Availability of mRNA Obtained from Peripheral Blood Mononuclear Cells for Testing Mutation Consequences in Dystrophic Epidermolysis Bullosa.

Dystrophic epidermolysis bullosa (DEB) is an inheritable blistering disease caused by mutations in COL7A1, which encodes type VII collagen. To address the issue of genotype-phenotype correlations in DEB, analyzing the consequences of COL7A1 mutations using mRNA is indispensable. Herein we established a novel method for testing the effect of mutations in DEB using COL7A1 mRNA extracted from peripheral blood mononuclear cells (PBMCs). We investigated the consequences of four COL7A1 mutations (c.6573 + 1G > C, c.6216 + 5G > T, c.7270C > T and c.2527C > T) in three Japanese individuals with recessive DEB. The novel method detected the consequences of two recurrent COL7A1 mutations (c.6573 + 1G > C, c.6216 + 5G > T) and a novel COL7A1 mutation (c.7270C > T) accurately. In addition, it detected aberrant splicing resulting from a COL7A1 mutation (c.2527C > T) which was previously reported as a nonsense mutation. Furthermore, we revealed that type VII collagen-expressing cells in PBMCs have similar cell surface markers as mesenchymal stem cells; they were CD105+, CD29+, CD45-, and CD34-, suggesting that a small number of mesenchymal stem cells or mesenchymal stromal cells are circulating in the peripheral blood, which enables us to detect COL7A1 mRNA in PBMCs. Taken together, our novel method for analyzing mutation consequences using mRNA obtained from PBMCs in DEB will significantly contribute to genetic diagnoses and novel therapies for DEB.

Multiple Milia as an Isolated Skin Manifestation of Dominant Dystrophic Epidermolysis Bullosa: Evidence of Phenotypic Variability.

We report a Japanese pedigree with dominant dystrophic epidermolysis bullosa (DDEB) harboring the p.G2251E mutation of COL7A1. The proband of this pedigree presented with multiple milia as an isolated skin manifestation without a history of blistering and subsequently developed generalized intractable blisters, suggesting that multiple milia could be a primary manifestation of DDEB. Her mother exhibited nail dystrophy and pruritic nodules and her elder sister was unaffected, despite having the same COL7A1 mutation. Inter- and intrafamilial clinical variability are often observed in DDEB, so we should be aware of this factor to provide appropriate genetic counselling.

Acquired cutis laxa secondary to acute generalized exanthematous pustulosis: A case report and mini-review of literature.

Cutis laxa presents as loose redundant skin folds and loss of dermal elastic tissue. Acquired cutis laxa (ACL) is characterized by later onset. It has been reported in association with various kinds of neutrophilic dermatoses, drugs, metabolic disorders, and autoimmune disorders. Acute generalized exanthematous pustulosis (AGEP) is usually classified as a severe cutaneous adverse reaction characterized by T cell-mediated neutrophilic inflammation. We previously reported a mild case of AGEP caused by gemcitabine in a 76-year-old man. Here, we report a case of ACL secondary to AGEP in this patient. He developed AGEP 8 days after gemcitabine administration. Four weeks after beginning chemotherapy, his skin had become atrophic, loose, and darkly pigmented in areas previously affected by AGEP. Histopathological examination revealed edema and perivascular lymphocytic infiltration but no neutrophilic infiltration in the upper dermis. Elastica van Gieson staining showed that the elastic fibers in all layers of the dermis were sparse and shortened. Electron microscopy showed elevated numbers of fibroblasts and altered elastic fibers with irregular surfaces. Finally, he was diagnosed with ACL secondary to AGEP. He was treated with topical corticosteroids and oral antihistamines. Skin atrophy decreased over 3 months. We summarize 36 cases (including our case) with ACL secondary to neutrophilic dermatosis. We discuss these clinical manifestations, causative neutrophilic disorders, treatments, and outcomes. The mean age of patients was 3.5 years. Five patients had an aortic lesion as systemic involvement. The most common causative neutrophilic disorders were Sweet syndrome (24 cases), followed by urticaria-like neutrophilic dermatosis (11 cases). There were no cases of AGEP except for our case. Although treatment for ACL secondary to neutrophilic dermatosis, such as dapsone, oral prednisolone, adalimumab, and plastic surgery were reported, ACL is generally refractory and irreversible. Our patient was considered reversibly cured due to the absence of continuous neutrophil-mediated elastolysis.

Upregulated expression of glucose transporter isoform 1 in invasive and metastatic extramammary Paget's disease.

Glucose transporter isoform 1 (GLUT1), which is upregulated in a variety of malignant tumors, facilitates cellular glucose uptake to boost rapid tumor growth and progression. In several types of cancer, inhibition of GLUT1 suppresses tumor proliferation and metastasis, indicating that GLUT1 is a potential target of anticancer therapy. The present study performed immunohistochemistry to analyze GLUT1 expression levels in 51 patients with extramammary Paget's disease (EMPD), including 23 with only intraepidermal lesions and 28 with dermal-invasive lesions. Of the 28 patients with dermal invasion, nine had available samples of lymph node metastasis. GLUT1 staining scores were significantly higher in dermal-invasive (P<0.0001) and metastatic lesions (P=0.0008) compared with in intraepidermal lesions. GLUT1 is upregulated during the transition from preinvasive to invasive or metastatic tumor in EMPD. Moreover, GLUT1 staining scores were statistically higher in intraepidermal tumor cells of dermal-invasive EMPD compared with tumor cells of only in situ EMPD (P=0.0338). GLUT1 is upregulated even during the preinvasive phase in patients with invasive EMPD, suggesting that GLUT1 immunostaining can predict the risk of dermal invasion. The present study provides novel evidence to pursue in vitro and in vivo studies to confirm that upregulated expression of GLUT1 enhances tumor aggressiveness in EMPD.

Case report: Exploring autosomal recessive woolly hair: genetic and scanning electron microscopic perspectives on a Japanese patient.

Woolly hair (WH) is a hair shaft anomaly characterized by tightly curled hair that typically stops growing at a few inches. Autosomal recessive WH (ARWH; OMIM no. 278150/604379/616760) has been reported to be caused by variants in genes coding lysophosphatidic acid receptor 6 (LPAR6), lipase H (LIPH), or keratin 25 (KRT25). In this study, we conducted a scanning electron microscopic (SEM) examination of the hair of a 3-year-old Japanese ARWH patient. The SEM revealed that her affected hair had an irregular and rough cuticle compared to her mother's hair. Many irregular small projections and longitudinal grooves were seen on the surface of the patient's hair shaft, and some free margins of the hair cortex were raised or serrated. Her hairs were oval-shaped on the cross-section. Mutation analysis revealed a homozygous pathogenic variant (c.736 T > A; Cys246Ser) in exon 6 in LIPH. In our clinic, we identified three additional cases with the homozygous Cys246Ser variant and one case with compound heterozygous variants in LIPH: Cys246Ser and c.671C > G (Pro224Arg). Consequently, genetic analyses, including genotype-phenotype correlation involving rare LIPH variants, have become more crucial in the Japanese population.

Dystrophic calcinosis cutis in a patient with cutaneous sarcoidosis in remission.

A 65-year-old Japanese woman was referred to our department because of a 5-month history of asymptomatic papules on the face. She was diagnosed with cutaneous sarcoidosis on the face 20 years ago. All of the lesions had completely disappeared with oral corticosteroids. Twenty years after the diagnosis of sarcoidosis, small papules developed in areas where the cutaneous sarcoidosis had been located. Physical examination revealed four yellow-white papules on the face. Dermoscopy revealed a homogenous, round, and yellow-white lesion. Serum levels of calcium and phosphorus were normal. Histopathology demonstrated calcium deposits in the dermis surrounded by inflammatory infiltrates without sarcoid granulomas. We made a diagnosis of calcinosis cutis. Basal cell carcinoma with calcinosis cutis, milia-like calcinosis cutis, and subcutaneous calcified nodule should be differentiated. Calcinosis cutis can be classified into four subtypes based on pathogenesis: dystrophic, metastatic, idiopathic, and iatrogenic. Dystrophic calcinosis cutis is caused by local tissue damage or abnormalities. Whereas, metastatic calcinosis cutis is often associated with hypercalcaemia, hyperphosphatemia, or hyperparathyroidism. There are reported cases of metastatic calcinosis cutis associated with sarcoidosis because patients with sarcoidosis often present with hypercalcaemia. However, dystrophic calcinosis cutis associated with sarcoidosis has been rarely reported. In the present case, systemic treatment for sarcoidosis may have degraded sarcoid granulomas and yielded necrotic tissue and dermal fibrosis, which might have induced ectopic calcification. Thus, we thought the present case consisted of dystrophic calcinosis cutis that developed in areas with cutaneous sarcoidosis in remission.

Two cases of cutaneous-type pemphigus vulgaris and a case of pemphigus foliaceus without mucosal involvement despite high anti-desmoglein 3 autoantibody levels.

Pemphigus is an autoimmune blistering disease with two major subtypes, pemphigus vulgaris (PV) and pemphigus foliaceus (PF). Although most patients with PV show oral lesions, cutaneous type PV (C-PV) is a rare subtype clinically characterized by predominant cutaneous involvement with no or subtle mucosal lesions. Patients with PF present with only skin involvement; they do not have mucosal lesions. Serologically, autoantibodies against desmoglein (Dsg) 3 and Dsg1 are observed in C-PV whereas PF is associated with anti-Dsg1 antibodies only. Herein, we describe three cases of pemphigus presenting with predominant skin lesions and no mucosal involvement despite high anti-Dsg 3 autoantibody levels in chemiluminescent enzyme immune assays (CLEIAs). In addition, anti-Dsg 1 autoantibodies were positive in patients 2 and 3, but negative in patient 1 based on CLEIAs. Histological examination of the skin showed suprabasal acantholysis in patients 1 and 2, and blister formation in the upper epidermis in patient 3. Histopathology of the oral membrane in patients 1 and 2 showed subtle acantholysis in the suprabasal layer. Thus, we diagnosed patients 1 and 2 as having cutaneous type PV and patient 3 as having PF. Ethylenediaminetetraacetic acid-treated enzyme-linked immunosorbent assay demonstrated a low proportion of anti-Dsg3 autoantibodies recognizing Ca2+ -dependent epitopes, antibodies against which are thought to be the main contributor to acantholysis. Thus, along with Dsg1 antibodies, weak anti-Dsg3 antibodies could induce acantholysis in the skin, but they are insufficient to induce mucosal lesions.

Case report: A case of epidermolysis bullosa acquisita with IgG and IgM anti-basement membrane zone antibodies relapsed after COVID-19 mRNA vaccination.

We report a case of autoimmune bullous disease (AIBD) with IgG and IgM autoantibodies against epidermal basement membrane zone (BMZ), which showed recurrence of mucocutaneous lesions after coronavirus disease 2019 (COVID-19) mRNA vaccination. A 20-year-old Japanese woman with a 4-year history of epidermolysis bullosa acquisita (EBA) presented to our clinic. She noticed fever and rash on the same day and visited at our hospital 2 days later. Physical examination revealed blisters, erosions and erythema on the face, shoulder, back, upper arms, and lower lip. A skin biopsy from the forehead showed subepidermal blister. Direct immunofluorescence showed linear depositions of IgG, IgM, and C3c in the epidermal BMZ. By indirect immunofluorescence of 1M NaCl-split normal human skin, circulating IgG autoantibodies were bound to the dermal side of the split at 1:40 serum dilution, and circulating IgM antibodies were bound to the epidermal side of the spilt. After the increase of prednisolone dose to 15 mg/day, the mucocutaneous lesions resolved in a week. The present case is the first case of possible EBA with IgG and IgM anti-BMZ antibodies, in which the mucocutaneous lesions were recurred after COVID-19 mRNA vaccination. Clinicians should be aware that bullous pemphigoid-like AIBDs, including EBA and IgM pemphigoid, might be developed after COVID-19 mRNA vaccination.

Pembrolizumab-Induced Lichen Planus on the Scalp of a Patient with Non-Small-Cell Lung Carcinoma.

A 67-year-old man with non-small-cell lung carcinoma was referred to our department because of a pruritic rash on his head and upper extremities. Prior to the development of the rash, he had received 4 cycles of combination therapy with pemetrexed, carboplatin, and pembrolizumab, followed by 2 cycles of pembrolizumab monotherapy. On physical examination, violaceous scaly erythema grouped on his scalp and upper extremities. Histologically, the scalp lesions demonstrated irregular acanthosis that formed a characteristic saw-tooth appearance with hypergranulosis and typical lichenoid tissue reaction. These findings suggested that the scalp lesions were lichen planus. Two-week administration of topical corticosteroid dramatically improved the rash. Immunotherapy with pembrolizumab, an anti-PD-1 antibody, can induce T-cell activation that results in various immune-related adverse effects such as lichenoid tissue reaction. However, lichen planus is generally found on the extremities and/or oral mucosa, and unlike in this case, the scalp is rarely affected. Although the exact mechanism underlying predominant scalp involvement is unknown, the present case indicates that anti-PD-1 therapy-induced lichen planus can develop not only on the extremities and oral mucosa but also on the scalp. Interestingly, the lesions were not induced by the combination of chemotherapy and pembrolizumab; rather, they occurred soon after initiation of pembrolizumab monotherapy. In the present case, pembrolizumab-induced T-cell activation which triggered lichenoid tissue reaction may have been suppressed by chemotherapy-induced immunosuppression. Dermatologists should have a thorough knowledge of the cutaneous lesions that manifest as irAEs of anti-PD-1 therapy.

Diversity of Mechanisms Underlying Latent TGF-ß Activation in Recessive Dystrophic Epidermolysis Bullosa.

Injury- and inflammation-driven progressive dermal fibrosis is a severe manifestation of recessive dystrophic epidermolysis bullosa-a genetic skin blistering disease caused by mutations in COL7A1. TGF-ß activation plays a prominent part in progressing dermal fibrosis. However, the underlying mechanisms are not fully elucidated. TGF-ß is secreted in a latent form, which has to be activated for its biological functions. In this study, we determined that recessive dystrophic epidermolysis bullosa fibroblasts have an enhanced capacity to activate the latent form. Mechanistic and functional assessment demonstrated that this process depends on multiple latent TGF-ß activators, including TSP-1, RGD-binding integrins, matrix metalloproteinases, and ROS, which act in concert, in a self-perpetuating feedback loop to progress fibrosis. Importantly, our study also disclosed keratinocytes as prominent facilitators of fibrosis in recessive dystrophic epidermolysis bullosa. They stimulate microenvironmental latent TGF-ß activation through enhanced production of the above mediators. Collectively, our study provides data on the molecular mechanism behind dysregulated TGF-ß signaling in recessive dystrophic epidermolysis bullosa, which are much needed for the development of evidence-based fibrosis-delaying treatments.

Abnormal keratinization and cutaneous inflammation in Mal de Meleda.

Mal de Meleda (MDM) is a rare, autosomal recessive form of palmoplantar keratoderma due to mutations in the gene, encoding for secreted lymphocyte antigen 6/urokinase-type plasminogen activator receptor related protein 1 (SLURP1). We report a four-year-old Taiwanese MDM female case whose biopsy specimen of hyperkeratotic lesions showed abnormal keratinization and cutaneous inflammation with characteristic transmission electron microscopic (TEM) findings and immunostaining results. The patient presented with pruritic and severely hyperkeratotic plaques on the bilateral palms and soles whichwere fringed with erythematous scaly areas. A homozygous c.256 G>A mutation, predicting a conversion of p.Gly86Arg, in SLURP1gene was detected. Histopathological examinations showed marked hyperkeratosis, acanthosis and hypergranulosis in the epidermis, accompanied by perivascular lymphocytic infiltrates in the dermis. The whole layers of the epidermis and perivascular infiltrates of the dermis were stained positive with anti-tumor necrosis factor alpha (TNFα) antibody in the biopsy specimen from the sole and the ankle. TEM examination of the biopsy specimen from the plantar hyperkeratotic plaque showed various-sized vacuoles surrounding nuclei of many keratinocytes in the spinous layer. In addition, there were numerous irregular keratohyaline granules in the granular layer. Several microorganisms and many lipid-like droplets were found in the thickened cornified layer. SLURP1 protein is known as a marker of late differentiation, predominantly expressed in the granular layer, and also known to have an inhibitory effect on TNFα release. Our results exhibited excessive TNFα expression in keratinocytes and perivascular infiltrates of the dermis, and several characteristic morphological observations of keratinocytes in MDM.

Epistatic connections between microphthalmia-associated transcription factor and endothelin signaling in Waardenburg syndrome and other pigmentary disorders.

Waardenburg syndrome (WS) is an inherited sensorineural deafness condition in humans caused by melanocyte deficiencies in the inner ear and forelock. Mutation of microphthalmia-associated transcription factor (MITF) is known to produce WS type IIA whereas mutations of either endothelin (EDN) or its receptor endothelin receptor B (EDNRB) produce WS type IV. However, a link between MITF haploinsufficiency and EDN signaling has not yet been established. Here we demonstrate mechanistic connections between EDN and MITF and their functional importance in melanocytes. Addition of EDN to cultured human melanocytes stimulated the phosphorylation of MITF in an EDNRB-dependent manner, which was completely abolished by mitogen-activated protein kinase kinase inhibition. The expression of melanocyte-specific MITF mRNA transcripts was markedly augmented after incubation with EDN1 and was followed by increased expression of MITF protein. Up-regulated expression of MITF was found to be mediated via both the mitogen-activated protein kinase-p90 ribosomal S6 kinase-cAMP response element-binding protein (CREB) and cAMP-protein kinase A-CREB pathways. In addition, EDNRB expression itself was seen to be dependent on MITF. The functional importance of these connections is illustrated by the ability of EDN to stimulate expression of melanocytic pigmentation and proliferation markers in an MITF-dependent fashion. Collectively these data provide mechanistic and epistatic links between MITF and EDN/EDNRB, critical melanocytic survival factors and WS genes.

Cross-sectional survey on disease severity in Japanese patients with harlequin ichthyosis/ichthyosis: Syndromic forms and quality-of-life analysis in a subgroup.

BACKGROUND: Congenital ichthyoses (CIs) adversely affect quality of life (QOL) in patients. However, the effects of CIs on patient QOL have not been studied sufficiently. OBJECTIVE: To investigate the association between disease severity and QOL in patients with harlequin ichthyosis (HI) and ichthyosis: syndromic forms (ISFs) METHODS: Clinical information of patients with HI and ISFs from 2010 to 2015 were obtained from 100 dermatology departments/divisions of principal institutes/hospitals throughout Japan. We examined the relationship between disease severity and QOL in patients with HI and ISFs. Patients who were aged 8 years or older and participated in a multicenter retrospective questionnaire survey in Japan were assessed by dermatology life quality index (DLQI, range of 0-30) and clinical ichthyosis score (range of 0-100). RESULTS: Netherton syndrome patients had a significantly higher risk of allergy to food or environmental allergens than patients with other phenotypes. Keratitis-ichthyosis-deafness (KID) syndrome patients showed a significantly higher risk of skin infections than patients with other phenotypes. Complete data on DLQI were obtained from 13 patients, whose median age was 21 (8-71) years. Nine patients were male, and 4 were female. Systemic retinoids were administrated to 2 of the 3 HI patients. The Spearman's correlation coefficient between the clinical ichthyosis score and DLQI was 0.611 (P < 0.05). CONCLUSION: We confirmed that Netherton syndrome and KID syndrome patients have a higher risk of allergy to food or environmental allergens and of skin infections, respectively. QOL impairment correlates with disease severity in HI and ISFs patients.