Ryosuke Hino

Tumor cell expression of programmed cell death‐1 ligand 1 is a prognostic factor for malignant melanoma

Melanoma tends to be refractory to various immunotherapies because of tumor‐induced immunosuppression. To investigate the mechanism underlining the immunosuppression of melanoma patients, the authors focused on programmed cell death‐1 (PD‐1)/PD‐1 ligand 1 (PD‐L1) interaction between tumor cells and T cells.

Augmented expression of programmed death‐1 in both neoplastic and non‐neoplastic CD4+ T‐cells in adult T‐cell leukemia/lymphoma

Adult T‐cell leukemia/lymphoma (ATL) is a CD4+CD25+ T‐cell malignancy infected with human T‐cell leukemia virus type‐I (HTLV‐I). HTLV‐I infection causes the T‐cell dysfunction, which contributes to the immunodeficient state of the patients. Programmed death‐1 (PD‐1) can negatively regulate T‐cell response, when its ligand, PD‐L1 or PD‐L2 mainly expressed on antigen presenting cells, binds to this B7 family receptor. We investigated whether PD‐1 is expressed on CD4+ neoplastic (and/or non‐neoplastic) cells or CD8+ cytotoxic cells in peripheral blood mononuclear cells from 11 patients with ATL. By flow cytometry, we found that the levels of PD‐1 expression on both CD4+CD25+ and CD4+CD25− T‐cell populations were increased in ATL patients compared to normal healthy volunteers, while PD‐1 levels on CD8+ T‐cells were comparable between the patients and normal subjects. In stimulation with anti‐CD3 antibody, the proliferation of PD‐1‐expressing T‐cells from ATL patients was weak when compared to that of PD‐1‐nonexpressing normal T‐cells. In addition to PD‐1, PD‐L1 was coexpressed on ATL cells in some patients, and PD‐L1 expression was enhanced by stimulation with anti‐CD3 antibody. Finally, the production of cytokines such as TNF‐α by ATL cells was restored by blockade of PD‐1/PD‐L1 interaction. These findings suggest that CD4+ T‐cells are the main PD‐1‐expressing cells rather than CD8+ T‐cells in ATL patients, and both neoplastic and normal CD4+ cells are exhausted as a result of PD‐1 expression, and additionally PD‐L1 expression on the neoplastic cell.

Type of skin eruption is an independent prognostic indicator for adult T-cell leukemia/lymphoma

Cutaneous involvement is seen in ~ 50% of adult T-cell leukemia/lymphoma (ATLL) patients. We investigated the association between skin eruption type and prognosis in 119 ATLL patients. ATLL eruptions were categorized into patch (6.7%), plaque (26.9%), multipapular (19.3%), nodulotumoral (38.7%), erythrodermic (4.2%), and purpuric (4.2%) types. When the T stage of the tumor-node-metastasis-blood (TNMB) classification of mycosis fungoides/Sézary syndrome was applied to ATLL staging, 16.0% were T1, 17.7% T2, 38.7% T3, and 4.2% T4, and the remaining 23.5% were of the multipapular and purpuric types. For the patch type, the mean survival time (median survival time could not be estimated) was 188.4 months. The median survival times (in months) for the remaining types were as follows: plaque, 114.9; multipapular, 17.3; nodulotumoral, 17.3; erythrodermic, 3.0; and purpuric, 4.4. Kaplan-Meier curves of overall survival showed that the erythrodermic type had the poorest prognosis, followed by the nodulotumoral and multipapular types. The patch and plaque types were associated with better survival rates. Multivariate analysis demonstrated that the hazard ratios of the erythrodermic and nodulotumoral types were significantly higher than that of the patch type, and that the eruption type is an independent prognostic factor for ATLL. The overall survival was worse as the T stage became more advanced: the multipapular type and T2 were comparable, and the purpuric type had a significantly poorer prognosis than T1.

A group of atopic dermatitis without IgE elevation or barrier impairment shows a high Th1 frequency: possible immunological state of the intrinsic type.

BACKGROUND Atopic dermatitis (AD) can be classified into the major extrinsic type with high serum IgE levels and impaired barrier, and the minor intrinsic type with normal IgE levels and unimpaired barrier. OBJECTIVE To characterize the intrinsic type of Japanese AD patients in the T helper cell polarization in relation to the barrier condition. METHODS Enrolled in this study were 21 AD patients with IgE<200kU/L (IgE-low group; 82.5±59.6kU/L) having unimpaired barrier, and 48 AD patients with IgE>500kU/L (IgE-high group; 8.050±10.400kU/L). We investigated filaggrin gene (FLG) mutations evaluated in the eight loci common to Japanese patients, circulating Th1, Th2 and Th17 cells by intracellular cytokine staining and flow cytometry, and blood levels of CCL17/TARC, IL-18, and substance P by ELISA. RESULTS The incidence of FLG mutations was significantly lower in the IgE-low group (10.5%) than the IgE-high group (44.4%) (normal individuals, 3.7%). The percentage of IFN-γ-producing Th1, but not Th2 or Th17, was significantly higher in the IgE-low than IgE-high group. Accordingly, Th2-attracting chemokine CCL17/TARC, was significantly lower in the IgE-low than the IgE-high group. There were no differences between them in serum IL-18 levels, or the plasma substance P levels or its correlation with pruritus. CONCLUSION The IgE-low group differed from the IgE-high group in that it had much less FLG mutations, increased frequency of Th1 cells, and lower levels of CCL17. In the intrinsic type, non-protein antigens capable of penetrating the unimpaired barrier may induce a Th1 eczematous response.

Increased circulating Th17 frequencies and serum IL-22 levels in patients with acute generalized exanthematous pustulosis.

Background/Objective  Acute generalized exanthematous pustulosis (AGEP) is a diffuse pustular disorder that usually begins in intertriginous folds with widespread erythema. The causes in the majority of the cases are drugs. T cells and interleukin (IL)‐8 play roles in the development of AGEP, but the mechanism remains to be elucidated. We investigated the involvement of Th17 cells and their cytokine IL‐22 in the pathogenesis.

Production of Thymus and Activation-Regulated Chemokine and Macrophage-Derived Chemokine by CCR4+ Adult T-Cell Leukemia Cells

Purpose: Adult T-cell leukemia/lymphoma (ATL) is a peripheral CD4+CD25+ T-cell malignancy caused by human T-cell leukemia virus type I. The tumor cells frequently infiltrate in the skin, lymph nodes and other organs and especially form prominent cutaneous masses. Recently, ATL cells have been shown to express Th2 chemokine receptor CCR4. The aim of this study is to investigate the possibility that CCR4 ligands, thymus and activation-regulated chemokine (TARC) and macrophage-derived chemokine (MDC), are produced by CCR4+ ATL cells per se. Experimental Design: CD4+ or CD4+CD14− cells were purified from peripheral blood mononuclear cells of 11 ATL patients with cutaneous involvement and normal healthy volunteers. Tissue-infiltrating cells were isolated from skin tumors. The expression of chemokine receptors on these cells were analyzed by flow cytometry. The production of chemokines and cytokines by the neoplastic cells was assessed by ELISA and reverse transcription-PCR after cultivation for 96 hours in the presence or absence of anti-CD3/CD28 monoclonal antibodies. Finally, TARC and CCR4 expressions were examined by immunohistochemistry. Results: ATL cells highly expressed CCR4 but did not necessarily exhibit the Th2 cytokine profile. The cells also produced TARC and MDC. The production level of MDC was higher in the skin tumor formation group than that in the nontumor group. Immunohistochemically, both CCR4 and TARC were expressed by the tumor cells in the lesional skin. Conclusions: ATL cells not only express CCR4 but also produce TARC and MDC. The skin tumor formation as well as the monoclonal integration of proviral DNA are the factors that are associated with the high production of Th2 chemokines by ATL cells.

Cholinergic Urticaria: Studies on the Muscarinic Cholinergic Receptor M3 in Anhidrotic and Hypohidrotic Skin

TO THE EDITOR Cholinergic urticaria (CU) is a sweating-associated, syringeal orifice-coincident wheal mediated by acetylcholine. CU is occasionally associated with depressed sweating, as reported under the name of anhidrosis (complete lack of sweating) or hypohidrosis (incomplete lack of sweating) (Itakura et al., 2000). There have been reported 29 patients with CU with anhidrosis and/or hypohidrosis (CUAH) (Kay and Maibach, 1969; Itakura et al., 2000; Yoshida et al., 2009). One hypothesis for the relationship between the wheal formation and the depressed sweating is that the patients are hypersensitive to unknown substance(s) in their sweat and develop wheals in response to sweat leaking from the syringeal ducts to the dermis possibly by obstruction of the ducts (Adachi et al., 1994; Kobayashi et al., 2002). In fact, some patients with common CU exhibit wheals to intradermal injection of the patients’ own diluted sweat as well as acetylcholine and histologically show sweat duct obstruction (Fukunaga et al., 2005). However, none of the reported patients with CUAH were positive to the intradermally injected autologous sweat, suggesting that ‘‘sweat hypersensitivity’’ is not responsible for CUAH. In addition, if sweat hypersensitivity is the mechanism, the anhidrotic area should be the predilection site for wheal, but such an observation has not been reported. The study design was approved by the review board of University of Occupational and Environmental Health. Measurements in this study were performed after informed consent had been obtained. The study was conducted according to the Declaration of Helsinki guidelines. To address its mechanism, we investigated four nonsmoker male CUAH patients, aged 23 (case 1), 24 (case 2), 50 (case 3), and 36 years (case 4). The exercise challenge and iodine–starch assessment for sweat induction (Kobayashi et al., 2002) revealed that the skin surfaces of all patients were divided into the anhidrotic and hypohidrotic areas (Figure 1a). The skin with normal sweating was not seen in any patient. As represented by case 1, the applied iodine–starch was discolored by sweat in the hypohidrotic but not anhidrotic area (Figure 1b, left), and following exercise challenge, the patients developed pinpoint wheals in only the hypohidrotic areas (Figure 1b, right). Intradermal injection of acetylcholine yielded wheal with sweating at only the hypohidrotic areas (Figure 1c). The injection of autologous sweat or serum did not produce wheal in either hypohidrotic or anhidrotic area. Histologically, a periglandular lymphocytic infiltrate was observed around eccrine glands in the anhidrotic but not hypohidrotic area of all patients (Figure 1d). There was no occlusion of sweat ducts. By immunohistochemistry, the infiltrating lymphocytes consisted of a mixture of CD4þ and CD8þ T cells (CD4/CD8 ratio, 1.21; Figure 1e). Skin sections were immunohistochemically stained with anti-cholinergic receptor muscarin 3 (CHRM3) antibody (H-210, 1:50; Santa Cruz Biotechnology, Santa Cruz, CA), which is the most important CHR for sweating

Inhibition of T helper 2 chemokine production by narrowband ultraviolet B in cultured keratinocytes

Background  Narrowband ultraviolet B (NB‐UVB) has recently been used for the treatment of various skin disorders. Its effects on the production of cytokines and chemokines by keratinocytes are unknown.

Ectopic extramammary Paget's disease: case report and literature review.

Extramammary Pagets disease that occurs in non-apocrine-bearing regions is referred to as ectopic and has been rarely reported. A 62-year-old man presented with a slowly progressive, asymptomatic light-brown plaque on his back. Histopathological examination revealed the presence of large pale cells with prominent nuclei, which proliferated diffusely and focally in the epidermis. Immuno-histochemically the tumour cells were positive for CK7, GCDFP-15, CEA, and p63. Based on these findings, we diagnosed the tumour as ectopic extramammary Pagets disease. We reviewed the English and Japanese literature and found 29 previously reported cases of ectopic extramammary Pagets disease, including our case, with a predominance of occurrence in the Asian population. The germinative milk line is known to be a possible site where extramammary Pagets disease occurs. Like-wise, some germinative apocrine-differentiating cells might exist on the trunk preferentially in Asians. Attention should be paid to the development of ectopic as well as triple or quadruple EMPD in Asians.

Formation of 8-hydroxy-2'-deoxyguanosine in the DNA of cultured human keratinocytes by clinically used doses of narrowband and broadband ultraviolet B and psoralen plus ultraviolet A.

Psoralen plus ultraviolet A (PUVA) and narrowband ultraviolet B (UVB) are widely used in skin disease phototherapy. Recently, the efficacy of UVB therapy has been greatly improved by narrowband UVB, compared to conventional broadband UVB. The objectives of the current study were to evaluate the influence of UVB‐induced and PUVA‐induced oxidative stress on cultured keratinocytes. We analyzed 8‐hydroxy‐2′‐deoxyguanosine (8‐OH‐dG) in human keratinocytes (HaCaT cell line) using a high‐performance liquid chromatography system equipped with an electrochemical detector. Non‐irradiated human keratinocytes contained a baseline of 1.48 ± 0.22 (mean ± SD) 8‐OH‐dG per 106 deoxyguanosine (dG) residues in cellular DNA, which increased linearly with higher doses of UVB. When their abilities to induce 8‐OH‐dG were compared to each other, based on the minimal erythemal and therapeutically used doses, by irradiating them with broadband UVB at 100 mJ/cm2, the amount of 8‐OH‐dG increased to 3.42 ± 0.46 residues per 106 dG, while a narrowband UVB treatment at 1000 mJ/cm2, with biological effects comparable to those elicited by 100 mJ/cm2 broadband UVB, increased it to 2.06 ± 0.31 residues per 106 dG. PUVA treatment, with 100 ng/mL 8‐methoxypsoralen and 5000 mJ/cm2 UVA, increased the 8‐OH‐dG level to 4.52 ± 0.42 residues per 106 dG. When HaCaT cells treated with 2000 mJ/cm2 narrowband UVB were cultured and the amount of 8‐OH‐dG was monitored in the living cells, 65.6% of the residues were repaired 24 h after treatment. Our study provides a warning that widely used narrowband UVB and PUVA induce cellular oxidative DNA damage at the therapeutically used doses, although to a lesser degree than broadband UVB with the same clinically effective dose. (Cancer Sci 2006; 97: 99  – 105)