthy controls were examined using a novel multispectral camera (MSC) that can simultaneously obtain the reflection intensity at 10-nm wavelength intervals from 400 to 760 nm of the photographed area. ∆Absorbance was calculated by subtracting the log of reflection intensity of the target area from that of a white reflection standard. RESULTS: Most RDIL lesions showed lower ∆Absorbance than healthy skin and higher ∆Absorbance than VV lesions between 400 and 550 nm. Statistical comparison of the maximum ∆Absorbance from 420 to 460 nm (Max∆Absorbance) for VV, RDIL and control skin showed that the Max∆Absorbance of RDIL was significantly higher than that of VV and lower than that of control skin. The comparison of ∆Absorbance of the same sites in RDIL lesions between the initial visit and 6 months later showed significant improvement after 6 months. CONCLUSIONS: These studies demonstrated quantitative changes in RDIL and its recovery phase and suggested the utility of a MSC in obtaining objective colour information of skin disorders.

Expression of CD39/Entpd1 on granuloma-composing cells and induction of Foxp3-positive regulatory T cells in sarcoidosis.

BACKGROUND: Regulatory T cells (Tregs), together with tolerogenic dendritic cells (tDCs) are involved in maintaining peripheral tolerance. A recent report suggested both Tregs and tDCs may be pathogenic in granulomatous skin disorders. AIM: To examined the expression of CD39 on granuloma-composing cells and Foxp3-positive Tregs in the skin in two representative granulomatous diseases, sarcoidosis and granuloma annulare (GA). METHODS: We immunohistologically examined expression of CD39 on granuloma-composing cells and expression of Foxp3 on CD4+ or CD25+ cells in fixed sections of lesional skin from 16 patients with sarcoidosis and five patients with GA. RESULTS: The granuloma-composing cells expressed CD39 in both sarcoidosis and GA. Significant numbers of CD4+ Foxp3+ Tregs were present diffusely throughout the granulomatous tissues in sarcoidosis, whereas Tregs in GA existed only at the peripheral lesion of palisading granulomatous tissue. CONCLUSIONS: There was infiltration of increased numbers of Foxp3+ Tregs around the CD39+ granuloma-composing cells in both GA and sarcoidosis.

Nuclear localization of activated STAT6 and STAT3 in epidermis of prurigo nodularis.

BACKGROUND: Prurigo nodularis (PN) is a chronic dermatitis characterized by discrete, raised, and firm papulonodules with intense pruritus. The pathogenesis still remains to be elucidated. OBJECTIVES: To clarify the role of Th1 and Th2 cytokines in the pathogenesis of PN. METHODS: We examined the cytokine signatures, such as phosphorylation of STAT1, STAT3 and STAT6, HLA-DR and hyaluronan accumulation, to reveal the Th1 and Th2 cytokine influence on the lesional epidermis of PN. RESULTS: We first optimized antigen retrieval methods to detect these signatures with antibodies for phospho-STAT1 (pSTAT1), phospho-STAT3 (pSTAT3), phospho-STAT6 (pSTAT6), HLA-DR and hyaluronic acid binding protein (HABP) on the formalin-fixed paraffin-embedded sections of psoriasis, lichen planus and atopic dermatitis biopsy samples. Activation of STAT1 and STAT6 in epidermis by Th1 and Th2 cytokines was further confirmed in a cultured skin equivalent model treated with interferon-γ or interleukin (IL)-4/IL-13. With the relevant immunostaining methods, we examined the cytokine signatures in 22 cases of PN. The results revealed that (i) the entire epidermis of 19 cases was stained with anti-pSTAT6 antibody, (ii) 21 cases demonstrated nuclear staining with anti-pSTAT3 antibody, (iii) the entire epidermis of 21 cases was stained with HABP, (iv) the epidermis of eight cases showed scattered staining with anti-pSTAT1 antibody, and (v) six cases were positive for HLA-DR membrane expression. CONCLUSIONS: These data indicated that Th2 cytokines related to STAT6 activation together with some unknown stimuli that activate STAT3 play a principal role in the pathogenesis of PN.

Functional analyses of the skin surface of the areola mammae: comparison between healthy adult male and female subjects and between healthy individuals and patients with atopic dermatitis.

BACKGROUND: Although the nipple and areola of the breast constitute a unique and prominent area on the chest, so far no study has been done on the functional properties of their skin surfaces. OBJECTIVE: To study the stratum corneum (SC) covering the areola using noninvasive methods. METHODS: Eighteen adult healthy subjects comprising nine men and nine women and 18 age- and sex-matched patients with atopic dermatitis (AD), none of whom had visible skin lesions, participated in the study. Transepidermal water loss (TEWL), skin surface hydration and skin surface lipid levels were measured on the areola and adjacent breast skin. The size of the skin surface corneocytes of these skin regions was assessed. RESULTS: All the healthy subjects showed significantly higher TEWL accompanied by smaller sized corneocytes on the areola than on the adjacent breast skin. Only female subjects revealed a significantly higher skin surface hydration state together with significantly increased skin surface lipid levels on the areola than on the adjacent breast skin. These sex differences were observed even in patients with AD. Comparison between healthy individuals and the patients with AD demonstrated higher TEWL, decreased skin surface hydration state and lower skin surface lipid levels associated with smaller sized corneocytes in the areola in the patients with AD, especially in male patients. CONCLUSIONS: In adults, the SC barrier function and SC water-binding capacity of the areola were functionally poorer than in the adjacent skin, being covered by smaller sized corneocytes and lower amounts of skin surface lipids, especially in men and in patients with AD.

Cutaneous accumulation of abnormal polyglutamine proteins of patients with dentatorubral-pallidoluysian atrophy.

BACKGROUND AND PURPOSE: Dentatorubral-pallidoluysian atrophy (DRPLA) is a hereditary spinocerebellar degeneration caused by expansion of a trinucleotide CAG repeat encoding a polyglutamine tract in a disease protein atrophin-1. The clinical features include ataxia, choreoathetosis, and dementia, which result from neural degeneration caused by the mutant atrophin-1. METHODS: We performed skin biopsy in two patients with DRPLA. RESULTS: We found multiple clear cells in the epidermis, which were positive for proteins containing an expanded polyglutamine stretches. The clear cells were p63 (+), S-100 (-), and cytokeratin 20 (-), showing that they were keratinocytes. Negative or weak signals of pan-cytokeratin were consistent with the finding of decreased tonofilaments at the electron microscopic level. CONCLUSIONS: The presence of clear keratincoytes showed that the mutant proteins interfered in cellular functions not only in neural cells but also in keratinocytes. The skin is accessible by biopsy, making it important in the diagnosis. Furthermore, the polyglutamine staining in the skin may be useful for evaluation of therapeutic modalities for DRPLA and other polyglutamine diseases.

Perilesional treatment of metastatic melanoma with interferon-beta.

Previous trials with various treatments have not shown satisfactory therapeutic results for cutaneous metastasis of malignant melanoma (MM). We report three patients who were treated with peritumoral injection of interferon (IFN)-beta for multiple skin metastases of MM. The metastatic tumours were infiltrated by significant numbers of CD8+ TIA+ cytotoxic lymphocytes, and the numbers of CD4+ cells and human leucocyte antigen-DR+ cells increased after IFN-beta injection. These results suggest that the peritumoral administration of IFN-beta enhanced the antitumour immune response against the MM, suggesting that it is a promising supportive treatment for skin metastasis of MM.

Pseudolymphomatous angiokeratoma: report of three cases and an immunohistological study.

BACKGROUND: Pseudolymphomatous angiokeratoma (PA), originally termed 'acral pseudolymphomatous angiokeratoma of children', is a disorder characterized clinically by development of red nodules on the extremities and histologically by a subepidermal dense lymphocyte infiltrate. METHODS: We report three cases of PA, with characteristically dense, nodular infiltrate composed predominantly of small lymphocytes, and thick-walled vessels. RESULTS: Immunohistochemical investigation revealed a dense accumulation of CD20+ cells with CD3+ cells in one case. Infiltrate in the other two cases was mainly composed of CD3+ cells and a mixture of CD4+ and CD8+ cells, with a few cells expressing CD20. CONCLUSION: Our immunohistological results reveal a wide spectrum of cellular infiltrate compositions ranging from T-cell to B-cell predominance.

Profiles of Foxp3+ regulatory T cells in eczematous dermatitis, psoriasis vulgaris and mycosis fungoides.

BACKGROUND: It is well known that regulatory T cells (Tregs), identified by their expression of CD4, CD25 and Foxp3, play a crucial role in maintaining peripheral tolerance. Recently, it has been demonstrated that a Treg population resides in normal human skin. However, only a few studies have demonstrated the presence of Foxp3+ Tregs in inflammatory skin disorders. OBJECTIVES: In this study, we immunohistologically examined the presence of CD4+ CD25+ Foxp3+ Tregs in the lesional skin of psoriasis vulgaris, mycosis fungoides and eczematous dermatitis. METHODS: We used immunohistochemistry to examine the presence of Foxp3+ Tregs in fixed sections of the lesional skin from 16 patients with psoriasis vulgaris, 17 patients with mycosis fungides and 18 patients with eczematous dermatitis in addition to 10 normal skin samples. RESULTS: In normal skin, epidermal and dermal Foxp3+ cells were rare. The psoriasis vulgaris, mycosis fungoides and eczematous dermatitis samples contained substantial numbers of epidermal and dermal CD3+, CD4+ and CD25+ Foxp3+ Tregs. The epidermis contained a higher percentage of CD3+, CD4+ and CD25+ Foxp3+ cells than the dermis. The percentage of Foxp3+ cells among CD3+ or CD4+ cells was significantly lower in eczematous dermatitis than in psoriasis vulgaris or mycosis fungoides, and that of dermal Foxp3+ cells was significantly lower in psoriasis vulgaris than in eczematous dermatitis or mycosis fungoides. CONCLUSIONS: The lower percentage of epidermal or dermal Foxp3+ cells in eczematous dermatitis or psoriasis vulgaris, respectively, might contribute to their pathogenesis.