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As a major intracellular degradation system, autophagy contributes to the maintenance of skin keratinocyte homeostasis. However, the precise role of autophagy in skin differentiation has not been fully investigated. To clarify whether autophagy plays a role in skin differentiation and maturation, autophagy-related gene 7 (Atg7)-deficient mice were generated. Atg7-deficient mice cannot survive for more than 24 h after birth. Therefore, the skins of Atg7-deficient mice and wild-type mice (as a control) were grafted onto severe combined immunodeficient mice. The resulting morphological and pathological changes were monitored for 28 days. Histopathological examination revealed acanthosis, hyperkeratosis, and abnormal hair growth in the skin grafts from the Atg7-deficient mice. Immune-density analysis of the skin grafts revealed reduced immunostaining of keratinization-related proteins, including loricrin, filaggrin, and involucrin, in the skin grafts from the Atg7-deficient mice. Furthermore, quantitative RT-PCR and Western blot analyses revealed the reduced expression of these three keratinization-related proteins in the skin grafts from the Atg7-deficient mice. Morphometric analysis using electron microscopy further revealed a reduction in the number and diameter of the keratohyalin and trichohyalin granules in these skin grafts. The differences were maintained for at least 1 month after transplantation. These results show that autophagy has a significant role in epidermal keratinization and hair growth until a certain stage of maturation.

Alopecia areata (AA) is an organ-specific and cell-mediated autoimmune disease involving hair loss, but its pathogenesis remains poorly understood. Many autoimmune diseases are genetically associated with alleles of the human leukocyte antigen (HLA) genes within the major histocompatibility complex. Associations between AA and HLA genes were previously observed in some different ethnic groups. However, the results were inconsistent, and a primary susceptibility HLA gene and/or region has not yet been assigned for AA. The aim of this study was to evaluate whether an allele of the HLA-C locus, HLA-C*07:04, which was strongly associated with AA in Chinese Hans, could be replicated in the Japanese population. The HLA-C locus was genotyped by the SSO method using 156 AA patients and 560 healthy controls. As a consequence, among the 17 alleles detected, only two alleles, C*04:01 (OR = 2.25, CI 95 % = 1.35-3.75, P = 1.84E-03) and C*15:02 (OR = 2.52, CI 95 % = 1.37-4.64, P = 2.90E-03), were significantly associated with AA after Bonferroni correction. Further, the stratification analysis suggested that C*04:01, C*07:02, and C*15:02 represented different AA genetic risk factors in each sub-phenotype.

Natural IgM can recognize apoptotic cells, but the molecular structure and the role in macrophage phagocytosis of apoptotic cells remain unclear.(1) To examine the binding of previously isolated natural IgM (3B4) to apoptotic cells and its effects on phagocytosis of apoptotic cells. (2) To characterize the molecular structure of 3B4.3B4 binding to apoptotic thymocytes was examined by flow cytometry. Polyreactivity of 3B4 was assayed by ELISA. PKH26-labeled Macrophages were incubated with PKH67-stained apoptotic cells in the presence of 3B4. Macrophages phagocytosis of apoptotic cell was evaluated by flow cytometry. The DNA segments of 3B V(H) and V(K) were sequenced and analyzed.3B4 IgM recognized late apoptotic cells. Polyreactive-recognitions of lysophosphatidylcholine (LPC) as well as some autoantigens were observed in 3B4. Phagocytosis of late apoptotic cells was increased in the presence of 3B4. The V(H) and V(K) genes of 3B4 showed a germline gene context, while N-sequences and nucleotide loss were observed in CDR3.3B4 promotes macrophage phagocytosis of late apoptotic cells in a complement-independent process. 3B4 has a germline configuration and is possibly ligand-selected. Out experiments suggest an independent role of natural IgM as opsonin in clearance of late apoptotic cells.

Mucosal immunity plays a pivotal role for body defense against infection and allergy. The aim of this study was to clarify the effects of 2,3,7,8-tetraclorodibenzo-p-dioxin (TCDD) on mucosal immunity in the gut.Fecal IgA level and oral tolerance induction were examined in TCDD-treated mice. Flow cytometric and histological analyses were also performed.Single oral administration of low dose 2,3,7,8-TCDD resulted in a marked decrease in IgA secretion in the gut without any effects on the cellular components of gut-associated lymphoid tissues (GALT) including Peyer's patches (PPs) and mesenteric lymph nodes (LNs). Decressed IgA secretion by TCDD was not observed in aryl hydrocarbon receptor (AhR)-deficient mice. Flow cytometric analysis revealed that IgA B cells in PPs and the mesenteric LNs remained unchanged in the TCDD-treated mice. An immunofluorescence study also demonstrated that a significant number of cytoplasmic IgA cells were present in the lamina propria of the gut in the TCDD-treated mice. Furthermore, oral tolerance induction by ovalbumin (OVA) was impaired in the TCDD-treated mice and OVA-specific T cell proliferation occurred in the peripheral lymphoid tissues including the spleen and LNs.These results suggest that a relatively low dose of TCDD impairs mucosal immunity in the gut and induces systemic sensitization by oral antigens.

DNA methylation is known to play an important role in gene transcription and alterations of methylation that contribute to the development of certain disorders such as cancer, immunodeficiency, and autoimmune diseases. We investigated the DNA methylation profiles in patients with atopic dermatitis (AD). Messenger RNA (mRNA) levels for DNA methyltransferase-1 (DNMT-1) in peripheral blood mononuclear cells (PBMC) were examined using a real-time quantitative polymerase chain reaction method. The levels of DNMT-1 mRNA were significantly lower in PBMC from the AD patients who had higher serum IgE levels compared with normal controls. Our observations suggest that suppression of DNMT-1 might be related to the pathogenesis of AD, especially in whom serum IgE level is high. This is the first report of DNMT-1 expression in AD patients.

Psoriasis is a common skin disease characterized by epidermal hyperproliferation, capillary dilatation, and the presence of acute and chronic inflammatory cells in both the dermis and epidermis [1, 2]. A unifying hypothesis that accounts for the wide range of clinical characteristics of psoriasis highlights the role of cytokines in the etio-pathophysiology of this disease [8]. Epidermal cells, particularly keratinocytes, may play a central role in transmitting pro-inflammatory signals by the secretion of cytokines and growth factors [7]. The production of these factors, in particular IL-1 is usually very low, but can be induced significantly by various injurious agents, such as endotoxin, virus particles, tumor promoter or UV light [6, 7]. Both IL-1a and c are expressed in human keratinocytes, although the major isotype secreted is IL-1a [6]. IL-1 and TNF-a in their turn act with an autocrine mechanism on keratinocytes to induce the release of a cascade of other cytokines with pro-inflammatory or chemotactic activity (IL-6, IL-8) [3]. All of these mentioned cytokines have been detected in psoriatic lesions. In addition to these observations, Takeda et al. [11] reported that SCCA2, the member of serine proteinase inhibitor family (serpin) is the major inducible SCCA in psoriatic skin. They found that SCCA2 was expressed in stratum corneum, subset of spinous layer cells and dermal infiltrated cells [11]. The level of SCCA2 is significantly increased in the sera of psoriatic patients, compared with non-psoriatic individuals [4], and probably be due, at least in part, to the production of cytokines in psoriatic tissues. Moreover, recent study suggested that the expression of SCCA in keretinocytes may be participated in the growth of keratinocytes and their differentiation [11]. Since keratinocytes can secrete TNF-a, IL-1a, and IL-6, and since these molecules have been detected in psoriatic skin, the goal of this study was to test whether the expression of SCCA2 in human keratinocytes plays a role in modulating expression of these cytokines. Commercially available human epidermal keratinocytes (Kurabo, Japan) were grown in supplemented low calcium medium and were sub-cultured at 60–70% confluence to avoid differentiation. A SCCA2 construct consisting of the SCCA2 coding region cloned in to the p3XFLAG-myc-CMV-26 vector (Sigma) has been described elsewhere [12]. The vector [p3XFLAG-mycCMV-26 (Sigma)] without SCCA2 was transfected into the keratinocytes as a control. The SCCA2 construct or the control vector was transfected into human keratinocytes using lipofectamine reagent (Sigma) according to the manufacturer’s instructions. TNF-a, IL-1a, and IL-6 concentrations in the culture media were measured by using ELISA kits (R&D Systems Inc., USA) according to the manufacturer’s instructions. SCCA2 expression in transfected cell was determined by Western blot analysis using anti-SCCA antibody (Sigma). The statistical analysis was performed using Student’s t test, and P

Generalized pustular psoriasis (GPP) is a rare but notoriously recalcitrant cutaneous diseases. Therefore, there have been few reports of more than ten patients with GPP who were treated at the same institution. The severity of this disease and its response to each therapeutic modality vary among patients. In some GPP is life-threatening, but in others it may show a benign, chronic course for a long period of time. Before starting treatment, a knowledge of the therapeutic efficacy and side effects of each drug used in the treatment of GPP is necessary. In our multicenter study, we compared the effectiveness of and adverse reactions to several systemically administered drugs. Following the development of a unique classification of the disease severity based on scoring the clinical symptoms and the laboratory findings, we propose here therapeutic guidelines for the treatment of GPP.

Hailey-Hailey disease (HHD, MIM 16960) is inherited in an autosomal dominant manner and characterized by persistent blisters and erosions of the skin1. Impaired intercellular adhesion and epidermal blistering also occur in individuals with pemphigus (which is due to autoantibodies directed against desmosomal proteins) and in patients with Darier disease (DD, MIM 124200), which is caused by mutations in a gene encoding a sarco/endoplasmic reticulum (ER)-Golgi calcium pump2. We report here the identification of mutations in ATP2C1, encoding the human homologue of an ATP-powered pump that sequesters calcium into the Golgi in yeast, in 21 HHD kindreds. Regulation of cytoplasmic calcium is impaired in cultured keratinocytes from HHD patients, and the normal epidermal calcium gradient is attenuated in vivo in HHD patients. Our findings not only provide an understanding of the molecular basis of HHD, but also underscore the importance of calcium control to the functioning of stratified squamous epithelia.

Keywords pso p27 AE Psoriasis AE Japanese descent AESquamous cell carcinoma antigenPsoriasis is a chronic inflammatory skin disease char-acterized by scaly plaques that result from epidermalhyperproliferation and infiltration of various inflam-matory cells. Although this disease was recently recog-nized as an autoimmune disorder [1], no promisingautoantigen candidates have yet been identified.Retrovirus-like particles were first identified in skinand urine from psoriatic patients in 1983 [2, 3]. In 1988,a mouse monoclonal antibody (mAb) was establishedagainst the 27-kDa detergent-soluble antigen derivedfrom the retrovirus-like particles in psoriatic patients(pso p27) [4]. We have demonstrated that the anti-psop27 mAb reacts with stratum corneum cells, some spi-nous cells and some tryptase-positive dermal infiltratingcells in skin biopsies from psoriatic patients of Europeandescent [5], and that this reactivity is diminished in skinbiopsies from patients who have undergone cyclosporineA therapy [7]. In addition, we have shown that pso p27participates in the formation of complement-activatingimmune complexes in psoriatic scales [8]. However, nostudies of the role of pso p27 had been carried out onJapanese patients with psoriasis. Moreover, in a proteindatabase search we found homology between apublished N-terminal protein sequence of pso p27 [6]and a squamous cell carcinoma antigen (SCCA)belonging to the serpin protease inhibitor family [9].In this study, we used immunofluorescent microscopyto examine the reactivity of the anti-pso p27 mAb withskin and peripheral mononuclear cells from Japanesepatients with psoriasis. We isolated recombinant SCCAfrom a cDNA library made using human keratinocyteRNA and examined the reactivity of the clones withanti-pso p27 antibody.Skin sections of 2–4