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Tumor necrosis factor-alpha (TNF-alpha) antagonists are effective for inflammatory diseases, such as Crohn's disease, rheumatoid arthritis (RA) and psoriasis. Although TNF-alpha antagonists are also useful for sarcoidosis, paradoxical occurrence of sarcoidosis or sarcoidal reaction may be observed. We report a Crohn's disease patient, who developed sarcoidosis during infliximab therapy. A 35-year-old man had been receiving infliximab for 7 months for Crohn's disease. He developed cough and fever, accompanied by an infiltrated erythematous plaque on his right knee. The chest radiography, skin biopsy and laboratory findings were all consistent with sarcoidosis.

A 64-year-old Japanese woman had a lightly brown-blackish pigmented macule (1.2 cm in diameter) on the left sole of her foot. She received surgical excision following a diagnosis of acral lentiginous melanoma (ALM), which was confirmed histopathologically. One month after the operation, a second melanoma lesion was noticed adjacent to the grafted site. Histopathologically, the two lesions had no continuity, but HMB-45 and cyclin D(1) double-positive cells were detected not only on aggregates of atypical melanocytes but also on single cells near the cutting edge of the first lesion. The unique occurrence of a sequential lesion of a primary melanoma might be caused by stimulated subclinical field cells during the wound healing process following the initial operation. This case warrants further investigation to establish the appropriate surgical margin of ALM lesions.

Psoriasis is a chronic inflammatory skin disease, which has been increasing during the last 50 years in Japan. The aim of the present study is to analyze psoriatic patients registered from 1983-2007 in Asahikawa Medical College Hospital, which is located in the northern part of Japan. A total of 607 cases were registered at the first inspection in the Department of Dermatology, Asahikawa Medical College. Men (403 cases, 66.4%) were predominant over women (204 cases, 33.6%). The clinical types of psoriasis were psoriasis vulgaris (91.5%), guttate psoriasis (4.2%), psoriasis arthropathica (2.8%), psoriatic erythroderma (0.6%), generalized pustular psoriasis (0.6%), localized pustular psoriasis (0.15%) and infantile psoriasis (0.15%). Topical corticosteroids (78.1 %) and vitamin D3 (18.1 %) products were the main previous topical agents. Previous systemic treatments included etretinate (7.7%), cyclosporine (1.5%) and methotrexate (0.3%). Use of topical vitamin D3 and cyclosporine therapies have been gradually increasing during the past 25 years. Regarding the previous phototherapy, topical psoralen and ultraviolet A therapy (PUVA) (4.9%) was predominant over ultraviolet B (0.9%), and systemic PUVA (0.7%). Use of ultraviolet B phototherapy has been increasing during the past 5 years. The results are essentially similar to those of a survey of psoriasis in Japan from 1982-2001. Although the incidence of psoriasis might be higher in Hokkaido Prefecture, there is essentially no variation in the disease profile of psoriatic patients.

Background Although the precise pathomechanism of psoriasis is still unknown, various cytokines and growth factors derived from T cells, dendritic cells or keratinocytes, are critically involved in this disease. There have been several studies determining the serum levels of cytokines in patients with psoriasis, but with conflicting results. The levels of various cytokines and growth factors were measured in the sera of patients with psoriasis and compared with those of healthy controls. The correlation with disease severity was also determined. Methods Sera were collected from 122 patients with psoriasis and 78 healthy controls for ELISA analysis to evaluate the levels of cytokines and growth factors. The severity of psoriasis was determined by the Psoriasis Area and Severity Index (PASI). Results Serum levels of tumour necrosis factor (TNF)-alpha, interferon (IFN)-gamma, interleukin (IL)-2, IL-6, IL-7, IL-8, IL-12, IL-17, IL-18 and vascular endothelial growth factor (VEGF) were significantly increased in patients with psoriasis compared with those of healthy controls. The serum levels of IL-2, soluble intercellular adhesion molecule-1, epidermal growth factor, hepatocyte growth factor and amphiregulin were not significantly different from those of healthy controls. Increased serum levels of TNF-alpha, IFN-gamma, IL-12, IL-17, IL-18 and VEGF correlated with PASI. Furthermore, these cytokine levels were decreased after psoriasis treatment. In contrast, serum levels of IL-10 were decreased in psoriasis and negatively correlated with PASI. Discussion Serum levels of TNF-alpha, IFN-gamma, IL2, IL-6, IL-7, IL-8, IL-12, IL-17, IL-18 and VEGF were positively correlated and that of IL-10 was negatively correlated with PASI in Japanese patients with psoriasis. These parameters might be useful for determining the disease activity of psoriasis.

Inherited disorders of elastic tissue represent a complex and heterogeneous group of diseases, characterized often by sagging skin and occasionally by life-threatening visceral complications. In the present study, we report on an autosomal-recessive disorder that we have termed MACS syndrome (macrocephaly, alopecia, cutis laxa, and scoliosis). The disorder was mapped to chromosome 20p11.21-p11.23, and a homozygous frameshift mutation in RIN2 was found to segregate with the disease phenotype in a large consanguineous kindred. The mutation identified results in decreased expression of RIN2, a ubiquitously expressed protein that interacts with Rab5 and is involved in the regulation of endocytic trafficking. RIN2 deficiency was found to be associated with paucity of dermal microfibrils and deficiency of fibulin-5, which may underlie the abnormal skin phenotype displayed by the patients.

Summary Background Epidermolysis bullosa acquisita (EBA) autoantibodies recognize epitopes predominantly within the N-terminal noncollagenous (NC)-1 domain of type VII collagen. Recently, some EBA cases with reactivity to other domains, i.e. the triple-helical (T-H) collagenous domain and the NC-2 domain, have been reported. Objectives To investigate the ultrastructural localization of epitopes for sera from five patients with EBA that were unreactive by immunoblotting with the NC-1, NC-2 and collagenous domains of type VII collagen. Methods Immunogold postembedding indirect immunoelectron microscopy was performed using normal human skin and type VII collagen-deficient skin as substrates. Results Postembedding indirect immunoelectron microscopy revealed that the five EBA sera showed immunoreactivity in the dermis, mainly located 0–400 nm below the lamina densa. IgG labelling was not observed in type VII collagen-deficient skin from a patient with recessive dystrophic epidermolysis bullosa. The distribution histogram found in this study was different from those of sera that reacted with the NC-1 and/or NC-2 domains, and was similar to those of sera reacting with the T-H collagenous domain. Conclusions Our results suggest that epitopes within the T-H collagenous domain of type VII collagen are recognized by IgG antibodies from some EBA sera. These antibodies appear to be found in patients with inflammatory-type EBA.

Skin lesions of porokeratosis consist of an atrophic center bordered by a peripheral grooved keratotic ridge that corresponds histopathologically to the cornoid lamella. Originally porokeratosis was named based on the assumption that the columns of parakeratosis emerge from the ostia of eccrine ducts. Despite this, it is generally accepted that the cornoid lamellae are rarely related to the acrosyringium. We recently encountered two cases of porokeratosis, where the cornoid lamellae were related to the follicular infundibulums. Therefore, we analyzed the location of the cornoid lamellae in 86 lesions of porokeratosis from 73 patients from our archives. We found that many cornoid lamellae are also detected at follicular infundibulum and acrosyringium. The existence of so many cornoid lamellae at follicular infundibulum and acrosyringium inside the plaques cannot be explained by pure coincidence and may be more than fortuitous.

The recessive dystrophic form of epidermolysis bullosa (RDEB) is a disorder of incurable skin fragility and blistering caused by mutations in the type VII collagen gene (Col7a1). The absence of type VII collagen production leads to the loss of adhesion at the basement membrane zone due to the absence of anchoring fibrils, which are composed of type VII collagen. We report that wild-type, congenic bone marrow cells homed to damaged skin, produced type VII collagen protein and anchoring fibrils, ameliorated skin fragility, and reduced lethality in the murine model of RDEB generated by targeted Col7a1 disruption. These data provide the first evidence that a population of marrow cells can correct the basement membrane zone defect found in mice with RDEB and offer a potentially valuable approach for treatment of human RDEB and other extracellular matrix disorders.

Lamellar bodies (LBs) are tubulovesicular secretory organelles of epithelial cells related to lysosomes. In the epidermis, they play a crucial role in permeability barrier homeostasis, secreting their contents, lipids, a variety of hydrolases, protease inhibitors, and antimicrobial peptides, in the upper keratinocyte layers. The identification of proteins transported in epidermal LBs is still far from complete, and the way their secretion is controlled unknown. In this study, we describe the first proteomics characterization by nano-LC-MS/MS of a fraction enriched in epidermal LBs. We identified 984 proteins, including proteins known or thought to be secreted by LBs. Moreover 31 proteins corresponded to lysosomal components further suggesting that LBs are a new class of secretory lysosomes. Many of the newly found proteins could play a role in the epidermal barrier and desquamation (one acid ceramidase-like protein, apolipoproteins, glycosidases, protease inhibitors, and peptidases) and in LB trafficking (e.g. Rab, Arf, and motor complex proteins). We focus here on CLIP-170/restin, a protein that mediates interactions between organelles and microtubules. Western blotting confirmed the presence of CLIP-170 and its known effectors IQGAP1 and Cdc42 in the LB-enriched fraction. We showed, by confocal microscopy analysis of skin cryosections, that CLIP-170 was expressed in differentiated keratinocytes, first at the periphery of the nucleus then with a granular cytoplasmic labeling evocative of LBs. It was preferentially co-localized with Cdc42 and with the known LB protein cathepsin D. CLIP-170 was also largely co-localized with Rab7. This study strongly suggests a new function for CLIP-170, its involvement together with Cdc42 and/or Rab7 in the intracellular trafficking of LBs, and provides evidence that nano-LC-MS/MS combined with monodimensional electrophoresis separation constitutes a powerful method for identifying proteins in a complex mixture such as subcellular structures.

Controlled proteolytic degradation of specialized junctional structures, corneodesmosomes, by epidermal proteases is an essential process for physiological desquamation of the skin. Corneodesmosin (CDSN) is an extracellular component of corneodesmosomes and, although considerable debate still exists, genetic studies have suggested that the CDSN gene in the major psoriasis-susceptibility locus ( PSORS1 ) may be responsible for susceptibility to psoriasis, a human skin disorder characterized by excessive growth and aberrant differentiation of keratinocytes. CDSN is also expressed in the inner root sheath of hair follicles, and a heterozygous nonsense mutation of the CDSN gene in humans is associated with scalp-specific hair loss of poorly defined etiology. Here, we have investigated the pathogenetic roles of CDSN loss of function in the development of skin diseases by generating a mouse strain with targeted deletion of the Cdsn gene. Cdsn -deficient mouse skin showed detachment of the stratum corneum from the underlying granular layer and/or detachment within the upper granular layers due to the disrupted integrity of the corneodesmosomes. When grafted onto immunodeficient mice, Cdsn -deficient skin showed rapid hair loss together with epidermal abnormalities resembling psoriasis. These results underscore the essential roles of CDSN in hair physiology and suggest functional relevance of CDSN gene polymorphisms to psoriasis susceptibility.

The clinical and histopathological similarities of nodular melanoma and Spitz nevus currently still make a definitive diagnosis difficult. We report here a case of nodular melanoma that was extremely difficult to diagnose both clinically and histopathologically. The primary tumor was a blackish nodule on the scalp and biopsy was performed for pathological diagnosis. Although our first impression was malignant melanoma, we asked two dermatopathologists for second opinions; however, one diagnosed a melanoma and the other a Spitz nevus. Faced with this clinical dilemma, an operation was performed with sentinel node biopsy. Only one sentinel node suggested a metastasis. Histopathological diagnosis to establish whether it was a melanoma metastasis or nodal nevi was also difficult, and we again asked for second opinions from another dermatopathologist in the USA. According to its clinical course and the histopathology of the sentinel lymph node with additional immunohistochemistry, this case was finally diagnosed as a nodular melanoma (T4aN1aM0, stage IIIA). To date, the patient has been given five courses of chemotherapy at 6-month intervals, with no local recurrence or distant metastases so far.

The cornified layer is a compacted lattice of lipid-embedded corneocytes that provides an organism's barrier to the external environment. Cornification is the final differentiative step for epidermal keratinocytes and involves dramatic cell condensation before death. Using conditional gene deletion in mice, we identified the transcriptional repressor Blimp-1 (B lymphocyte-induced maturation protein-1) as an important regulator of keratinocyte transition from the granular to the cornified layer. More than 250 genes are misregulated in conditional knockout epidermis, including those encoding transcription factors, signal transduction components, proteinases, and enzymes involved in lipid metabolism. Steady-state mRNA and ChIP analyses of a subset of these genes provide evidence that nfat5 , fos , prdm1 , and dusp16 are novel direct targets of Blimp-1. Identifying nfat5 as a target of Blimp-1 repression indicates that cornification involves suppression of normal osmotic regulation in granular cells. Consistently, conditional knockout mice have delayed barrier formation as embryos, enlarged granular layer cells and corneocytes, and a morphologically abnormal cornified layer. These studies provide insight into cornification, identifying transcriptional regulatory circuitry and indicating the importance of blocking osmotic homeostasis.

Summary We describe a case of Hutchinson–Gilford progeria syndrome (HGPS) with long-term follow-up. A 1-month-old girl with marked sclerodermatous skin changes developed various symptoms of HGPS during follow-up. These included sclerotic skin, pigmentation, skin atrophy with translucent veins, wispy hair and alopecia, nail dystrophy and decreased sweating. Marked skin calcinosis was observed over almost the entire body, a symptom that has apparently been ignored in the literature. At 16 years old, the girl underwent surgery for a skull fracture and subdural haematoma, which was followed by chronic ulceration. Wet dressing with insulin-like growth factor was used with considerable effect. Mutation of the lamin A/C (LMNA) gene mutation, which encodes nuclear lamin A and C, has been reported to be the cause of HGPS. Our case showed the mutation G608G (GGCGGT), which resulted in a cryptic splice site and consequently in a truncated lamin A/C protein.

Although antitumor necrosis factor (anti-TNF)-alpha therapy provides beneficial effects on various chronic inflammatory skin diseases including psoriasis, cutaneous side-effects have also been reported. We describe four patients who showed psoriasiform eruption following anti-TNF-alpha therapy (infliximab) for Crohn's disease. In all patients, Crohn's disease had responded well to infliximab. Three patients developed plaque-type psoriasiform eruption on the trunk and four extremities, while one patient showed pustular eruption on the palms and soles accompanied with plaque-type psoriasis. In all these patients, skin lesions subsided with topical application of corticosteroid ointment or psoralen plus ultraviolet A treatment.

Summary Background Cystatin A is a cysteine proteinase inhibitor abundantly expressed in keratinocytes. Although cystatin A is one of the cornified cell envelope constituents and expressed in the upper epidermis, its precise function is still unknown. Ultraviolet B irradiation (UVB) induces apoptosis accompanied with the activation of cysteine proteinases, caspases. Objective We investigated the effect of cystatin A on UVB-induced apoptosis of keratinocytes. Methods We assessed the caspase activities and apoptotic cell numbers induced by UVB ittadiation in cystatin A gene transfected keratinocytes. Results UVB-induced pro-caspase 3 cleavage and caspase 3 activation were suppressed in cystatin A expression vector-transfected SV40-transformed human keratinocytes (SVHK). Furthermore, the transfected SVHK cells were resistant to UVB-induced apoptosis. In contrast neither caspase 8 nor caspase 9 activities were affected by UVB irradiation in cystatin A-transfected SVHK cells. The effects were also observed in cystatin A expression adenovirus vector-transfected cultured normal human keratinocytes (NHK). Conversely knockdown of cystatin A by si-RNA induced marked apoptosis of NHK cells following UVB irradiation accompanied with increased caspase 3 activity. In order to confirm the antiapoptotic effect of cystatin A in vivo UVB irradiation was performed on cystatin A transgenic mice (cystatin A-tg). The epidermis from cystatin A-tg was resistant to UVB-induced apoptosis compared to control mice epidermis. Conclusion These results indicate that cystatin A suppresses UVB-induced apoptosis of keratinocytes by the inhibition of caspase 3 activation.

Bcl-2 is a member of the large Bcl-2 family and protects cells from apoptosis. Ultraviolet B (UVB) irradiation induces apoptosis of keratinocytes that is known as "sunburn cells." Previously we reported that UVB irradiation induces apoptosis accompanied by sequential activation of caspase 8, 3 and 1 in keratinocytes, and that the process is inhibited by various caspase inhibitors. Using bcl-2-expressing adenovirus vector we investigated the effect of Bcl-2 on UVB-induced apoptosis. Adenovirus vector efficiently introduced bcl-2 gene in cultured normal mouse keratinocytes (NMK cells); almost all NMK cells (1 x 10(6)) were transfected at 1 x 10(8) plaque-forming unit (PFU)/mL. Bcl-2-transfected NMK cells were significantly resistant to UVB-induced apoptosis with the suppressive effect dependent on the Bcl-2 expression level. Following UVB irradiation caspase 8, 3 and 9 activities were stimulated in NMK cells, whereas in bcl-2-transfected cells only caspase 8, but not caspase 3 or 9, activity was stimulated. In order to investigate the effect of Bcl-2 in vivo topical application of Ad-bcl-2 on tape-stripped mouse skin was performed. Following the application Bcl-2 was efficiently overexpressed in almost all viable keratinocytes. The expression was transient with the maximal expression of Bcl-2 on the first day following the application of 1 x 10(9) PFU in 200 microL. The introduced Bcl-2 remained at least for 6 days. UVB irradiation (1250 J/m2) induced apoptosis within 12 h and the maximal effect was observed at 24 h in control mouse skin. Both bcl-2-transfected and topical caspase 3 inhibitor-treated mice skin were resistant to UVB-induced apoptosis. The suppressive effect of Bcl-2 was more potent than that of caspase 3 inhibitor application. Topical application of empty adenovirus vector alone had no effect on Bcl-2 expression or UVB-induced apoptosis. These results indicate that adenovirus vector is an efficient gene delivery system into keratinocytes and that Bcl-2 is a potent inhibitor of UVB-induced apoptosis both in vitro and in vivo.

In this article, we describe a novel autosomal recessive ichthyosis with hypotrichosis syndrome, characterized by congenital ichthyosis associated with abnormal hair. Using homozygosity mapping, we mapped the disease locus to 11q24.3-q25. We screened the ST14 gene, which encodes matriptase, since transplantation of skin from matriptase(-/-)-knockout mice onto adult athymic nude mice has been shown elsewhere to result in an ichthyosislike phenotype associated with almost complete absence of erupted pelage hairs. Mutation analysis revealed a missense mutation, G827R, in the highly conserved peptidase S1-S6 domain. Marked skin hyperkeratosis due to impaired degradation of the stratum corneum corneodesmosomes was observed in the affected individuals, which suggests that matriptase plays a significant role in epidermal desquamation.

Kallikrein type serine proteases, KLK8/neuropsin, KLK6, and KLK7, have been implicated in the proliferation and differentiation of epidermal keratinocytes and in the pathogenesis of psoriasis. However, their mechanistic roles in these processes remain largely unknown. We applied 12-O-tetradecanoylphorbol-13-acetate on the wild type (WT) and the Klk8 gene-disrupted (Klk8(-/-)) mouse skin, inducing keratinocyte proliferation similar to the human psoriatic lesion. Klk8 mRNA as well as Klk6 and Klk7 mRNA were up-regulated after 12-O-tetradecanoylphorbol-13-acetate application in the WT mice. In contrast, Klk8(-/-) mice showed minimum increases of Klk6 and Klk7 transcripts, the proteins, and enzymatic activities. Relative to the WT, the Klk8(-/-) skin showed less proliferation and an increase in the number of cell layers in the stratum corneum. However, overexpression of Klk8 by adenovirus vector in knock-out keratinocytes did not result in an increase in Klk6 or Klk7 mRNA. The inefficient cleavage of adhesion molecules DSG1 and CDSN in Klk8(-/-) skin contributes to a delay in corneocyte shedding, resulting in the hyperkeratosis phenotype. We propose that in psoriatic lesion, KLK8 modulates hyperproliferation and prevents excessive hyperkeratosis by shedding the corneocytes.

Contains fulltext : 51942.pdf (Publisher’s version ) (Closed access) Cystatin M/E is a cysteine protease inhibitor with two distinct binding sites for papain-like cysteine proteases (family C1) and the asparaginyl endopeptidase (AEP) legumain of family C13. We have previously demonstrated that deficiency of cystatin M/E in mice causes ichthyosiform skin changes and barrier disruption, which could be caused by unrestrained AEP activity. Recently, we provided biochemical evidence that human cathepsin V (CTSV) and cathepsin L (CTSL) are additional biological targets for human cystatin M/E. To address the possible role of these three proteases and their inhibitor in epidermal differentiation, we investigated the localization of these proteins in normal human skin. Whereas CTSL and AEP were broadly expressed in epithelial cells of the skin, we found a specific colocalization of cystatin M/E and CTSV in the stratum granulosum and in the root sheets of the hair follicle, using immunofluorescence microscopy. Immunoelectron microscopy revealed that cystatin M/E and CTSV are separately transported within the lamellar granules. Cystatin M/E was also found in the extracellular space in the stratum corneum associated with corneodesmosomes, where it was closely associated with CTSV. Based on the striking stratum-specific colocalization of cystatin M/E and CTSV, we propose that these molecules could have an important role in epidermal differentiation and desquamation.

Erythrokeratodermia variabilis et progressiva (EKVP) is a rare inherited skin disease characterized by fixed hyperkeratotic plaques and transient erythematous patches. EKVP is most often transmitted in an autosomal dominant manner. Causal mutations were found in the GJB3, GJB4 and GJA1 genes encoding connexins 31, 30.3 and 43, respectively. Approximately 50% of affected individuals develop palmoplantar keratoderma. Connexins are components of gap junctions, which are intercellular channels that are found in almost all tissues including the skin. Treatment of EKVP usually involves use of topical keratolytics and emollients resulting in some improvement in hyperkeratosis. Low-dose systemic retinoid may be beneficial. Novel therapies targeting connexin hemichannels and gap junctions may become available in the future.

Background Hypohidrosis/anhidrosis are congenital or acquired sweating impairments. Among them, acquired idiopathic generalized anhidrosis/hypohidrosis (AIGA) is the most common, and characterized by favourable response to systemic corticosteroid, however, no clinical markers for disease severity or activity have been developed. Objective Our aim was to verify the usefulness of serum carcinoembryonic antigen (CEA) level monitoring as a clinical marker for disease activity of AIGA. Methods Ten cases of AIGA diagnosed at Asahikawa Medical University, from 1980 to 2014 were included in the study. CEA and/or CEACAM1 expression level was analysed using immunohistochemistry and enzyme-linked immunosorbent assay. Result CEA expression was restricted to the apical membrane of glandular cells in eccrine sweat glands in most of the three types of cases we examined [healthy control, patients with atopic dermatitis (AD) or urticaria]. However, CEA expression was detected diffusely and much more intensively in eight of the 10 AIGA cases included in this study. CEACAM1-expression was much more restricted on the apical membrane of glandular cells of both the AIGA cases and the other control subjects. While serum CEA levels increased in all five AIGA cases examined (5.8–43.2 ng/mL), it remained within normal limits in all control subjects: nine healthy individuals; 10 cases of AD; 10 cases of idiopathic urticaria; four cases of normohidrotic cholinergic urticaria (Mann–Whitney's U-test, P < 0.05). The increased serum CEA levels in AIGA decreased in conjunction with improved sweating during methyl prednisolone pulse therapy or repeated bathing. Conclusion Serum CEA level may serve as a clinical marker for AIGA activity.

Hypohidrosis and anhidrosis are congenital or acquired conditions which are characterized by inadequate sweating. Acquired idiopathic generalized hypohidrosis/anhidrosis (AIGA) includes idiopathic pure sudomotor failure (IPSF), which has the following distinct features: sudden onset in youth, increased serum immunoglobulin E and responds favorably to systemic corticosteroid. No clinical markers reflecting the disease severity or activity have been established. Here, we report a case of AIGA in a Japanese patient successfully treated with repeated methylprednisolone pulse therapy. In this case, serum carcinoembryonic antigen (CEA) levels increased up to 19.8 ng/mL along with aberrant CEA immunoreactivity of eccrine sweat glands. Interestingly, the serum CEA level normalized as sweating improved with repeated methylprednisolone pulse therapy. Therefore, serum CEA level may serve as a useful clinical marker of hypohidrosis or anhidrosis.

Herpes zoster (HZ), a common vesiculo-erythematous skin disease associated with reactivation of varicella zoster virus in the cranial nerve, dorsal root, and autonomic ganglia, is accompanied by several related symptoms represented by postherpetic neuralgia. Among them, involvement of vesicorectal dysfunction is relatively rare. The vesicorectal symptom can usually be recovered in transient course, but is quite important in terms of impaired quality of life. Male individuals affected with HZ and skin lesions on sacral dermatome have been reported as independent risk factors of zoster-related voiding dysfunction. In this study, urinary symptoms were focused upon and six patients with zoster-related voiding dysfunction at a single faculty of dermatology in Japan from 2009 to 2014 were retrospectively analyzed. All patients showed HZ lesions on the sacral area and the urinary symptom recovered in approximately 2 months (14 days to 7 months). The term of treatment for zoster-associated urinary dysfunction was positively correlated with that for zoster-related pain without significance (r = 0.661, P = 0.153). Average treatment term for pain relief of sacral HZ accompanied by voiding dysfunction (91.3 ± 76.44 days) was significantly longer than that of sacral HZ without urinary symptom (18.9 ± 20.42 days) (P = 0.032). These results suggested that zoster-related voiding dysfunction would mainly be involved in sacral HZ and closely associated with severity of zoster-related pain. Dermatologists should be aware that severe zoster-related pain accompanied by sacral HZ, which is related to prolonged treatment of pain relief, can be a predictive factor of voiding dysfunction.

Naegeli-Franceschetti-Jadassohn syndrome (NFJS) and dermatopathia pigmentosa reticularis (DPR) are two closely related autosomal dominant ectodermal dysplasia syndromes that clinically share complete absence of dermatoglyphics (fingerprint lines), a reticulate pattern of skin hyperpigmentation, thickening of the palms and soles (palmoplantar keratoderma), abnormal sweating, and other subtle developmental anomalies of the teeth, hair, and skin. To decipher the molecular basis of these disorders, we studied one family with DPR and four families with NFJS. We initially reassessed linkage of NFJS/DPR to a previously established locus on 17q11.2-q21. Combined multipoint analysis generated a maximal LOD score of 8.3 at marker D17S800 at a recombination fraction of 0. The disease interval was found to harbor 230 genes, including a large cluster of keratin genes. Heterozygous nonsense or frameshift mutations in KRT14 were found to segregate with the disease trait in all five families. In contrast with KRT14 mutations affecting the central alpha -helical rod domain of keratin 14, which are known to cause epidermolysis bullosa simplex, NFJS/DPR-associated mutations were found in a region of the gene encoding the nonhelical head (E1/V1) domain and are predicted to result in very early termination of translation. These data suggest that KRT14 plays an important role during ontogenesis of dermatoglyphics and sweat glands. Among other functions, the N-terminal part of keratin molecules has been shown to confer protection against proapoptotic signals. Ultrastructural examination of patient skin biopsy specimens provided evidence for increased apoptotic activity in the basal cell layer where KRT14 is expressed, suggesting that apoptosis is an important mechanism in the pathogenesis of NFJS/DPR.

Recent study revealed that photodynamic therapy (PDT) with a novel photosensitizer (ATX-S10(Na)) shows more potent effects for various skin diseases than ALA-PDT. The effect of ATX-S10(Na)-PDT on dermal fibroblasts is still unknown. Using dermal fibroblasts derived from normal and scleroderma patients, and mouse skin in vivo, we compared the effects of ATX-S10(Na)-PDT and ALA-PDT. Fibroblasts from normal, scleroderma patients or mice skin were treated with ATX-S10(Na)-PDT or ALA-PDT. After the PDT treatments, the expression of matrix metalloproteinases (MMPs) Tissue inhibitors of metalloproteinases (TIMPs) and collagen synthesis was assayed using ELISA and reverse transcription-PCR (RT-PCR). The expression of MMP-1 and MMP-3 was slightly decreased and collagen I mRNA was significantly increased in scleroderma fibroblasts compared with normal fibroblasts. Both ATX-S10(Na)-PDT and ALA-PDT increased the expression of MMP-1 and MMP-3 in protein and mRNA levels in both normal and scleroderma fibroblasts with more potent effect by ATX-S10(N)-PDT. Collagen I synthesis was markedly decreased by ATX-S10(Na)-PDT and by ALA-PDT again with more potent effect by ATX-S10(Na)-PDT in both normal and scleroderma fibroblasts. In mice skin the effect of PDT for MMPs and collagen I was also detected and the effect was more potent in ATX-S10(Na)-PDT. In contrast, MMP-2, TIMP-1, TIMP-2, and collagen III expression was not affected by the ATX-S10(Na)-PDT or ALA-PDT treatment. ATX-S10(Na)-PDT is more potent modulator for dermal matrix components than ALA-PDT and might be useful for scleroderma patients.

In the course of a large scale analysis of late-expressed genes in the human epidermis, we identified a new member of the alpha(2)-macroglobulin (alpha2M) protease inhibitor family, A2ML1 (for alpha(2)-macroglobulin-like 1). Like A2M and PZP, A2ML1 is located on chromosome 12p13.31. A2ML1 encodes a protein of 1454 amino acids, which fits the characteristics of alpha2Ms: 1) strong conservation in amino acid sequence including most of cysteine positions with alpha2M; 2) a putative central bait domain; 3) a typical thiol ester sequence. Northern blot and reverse transcriptase-PCR studies revealed a single 5-kb A2ML1 mRNA, mainly in the epidermis granular keratinocytes. A2ML1 is also transcribed in placenta, thymus, and testis. By Western blot analysis, alpha2ML1 is detected as a monomeric, approximately 180-kDa protein in human epidermis. In vitro keratinocyte differentiation is associated with increased expression levels. By immunohistochemistry, alpha2ML1 was detected within keratinosomes in the granular layer of the epidermis, and as a secreted product in the extracellular space between the uppermost granular layer and the cornified layer. Recombinant alpha2ML1 displayed inhibitory activity toward chymotrypsin, papain, thermolysin, subtilisin A, and to a lesser extent, elastase but not trypsin. Incubation with chymotrypsin and the chymotrypsin-like kallikrein 7 protease indicated that alpha2ML1 binds covalently to these proteases, a feature shared with other members of the family. Therefore, alpha2ML1 is the first alpha2M family member detected in the epidermis. It may play an important role during desquamation by inhibiting extracellular proteases.

Neurocutaneous syndromes represent a vast, largely heterogeneous group of disorders characterized by neurological and dermatological manifestations, reflecting the common embryonic origin of epidermal and neural tissues. In the present report, we describe a novel neurocutaneous syndrome characterized by cerebral dysgenesis, neuropathy, ichthyosis, and keratoderma (CEDNIK syndrome). Using homozygosity mapping in two large families, we localized the disease gene to 22q11.2 and identified, in all patients, a 1-bp deletion in SNAP29, which codes for a SNARE protein involved in vesicle fusion. SNAP29 expression was decreased in the skin of the patients, resulting in abnormal maturation of lamellar granules and, as a consequence, in mislocation of epidermal lipids and proteases. These data underscore the importance of vesicle trafficking regulatory mechanisms for proper neuroectodermal differentiation.

Nature Publishing Group, Journal of Investigative Dermatology, 124, 2, 2005, 360-366 author, Lympho-epithelial Kazal-type-related inhibitor (LEKTI) is a putative serine protease inhibitor encoded by serine protease inhibitor Kazal-type 5 (SPINK5). It is strongly expressed in differentiated keratinocytes in normal skin but expression is markedly reduced or absent in Netherton syndrome (NS), a severe ichthyosis caused by SPINK5 mutations. At present, however, both the precise intracellular localization and biological roles of LEKTI are not known. To understand the functional role of LEKTI, we examined the localization of LEKTI together with kallikrein (KLK)7 and KLK5, possible targets of LEKTI, in the human epidermis, by confocal laser scanning microscopy and immunoelectron microscopy. In normal skin, LEKTI, KLK7, and KLK5 were all found in the lamellar granule (LG) system, but were separately localized. LEKTI was expressed earlier than KLK7 and KLK5. In NS skin, LEKTI was absent and an abnormal split in the superficial stratum granulosum was seen in three of four cases. Collectively, these results suggest that in normal skin the LG system transports and secretes LEKTI earlier than KLK7 and KLK5 preventing premature loss of stratum corneum integrity/cohesion. Our data provide new insights into the biological functions of LG and the pathogenesis of NS.

Summary Background: Epidermolysis bullosa simplex associated with muscular dystrophy is caused by plectin deficiency. Objective To report clinical, immunohistochemical, ultrastructural and molecular features of a 52-year-old Japanese patient affected with this disease, whose muscular disease had been followed-up for 27 years. Methods: We performed histopathological study, immunofluorescence, electron microscopic study and mutation detection analysis for plectin. Results: The patient developed blisters and erosions followed by nail deformity on the traumatized regions from birth. The skin lesions were continuously developed to date. The histopathological study showed subepidermal blister. Electron microscopic study showed blister formation inside the basal cells at the level just above the attachment plaque of hemidesmosome. Immunofluorescence showed complete loss of staining to plectin. The mutation analysis using protein truncation test and DNA sequencing revealed a C-to-T transition at nucleotide position 7006 of the plectin cDNA sequence, which lead a novel homozygous nonsense mutation (R2319X). Conclusion: From the above results, the diagnosis of epidermolysis bullosa simplex associated with muscular dystrophy was made. Slight muscular dystrophy was noticed at the age of 25 years. The muscular dystrophy gradually progressed and she could not walk at the age of 46 years. However, she can still breathe and swallow by herself. This is the patient of this disease with the longest follow-up, and may indicate the slow progress of muscular condition of this disease.

Summary Background: Epidermal keratinization involves various post-translational modifications including the deimination of arginine residues. Major deiminated proteins are derived from keratin K1. Two preferred deimination sites were identified in the V subdomain of mouse K1. An antibody against one of the deiminated peptide sequences (ACP) recognized deiminated mouse and human K1, and stained the cornified layers of human and infant mouse epidermis. ACP also stained the outermost layer of mouse embryonic epidermis. Western blotting revealed minor proteins showing strong ACP-positive signals in the mouse embryonic epidermal extract in which deiminated K1 derivatives were hardly detected. Objective: To characterize ACP-positive proteins expressed in mouse embryonic epidermis. Methods: ACP-positive proteins were isolated by preparative gel electrophoresis for N-terminal sequencing followed by blast searches for matching sequences in the protein and nucleotide database. Results: We obtained N-terminal sequences of two ACP-positive proteins. A cDNA clone in the est_mouse database has an open reading frame for 202 amino acid residues containing both sequenced peptides. The deduced sequence shows typical features of the N-terminal portion of type II keratins. A virtually identical sequence to this reading frame is present in a genomic contig of chromosome 15 on which keratin type II genes are clustered. Sequential searches for overlapping cDNA clones in the est_mouse database along with similar searches in the genomic contig formulated a hypothetical cDNA sequence encoding a putative protein of 572 amino acid residues tentatively called K1-emb. Conclusion: We predicted a sequence of novel type II keratin site-specifically deiminated in embryonic mouse epidermis.

Summary Although many antihistamines are now in clinical use, few studies directly compare their pharmacodynamic and sedative activities in humans in vivo. We designed a double-blind, placebo-controlled, crossover study to compare the inhibitory effects of bepotastine, cetirizine, fexofenadine, and olopatadine on histamine-induced flare-and-wheal response. Systemic sedative effects and impaired psychomotor activities by these drugs were also evaluated. Bepotastine (10 mg twice a day), cetirizine (10 mg once a day), fexofenadine (60 mg twice a day), and olopatadine (5 mg twice a day) or placebo was given in a double-blind manner to seven healthy volunteers before histamine challenge by iontophoresis. At 0, 1, 2, 4, 8, 12, and 24 h following the oral administration of these drugs, histamine iontophoresis-induced wheal-and-flare response was measured. Sedative effects by the drugs were also evaluated by a visual analogue scale for subjective sedation, and by word processor test for psychomotor activity. Each volunteer was tested with all of the drugs (including placebo), administered in a random order with a washout period of at least 1 week. Histamine iontophoresis induced marked wheal-and-flare response in all participants. Bepotastine, cetirizine, fexofenadine, and olopatadine yielded significant reduction of histamine-induced wheal-and-flare response compared to placebo (P

Epidermal remodeling is the concept that epidermal architecture is determined by a simple self-organizing mechanism; epidermal hyperproliferation constructs typical psoriatic architecture. This is based on the assumption that the enlargements in both the two-dimensional proliferative compartment (basal cell layer) and three-dimensional whole epidermal volume coexist. During this process, the dermal papillae become markedly, but passively, expanded by enlargement of the proliferative compartment. This creates a considerable shrinkage force against the crowded basal cell layer, which is forced to lose adherence to the dermal extracellular matrix (ECM). This results in anoikis, a type of apoptosis characterized by cell detachment, and, consequently, a markedly diminished epidermal turnover time in psoriasis. The papillary shrinkage force also explains the fact that dermal papillary height does not exceed a certain limit. At the cessation of hyperproliferation a normalisation remodeling takes place toward normal tissue architecture. Thus the concept of epidermal remodeling explains the self-organizing mechanism of the architectural change in psoriasis, which is essentially a reversible disorder depending on epidermal hyperproliferation.

The possible functional abnormalities in three different Darier disease-causing Ca(2+)-ATPase (SERCA2b) mutants, Ile(274) --> Val at the lumenal end of M3, Leu(321) --> Phe on the cytoplasmic part of M4, and Met(719) --> Ile in P domain, were explored, because they exhibited nearly normal expression and localization in COS-1 cells and the high ATPase and coupled Ca(2+) transport activities that were essentially identical (L321F) or slightly lower (I274V by approximately 35% and M719I by approximately 30%) as compared with those of the wild type. These mutations happened to be in Japanese patients found previously by us. Kinetic analyses revealed that each of the mutants possesses distinct types of abnormalities; M719I and L321F possess the 2-3-fold reduced affinity for cytoplasmic Ca(2+), whereas I274V possesses the normal high affinity. L321F exhibited also the remarkably reduced sensitivity to the feedback inhibition of the transport cycle by accumulated lumenal Ca(2+), as demonstrated with the effect of Ca(2+) ionophore on ATPase activity and more specifically with the effects of Ca(2+) (up to 50 mm) on the decay of phosphoenzyme intermediates. The results on I274V and M719I suggest that the physiological requirement for Ca(2+) homeostasis in keratinocytes to avoid haploinsufficiency is very strict, probably much more than considered previously. The insensitivity to lumenal Ca(2+) in L321F likely brings the lumenal Ca(2+) to an abnormally elevated level. The three mutants with their distinctively altered kinetic properties will thus likely cause different types of perturbation of intracellular Ca(2+) homeostasis, but nevertheless all types of perturbation result in Darier disease. It might be possible that the observed unique feature of L321F could possibly be associated with the specific symptoms in the pedigree with this mutation, neuropsychiatric disorder, and behavior problems. The results also provided further insight into the global nature of conformational changes of SERCAs for ATP-driven Ca(2+) transport.

The members of the plakin family of proteins serve as epidermal cytolinkers and components of cell-cell and cell-matrix adhesion complexes, i.e., desmosomes and hemidesmosomes, respectively. Periplakin is a recently characterized member of this family. Human and mouse periplakin genomic loci are conserved, and the proteins are highly homologous, suggesting a role for periplakin in vertebrate physiology. In order to evaluate the functional role of periplakin, we generated periplakin null mice through targeted homologous recombination of mouse embryonic stem cells, followed by development of Ppl(-/-) mice. Mice homozygous for the targeted allele were born in the expected Mendelian frequency, developed normally, possessed grossly normal epidermis and hair, and were healthy and fertile. The epidermal barrier appeared to develop normally during fetal days E15.5 to E16.5, and the cornified envelope and desmosomes in the newborn mice were ultrastructurally normal. No compensatory increase in the expression of other epithelial proteins was detected in the neonatal mouse epidermis lacking periplakin. Consequently, the primary role of periplakin may not relate to the physiology of the cornified cell envelope in epidermal keratinocytes but may reside in the challenges, which normal laboratory mice do not encounter.

Summary Background The sera of epidermolysis bullosa acquisita (EBA) react with type VII collagen, a major component of anchoring fibrils, in which the major epitopes have been considered to be present in the N-terminal noncollagenous (NC) 1 domain. Objectives To determine whether there are also epitopes in the C-terminal NC2 domain, and to determine their ultrastructural localization. Methods Immunoblotting using recombinant proteins of the NC1 and NC2 domains of type VII collagen, and postembedding immunoelectron microscopy. Results Twenty of 28 EBA sera tested reacted with the NC1 domain and eight sera reacted with the NC2 domain. The sera that reacted with the NC1 domain showed immunoreactivity within the lamina densa and the sera that reacted with the NC2 domain showed immunoreactivity in the dermis 300–360 nm below the lamina densa. Conclusions This study clearly identified the presence of epitopes in the NC2 domain, and showed that the epitope in the NC1 domain is present in the lamina densa and that the epitope in the NC2 domain is in the dermis below the lamina densa.

Nature Publishing Group, Journal of Investigative Dermatology, 122, 5, 2004, 1137-1144 author, Lamellar granules (LG) of the epidermis appear as discrete round or oblong shaped granules in classical transmission electron micrographs, but a recent cryo-transmission electron microscopy study has claimed that LG are in fact branched tubular structures. LG contain various cargoes including lipids, hydrolytic enzymes, and several other proteins. It is not known whether there are any differences in the timing of expression among them and whether they are sorted into the granules individually or collectively. In order to address these questions, we studied the expression of glucosylceramides (GlcCer), cathepsin D (CatD), corneodesmosin (Cdsn), kallikrein (KLK)7, and KLK8 in normal human epidermis using confocal laser scanning microscopy and immunoelectron microscopy. The results were consistent with the model that LG are parts of a branched tubular structure. In this structure, all the components were shown to be distributed as separate aggregates. In the trans-Golgi network (TGN), bulbous protrusions containing GlcCer, Cdsn, KLK7 and KLK8, and small CatD-positive vesicles were observed. The molecules were shown to be delivered to the apical region of granular keratinocytes. This study provides strong evidence for the sequential synthesis and independent trafficking of various LG cargoes, including for the first time CatD and KLK8, from TGN.

The keratinization process in psoriasis is a unique phenomenon. We have proposed an organized system for keratinization in psoriasis based on the recognition of early and late differentiation markers combined with premature cell death. The early differentiation markers, such as involucrin, small proline-rich proteins (SPRR), cystatin A and transglutaminase l, are more conspicuously expressed in psoriasis, while the late differentiation markers, such as profilaggrin and loricrin, are abolished. Keratinization markers that are not observed in the normal epidermis are also detected; these include SKALP/elafin as well as K6 and K16. With a markedly diminished turnover time, the psoriatic epidermis rapidly synthesizes differentiation markers that are mostly under the control of the protein kinase C-AP1 transcriptional control system. Because of the premature cell death, however, the late differentiation markers are not expressed. During the improvement of the lesion and the therefore longer turnover time, the late differentiation markers rapidly catch up to reveal their expression. This explains the rapid appearance of keratohyalin granules (profilaggrin) in the healing lesion of psoriasis. Thus the keratinization process in psoriasis can be explained by the accelerated keratinization combined with premature cell death. The keratinization process in psoriasis is unique, because both accelerated keratinization and premature cell death co-exist, resulting in the disappearance of late differentiation markers such as profilaggrin and loricrin. It is interesting to note that the premature cell death is also under the control of protein kinase C signaling.

Background: In addition to their antimicrobial action, roxithromycin (RXM), a new 14-membrane macrolide antibiotics, have immunomodulatory, anti-inflammatory and anti-oxidative activity. Ultraviolet B (UVB) irradiation induces reactive oxygen intermediates and apoptosis of keratinocytes. Objective : To examine the anti-apoptotic and anti-oxidative effect of RXM on UVB-irradiated keratinocytes. Methods : UVB-induced apoptosis was determined by cell death assay using crystal violet staining, and DNA fragmentation assay. Superoxide dismutase (SOD), glutathione peroxidase (GPx), glutathione reductase (GR), and calatase activities were measured in UVB-irradiated SV40-trasnformed human keratinocytes (SVHK cells). Detection of superoxide was performed histologically using hydroethidine and colorimetric quantitative assay using ferrous irons. H 2 O 2 was measured by colorimetrical assay. Results : RXM suppressed UVB-induced apoptosis of SVHK cells. UVB-irradiated SVHK cells showed decreased SOD, GPx, GR, and catalase activities. RXM pretreatment suppressed the decrease in these enzyme activities with the maximal effect detected at 10 μM of RXM. The effect was associated with suppression of UVB-induced superoxide and H 2 O 2 production. Conclusion : The present study demonstrated that RXM has anti-apoptotic and anti-oxidative effects against UVB-irradiated keratinocytes.