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Pemphigus vulgaris (PV) and bullous pemphigoid (BP) are autoimmune blistering diseases, and substantial numbers of CD163+ tissue-associated macrophages (TAMs) are detected in both diseases. PV and BP possess different subsets of helper T cells, suggesting that the cytokine profiles of PV and BP might be different. The purpose of this study was to investigate the microenvironment of lesional skin and serum of PV and BP patients, focusing on the immunomodulatory factors related to TAMs, such as periostin (POSTN), chemokines, cytokines and matrix metalloproteinases (MMPs). We first performed immunohistological staining of POSTN in PV and BP lesions. POSTN was prominent in the superficial dermis in both PV and BP lesions. Next, to validate the activation of CD163+ TAMs in PV and BP patients, we examined the serum levels of soluble (s)CD163. The serum sCD163 levels in PV and BP patients are significantly higher than in healthy controls. To further elucidate the molecular mechanisms of the effects of POSTN on CD163+ TAMs in PV and BP, we examined chemokines, MMPs and cytokines selected by DNA microarray database. The serum CXCL5 levels from PV patients are significantly higher than those in BP patients and healthy controls. The IL-36γ expression on infiltrating macrophages was prominent only in the lesional skin of PV, while the MMP12 deposition was detected in both PV and BP lesions. Our results shed light on the novel pathogenesis of PV through CD163+ TAMs.

SummaryBackground Extramammary Paget disease (EMPD) is a skin adenocarcinoma of apocrine gland origin, in which Paget cells express receptor activator of nuclear factor kappa B (RANK) ligand (RANKL) and matrix metalloproteinase (MMP)-7, and release soluble (s)RANKL into the tumour microenvironment. We previously reported that about 60% of the RANK+ cells among the stromal cells are M2 macrophages, but the identity of the remaining population of RANK+ cells is still unknown. Objectives To investigate the unknown subpopulation of RANK-expressing cells in EMPD. Methods The main population of RANK-expressing cells in the epidermis was composed of epidermal Langerhans cells (LCs). To explore the effects of RANKL on LCs, we stimulated LCs generated from human CD34+ hematopoietic progenitor cells with graded concentrations of sRANKL. To further examine the correlation between LCs and regulatory T cells (Tregs) in EMPD, we employed immunohistochemical staining. Results sRANKL stimulation was shown to augment the production of C-C motif chemokine ligand 17 (CCL17) from LCs. We additionally demonstrated CCL17 expression by CD1a+ LCs in EMPD in an immunofluorescence study. Spearman's rank correlation test confirmed a correlation between the number of LCs and the number of Foxp3+ Tregs in the lesional skin of invasive EMPD. In addition, the numbers of Foxp3+ Tregs in the sentinel lymph nodes of metastatic EMPD were significantly higher than those of metastatic melanoma, which did not express RANKL. Conclusions The findings suggest that the RANKL/RANK pathway in EMPD might contribute to the recruitment of Tregs and to maintenance of the tumour microenvironment.

Background Tumor-associated M2 macrophages (TAMs) produce chemokines that affect the formation of cutaneous T-cell lymphoma (CTCL) by stromal factors. Since IFNs are an effective treatment for advanced-stage mycosis fungoides (MF), we hypothesized that IFNs might modulate M2 macrophages. Objective To prove our hypothesis, we stimulated monocyte-derived M2 macrophages with IFN-α2a or IFN-γ and examined the mRNA expression of chemokines. Methods By using a microarray, we selected a series of chemokines and MMPs that were strongly connected with the IL-4 stimulation. Then, we investigated the effects of IFN-α2a and IFN-γ on these chemokines. Results IFN-α2a and IFN-γ decreased the expression and production of CCL17 and CCL18 and increased those of CXCL10 and CXCL11. Moreover, the subcutaneous administration of IFN-α2a increased the CXCL11-producing cells in the lesional skin of patients with advanced MF. Conclusion Our data suggest one possible mechanism of the therapeutic effects of IFNs through TAMs for the treatment of advanced-stage MF.

Immunosuppressive tumor-associated macrophages (TAMs) promote an immunosuppressive environment in the tumor-bearing host, together with regulatory T cells (Tregs). TAMs compose cancer stroma in skin cancers including melanomas and non-melanomas. The majority of tumor-associated macrophages (TAMs) are alternatively activated M2 macrophages that favor tumor development, and they comprise one of the main populations of inflammatory cells in skin cancers. On the other hand, TAMs could be modulated into M1-type macrophages that suppress tumor growth by stimulating and recruiting Th1 and effector cells in the tumor sites. Therefore, TAMs are a target for immunotherapy in various cancers. In this review, we discuss the definition and suppressive mechanisms of TAMs, as well as their biological activities in tumor-bearing hosts to assess potential therapeutic strategies.

Since bacillus Calmette-Guérin (BCG) is an attenuated strain of Mycobacterium bovis, lupus vulgaris (LV) is reported as one of the rare complications after BCG vaccination, correlating with immunosuppression in the lesional skin. In this report, we describe a case of LV arising from the BCG vaccination site 22 years after vaccination. Interestingly, in the present case, granuloma cells were composed of M2-polarized macrophages. Our case might explain the contribution of M2-polarized macrophages to the biology of LV arising from a BCG vaccination site.

Background Pagetoid squamous cell carcinoma in situ (SCCIS) is a histopathologic variant of SCCIS composed of cells that display an abundant, pale-staining cytoplasm in a pagetoid distribution within the epidermis. As pagetoid SCCIS is sometimes difficult to differentiate from extramammary Paget disease (EMPD) histopathologically, specific markers for pagetoid SCCIS or EMPD are needed by dermatopathologists. Methods In this report, we employed immunohistochemical staining for receptor of activated nuclear factor kappa ligand (RANKL) and programmed death-ligand 1 (PD-L1) in six cases each of pagetoid SCCIS and EMPD. Results The Paget cells strongly expressed RANKL in EMPD, whereas the atypical keratinocytes did not express RANKL in any of the six cases of pagetoid SCCIS. In all cases of pagetoid SCCIS, atypical keratinocytes expressed PD-L1. In EMPD, Paget cells expressed PD-L1 in half of the cases at a lower level of expression than was seen in the surrounding keratinocytes. Conclusion This study suggested that RANKL, but not PD-L1, could be a marker to differentiate between pagetoid SCCIS and EMPD.

Ectopic extramammary Paget’s disease (EMPD) is a rare variant of EMPD that develops in nonapocrine regions. Since reports about ectopic EMPD are limited, little is known about the biological and immunological background of ectopic EMPD. In this report, we present a case of ectopic EMPD on the lower abdomen that expressed RANKL but lacked the expression of MMP7. As we previously reported, Paget’s cells express RANKL and MMP7, release soluble RANKL in the tumor microenvironment, and stimulate tumor-associated macrophages to produce tumor-loading factors in conventional EMPD. In our present case, both CCL5-expressing cells and MMP25-bearing cells were lacking, whereas substantial numbers of CCL5-expressing cells and MMP25-bearing cells were found in conventional EMPD. Our case suggested that the lack of MMP7 on Paget’s cells might be one of the possible explanations for the biology of ectopic EMPD.

Mycosis fungoides palmaris et plantaris (MFPP) is a rare variant of mycosis fungoides limited to the palms and soles. Although little is known about the pathogenesis of MFPP, this variant of mycosis fungoides presents a relatively good prognosis. In this report, we describe an 85-year-old Japanese man with MFPP. Immunohistochemical staining revealed the dense deposition of periostin in the cancer stroma, as well as infiltration of CD163+CD206- tumor-associated macrophages (TAMs), which suggested the phenotypes of TAMs were not polarized to the M2 phenotype in the lesional skin of MFPP. Our present case might suggest one of the possible reasons for the good prognosis of MFPP.

Mycosis fungoides (MF) starts as an indolent disease, progresses from a patch stage to confluent plaques and ultimately develops skin tumors. Tumor-associated macrophages (TAMs) play roles in maintaining the tumor microenvironment in MF. The purpose of this study was to elucidate the involvement of TAMs in the lesional skin of different stages of MF. First, we immunohistologically examined the percentage of CD163+ macrophages and CD206+ cells, as well as the levels of periostin and IL-4 in cancer stroma. The percentage of CD206+ cells increased in parallel with tumor progression, while there was no significant difference in the percentage of CD163+ cells. Periostin was prominent in the stromal area at the patch and plaque stages but decreased at the tumor stage. In contrast, IL-4 was prominently stained at both plaque and tumor stages. To further elucidate the molecular mechanisms of the effects of these stromal factors on TAMs, we examined their effects on mRNA expression in monocyte-derived macrophages in vitro. Based on microarray analysis and gene ontology, we examined a series of chemokines and MMPs whose expression was strongly connected with periostin stimulation. The DNA microarray results were verified in M2 macrophages using real-time PCR. We further examined the mRNA expression of these chemokines and MMPs in the presence of periostin and IL-4 to simulate the advanced stages of MF and validated their protein expression by ELISA. Our present report suggests possible roles of periostin on TAMs in establishing the tumor microenvironment in MF.

Langerhans cell histiocytosis (LCH) is characterized by the clonal proliferation of Langerhans cells; it is categorized as a single-system disease with single or multifocal lesions, and as a multi-system disease with or without the risk of organ involvement. Although the skin is not categorized as a risk organ, the precise diagnosis of skin lesions is necessary to determine the protocol for the treatment of LCH. In this report, we describe a 28-year-old Japanese man with adult onset of BRAFV600E-mutated LCH with cutaneous involvement successfully diagnosed by immunohistochemical staining. Our report suggests that immunohistochemical staining for the BRAFV600E gene could be a diagnostic tool to determine the clinical type of LCH.

Abatacept is a biological immune modifier that is used for the treatment of rheumatoid arthritis. Although psoriasiform drug eruption is reported as one of the cutaneous adverse effects of abatacept, the precise mechanisms are not fully understood. In this report, we describe a 65-year-old Japanese man with psoriasiform drug eruption caused by abatacept. Interestingly, immunohistochemical staining revealed that the epidermal keratinocytes in the basal layer and lower layers of the stratum spinosum were positive for pSTAT3, partially positive for pSTAT1 and negative for pSTAT6, which is similar to conventional psoriasis vulgaris. Our present study suggests that psoriasiform drug eruption caused by abatacept might develop by similar immunological mechanisms as those of psoriasis vulgaris.

Lipodystrophy is a group of metabolic disorders, possibly caused by autoimmune disease. In this report, we describe a case of adult-onset acquired partial lipodystrophy accompanied by rheumatoid arthritis without a family history. Interestingly, immunohistochemical staining revealed dense infiltration of IL-27-producing cells as well as MMP-7- and MMP-28-expressing cells, both of which have been reported to facilitate the development of autoimmune disease. Our present case might suggest possible mechanisms for acquired partial lipodystrophy. © 2015 S. Karger AG, Basel

Hypopigmented mycosis fungoides (HMF) is a clinical variant of MF presenting a good prognosis. In this report, we describe a case of HMF with dense infiltration of tumor-infiltrating leukocytes that bear several cytotoxic molecules, such as granulysin and T-cell intracellular antigen-1. In addition, our immunohistochemical study revealed that the ratio of Foxp3 + regulatory T cells to the total CD4 + cells is decreased in the lesional skin of HMF, compared with that of conventional MF. Our case suggested possible mechanisms for the hypopigmentation and good prognosis of this type of MF.

Rhododendrol (RD), 4-(4-hydroxyphenyl)-2-butanol, inhibits melanin synthesis and has been used for skin-whitening cosmetic products. RD has been very effective in lightening skin pigmentation, but some persons have developed so-called RD vitiligo, in which vitiligo starts on the face, neck and hands where topical RD has been applied and even extended over skin areas where RD has not been applied. RD vitiligo lesions in some patients have lasted for years and have been resistant to conventional vitiligo treatments. We examined the effects of cholecalciferol on RD vitiligo in a blinded randomized clinical trial. Forty-eight female RD vitiligo patients were recruited for the trial and were randomized into two groups: the vitamin D (VD)-intervention group that received daily 5000 IU cholecalciferol for 5 months and the control group. Three blinded investigators scored vitiligo improvement by comparing photographic images of baseline and at 5-month observation. Serum 25(OH)D3 of RD vitiligo patients was not significantly different from age-matched healthy volunteers. Twenty-two in the VD-intervention group and 23 in the control group completed the 5-month observation. Serum 25(OH)D3 levels were significantly increased after the 5-month VD intervention, while the control group did not change. The improvement scores were significantly higher in the VD-intervention group than the control group. The improvement scores were positively correlated with the serum 25(OH)D3 levels after the 5-month intervention period but not before the treatment. This blinded randomized clinical trial showed favor in administrating 5000 IU cholecalciferol daily to RD vitiligo patients.

Together with regulatory T cells (Tregs), tumor-associated macrophages (TAMs) play roles in maintaining the tumor microenvironment. Although cytotoxic antimelanoma drugs such as dacarbazine (DTIC), nimustine hydrochloride (ACNU) and vincristine (VCR) have been used for the treatment of malignant melanoma as adjuvant therapy in Japan, the detailed mechanisms of their immunomodulatory effects are not fully understood. As the majority of TAMs are alternatively activated M2 macrophages that favour tumor development, the aim of this study was to elucidate the immunomodulatory effects of these reagents on human monocyte-derived M2 macrophages. First, mRNA expressions and protein production of immune checkpoint molecules, PD-L1 and chemokines by CD163+ CD206+ M2 macrophages derived from peripheral blood mononuclear cells were investigated to determine the immunomodulatory effects of DTIC, ACNU, and VCR. DTIC and VCR significantly decreased PD-L1 mRNA expression, which was confirmed by flow cytometry. Moreover, the mRNA expression and production of CCL22 were significantly decreased by DTIC, which suggested that DTIC might suppress the recruitment of Tregs in the tumor site. Furthermore, the decreased expression of PD-L1 and production of CCL22 were validated in vivo, using the B16F10 mouse melanoma model, leading to abrogation of the suppressive function of T-cell proliferation. The present report suggests one of the possible antimelanoma mechanisms of DAV combination chemotherapy for melanoma patients.

Keratoacanthoma (KA) is a benign keratinocytic neoplasm that spontaneously regresses after 3-6 months and shares features with well-differentiated squamous cell carcinoma (SCC). An increased incidence of both KA and non-melanoma skin tumor, including SCC, is seen among immunosuppressed, organ-transplant recipients. In this report we describe a case of KA accompanied by multiple lung SCCs developing in a renal transplant recipient.

Background: M2 macrophages play a critical role in the recruitment of T helper 2 (Th2) regulatory T cells (Treg). Objectives To study the role of M2 macrophages and Treg cells in eosinophilic celulitis. Material and Methods: We employed immunohistochemical staining for CD163 and CD206 (macrophages) as well as FoxP3 (Treg), in lesional skin of four cases of eosinophilic cellulitis. Results: CD163+ CD206+ M2 macrophages, which were previously reported to produce CCL17 to induce Th2 cells and Treg cells, were predominantly infiltrating the subcutaneous tissues and interstitial area of the dermis. M2 macrophages derived from PBMC showed significantly increased expression of CCL11, CCL17, CCL24 and CCL26 mRNA and production of CCL17 and CCL24, when stimulated by IL-4 or IL- 13. In addition, CCL17-producing cells and CCL24-producing cells were prominent in the lesional skin of EC. Conclusion: Our study sheds light on one of the possible immunological mechanisms of eosinophilic cellulitis.

Th17 cells, characterized by IL-17 production, play a critical role in the pathogenesis of autoimmune diseases, including autoimmune bullous disorders and aplastic anemia (AA). In this report, we describe a 58-year-old Japanese man with bullous pemphigoid (BP) accompanied by AA. Interestingly, immunohistochemical staining revealed the existence of IL-17-producing cells in the skin biopsy specimens from BP. Our findings might suggest relationships between IL-17 and the pathogenesis of these autoimmune diseases, and, to our knowledge, this is the first English report of BP accompanied by AA.

Pseudolymphomatous folliculitis (PLF), which sometimes mimicks cutaneous lymphoma, is a rare manifestation of cutaneous pseudolymphoma and cutaneous lymphoid hyperplasia. We describe a 57-year-old Japanese woman with PLF on the nose that resembled cutaneous lymphoma clinically. The biopsy specimen revealed dense lymphocytes, especially CD1a+ cells, infiltrated around the hair follicles. Without any additional treatment, her nodule rapidly decreased before we performed a second biopsy for analysis of the clonal gene rearrangement. Though PLF typically behaves as benign lymphohyperplasia, differentiation from cutaneous lymphoma is necessary.

CD30+ lymphoproliferative disorders represent a spectrum of diseases from lymphomatoid papulosis (LyP) to primary cutaneous anaplastic large-cell lymphoma (PCALCL) (1, 2). LyP is characterized by recurrent papulonodular lesions, which undergo spontaneous regression (1, 2). Survival is unaffected by LyP, but patients are at risk of a second cutaneous or lymphoid malignancy (1). PCALCL has a favourable prognosis with a 5-year survival rate between 76% and 96%. In this report, we used immunohistochemical (IHC) staining of LyP type A and PCALCL specimens to examine for macrophage markers as well as dendritic cell (DC)-specific markers and their functional markers.

Because nivolumab significantly prolongs survival in patients with metastatic melanoma, enhancing its antitumor immune response is of great interest to dermato-oncologists. In this report, we describe two cases of metastatic melanoma successfully treated with nivolumab in combination with contact immunotherapy, using contact sensitizing agents, such as squaric acid dibutylester and diphencyprone. In addition, immunohistochemical staining supported one of the possible mechanisms of this combination therapy. Our present cases suggested a possible therapy for metastatic melanoma using nivolumab in combination with contact immunotherapy.

We describe a 34-year-old Japanese man with syringotropic CD8+ mycosis fungoides (MF) accompanied by hypohidrosis who was treated with vorinostat and retinoids. Interestingly, immunohistochemical staining for dermcidin revealed a decrease of sweat in the eccrine glands, and a sweat test by the iodine starch method proved hypohidrosis in the MF-affected areas. Six months after treatment with this combination therapy, the patient's advanced MF was under control.