Hiroaki Iwata

Applicability of radiocolloids, blue dyes and fluorescent indocyanine green to sentinel node biopsy in melanoma

Patients with primary cutaneous melanoma underwent sentinel node (SN) mapping and biopsy at 25 facilities in Japan by the combination of radiocolloid with gamma probe and dye. Technetium‐99m (99mTc)‐tin colloid, 99mTc‐phytate, 2% patent blue violet (PBV) and 0.4% indigo carmine were used as tracers. In some hospitals, 0.5% fluorescent indocyanine green, which allows visualization of the SN with an infrared camera, was concomitantly used and examined. A total of 673 patients were enrolled, and 562 cases were eligible. The detection rates of SN were 95.5% (147/154) with the combination of tin colloid and PBV, 98.9% (368/372) with the combination of phytate and PBV, and 97.2% (35/36) with the combination of tin colloid or phytate and indigo carmine. SN was not detected in 12 cases by the combination method, and the primary tumor was in the head and neck in six of those 12 cases. In eight of 526 cases (1.5%), SN was detected by PBV but not by radiocolloid. There were 13 cases (2.5%) in which SN was detected by radiocolloid but not by PBV. In 18 of 36 cases (50%), SN was detected by radiocolloid but not by indigo carmine. Concomitantly used fluorescent indocyanine green detected SN in all of 67 cases. Interference with transcutaneous oximetry by PVB was observed in some cases, although it caused no clinical trouble. Allergic reactions were not reported with any of the tracers. 99mTc‐tin colloid, 99mTc‐phytate, PBV and indocyanine green are useful tracers for SN mapping.

Characterization of NLRP3 Variants in Japanese Cryopyrin-Associated Periodic Syndrome Patients

The etiology of cryopyrin-associated periodic syndrome (CAPS) is caused by germline gene mutations in NOD-like receptor family, pryin domain containing 3 (NLRP3)/cold-induced autoinflammatory syndrome 1 (CIAS1). CAPS includes diseases with various severities. The aim of this study was to characterize patients according to the disease severity of CAPS. Five Japanese patients with four kinds of gene variations in NLRP3 were found and diagnosed as CAPS or juvenile idiopathic arthritis. Two mutations in NLRP3, Y563N and E688K, found in CAPS patients exhibit significant positive activities in the nuclear factor-κB reporter gene assay. Increased serum interleukin (IL)-18 levels were only observed in severe cases of CAPS. In mild cases of CAPS, the serum IL-18 levels were not increased, although lipopolysaccharide- or hypothermia-enhanced IL-1β and IL-18 production levels by their peripheral blood mononuclear cells were detectable. This series of case reports suggests that a combination of in vitro assays could be a useful tool for the diagnosis and characterization of the disease severity of CAPS.

A case of anti-p200 pemphigoid: evidence for a different pathway in neutrophil recruitment compared with bullous pemphigoid

SIR, Anti-p200 pemphigoid was first described in 1996 by Zillikens et al. as a subepidermal blistering disease that involves autoantibodies directed against a 200-kDa component (p200) of the basement membrane zone (BMZ). Although p200 is known to be an acidic noncollagenous N-linked glycoprotein, which localizes to the lower lamina lucida of the BMZ, the nature of this molecule had not been elucidated for a long time. Currently, it is reported that patient sera react with laminin c1, which is a component of the lamina densa, suggesting that p200 may be laminin c1. In patients with anti-p200 pemphigoid, histopathological examination reveals subepidermal blisters and infiltration predominantly by neutrophils, but occasionally also by eosinophils. Interleukin (IL)-8 is a major chemotactic cytokine for neutrophils and has been implicated in the inflammatory process of bullous pemphigoid (BP). A 75-year-old Japanese man presented with bullae and pustules on his palms, which had arisen suddenly, then spread over the whole body (Fig. 1a). Mucous membranes, including the oral mucosa and conjunctiva, were also affected. Both flaccid and tense blisters were found. Nikolsky’s sign was negative. Milia arose after blister healing in some cases. Blood tests showed a white blood count of 11Æ85 · 10 L (normal 3Æ3– 9Æ2 · 10) and a neutrophil ratio of 81Æ2% (normal 45Æ3– 73Æ2%). Complement C3 and C4 and immunoglobulin (including IgG and IgA) levels were within the normal range. Histopathological examination of a skin biopsy specimen from the neck revealed the presence of subepidermal bullae containing many neutrophils and some eosinophils (Fig. 1b). Direct immunofluorescence (DIF) revealed linear deposits of IgG and C3 at the BMZ. Indirect immunofluorescence (IIF) using 1 mol L salt-split skin showed that IgG antibody bound on the dermal side (Fig. 1c). Immunoblotting with dermal extracts showed that the patient’s IgG autoantibodies reacted with a 200-kDa protein (Fig. 1d). The titre of autoantibody determined by IIF was 1 : 160. IgG subclass of autoantibodies against the BMZ was determined by DIF and IIF. The results of DIF showed linear deposits of IgG2, but not IgG1, IgG3 and IgG4 (Fig. 1e). The deposition of IgG2 autoantibody, but not IgG1, IgG3 and IgG4, was detected on the basement membrane by IIF (Fig. 1e). An indirect complement fixation test with patient serum revealed weak C3 deposits along the basement membrane (Fig. 1f). Enzyme-linked immunosorbent assay (ELISA) revealed that the concentrations of antibodies against BP180, BP230 and desmoglein 1 and 3 were all within the normal range. The patient was diagnosed as having antip200 pemphigoid and was initially treated with oral prednisolone 10 mg daily, but serum levels of autoantibodies, as determined by IIF, continued to increase, and the clinical symptoms worsened. In response to a treatment regimen of prednisolone 30 mg daily, dapsone 50 mg daily and azathioprine 100 mg daily, the blisters gradually disappeared. As the clinical symptoms resolved, the autoantibody titre against p200, as determined by IIF, also decreased gradually. At 7 months after onset, prednisolone therapy was tapered to 20 mg daily, but at this time the patient died due to sepsis, which was probably caused by aspiration pneumonia. In the present case, we speculate that autoantibody against p200 upregulated the release of IL-8 from keratinocytes and condensed it at the BMZ, because of the massive infiltration of neutrophils into the blisters, although the serum level of IL-8 in the patient was under the cut-off value (Quantikine; R&D Systems, Minneapolis, MN, U.S.A.; cut-off value 3Æ5 pg mL). Therefore, in order to examine this speculation, the IL-8 concentration in the growth media of normal human epidermal keratinocytes (NHEK) or human skin (thickness about 400 lm) was measured by ELISA after the cells and the skin were stimulated with IgG from this patient. NHEK culture conditions were as reported previously. NHEK were grown to 70–80% confluence in 24-well plates. Human skin was obtained from a healthy volunteer, and cut into 1 mm specimens. Eight pieces of skin were cultured in a 24-well plate. Cells were starved in culture medium without hydrocortisone 12 h prior to stimulation. Cultured NHEK and skin tissues were treated with IgGs for 12 h, which were purified with protein A–Sepharose column from sera of the present case, a patient with BP and a healthy volunteer, respectively. Culture supernatants were subjected to IL-8 ELISA for determination of the amount of IL-8 secreted. The IL-8 concentration in the growth medium of the cells increased in a dose-dependent manner in response to BP IgG treatment, but not in response to p200 IgG, for both cultured NHEK and skin tissue (Fig. 2). IL-8 release from keratinocytes is known to be increased in patients with BP and psoriasis; however, the pathways of IL-8 release differ between the two conditions. In psoriasis, tumour necrosis factor-a, which is produced by both dermal dendrocytes and epidermal keratinocytes, induces the release of IL-8 from epidermal keratinocytes. In contrast, antibodies to BP180 have been found to mediate, in a doseand timedependent manner, the release of IL-8 from cultured NHEK. These observations suggest that the binding of BP IgG to NHEK triggers an intracellular signal-transducing event that leads to increased production and release of IL-8. Keratinocyte-derived IL-8 recruits neutrophils, which are essential for blister formation both in in vitro models and in passive transfer animal models of BP. In most subepidermal autoimmune blistering conditions, effector mechanisms triggered by immunoglobulin comprise direct inhibition of protein–protein adhesion, signal transduction and induction of the innate immune system, including activation of the complement system and inflammatory cells. Direct association of variable regions to their target antigens causes primary effects of autoantibodies, which are direct inhibition and signal transduction. The Fc region of

Interleukin-17 expression in the urticarial rash of familial cold autoinflammatory syndrome: a case report.

and IL-10-producing regulatory CD4+ T cells in fixed drug eruption. J Allergy Clin Immunol 2003; 112:609–15. 6 Kupper TS, Fuhlbrigge RC. Immune surveillance in the skin: mechanisms and clinical consequences. Nat Rev Immunol 2004; 4:211–22. 7 Nograles KE, Zaba LC, Guttman-Yassky E et al. Th17 cytokines interleukin (IL)-17 and IL-22 modulate distinct inflammatory and keratinocyte-response pathways. Br J Dermatol 2008; 159:1092– 102. 8 Nestle FO, Kaplan DH, Barker J. Psoriasis. N Engl J Med 2009; 361:496–509. 9 Kabashima R, Sugita K, Sawada Y et al. Increased circulating Th17 frequencies and serum IL-22 levels in patients with acute generalized exanthematous pustulosis. J Eur Acad Dermatol Venereol 2010; Jun 21 [Epub ahead of print]. 10 Zaba LC, Fuentes-Duculan J, Eungdamrong NJ et al. Psoriasis is characterized by accumulation of immunostimulatory and Th1 ⁄Th17 cell-polarizing myeloid dendritic cells. J Invest Dermatol 2009; 129:79–88.

Spindle cell squamous cell carcinoma showing epithelial–mesenchymal transition

© 2008 The Authors JEADV 2009, 23, 169–243 Journal compilation

Overexpression of monocyte chemoattractant protein-1 in the overlying epidermis of multicentric reticulohistiocytosis lesions: a case report.

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False‐negative sentinel lymph node biopsy resulting from obstruction of lymphatic basin by nodal metastasis: A case report of malignant melanoma

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Three siblings with systemic lupus erythematosus.

We present the cases of three siblings with systemic lupus erythematosus (SLE). The diagnosis was made when the sisters were of age 21, 25 and 28u2003years. They shared some clinical features, including typical facial rash, photosensitivity and Raynaud’s phenomenon, and tested positive for antinuclear antibodies. However, their symptoms and clinical courses varied. Human leukocyte antigen (HLA) typing revealed that DR4 and A2 were present in all three sisters, while HLA type A11, B35 and B54 were each found in two of the three sisters. The two elder sisters developed lupus glomerulonephritis 8 and 11u2003years after the onset of SLE. It is suggested that there are genes responsible for the onset of the disease and also unknown regulatory genes other than HLA result in different phenotypes.

Extramammary Paget's disease: unique pathological characteristics showing epidermal proliferation of squamoid and basaloid cells

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Autopsy Cases of Extramammary Paget's Disease

乳房外パジェット病は多くの場合表皮内癌として緩徐に進行する一方で, 一度転移を来した例では治療に難渋し, その多くは腫瘍死することとなる。今回, 後者のような進行期乳房外パジェット病で腫瘍死し病理解剖検査を施行し得た1例を報告するとともに, 当院剖検例3例を加え, 本症の転移・進展様式について検討した。本症の転移・進展様式には特徴があり, 血行性転移もリンパ行性転移も来し得るが, 当院剖検例からは後者のほうがより優位と思われた。またその際には, 高度の腫瘍浸潤にもかかわらず画像上は腫瘍そのものを確認することができず, 水腎症や癌性リンパ管症による胸水貯留といった腫瘍浸潤や腫瘍塞栓による結果のみが確認できることもある。これらは本症の進行期症例を診療する上で, また本症の転移・進展様式を理解する上で有用な所見と考えたため報告する。