Wataru Fujimoto

Confocal laser scanning microscopic observation of glycocalyx production by Staphylococcus aureus in skin lesions of bullous impetigo, atopic dermatitis and pemphigus foliaceus

Summary Background Glycocalyx collapses during dehydration to produce electron‐dense accretions. Confocal laser scanning microscopy (CLSM) may be used to visualize fully hydrated microbial biofilms.

Expression of keratins (K10 and K17) in steatocystoma multiplex, eruptive vellus hair cysts, and epidermoid and trichilemmal cysts.

We compared the patterns of keratin 10 (K10) and keratin 17 (K17) expression in epidermoid cysts, trichilemmal cysts, eruptive vellus hair cysts, and steatocystoma multiplex. Epidermoid cysts expressed K10 and eruptive vellus hair cysts expressed K17, whereas trichilemmal cysts and steatocystoma multiplex showed expression of both K10 and K17. Our findings support the opinion that eruptive vellus hair cysts, which stained negative for K10, and steatocystoma multiplex are distinct entities and not variants of one disorder.

Bullous Systemic Lupus Erythematosus as an Initial Manifestation of SLE

Bullous systemic lupus erythematosus (BSLE) is a rare subset of systemic lupus erythematosus that is often associated with autoimmunity to type VII collagen. We describe a 45‐year‐old woman with BSLE who presented with vesiculobullous lesions as an initial manifestation of SLE. The patient first noticed a widespread urticarial, erythematous eruption associated with tense blisters, erosions, and crusting. She was diagnosed with bullous pemphigoid and underwent a one‐month course of treatment with betamethazone. Because of the appearance of marked proteinuria, a subsequent renal biopsy, and serological tests, the patient was diagnosed with rapidly progressive glomerulonephritis and systemic lupus erythematosus. The patients IgG circulating antibodies labeled the dermal floor of salt‐split skin and recognized type VII collagen in immunoblot studies. Although methylprednisolone pulse therapy for glomerulonephritis did not alleviate the vesicullobullous eruption, treatment with dapsone resulted in dramatic disappearance of the lesions. Cessation of dapsone therapy due to hemolysis with Heinz‐body formation did not aggravate the bullous disease. Our case illustrates that a generalized vesiculobullous eruption can be the sole presenting manifestation of SLE. It also emphasizes the close temporal relationship between BSLE and lupus nephritis.

Environmental factors producing autoimmune dysregulation - Chronic activation of T cells caused by silica exposure

Autoimmune disorders are induced by various environmental and occupational substances. Among the most typical factors involving these substances, it is well known that silica exposure causes not only pulmonary fibrosis known as silicosis, but also induces autoimmune diseases such as rheumatoid arthritis known as Caplans syndrome, systemic sclerosis, systemic lupus erythematosus, and anti-neutrophil cytoplasmic autoantibody (ANCA)-related vasculitis/nephritis. To investigate the immunological effects of silica, a focus on the occurrence of autoimmune dysfunction may clarify these autoimmune diseases and develop effective tools for observing silicosis patients (SIL). In this review, our investigation concerns the autoantibodies found in SIL, alteration of CD95/Fas and related molecules in SIL, case-oriented and in vitro analyses of silica-induced activation of responder and regulatory T cells, and supposed mechanisms of reduction of CD4+25+FoxP3+ regulatory T cells (T(reg)) in SIL. Further studies are required to investigate Th17 and the interaction with T(reg) in SIL to understand the cellular and molecular mechanisms of environmental and occupational autoimmune disorders.

Reductive alteration of the regulatory function of the CD4(+)CD25(+) T cell fraction in silicosis patients.

Causal links have been documented between silica and rheumatoid arthritis, lupus erythematosus, systemic sclerosis and glomerulonephritis. Two different effects of silica have been suggested, an enhanced inflammatory response in the pulmonary region (e.g. activation of alveolar macrophages) and dysregulation of autoimmunity. Based on our previous reports showing in vitro activation of peripheral T cells by silica and reduced regulatory function of the peripheral CD4+CD25+ fraction in which FoxP3+ regulatory T cells (Treg) are located, reconstitution of the CD4+CD25+ fraction in silicosis patients (SILs) was investigated. Since T cells in peripheral CD4+CD25+ and CD4+CD25− (effector T cells; Teff) fractions from SILs showed higher expression of pd-1 (a marker gene for T cell activation) in comparison to that of healthy donors (HDs), chronic T cell activation was considered to have occurred in SILs. In this study, a higher expression of the CD95/Fas molecule in Treg was recorded from silicosis patients (SILs) compared to healthy donors (HDs), and excess loss of FoxP3+ Treg in freshly isolated peripheral blood mononuclear cells (PBMCs) from SILs relative to HDs was demonstrated when these cells were cultured with silica ex vivo, whereas CD25+ cells were not reduced due to contamination of activated Teff in the CD4+CD25+ fraction. The activation of both Teff and Treg results in reconstitution of the peripheral CD4+ CD25+ fraction, loss of Treg and contamination of activated Teff, resulting in reduction of the number and function of Treg. These results contribute to our understanding of the development of autoimmune diseases found in SILs.

Sentinel lymph node detection in skin cancer using fluorescence navigation with indocyanine green

Figure 1. (a) Indocyanine green injection around the left big toe. (b) Fluorescence at the injected site: The lymphatic vessels were clearly observed. Dear Editor, Sentinel lymph node (SLN) biopsy has been used to determine the regional lymph node status of patients with various cancers. Two methods are available for detecting an SLN, dye and ⁄or an isotope with gamma probe. Patent blue has often been used in the dye-guided method of SLN biopsy, although it is not approved as a diagnostic reagent because of its possible carcinogenesis. In the area of breast cancer surgery, indocyanine green (ICG) is often used due to its safety profile. Injected ICG binds to albumin and generates intensive fluorescence when excited with near-infrared wavelength light. Recently, SLN biopsy in skin cancer patients using real-time fluorescence navigation with ICG has been reported. We report our experience of SLN detection with this new method. Six skin cancer patients (four with malignant melanoma, one with squamous cell carcinoma and one with extramammary Paget’s disease) were studied. A total of 1 mL of ICG (5–8 mg ⁄mL) was injected i.d. at various sites around the tumor, and fluorescence images were obtained using the ICG fluorescence system (PDE; Hamamatsu Photonics, Hamamatsu, Japan). In all cases, the gamma probe-guided method with technetium-m-stannous phytate was used in combination with the ICG. The SLN were successfully identified in all patients using fluorescence navigation with ICG. A case of a malignant melanoma is presented in Figures 1 and 2. All the lymph nodes with ICG fluorescence corresponded well to the isotope-positive lymph nodes. Conversely, all the SLN identified by the gamma probe-guided method yielded ICG fluorescence. On the other hand, no lymph nodes were detected with ICG fluorescence alone. There were no metastases in any of the SLN of the six patients. No adverse effects were observed.

Detection, epitope-mapping and function of anti-Fas autoantibody in patients with silicosis

Dysregulation of apoptosis through the Fas–Fas ligand pathway is associated with the onset of autoimmune disease. Since autoantibodies directed against unknown antigens are present in the sera of these patients, sera samples were examined for the presence of autoantibodies directed against the Fas molecule. Using Western blotting and a ProteinChip analysis, autoantibodies against Fas were detected in patients with silicosis, systemic lupus erythematosus (SLE) and systemic sclerosis (SSc), and weakly detected in healthy individuals. Using epitope mapping employing 12‐amino‐acid polypeptides with the SPOTs system, a minimum of four epitopes and a maximum of 10 epitopes were found. Several amino acid residues involved in binding FasL, such as C66, R87, L90, E93 and H126, were presented within the epitopes. Serum containing a large amount of anti‐Fas autoantibody from silicosis patients inhibited the growth of a Fas‐expressing human cell line, but did not inhibit the growth of a low Fas‐expresser nor a Fas‐expresser in which the Fas gene had been silenced by small interference RNA. All epitopes in the intracellular region of Fas were located in the death domain. The possible roles of anti‐Fas autoantibody detected in healthy volunteers and patients with silicosis or autoimmune diseases are discussed here.

Expression of retinoic acid receptor genes in keratinizing front of skin

We found, by an in situ hybridization method with riboprobes synthesized from human cDNA of the retinoic acid receptor (RAR), that the RAR genes (predominantly γ‐subtype) are intensively expressed in the epidermis of normal and psoriasic human skins, and also in keratinizing fronts of 4‐day‐old mouse skins, nail matrices and hair follicles. Thus, target cells of retinoic acid in the skins are concluded to be keratinocytes, which is quite consistent with the fact that retinoic acid regulates keratinization of epidermis in vivo and also modulates expression of the keratin gene in vitro.

Amicrobial pustular dermatosis in two patients with immunological abnormalities

Summary We report two patients with severe amicrobial pustular dermatosis with immunological abnormalities: a 63‐year‐old woman with a 30‐year‐history of discoid lupus erythematosus and sicca syndrome, and a 35‐year‐old woman with high levels of γ‐globulinemia and positive antinuclear antibodies. Both patients presented with crusty and eroded erythematous plaques studded with aseptic pustuleson the back, face, and scalp. Histological examination showed acanthosis, neutrophilic exocytosis to the epidermis, and neutrophilic and lymphocytic infiltration with nuclear dust in the dermis. These patients were diagnosed as having ‘amicrobial pustulosis associated with autoimmune diseases’. The eruptions improved with combination treatment of oral prednisolone with cyclosporin A or diaminodiphenylsulphone. Although the pathogenesis remains unclear, amicrobial pustular dermatosis might be one of the cutaneous complications in autoimmune diseases.

Silica exposure and altered regulation of autoimmunity

Silica particles and asbestos fibers, which are known as typical causatives of pneumoconiosis, induce lung fibrosis. Moreover, silicosis patients often complicate with autoimmune diseases, and asbestos-exposed patients suffer from malignant diseases such as pleural mesothelioma and lung cancer. We have been conducting experimental studies to investigate altered regulation of self-tolerance caused by silica exposure, including analyses using specimens such as plasma and immunocompetent cells obtained from silicosis patients, as a means of examining the supposition that silica exposure induces molecular and cellular biological alterations of immune cells. These approaches have resulted in the detection of several specific autoantibodies, alterations of CD95/Fas and its related molecules, and evidence of chronic activation of responder T cells and regulatory T cells following silica exposure. In this review, we present details of our investigations as an introduction to scientific approaches examining the immunological effects of environmental and occupational substances.