Hayato Yamaguchi

TSLP Directly Interacts with Skin-Homing Th2 Cells Highly Expressing its Receptor to Enhance IL-4 Production in Atopic Dermatitis

Thymic stromal lymphopoietin (TSLP) is overtly expressed on skin lesions of atopic dermatitis (AD), and the initiative role of TSLP-activated DCs in AD has gained much attention in the past few years, while its actions on other immune cells such as T cells have been given less notice. We aimed to clarify whether TSLP receptor (TSLPR) is expressed on certain populations of T cells and whether TSLP possesses the capability to directly interact with T cells from AD patients. Peripheral lymphocytes from 51 AD patients are analyzed by flow cytometry, and ex vivo experiments using peripheral blood and lesional skin-derived T cells were conducted. TSLPR expression was defined to CD4+ T cells, and CD4+CCR4+CXCR3-CCR7-CCR10+CLA+ T cells in AD patients exhibited enhanced TSLPR expression. The frequency of TSLPR+CD4+ T cells correlated with disease activity. CD4+ T cells from AD patients directly interacted with TSLP to produce a higher amount of IL-4 than those from normal subjects, and this action was attenuated with anti-TSLPR antibody. The importance of IL-4 in the induction of TSLPR expression was found in AD T cells. Our findings indicate that T cells from AD patients possess strong potential to directly interact with TSLP to promote Th2 response.

High frequencies of positive nickel/cobalt patch tests and high sweat nickel concentration in patients with intrinsic atopic dermatitis

BACKGROUND Atopic dermatitis (AD) is classified into extrinsic AD with high serum IgE levels and impaired barrier, and intrinsic AD with low serum IgE levels and unimpaired barrier. Intrinsic AD has a lower frequency of FLG mutations and a higher frequency of circulating Th1 cells, implying that non-protein antigens, represented by metals, may be an exacerbation factor in intrinsic AD. OBJECTIVE To investigate metal allergy in intrinsic AD. METHODS Enrolled in this study were 86 Japanese AD patients seen in three university hospitals, consisting of 55 extrinsic and 31 intrinsic AD patients. Patch testing was performed, focusing on nickel, cobalt, and chrome, in parallel with other 14 metals. FLG mutations were analyzed in 49 patients (extrinsic, 29; intrinsic, 20). In 17 patients (extrinsic, 12; intrinsic, 5), sweat was collected from the forearms by exercise, and the concentration of nickel was fluorometrically measured. RESULTS Nickel, cobalt, and chrome were the major positive metals. Intrinsic AD showed significantly higher percentages of positive reactions than extrinsic AD to nickel (intrinsic 41.9% vs extrinsic 16.4%, P=0.019) and cobalt (38.7% vs 10.9%, P=0.005). There was no significant difference between FLG mutation-bearing and non-bearing patients. The concentration of nickel was higher in the sweat of intrinsic AD than extrinsic AD patients (333.8 vs 89.4ng/g, P=0.0005) and inversely correlated with serum IgE levels. CONCLUSIONS Nickel and cobalt allergy may be involved in intrinsic AD. Given that the metals are excreted through sweat, intrinsic AD might be exaggerated by highly metal-containing sweat.

Proteome analysis of stratum corneum from atopic dermatitis patients by hybrid quadrupole-orbitrap mass spectrometer

may also be anti-inflammatory by reducing nuclear factor kappa B gene expression. Similarly, in addition to their inflammatory role, Langerhans cells are thought to have immunoregulatory functions. Indeed, we see that langerin cells are largely reduced in lesional than in nonlesional AD skin at baseline, and significantly increase on clinical reversal after 12 weeks of CsA treatment (Fig 1, E, and Table E3). In addition to the RDGP, other possible mechanisms for disease recurrence in the same areas need to be considered, including (1) regional differences (increased humidity/friction, transepidermal water loss, pH, and lipids) that allow increased antigen penetration, (2) epigenetic modifications, and (3) microbiome differences. In summary, we have demonstrated that although the CsA RDGP is much smaller than the NB-UVB RDGP, important structural defects and residual inflammation remain and the overall size of the RDGP does not predict relapse kinetics. Given that NB-UVB and CsA have different courses of disease maintenance on discontinuing therapy, some elements in the RDGP of each treatment might explain relevant treatmentand disease-specific mechanisms. Mariya Rozenblit, BA Mayte Suarez-Farinas, PhD Avner Shemer, MD Saakshi Khattri, MD Patricia Gilleaudeau, NP Mary Sullivan-Whalen, NP Xiuzhong Zheng, MSc Hui Xu, MSc Irma Cardinale, MSc James G. Krueger, MD, PhD Emma Guttman-Yassky, MD, PhD

Intrinsic atopic dermatitis shows high serum nickel concentration

Atopic dermatitis (AD) can be divided into two types,1 serum IgE-high extrinsic AD (EAD), occupying approximately 80% of total AD, and serum IgE-normal intrinsic AD (IAD). We have investigated various clinical and immunological aspects of IAD.2e5 Female predominate is noted in IAD. While Th2 cells are increased in the peripheral blood of EAD, the frequency of IFN-g-producing Th1 cells is high in IAD. ThemRNA expression of Th1 cytokines, such as interferon (IFN)-g, in lesional and nonlesional skin from IAD patients are significantly higher than EAD.6 IAD patients often suffer frommetal allergy. Given that protein antigens and metals/haptens give rise to Th2 and Th1 responses, respectively, metals are a candidate of causes for relatively Th1-skewing IAD. We have previously shown that the frequencies of positive patch tests to nickel, cobalt and chrome in IAD patients are 41.9%, 38.7% and 22.6%, respectively, and 61.3% of IAD patients show positive reactions to at least one of the three metals.7 This is 2.4-fold higher than that of EAD. A high nickel concentration in sweat of IAD patients was also found.7 Here, we measured nickel concentration of sera in patients with IAD and EAD, and healthy control (HC). We performed a dietary nickel loading study using a chocolate containing a high nickel dose. This study was approved by the humanmedical and ethics committees of both Hamamatsu University School of Medicine and Tohoku University. Patients gave written informed consent. Enrolled in this study were 17 AD patients and 17 HC volunteers. Leukocyte counts, eosinophil percentage, serum IgE, serum specific IgE to Dermatophagoides pteronyssinus (DP), serum CCL17/TARC were measured in AD patients. There is no uniform criteria for AD categorization. As previously reported, we classified AD into IAD and EAD by using serum IgE and IgE specific to DP.7 IAD was defined as serum IgE levels 200 kU/L or 200 < IgE 400 plus class 0 or 1 of IgE specific to DP and EAD was defined as 400 < IgE levels or 200 < IgE 400 plus class 2 or more of the specific IgE. Severity Scoring of AD index (SCORAD) and 100 mm visual analog scale (VAS) for pruritis were used for evaluation of clinical severity of AD. Eight filaggrin gene (FLG) mutations common to Japanese AD patients were analyzed as described previously.2,7 The nickel loads were applied by using chocolate bar containing a high dose nickel (Lindt chocolate excellence 85% cocoa, Lindt &Sprungli, Kilchberg, Switzerland). This chocolate contains 470 mg nickel per bar (National consumer affairs center of Japan, 2008). Participants ingested one-half chocolate bar containing 235 mg nickel per day for 4 consecutive days. The dose of

Psychological aspects of patients with intrinsic atopic dermatitis.

Atopic dermatitis (AD) is classified into the IgE-high extrinsic and IgE-normal intrinsic types. While extrinsic AD is the classical type with high prevalence, intrinsic AD occupies approximately 20% of total AD with female predominance [1]. The skin barrier is perturbed in the extrinsic, but not intrinsic type [1], and thus filaggrin gene mutations are not a feature of intrinsic AD [2]. While the two types have comparable frequencies of circulating Th2 cells, intrinsic AD shows a significantly increased [...]

Second report of FLG R501X mutation in Japanese patients with atopic dermatitis.

1 Barthelmes L, Goyal A, Newcombe RG, McNeill F, Mansel RE. Adverse reactions to patent blue V dye The NEW START and ALMANAC experience. Eur J Surg Oncol 2010; 36: 399–403. 2 B ezu C, Coutant C, Salengro A, Daraı̈ E, Rouzier R, Uzan S. Anaphylactic response to blue dye during sentinel lymph node biopsy. Surg Oncol 2011; 20: e55–e59. Epub 2010 Nov 11. 3 Uhara H, Yamazaki N, Takata M et al. Applicability of radiocolloids, blue dyes and fluorescent indocyanine green to sentinel node biopsy in melanoma. J Dermatol 2012; 39: 336–8. 4 Longley J, Duffy T, Kohn S. The mast cell and mast cell disease. J Am Acad Dermatol 1995; 32: 545–561.

Sensitive skin is highly frequent in extrinsic atopic dermatitis and correlates with disease severity markers but not necessarily with skin barrier impairment

BACKGROUND Sensitive skin is a condition of cutaneous hypersensitivity to environmental factors. Lactic acid stinging test (LAST) is commonly used to assess sensitive skin and composed of four distinct sensations (pain, burning sensation, itch, and crawly feeling). A link between sensitive skin and barrier dysfunction has been proposed in atopic dermatitis (AD) patients. However, clinical and laboratory factors that are associated with sensitive skin remain unelucidated. OBJECTIVE To investigate relationship between sensitive skin and AD-associated markers. METHODS Forty-two Japanese AD patients and 10 healthy subjects (HS) were enrolled. AD patients were divided into extrinsic (EAD; high IgE levels) and intrinsic (IAD; normal IgE levels) types. We conducted 1% LAST by assessing the four distinct sensations and calculated the frequencies of sensitive skin in EAD, IAD, and HS. We also performed clinical AD-related tests, including transepidermal water loss (TEWL), visual analogue scale (VAS) of pruritus, and quality of life, and measured laboratory markers, including blood levels of IgE, CCL17/TARC, lactate dehydrogenase (LDH) and eosinophil counts, and concentration levels of serum Th1/Th2 cytokines. Filaggrin (FLG) mutations were examined in 21 patients. These values were subjected to correlation analyses with each of the four sensation elements. RESULTS According to the standard criteria for LAST positivity, the frequencies of LAST-positive subjects were 54.8% and 10.0% in AD and HS, respectively (P=0.014). EAD patients showed a significantly (P=0.026) higher frequency of positive LAST (65.6%) than did IAD patients (20.0%). Among the four LAST sensation elements, the crawly feeling and pain scores positively correlated with VAS of pruritus, total serum IgE, mite-specific IgE, CCL17/TARC, and/or LDH. There was no association of the LAST scores with serum Th1/Th2 cytokine levels. Notably, neither TEWL nor FLG mutations correlated with LAST positivity or any sensation scores. CONCLUSIONS The frequency of sensitive skin is higher in EAD than in IAD. Sensitive skin is associated with AD severity, but not necessarily with barrier condition.

Leukoderma in patients with atopic dermatitis

Atopic dermatitis (AD) is occasionally associated with vitiligo, however, the incidence and conditions of vitiligo or leukoderma, and the characteristics of concurrent AD, remain unclear. We conducted a prospective observational study to investigate the leukoderma‐related clinical manifestations and bioparameters of AD. Because vitiligo in AD lesions is occasionally associated with inflammation, we used leukoderma in this study. Enrolled were all AD patients who had been followed up in our AD outpatient clinic and visited within the previous 4 months. During this period, we carefully inspected whether the patients had leukoderma. Eight of 52 patients had leukoderma (15.4%) and were designated as the leukoderma group, and the remaining 44 patients comprised the non‐leukoderma group. While the ages were statistically not different between the two groups, female preponderance was significantly observed in the leukoderma group. The leukoderma patients tended to have higher values of SCORAD, CCL17/thymus and activation regulated chemokine and lactate dehydrogenase than the non‐leukoderma patients. The leukoderma group was also characterized by a lower frequency of allergic rhinitis and a higher frequency of prurigo lesions. Thus, despite the possession of high AD severity, the leukoderma patients may possibly retain a relatively T‐helper 1‐skewing state in relation to the development of leukoderma and less association with rhinitis.

Gross Cystic Disease Fluid Protein 15 in Stratum Corneum Is a Potential Marker of Decreased Eccrine Sweating for Atopic Dermatitis

It is well known that eccrine sweating is attenuated in patients with atopic dermatitis (AD). We have reported by using proteome analysis that gross cystic disease fluid protein 15 (GCDFP15), a substance secreted from eccrine sweat glands, is decreased in tape-stripped stratum corneum (SC) samples from AD patients. The aim of this study was to evaluate GCDFP15 production by eccrine glands with SC samples and to assess sweating in AD. SC samples were obtained from 51 healthy control (HC) and 51 AD individuals. Sweat samples were from 18 HC and 12 AD subjects. GCDFP15 was quantified by ELISA. By immunohistochemistry, the expression of GCDFP15 in eccrine glands was examined in normal and AD skin specimens. To identify GCDFP15-producing cells, double immunofluorescence staining for GCDFP15 and S100 protein was performed in frozen sections. To address the mechanism underlying the decreased eccrine sweating in AD patients, we examined the expression of cholinergic receptor M3 (CHRM3), a receptor for acetylcholine-induced sweating, in eccrine sweat glands. The amounts of GCDFP15 in the SC extracts were significantly lower in AD than HC (P < 0.0001). The sweat samples from AD patients also had lower levels of GCDFP15 concentration (P < 0.05). Immunohistochemistry showed positive GCDFP15 staining in the eccrine gland secretory cells and the ductal and acrosyringial lumen in normal skin, but AD lacked clear staining. Immunofluorescence staining revealed that GCDFP15 was co-expressed with S100 protein, suggesting that the clear cell of eccrine glands produces GCDFP15. Finally, we found that the expression of CHRM3 was depressed in AD, suggesting contribution to the low sweating. The SC of AD patients contains a low amount of GCDFP15 due to both low sweating and low GCDFP15 concentration in the sweat. GCDFP15 in SC is a potential marker for dysregulated sweating in AD.