Cutaneous Lymphocyte-associated Antigen Research Articles

Immunohistochemical Findings in Active Vitiligo Including Depigmenting Lesions and Non-Lesional Skin

Background: The evidence that vitiligo is an autoimmune disease is supported by its association with autoimmune conditions, the presence of activated cytotoxic T lymphocytes in the lesions and melanocyte-specific circulating auto-antibodies. Some studies have indicated the normal-appearing skin being immune-targeted for latent melanocyte disappearance. Method: We aimed to characterize immunohistologically the cellular infiltrate and to identify the distribution pattern of T and Langerhans cells (LCs) in the active border of depigmenting lesions and clinically normal skin of 22 patients with progressive non-segmental vitiligo, besides the cellular expression for the cutaneous lymphocyte-associated antigen (CLA), comparing to 10 controls. Results: Immunohistochemical analyses showed an overall reduction of the number of CD1a+ cells, dermal increase of CD8+ T cells, and increase of the epidermal CD3+ T cells number in the active border of lesions. Surprisingly, those cellular changes were also observed in clinically pigmented skin. However, a significant intra-epidermal infiltration of CD8+ T cells was evident only within active border biopsies. The CLA+ cells were not significantly increased in the patients' skin. Conclusions: The findings of this study suggest an extensively immune-committed skin in active vitiligo, mainly characterized by overall scarcity of CD1a+ cells and dermal increase of CD8+ cells even in apparently normal skin, in addition to the epidermal infiltrate of CD8+ T cells in the depigmenting areas. This report further supports that some CD8+ T and LCs cells play a pivotal role in the process of melanocyte loss, and strengthens an auto-immune hypothesis for vitiligo.

T-cell Large Granular Lymphocytic Leukaemia with an Uncommon Clinical and Immunological Phenotype

A 39-year-old man presented with a rapidly growing unilateral painless nodule on the right cheek. Histopathological examination and peripheral blood analysis both showed a population of T-cell large granular lymphocytes, which were CD1+, CD2+, CD5+, CD7+ and CD16+, with expression of cutaneous lymphocyte-associated antigen. Further laboratory examination revealed severe neutropaenia, relative lymphocytosis and a clonally rearranged T-cell receptor. The cutaneous manifestation of T-cell large granular lymphocytic leukaemia is very rare. In this particular patient, however, it was instrumental in establishing the diagnosis and may have been enabled by the expression of cutaneous lymphocyte-associated antigen on the cell surface.

Cutaneous lymphocyte antigen expression in benign and neoplastic cutaneous B‐ and T‐cell lymphoid infiltrates

Cutaneous lymphocyte-associated antigen (CLA) is expressed in resident cutaneous T lymphocytes, high endothelial venules, peripheral monocytes, granulocytes and a small percentage of memory B cells. It has been postulated to be an important factor in homing of lymphocytes to the skin because of its function as a ligand for E-selectin expressed on cutaneous endothelial cells. We investigated the expression of CLA using the HECA-452 antibody on paraffin-embedded, formalin-fixed tissue in a variety of reactive, neoplastic and preneoplastic cutaneous lymphoid infiltrates of T- and B-cell derivation. CLA was expressed at high levels in various types of inflammatory dermatoses with the exception of lupus erythematosus. High levels of CLA expression were seen in epidermotropic T-cell dyscrasias and epidermotropic T-cell lymphomas including mycosis fungoides (MF) and adult T-cell leukemia/lymphoma (ATCLL). A loss of CLA in tumors normally positive for CLA was a feature of disease progression best exemplified by tumor-stage MF and acute ATCLL. There was a lack of CLA expression in those lymphocytic infiltrates manifesting subcutaneous localization including lupus profundus, panniculitis-like T-cell lymphoma and atypical lymphocytic lobular panniculitis. CLA expression was not only observed in primary cutaneous anaplastic large cell lymphoma but also seen in cases of nodal anaplastic large cell lymphoma secondarily involving the skin and was negative in cases of nodal anaplastic large cell lymphoma without any established skin involvement. An oligodot pattern defined a novel reaction pattern in those aggressive systemic dyscrasias with a proclivity to involve the skin. CLA was negative in the majority of B-cell lymphomas. CLA plays a role in the pattern of T-cell lymphocyte migration in the skin and subcutis in both reactive and neoplastic states. An alteration in the expression of this marker, whether it is in the context of the acquisition of expression in a cell that is normally CLA negative or its loss of expression, may define a key event in determining cutaneous and extracutaneous hematopoietic cell distribution.

Staphylococcal toxin-induced T cell proliferation in atopic eczema correlates with increased use of superantigen-reactive Vbeta-chains in cutaneous lymphocyte-associated antigen (CLA)-positive lymphocytes.

Staphylococcal superantigens have been implicated in the pathogenesis of atopic dermatitis (AD). This may occur through superantigenic activation of T lymphocytes and their subsequent induction of the skin homing receptor CLA on activated cells. We investigated the proliferative responses of peripheral blood mononuclear cells (PBMC) from 10 patients with an infective exacerbation of AD and six normal controls to the staphylococcal superantigens, staphylococcal enterotoxin A and B (SEA, SEB) and toxic shock syndrome toxin-1 (TSST-1), and the mitogens phytohaemagglutinin (PHA) and concanavalin A (Con A). We also assessed CLA and T cell receptor (TCR) Vbeta-chain expression by immunofluorescence and flow cytometry before and after stimulation. PBMC from AD patients showed two-fold increased proliferation to SEA and SEB (P < 0.01) compared with normals, whereas the response to mitogenic stimulation was identical. Analysis of (TCR) Vbeta-chain expression demonstrated increased use of superantigen-reactive Vbeta families in freshly isolated PBMC in AD patients compared with controls. This pattern of Vbeta-chain expression was only observed in the CLA+ but not the total population of T cells. Furthermore, there was a positive correlation between the enhanced PBMC proliferative response and increased expression of superantigen-reactive Vbeta families in atopic patients. These data support the concept that superantigens are important in the pathogenesis of this common condition, and also provide evidence that the increased use of certain Vbeta families in circulating, CLA+, skin homing lymphocytes is of functional significance.

Elevated serum soluble E‐selectin levels in Korean children with measles

Measles is a highly infectious, acute viral illness characterized by high fever and generalized skin rash of which pathogenesis is uncertain. E-selectin, a ligand of the cutaneous lymphocyte-associated antigen (CLA)+ T lymphocytes is highly expressed on vascular endothelium in atopic dermatitis and psoriasis, but in the past there has been a lack of statistical data on its impact on patients with measles. The aim of the present study was to investigate whether the serum soluble E-selectin (sE-selectin) levels are elevated in children with measles. Twenty-five children with measles, 33 children with atopic dermatitis, 20 with atopic asthma and 20 healthy controls were enrolled. Serum sE-selectin and tumor necrosis factor (TNF)-alpha levels were measured using sandwich enzyme-linked immunosorbent assay. Serum levels of sE-selectin were found to be significantly higher in children with measles than in children with atopic dermatitis, atopic asthma and healthy controls. But it was not significantly correlated with the duration of rash or with the presence of a complication in children with measles. Serum TNF-alpha levels were higher in children with measles than in children with atopic dermatitis. There was no correlation between sE-selectin and TNF-alpha levels. sE-selectin levels are elevated in children with measles, which may imply that CLA+ T lymphocytes are associated with the development of measles rash.

Mast Cells Regulate the Magnitude and the Cytokine Microenvironment of the Contact Hypersensitivity Response

The role that mast cells play during contact hypersensitivity (CS) response is unclear because some studies have shown that mast cell-deficient mice have relatively intact CS responses whereas others have shown opposing results. Mast cells secrete a wide range of immunomodulatory mediators and can potentially influence the type of immune response generated in the skin during CS. Therefore, we examined the type of microenvironment generated during CS in both W/Wv mast cell-deficient and wild-type mice in response to different immunizing doses of hapten (oxazolone). The CS response elicited after low-dose oxazolone was significantly diminished in W/Wv mice compared with wild-type mice. Unexpectedly, the CS response elicited in W/Wv mice immunized with high-dose oxazolone was more severe compared with wild-type mice. In addition, after immunization with high-dose oxazolone, the granulocyte infiltrate in W/Wv mice was increased by twofold compared with wild-type mice. A shift in the cytokine milieu toward the expression of type-1 cytokines as well as a significant increase in the local adhesion of neutrophils and CD4 T cells in the microvasculature of the skin was observed after hapten challenge in W/Wv mice immunized with high-dose oxazolone compared with wild-type mice. These results suggest that mast cells can act as regulators and inducers of the inflammatory response depending on immunizing stimulus strength.

Monitoring non-immediate allergic reactions to iodine contrast media

Non-immediate reactions to iodine contrast media (ICM) affect 2-5% of patients receiving these agents. We studied the immunological mechanisms involved in patients with a confirmed non-immediate reaction, maculopapular exanthema, after administration of ICM. The diagnosis was carried out by skin testing or drug provocation test. The immunological study was performed in sequential peripheral blood mononuclear cells taken from the onset of the reaction by flow cytometry and in skin biopsy by immunohistochemistry, with specific recognition by the lymphocyte transformation test (LTT) with different ICM. Flow cytometry showed an increase in the different activation markers [CD69, CD25 and human leucocyte antigen D-related (HLA-DR)] and the skin homing receptor [cutaneous lymphocyte-associated antigen (CLA)] in CD4 lymphocytes, whereas perforin was higher in the CD8 lymphocytes. The skin biopsy showed a perivascular mononuclear infiltrate composed of CD4 lymphocytes, expressing CD25, HLA-DR and CLA, with eosinophils. Intradermal skin tests and the LTT were positive to several ICM, including the culprit agent in four and three patients, respectively, with negative results in all 10 tolerant controls. We showed that a specific immunological mechanism was implicated in patients with non-immediate reactions to ICM. Moreover, the positive results in skin tests and lymphocyte proliferation tests indicated that an important cross-reactivity exists.

Regulation of P‐selectin Ligand Expression by Activated Cutaneous Lymphocyte‐Associated Antigen (CLA) T Cells

T cells utilize E‐ and P‐selectin to enter inflamed skin; however, these C‐type lectins require different glycan modifications on T cells for binding. Sialyl‐Lewis x is sufficient for E‐selectin binding, whereas a core‐2 O‐glycan terminated with sLex (C2‐O‐sLex) is required for P‐selectin binding. We have characterized a mAb CHO‐131 specific to C2‐O‐sLex, which detects a subset of CLA+ T cells. A direct comparison of the selectin binding of CHO‐131+ and CHO131−CLA+ T cells from peripheral blood revealed a significantly greater P‐selectin binding activity by the former subset. Interestingly, we observed that the in vitro activation of CLA+ T cells transiently upregulated the CHO‐131 epitope and the mRNA level of the core‐2 O‐glycan branching enzyme C2GnT‐I. Moreover, the activated CLA+ T cells uniformly were reactive with P‐selectin/Fc in contrast to resting cells. PSGL‐1 isolated from activated CLA+ T cells revealed increased reactivity with CHO‐131, indicating an increased decoration of PSGL‐1 by P‐selectin glycan ligands following activation. Our findings suggest that P‐selectin ligand expression is upregulated in a transient manner by recently activated CLA+ T cells.

Analysis of CD25 hiCD4 + “regulatory” T-cell subtypes in atopic dermatitis reveals a novel T H2-like population

It is unresolved whether circulating CD25hiCD4+ T cells in patients with atopic dermatitis who have elevated IgE (IgE(high)) are regulatory or effector in nature. To analyze the properties of CD25hi T-cell subtypes in IgE(high) atopic dermatitis. The phenotype of circulating CD25hi T cells was analyzed by flow cytometry using PBMCs from patients with atopic dermatitis (total IgE > 250 IU/mL). Cytokines induced in CD25hi subtypes were analyzed after activation with anti-CD3 mAb (+/-IL-2) and in the presence of activated autologous effector T cells (CD25negCD4+). Reactivity to bacterial superantigen derived from the skin-colonizing organism Staphylococcus aureus was also evaluated. CD25(hi) T cells expressing regulatory T-cell markers (Foxp3, CCR4, cutaneous lymphocyte-associated antigen) were increased in atopic dermatitis compared with IgE(low) controls. This phenomenon was linked to disease severity. Two subtypes of CD25hi T cells were identified on the basis of differential expression of the chemokine receptor CCR6. Although the ratio of CCR6+ and CCR6neg subtypes within the CD25hi subset was unaltered in atopic dermatitis, each subtype proliferated spontaneously ex vivo, suggesting in vivo activation. Activated CCR6neg cells secreted T(H)2 cytokines, and coculture with effector T cells selectively enhanced IL-5 production. Moreover, induction of a T(H)2-dominated cytokine profile on activation with bacterial superantigen was restricted to the CCR6neg subtype. Despite a regulatory phenotype, activated CD25hi T cells that lack expression of CCR6 promote T(H)2 responses.

Primary cutaneous anaplastic large cell lymphoma with fatal leukemic outcome in association with CLA and CCR4-negative conversion

A 70-year-old Japanese male presented with a 1-year history of skin tumors, which were diagnosed as primary cutaneous anaplastic large cell lymphoma (ALCL) because of the CD3(low+), CD4(+), CD25(+), CD30(+), CD45RO(+), CD71(+), HLA-DR(+), CD8(-), CD56(-), and NPM/ALK(-) phenotype and monoclonal T-cell receptor-rearranged property of tumor cells as well as the absence of systemic involvement. At this time, the tumor cell was positive for cutaneous lymphocyte-associated antigen (CLA) and TH(2) chemokine receptor CCR4. The eruption had repeatedly appeared and spontaneously regressed or regressed by virtue of several therapeutic modalities, including radiotherapy, interferon-alpha and chemotherapy, until the tumor cell invaded the gastric mucosa and spread to the peripheral blood 5 years later. Upon progression to the fatal leukemic change, the skin lesions inversely disappeared. Flow cytometric monitoring of the phenotype of peripheral blood and skin-infiltrating lymphocytes disclosed that the expression of CLA and CCR4 on the tumor cells was converted from positive to negative in association with the leukemic change. The altered expression of skin-homing receptors might change its clinical behavior.

Vitamins A and D are potent inhibitors of cutaneous lymphocyte-associated antigen expression

Cutaneous lymphocyte-associated antigen (CLA) is a surface glycoprotein expressed by skin-homing T cells. This carbohydrate moiety expressed on mucin-like surface glycoproteins, including P-selectin glycoprotein ligand 1 and CD43, confers binding activity to dermal endothelial E-selectin and is critical for T-cell recruitment to the skin. Vitamin A (retinoic acid [RA]) and the active form of vitamin D3 (1,25 dihydroxyvitamin D3 [1,25D(3)]) have been used to treat certain T cell-mediated inflammatory skin diseases, as well as cutaneous T-cell lymphomas; however, their effect on CLA expression has not been studied. We analyzed the effects of RA and 1,25D(3) on expression of CLA and other lymphocyte-homing receptors on human T cells. We cultured human T cells with 1,25D(3) and RA and analyzed the expression of CLA and other homing receptors. We also pretreated mice with either vitamin and then induced an antigen-dependent contact hypersensitivity response. Both RA and 1,25D(3) downregulated expression of the CLA and, in parallel, functional E-selectin ligand. Whereas RA increased expression of the gut-homing receptor alpha4beta7 and reduced L-selectin expression, 1,25D(3) had no effect on other homing receptors. In an in vivo assay treatment with RA or 1,25D(3) downregulated the skin infiltration of effector CD4+ T cells. These findings suggest that 1,25D(3) can selectively downregulate CLA expression without influencing lymphocyte migration patterns to other tissues.

Bexarotene blunts malignant T‐cell chemotaxis in Sezary syndrome: Reduction of chemokine receptor 4‐positive lymphocytes and decreased chemotaxis to thymus and activation‐regulated chemokine

The malignant cells in Sezary syndrome express the skin trafficking molecules' cutaneous lymphocyte associated antigen (CLA) and chemokine receptor 4 (CCR4). High levels of the CCR4 ligand, thymus, and activation-regulated chemokine (TARC), have been reported in the blood and skin of patients. The rexinoid X-receptor specific retinoid, bexarotene, has contributed to the resolution of cutaneous disease among patients. To evaluate the effects of bexarotene on skin trafficking molecule expression and chemotaxis, peripheral blood mononuclear cells from Sezary syndrome patients and healthy controls were treated with bexarotene in vitro. CCR4 and CLA expression levels and chemotaxis in response to TARC (6.25 ng/ml) were evaluated among lymphocytes before and after treatment with bexarotene (10 microM). Flow cytometric analysis was performed to evaluate CD4, CD26, CLA, and CCR4 cell surface expression. Transwell migration assays were performed to evaluate chemotaxis to TARC. Prior to treatment, malignant cells exhibited higher CCR4 expression (45-90%) and greater than four times more chemotaxis to TARC compared with healthy controls. After treatment with bexarotene for 36-96 hr, a 28% reduction in CCR4 expression was noted (P < 0.05) among the malignant population with an associated 9% decrease in chemotaxis to TARC (P < 0.05). Our results show that bexarotene may inhibit malignant cell trafficking to the skin through an ability to suppress CCR4 expression among Sezary syndrome lymphocytes.

Changes in circulating lymphocyte subpopulations following administration of the leucocyte function-associated antigen-3 (LFA-3)/IgG1 fusion protein alefacept

Alefacept, a recombinant leucocyte function-associated antigen-3 (LFA-3)/IgG1 fusion protein approved for the treatment of psoriasis, is reported to reduce selectively the numbers of circulating CD4(+) CD45RO(+) and CD8(+) CD45RO(+) T cells, while sparing the naive cells. The purpose of the present study was to elucidate further the effect of alefacept on various circulating lymphocyte subsets. Sixteen patients, 12 with chronic plaque psoriasis and four with pustular psoriasis, received alefacept 7.5 mg once weekly for 12 weeks. Blood samples collected at study entry and after 12 weeks of treatment were analysed by four-colour flow cytometry. There were statistically significant reductions in the total number of conventional memory (CD45RA(-) CD27(+)) and effector (CD45RA(-) CD27(-) or CD45RA(+) CD27(-)) T cells, including CD4(+) and CD8(+) T cells expressing CD161 and CD8(+) T cells expressing cutaneous lymphocyte-associated antigen (CLA). Natural killer (NK) T cells were also reduced significantly, while no statistically significant changes were seen in NK cells and CD4(+) CD25(high) cells. The affected subpopulations were all characterized by a high expression of CD2. However, CD4(+) CD25(low), and CD4(+) CLA(+) cells, which also expressed relative high levels of CD2, were not reduced significantly. Our results suggest a heterogeneous effect of alefacept on the circulating memory T cell population, indicating that high expression of CD2 may not, by itself, be sufficient to explain the reduction in cell count for a specific subpopulation.

Assessing function of skin homing T cell selectin-ligands via selective knockdown with siRNA (B62)

Abstract The recruitment of T cells to skin is a key feature of inflammatory diseases such as psoriasis and eczema. Skin homing T cells express cutaneous lymphocyte-associated antigen (CLA), a carbohydrate epitope that binds to E-selectin on postcapillary venules and mediates lymphocyte tethering and rolling in shear flow. E-selectin ligand (ESL) activity and CLA epitope expression have been described on P-selectin glycoprotein ligand-1 (PSGL-1) and leukosialin (CD43). CLA+ PSGL-1 serves as both a P-selectin ligand (PSL) and an ESL, whereas CLA+ CD43 is only an ESL. Although their ESL functions appear redundant, the true relative contribution of PSGL-1 and CD43 to ESL activity in human CLA+ T cells has not yet been evaluated. Using siRNA, we inhibited expression of PSGL-1, CD43, or both on CLA+ T cells by 70–80%, as measured by flow cytometry. PSGL-1 knockdown resulted in a 70% reduction of PSL but relatively unchanged ESL activity, as assessed by shear flow binding assays. CD43 knockdown did not signficantly impact either PSL or ESL activity. Surprisingly, knockdown of both PSGL-1 and CD43 did not noticeably reduce ESL activity nor CLA expression. As predicted from animal models, PSGL-1 is the major PSL on human CLA+ T cells but serves a redundant role as an ESL. The high residual amounts of CLA expression and ESL activity remaining after knockdown of both PSGL-1 and CD43 suggest that an uncharacterized ESL distinct from PSGL-1 and CD43 exists and may represent a signficant pool of ESL function on skin homing T cells.

Homing molecule expression on skin tropic herpes simplex virus type-2 (HSV-2)-specific CD8+ T-cells: role of fucosyltransferase-VII (B68)

Abstract Homing of pathogen specific T-cells is an important component of the host immune response. HSV-2-specific T-cells express high levels of cutaneous lymphocyte-associated antigen (CLA) and the related E-selectin ligand (ESL). Control CMV- or EBV-specific T-cells express low levels of CLA/ESL. CLA/ESL expression and lymphocyte homing to skin are dependent on fucosyltransferases-IV and –VII (FT-IV and FT-VII). We used real-time PCR to compare levels of FT-VII mRNA in isolated CD8-pos T-cells specific for HSV-2, EBV, and CMV from un-manipulated blood using tetramer (tet) staining and cell sorting. Within a group of subjects, circulating HSV-2-specific cells had higher levels of FT-VII mRNA than did EBV-specific cells (p=0.02) but not CMV-specific cells (p=0.29). Using within-subject comparisons, HSV-2 tet-pos cells had higher levels of FT-VII mRNA than did CD8-pos tet-neg bystander cells (p=0.02). In contrast, FT-VII mRNA levels within EBV or CMV tet-pos cells were not higher than levels in CD8 tet-neg bystanders (p=0.31 and p=0.44, respectively). Ongoing studies using HSV-2-specific cells to study skin-homing at the protein and mRNA levels are in progress. Research support provided by: STD/AIDS Research Training Grant NIHT32 AI07140 and UW Royalty Research Fund 65-0875

Varied levels of reactivity by different E-selectin/Fc constructs with cutaneous lymphocyte-associated antigen (CLA) + CD4 + T cells

T cells utilize the vascular adhesion molecule E-selectin to enter inflamed skin. T cells identified by the mAb HECA-452 [cutaneous lymphocyte-associated antigen (CLA) T cells] are enriched in E-selectin ligand expressing cells. However, the proportion of CLA + T cells reactive with an E-selectin/Fc chimeric construct, as determined by flow cytometry, can vary considerably between studies. One important variable in these studies has been the E-selectin/Fc chimera used to assess ligand expression. We therefore compared the reactivity of mouse, rat, and human E-selectin/Fc from the same widely used commercial source with peripheral blood CLA + CD4 + T cells, neutrophils, and the promyelocytic cell line HL-60 by flow cytometry and by shear flow assays. We observed that the binding activities of the different E-selectin/Fc chimeras were considerably different. Mouse E-selectin/Fc demonstrated the highest binding activity with human neutrophils and HL-60 cells by both assay approaches, whereas human E-selectin/Fc demonstrated the lowest binding activity. In addition, mouse E-selectin/Fc binding increased essentially in a linear manner with increasing expression of CLA by CD4 + T cells, whereas human and rat E-selectin/Fc binding occurred with only a subset of CLA + CD4 + T cells. We conclude that there is substantial variability in the reactivity of different E-selectin/Fc constructs, thus caution should be used when assessing E-selectin ligand expression with these reagents. For instance, the discordance in expression of CLA and E-selectin ligands by T cells may in part be due to the E-selectin/Fc construct being used.

Interleukin-4 promotes human CD8 T cell expression of CCR7.

Despite strong evidence supporting a pathway of human T cell differentiation characterized by changes in the expression of CCR7, CD28, CD27 and CD62L, few studies have addressed the mechanisms of pathway regulation. Cutaneous lymphocyte-associated antigen (CLA)-positive skin-homing CD8(+) T cells expressed significantly elevated levels of activation markers compared with CLA(-) CD8(+) T cells in individuals (n = 27) with cutaneous atopic disease. Despite such an activated phenotype, CLA(+) T cells expressed significantly higher levels of CCR7 than a CLA(-) T cell subset. Interleukin (IL)-4 was found to dramatically promote CCR7 expression by antigen-specific CD8(+) cells. Furthermore, skin-homing CD8(+) T cells from individuals with severe disease produced significantly less IL-10 than those derived from mildly affected atopic subjects. Thus in a T-helper 2 dominated disease, tissue-specific CD8(+) T cells show altered CCR7 expression and cytokine production, which may contribute to continued lymph node homing, antigen presentation and disease. IL-4 promotes expression of CCR7, a marker linked to existing models of CD8(+) T cell differentiation.

The MAb CHO-131 Identifies a Subset of Cutaneous Lymphocyte-Associated Antigen T cells Enriched in P-Selectin Binding Cells

The MAb CHO-131 Identifies a Subset of Cutaneous Lymphocyte-Associated Antigen T cells Enriched in P-Selectin Binding Cells

The role of skin-homing T cells in extrinsic atopic dermatitis

T cells that express Cutaneous Lymphocyte-Associated antigen (CLA) have the potential of migrating to the skin, and are hypothesized to play a role in cutaneous atopic disease. To investigate the immune phenotype and cytokine responses to Der p 1 stimulation of CLA+ T cells in extrinsic atopic dermatitis (EAD). In vitro testing, with controls. Peripheral blood mononuclear cells (PBMC) were obtained from EAD patients (n=27) and non-atopic healthy individuals (n=22). Phenotypic analysis of naive, CLA+ and non-CLA+ memory/effector CD4+ and CD8+ T cells used markers of cell activation, differentiation, adhesion, apoptosis and chemokine receptor expression. Cytokine responses in these cells were studied following Der p 1 stimulation. CLA+ T cells from EAD patients expressed significantly higher levels of CD25, HLA-DR, CD38, CD71, CXCR1, CXCR2 and lower levels of bcl2, CCR5, CCR7, CXCR3, and CD62L (p<0.05). In EAD patients, CLA+ T cells express increased levels of markers associated with activation, adhesion and apoptosis, show differences in the level of expression of differentiation markers and display a distinct chemokine receptor preference, compared with cells from healthy controls. These data suggest a significant role for CLA+ T cells in the pathogenesis of cutaneous atopic disease.

Frequency, function and CLA expression of CD4+CD25+FOXP3+ regulatory T cells in bullous pemphigoid

Bullous pemphigoid (BP) is an autoimmune blistering skin disease associated with autoantibodies to collagen XVII and tissue-separation along the dermo-epidermal junction. We addressed the question whether the loss of tolerance in BP patients is associated with a reduction and/or functional impairment of CD4+CD25+FOXP3+ regulatory T cells, which are essential for the active maintenance of self tolerance. The relative and absolute frequency of CD4+CD25+ and CD4+CD25(high) regulatory T cells in the peripheral blood of newly diagnosed, untreated patients was similar to that of healthy controls. Interestingly, more than 50% of circulating CD4+CD25(high) regulatory T cells from both patients as well as healthy controls expressed cutaneous lymphocyte-associated antigen. Considerable numbers of FOXP3+ cells were detected in lesional skin of patients. CD4+CD25+ regulatory T cells of patients were functionally intact as assessed by their ability to suppress allogeneic as well as antigen-specific T-cell proliferation. These data argue against a general defect of CD4+CD25+FOXP3+ regulatory T cells in patients with BP.