Establishment of murine model of allergic photocontact dermatitis to ketoprofen and characterization of pathogenic T cells

BACKGROUND: Ketoprofen is well known to evoke the allergic type of photocontact dermatitis when it is applied to the skin and irradiated with ultraviolet A (UVA) light. OBJECTIVE: We aimed to establish a murine model of this photosensitivity and to characterize pathogenic T cells concerned with the sensitivity. METHODS: Various strains of mice were sensitized on two consecutive days by application of ketoprofen to the shaved abdomen and irradiation of the skin with UVA. Five days later, they were elicited with ketoprofen plus UVA on the earlobes. Immune lymph node cells and epidermal cells from the challenged sites were analyzed by RT-PCR. RESULTS: Mice were successfully sensitized and challenged with 4% and 2% ketoprofen, respective, plus UVA at 20J/cm2. The responses in H-2k mice were higher than those in the other strains examined. Immune lymph node CD4+ or CD8+ cells from ketoprofen-photosensitized H-2k mice were transferred i.v. to na?ve syngeneic recipients. Mice receiving CD4+ but not CD8+ cells exhibited ketoprofen photosensitivity, but transference of both CD4+ and CD8+ cell populations was more effective. Lymph node cells from photosensitized mice expressed high levels of mRNA for Th2 cytokine (IL-4) and Th2 chemokine receptor (CCR4) as well as Th1 cytokine (IFN-gamma) and Th1 chemokine receptor (CXCR3), as assessed by RT-PCR. In addition, epidermal cells from challenged earlobes expressed increased levels of both Th1 (TARC) and Th2 (Mig) chemokines. CONCLUSION: It is considered that not only Th1 but also Th2 cells participate in the pathogenesis of murine photocontact dermatitis to ketoprofen.     

The CXCR3 activating chemokines IP-10, Mig, and IP-9 are expressed in allergic but not in irritant patch test reactions.

Differentiation between allergic and irritant contact dermatitis reactions is difficult, as both inflammatory diseases are clinically, histologically, and immunohistologically very similar. Previous studies in mice revealed that the chemokine IP-10 is exclusively expressed in allergic contact dermatitis reactions. In the present study, we investigated whether the mRNA expression of IP-10 and the related CXCR3 activating chemokines, Mig and IP-9 are also differentially expressed in human allergic contact dermatitis and irritant contact dermatitis reactions. Skin biopsies from allergic (13 cases) and sodium lauryl sulfate-induced irritant patch test reactions (13 cases), obtained 1-72 h after patch testing, were studied by means of an in situ hybridization technique. Results of chemokine mRNA expression were correlated with clinical scoring, histology, and immunohistochemical data including the proportion of inflammatory cells expressing CXCR3, the receptor for IP-10, Mig, and IP-9, and ICAM-1 and HLA-DR expression on keratinocytes. IP-10, Mig, and IP-9 mRNA were detected in seven of nine allergic contact dermatitis reactions after 24-72 h, but not in sodium lauryl sulfate-induced irritant contact dermatitis reactions. ICAM-1 expression by keratinocytes was only found in allergic contact dermatitis reactions and correlated with chemokine expression. Moreover, up to 50% of the infiltrating cells in allergic contact dermatitis expressed CXCR3, in contrast to only 20% in irritant contact dermatitis reactions. In conclusion, we have demonstrated differences in chemokine expression between allergic contact dermatitis and irritant contact dermatitis reactions, which might reflect different regulatory mechanisms operating in these diseases and may be an important clue for differentiation between allergic contact dermatitis and irritant contact dermatitis reactions.     

Human IP-9: A keratinocyte-derived high affinity CXC-chemokine ligand for the IP-10/Mig receptor (CXCR3)

Chemokines and their receptors play a crucial part in the recruitment of leukocytes into inflammatory sites. The CXC chemokines IP-10 and Mig are selective attractants for activated (memory) T cells, the predominant cell type in skin infiltrates in many inflammatory dermatoses. The selectivity for activated T cells can be explained by the fact that both chemokines exert their effects through a common receptor, CXCR3, which is nearly exclusively expressed on activated T cells. The aim of this study was to identify biologically active CXCR3 ligands produced by keratinocytes. To that end, Chinese hamster ovary cells expressing a cDNA encoding CXCR3 were challenged with proteins obtained from interferon-gamma stimulated keratinocytes and subsequently monitored for effects on second messenger systems. By this approach we were able to isolate IP-10 and Mig, and in addition identified a novel highly potent ligand for the CXCR3 receptor, designated interferon-gamma-inducible protein-9, which proved to be chemotactic for activated T cells expressing CXCR3. Protein sequence and mass spectrometric analysis followed by molecular cloning of the cDNA encoding interferon-gamma-inducible protein-9, revealed that interferon-gamma-inducible protein-9 is a CXC chemokine with a molecular mass of 8303 Da. From a GenBank database query it became clear that interferon-gamma-inducible protein-9 is in fact the protein encoded by the cDNA sequence also known as beta-R1, H174 or I-TAC. In situ hybridization experiments showed that interferon-gamma-inducible protein-9 mRNA is expressed by basal layer keratinocytes in a variety of skin disorders, including allergic contact dermatitis, lichen planus, and mycosis fungoides suggesting a functional role for this chemokine in skin immune responses.     

镍接触性皮炎皮损趋化因子及其受体的研究

目的 探讨趋化因子及其受体在镍接触性皮炎发病中的作用.方法 留取镍接触性皮炎患者直接接触部位皮损和部分患者非直接接触部位皮损进行趋化因子受体CXCR3和CCR4免疫组化染色,同时采用RT-PCR分析HaCaT细胞经硫酸镍刺激后γ干扰素诱导蛋白-10(IP-10)、干扰素诱导T细胞趋化因子α(I-TAC)、胸腺活化调节的趋化因子(TARC)和巨噬细胞来源趋化因子(MDC)在mRNA水平的表达情况.结果 镍接触性皮炎患者皮损中浸润细胞以CD45Ro⁺CD4⁺T细胞为主,CD8⁺细胞比例小于25%;直接接触与非直接接触部位皮损浸润细胞中50%以上CXCR3染色阳性,CCR4阳性细胞少于20%.HaCaT细胞经硫酸镍刺激后主要表达趋化因子IP-10和I-TAC,48～72 h最明显.结论 镍所致系统性接触性皮炎患者非直接与直接接触部位皮损在T细胞免疫反应的效应阶段炎症反应类型基本相同.     

The role of chemokines in allergic contact dermatitis

Chemokines are important mediators of immune-mediated skin diseases. Allergic contact dermatitis (ACD) is the most thoroughly investigated T cell-mediated disorder because of the ability to easily reproduce the lesions in humans and the availability of an excellent mouse model. Migration of dendritic cells from the skin to lymph nodes is absolutely required for induction of hapten sensitization, and depends upon expression of CCR7 by mature dendritic cells and SLC in the lymph nodes. During expression of ACD, recruitment of T lymphocytes is driven by chemokines exposed on the surface of endothelial cells or released by activated resident skin cells such as mast cells, fibroblasts and keratinocytes. Chemokines are produced in a coordinated and sequential manner, with IL-8 and RANTES induced by TNF-alpha during early stages, and MCP-1, IP-10, Mig, I-TAC, I-309 and MDC induced by IFN-gamma during later stages. Infiltrating monocytes, dendritic cells and T cells are additional sources of chemokines for further leukocyte accumulation. Distinct T cell subsets express different chemokine receptors, with type 2 cells mostly attracted by eotaxin, MDC, TARC and I-309, and type 1 cells sensitive to IP-10, Mig, I-TAC, RANTES and MIP-1beta. MCP-1 is effective on both subsets. T regulatory cells, which inhibit dendritic cell function and are probably involved in the termination of ACD, are sensitive to MCP-1, MIPs and TARC, but express high levels of CCR8 and are more specifically attracted by I-309. Targeting chemokines and chemokine receptors may offer new opportunities for therapeutic interventions in ACD and other chronic inflammatory skin diseases.     

CC chemokine receptor 4 expression on peripheral blood CD4+ T cells reflects disease activity of atopic dermatitis

Recent studies indicate that Th1 and Th2 cells differ in their chemokine receptor expression and their responsiveness to various chemokines. Therefore, selective Th2 cell recruitment in Th2-predominant inflammatory diseases such as atopic dermatitis may be under the influence of some chemokines. It is reported that CC chemokine receptor (CCR) 4 is selectively expressed on Th2 cells whereas CXC chemokine receptor (CXCR) 3 is selectively expressed on Th1 cells. In this study we examined CCR4 and CXCR3 expression on peripheral blood CD4+ and CD8+ T cells obtained from adult atopic dermatitis subjects, and compared the results with those from patients with psoriasis vulgaris and healthy controls. CCR4 was preferentially expressed on CD4+ T cells from atopic dermatitis subjects and CXCR3 was preferentially expressed on CD4+ T cells from psoriasis vulgaris subjects. This CCR4 expression was prominent especially in severe atopic dermatitis subjects. CCR4 expression on CD4+ T cells in severe atopic dermatitis subjects decreased on improvement of disease activity. CD25 was preferentially expressed on CCR4+CD4+ T cells but not on CXCR3+CD4+ T cells in atopic dermatitis subjects. Cutaneous lymphocyte-associated antigen was also preferentially expressed on CCR4+CD4+ T cells but not on CXCR3+CD4+ T cells in atopic dermatitis subjects. CD4+ T cells in atopic dermatitis skin lesions were predominantly CCR4+ cells. Taken together, this study strongly indicates that CCR4+CD4+ T cells reflect disease activity and suggests that CCR4 expression is important for T cell infiltration into atopic dermatitis lesions. Thus, CCR4 may be a possible target for therapy of atopic dermatitis in the future.     

Cutaneous hypersensitivities to hapten are controlled by IFN-gamma-upregulated keratinocyte Th1 chemokines and IFN-gamma-downregulated langerhans cell Th2 chemokines

There are immediate, late-phase, and delayed-type reactions to exogenous agents. In IFN-gamma-knockout (IFN-gamma(-/-)) and wild-type B6 mice, we examined the response to picryl chloride (PCl) for assessing delayed-type reactions, and the responses to repeatedly challenged FITC for immediate and late-phase reactions. The delayed-type hypersensitivity was depressed in IFN-gamma(-/-) mice, and the immediate and late-phase reactions were enhanced in IFN-gamma(-/-) mice. As skin-infiltrating lymphocytes were scarce at the PCl-challenged site of IFN-gamma(-/-) mice, we investigated chemokine production by keratinocytes and Langerhans cells (LCs). A real-time PCR analysis demonstrated that Th1 chemokines (CXCL9 and CXCL10) and Th2 chemokines (CCL17 and CCL22) were derived mainly from keratinocytes and LCs, respectively. Challenge with PCl or FITC augmented keratinocyte expression of Th1 chemokines in wild-type but not in IFN-gamma(-/-) mice, and Th2 chemokine production by LCs was induced by repeated FITC in IFN-gamma(-/-) mice. Finally, transfer of carboxyfluorescein diacetate succinimidyl ester (CFSE)-labeled draining lymph node cells from hapten-sensitized B6 mice or lymph node cells from sensitized green fluorescent protein (GFP) mice to naive IFN-gamma(-/-) mice revealed less infiltration of CFSE(+) or GFP(+) lymphocytes at the challenged site. Our study suggests that one of the crucial actions of IFN-gamma is upregulation of keratinocyte production of Th1 chemokines and downregulation of LC production of Th2 chemokines.     

蕈样肉芽肿中恶性T细胞亲表皮性机制研究进展

皮肤T细胞淋巴瘤中最常见的蕈样肉芽肿,早期可见不同程度的亲表皮现象,即恶性T细胞不同程度侵入表皮,对诊断有较大提示意义,当病情进展到肿瘤期时可逐渐消失.目前对此现象产生和维持的机制研究主要集中在以下两个方面,一为角质形成细胞表达的趋化因子及淋巴细胞表面的趋化因子受体相互作用,如IP-10/CXCR3、SDF-1/CXCR4、TARC/CCR4等,二为表皮内的朗格汉斯细胞及调节性T细胞的相互作用.

Abstract：

Mycosis fungoides is the most common cutaneous T cell lymphoma.Epidermotropism,manifested as the infiltration of epidermis with atypical CD4+ T lymphocytes, may be observed at the early stage of mycosis fungoides, and disappear at the tumor stage.Current studies concerning epidermotropism in mycosis fungoides are mainly focused on the following two aspects, i.e., the interaction of chemokines expressed by keratinocytes with their receptors on the surface of lymphocytes, such as IP-10/CXCR3, SDF-1/CXCR4, TARC/CCR4, as well as the interaction between intraepithelial Langerhans cells and regulatory T cells.     

IL-13-stimulated human keratinocytes preferentially attract CD4+CCR4+ T cells: possible role in atopic dermatitis

Skin inflammation in atopic dermatitis (AD) is characterized by the predominant infiltration of T-helper (Th)2-cells in lesional skin. However, the mechanism of recruitment of these cells in lesional skin of AD is not yet fully elucidated. In this study, we investigated the role of IL-13-stimulated human primary keratinocytes (HPKs) in the recruitment of lymphocytes and further delineated the mechanism of enrichment of these cells. In the migration assays, we observed preferential enrichment of CD4(+)CCR4(+) T cells towards IL-13-stimulated HPKs. Interestingly, CD4(+)CCR4(+) T cells from AD showed a higher chemotactic response than those from healthy individuals. We observed a significant increase in the expression of CCL22 in IL-13-stimulated HPKs as compared to unstimulated cells. Blocking of CCL22 in IL-13-stimulated HPKs by a neutralizing antibody resulted in 70-90% inhibition in migration of CD4(+)CCR4(+) T cells. Moreover, IL-13 upregulated IFN-gamma-induced chemokines, CCL2 and CCL5, in HPKs. Taken together, our data suggest that IL-13-stimulated HPKs participate in a positive feedback loop by preferentially enriching Th2-cells in lesional skin of acute AD patients. However, in chronic phase, IL-13 may act in synergy with IFN-gamma resulting in lymphocytes recruitment of a mixed phenotype at the site of inflammation, thus contributing to the chronification of eczema.     

Expression pattern of chemokine receptors and chemokine release in inflammatory erythroderma and Sézary syndrome

BACKGROUND: Erythroderma can be caused by inflammatory dermatoses or cutaneous T-cell lymphoma. Even if chemokines and their receptors are involved in the skin-selective lymphocyte recruitment, their role in inflammatory erythroderma is yet unclear. OBJECTIVE: To evaluate the chemokine release (TARC, MDC, IP-10) and to define the expression pattern of Th1- (CCR5, CXCR3) and Th2-related (CCR4) chemokine receptors in inflammatory erythroderma and Sézary syndrome (SS). MATERIALS AND METHODS: Flow cytometry has been carried out on both circulating and skin-infiltrating T lymphocytes; serum chemokine levels have been evaluated using ELISA techniques. RESULTS: CCR4, CCR5 and CXCR3 were expressed on about 40% of peripheral blood lymphocytes and on the majority of skin-infiltrating lymphocytes in the inflammatory erythroderma patients, whereas the leukemic CD4+CD26- subpopulation in SS was characterized by a high CCR4 expression without a concurrent increase in CCR5 or CXCR3. TARC, MDC and IP-10 serum levels were significantly increased in both erythrodermic and SS patients. CONCLUSIONS: Our results confirm that SS is a Th2 disorder with a selective expression of CCR4, whereas inflammatory erythroderma shares an overexpression of both Th1- and Th2-related chemokine receptors, suggesting an activation of different pathways driving reactive lymphocytes to the skin.     

Regulatory effects of antihistamines on the responses to staphylococcal enterotoxin B of human monocyte-derived dendritic cells and CD4+ T cells

BACKGROUND: Antihistamines are widely used for the treatment of allergic diseases, such as urticaria and allergic rhinitis. They are also effective for the treatment of diseases in which T cells are mainly involved in the pathogenesis, such as atopic dermatitis (AD) and contact dermatitis. Dendritic cells (DCs) drive polarization of naive T cells into Th1 or Th2 subsets, and are also likely to be involved in AD pathogenesis. OBJECTIVES: The aim of this study was to determine the effects of antihistamines on DCs and CD4(+) T cells. METHODS: Human monocyte-derived DCs (MoDCs) and autologous CD4(+) T cells were obtained from healthy subjects, and cultured together or independently in the presence of antihistamines. As a stimulant, we used staphylococcal enterotoxin B or the combination of anti-CD3 monoclonal antibody (mAb) and anti-CD28 mAb. The concentrations of cytokines and chemokines in culture supernatants were measured by ELISA. The expression of surface molecules on MoDCs was measured by flow cytometry. Cell proliferation in the cocultures of MoDCs and CD4(+) T cells (DC-T cocultures) was measured by a [(3)H] thymidine incorporation assay. RESULTS: Antihistamines inhibited the production of IFN-gamma, and enhanced the production of IL-4 in DC-T cocultures. Antihistamines inhibited the production of TNF-alpha, TARC, MDC, IP-10, and Mig from MoDCs. Epinastine, one of antihistamines, suppressed the expression of ICAM-1 (CD54) on MoDCs. Epinastine also inhibited the proliferation of CD4(+) T cells cocultured with MoDCs. CONCLUSIONS: Our findings show that antihistamines regulate immune responses by affecting the interaction between DCs and CD4(+) T cells, and further DCs and CD4(+) T cells independently, which may partially contribute to the control of allergic diseases such as AD and contact dermatitis.     

Essential role of CCR6 in directing activated T cells to the skin during contact hypersensitivity

CCR6 is expressed in a number of dermatological inflammatory diseases. Here, we report that mice sensitized with the hapten oxazolone had increased numbers of CCR6+ T cells in the draining lymph nodes. Using CCR6-/- mice, we assessed the role of CCR6 on the development of contact hypersensitivity. After hapten sensitization and re-challenge, ear swelling in CCR6-/- animals was reduced 80% as compared with wild-type (WT) control mice. This decreased level of inflammation was not related to an inhibition in T-cell activation, because CCR6-/- lymph node cells from sensitized mice produced threefold higher levels of IFN-gamma in culture than cells from sensitized WT mice and, when these cells were directly injected into the site of hapten challenge, induced a robust inflammatory response. However, intravenous injection of CCR6-/- lymph node cells from sensitized mice were unable to prime naive mice to re-challenge whereas cells from primed WT mice were able to sensitize animals. These results suggest that CCR6 plays an important role in directing the trafficking of activated T cells into the skin and suggests that a CCR6 antagonist could be useful to treat skin-mediated inflammatory reactions.     

Role of chemokines and the corresponding receptors in vitiligo: A pilot study

To determine the levels and sources of chemokines in the serum and epidermis of vitiligo patients, we examined 80 active patients, 80 stable patients and 40 healthy controls. First, the serum levels of candidate chemokines were measured by Luminex assay, and levels of CCR5, CXCR1 and CXCR3 were measured in peripheral mononuclear cells (PMBC) by flow cytometry. Then, the local epidermis levels of elevated chemokines in vitiligo were tested by Luminex. Finally, the mRNA and protein expression levels of elevated chemokines in HaCaT cells stimulated with interferon (IFN)‐γ or tumor necrosis factor (TNF)‐α were tested by quantitative real‐time polymerase chain reaction and Luminex. The serum levels of CCL5, CXCL8 and CXCL10 in active vitiligo were significantly elevated compared with those in stable vitiligo patients. Furthermore, the levels of CCL3 and CCL4 had weak and positive correlations with the Vitiligo Area Scoring Index. In the peripheral blood of active vitiligo patients, the percentages of CD3⁺CD8⁺CCR5⁺ and CD3⁺CD8⁺CXCR3⁺ T cells were significantly increased compared with those in stable vitiligo and healthy controls. In the epidermis of lesions, the expression levels of CCL5 and CXCL10 in active vitiligo were significantly increased. In addition, the mRNA and protein levels of CCL5, CXCL8 and CXCL10 were significantly elevated in HaCaT cells after stimulation with TNF‐α or IFN‐γ. The CCR5/CCL5 and CXCR3/CXCL10 axes may play an important role in the progression and maintenance of vitiligo. Moreover, keratinocytes stimulated with TNF‐α and IFN‐γ may be a primary source of CCL5 and CXCL10.     

FS02.2CD4+ CCR10+ effector T cells reside at former allergic contact dermatitis sites

Local skin memory (LSM) describes the clinical phenomenon of an accelerated and increased inflammatory allergic contact dermatitis (ACD) response upon renewed allergen exposure. This has been ascribed to the local persistence of few, but allergen‐specific, T cells. Here, firstly, we characterized effector T cells, and, subsequently, studied which of these cell populations are present at former challenge sites and might contribute to LSM. Peripheral blood T cells were stimulated with nickel sulphate. Cellular phenotypes and chemokine receptor expression profiles were analysed by FACS‐staining: CLA together with CD4/CD8, CD45R0/RA, CXCR3, CCR4, CCR6 and CCR10. Skin biopsies were taken at 0, 3 and 21 days after allergen application and analysed for the same markers. Upon nickel‐stimulation, amount of CD4+ CLA+ CD45R0+ T cells was increased. Together with CLA, CXCR3, CCR4 and, mainly, CCR10 expression was augmented. CCR6 expression was negative on CLA+ cells. In biopsies from patch tests, cellular infiltrates were present at 3 and 21 days after allergen application. Interestingly at day 21, residing cells were localized at the perivascularity and were characterized as CD4+ CD8− CCR10+ T cells. In conclusion, nickel‐activated effector T cells can be characterised as CD4+ CD8− CLA+ memory T cells. They express CXCR3, CCR4 and, in particular, CCR10. After clinical recovery from an ACD reaction, CD4+ CCR10+ memory T cells apparently reside locally. The persistence of these CCR10+ T cells, expressing the appropriate receptor of the skin specific chemokine CCL27, can explain clinically important phenomena such as LSM and flare up reactions.     

Upregulation of the IFN-gamma-stimulated genes in the development of delayed pigmented spots on the dorsal skin of F1 mice of HR-1 x HR/De

DNA microarray hybridization was used to measure the changes of mRNA levels over time during the development of delayed pigmented spots on the dorsal skin of F1 mice of HR-1 x HR/De. Upregulation of a number of interferon (IFN)-gamma-stimulated genes was detected in delayed pigmented lesions, suggesting that IFN-gamma may play a pivotal role in the development of delayed pigmented spots in this model. Upregulation of these genes was further supported by the increased protein expression level of IFN-gamma in the lesions. Epidermal infiltration of CD8(+) T lymphocytes and mast cell accumulation in the dermis were observed in delayed pigmented spots. Genes encoding chemokines such as monocyte chemoattractant protein-2 (MCP-2), IFN-inducible protein 10 (IP-10), and monokine induced by IFN-gamma (MIG) were among those upregulated by IFN-gamma. We hypothesize that chemokines produced in the epidermis induce migration of inflammatory cells, such as T lymphocytes, mast cells, and macrophages, to the vicinity of melanocytes. Keratinocytes, T lymphocytes, mast cells, and macrophages would become involved in an interactive network, providing a suitable local environment for melanocyte activation. In this environment, melanocytes are exposed to an extensive array of secreted mediators. Reciprocal activation among these cells to maintain this interactive network results in constitutive melanocyte activation and chronic melanin synthesis in delayed pigmented lesions.     

In vivo elimination of CD25+ regulatory T cells leads to tumor rejection of B16F10 melanoma, when combined with interleukin-12 gene transfer

CD4(+)CD25(+) T cells are an important population that plays a crucial role in the maintenance of peripheral self-tolerance. Recently, it was shown that the elimination of these cells by in vivo administration of anti-CD25 monoclonal antibody (mAb) caused the regression of highly immunogenic tumors in syngeneic mice. In this study, we examined whether B16F10 melanoma cells regressed with the elimination of CD25(+) regulatory T cells. We found the melanoma cells were not affected at all by in vivo anti-CD25 mAb administration alone but tumor rejection resulted in all mice when the administration was combined with IL-12 gene transfer to tumor cells. In vivo, depletion of natural killer (NK) cells or CD8(+) T cells cancelled the tumor rejection. NK-cell depletion allowed IL-12-transfected B16F10 melanoma (B16/IL-12) to grow from an early stage and resulted in a more rapid tumor growth of B16/IL-12 than that in mice without administration of anti-CD25 mAb. On the other hand, CD8(+) T-cell depletion did not affect the tumor growth in the early phase but allowed B16/IL-12 to grow in rather a late phase and resulted in almost the same degree of tumor growth as in mice without administration of anti-CD25 mAb. In a previous study, we showed that the elimination of CD4(+) T cells enhanced the antitumor effect of B16/IL-12 and induced vitiligo-like coat color alteration. Therefore, we also examined the frequency of the change to a vitiligo-like coat color in mice showing tumor rejection caused by CD25(+) T-cell elimination to compare with the mechanism enhancing the antitumor effects by cell elimination. The elimination of CD25(+) T cells did not induce vitiligo-like coat color changes, though that of CD4(+) T cells induced the change in 60% of mice. Furthermore, we confirmed that elimination of CD25(+) T cells did not affect the T-helper (Th) 1/Th2 cytokine profile, while that of CD4(+)T cells abrogated the Th2 cytokines (IL-4 and IL-10) and resulted in a Th1-dominant cytokine profile in the tumor-draining lymph nodes (TDLNs) of B16/IL-12-bearing mice. These results indicate that in vivo depletion of CD25(+) regulatory T cells is a potent useful adjuvant in immunotherapy of B16F10 melanoma, when combined with IL-12 gene transfer and that the enhancement of the antitumor effect by CD25(+) T-cell depletion is mediated through CD8(+) T cells and may differ from the enhancing mechanism caused by CD4(+) T-cell depletion.     

Nickel-responding T cells are CD4+ CLA+ CD45RO+ and express chemokine receptors CXCR3, CCR4 and CCR10

BACKGROUND: Whereas T lymphocytes are widely accepted as effector cells determining the pathogenesis of allergic contact dermatitis, contradictory results have been found regarding the roles of different T-cell subsets. The use of various experimental models, involving long-term cultured T-cell lines or clones, may explain these contradictory results. OBJECTIVE: To investigate the involvement of distinct T-cell subsets in patients with nickel contact allergy. METHODS: Different T-cell subsets were directly isolated from peripheral blood mononuclear cells (PBMCs) of nickel-allergic patients, and their proliferative capacity, type-1 or type-2 cytokine secretion [measured by interferon (IFN)-gamma or interleukin (IL)-5 release] and phenotypical marker expression were analysed after stimulation with nickel. RESULTS: Only CD4+ CLA+ CD45RO+ and not CD8+ T cells proliferate and produce both type-1 (IFN-gamma) and type-2 (IL-5) cytokines in response to nickel. Moreover, cells expressing the marker CLA in combination with CD4, CD45RO or CD69 are increased after nickel-specific stimulation. Interestingly, in addition, CD45RA+ CLA+ cells showed an increased frequency after allergen-specific stimulation. Analysis of nickel-reactive T cells for expression of distinct chemokine receptors showed that both proliferative capacity and cytokine production are restricted to subsets expressing CXCR3, CCR4 but not CCR6. Fluorescence-activated cell sorting analysis of chemokine receptors expressed on nickel-stimulated T cells confirmed these results; a subset of T cells expressing CLA and CXCR3, CCR4 and, most importantly, CCR10 increased in response to allergen, while these CLA+ nickel-reactive T cells were all negative for CCR6. CONCLUSIONS: These findings demonstrate that freshly isolated nickel-reactive T cells can be characterized as CD4+ CLA+ memory T cells which express the chemokine receptors CXCR3, CCR4 and CCR10, but not CCR6.