Natural killer T cells expressing IFN-γ and IL-4 in lesional skin of atopic eczema

Background:  The inflammation of atopic eczema (AE) is orchestrated not only by T cells predominantly but also B cells, eosinophils and dendritic cells. Recently, a role of invariant natural killer T (NKT) cells has been reported in bronchial asthma and allergy. Natural killer T cells express a restricted repertoire of T-cell receptor α/β and produce interferon (IFN)-γ and/or interleukin (IL)-4 upon activation.
Aim of the study:  To determine the presence of NKT cells in lesional AE skin in comparison with other eczematous disorders and to analyse their cytokine expression.
Methods:  Immunofluorescence stainings were carried out using antibodies recognizing NKT cells, CD3+ and CD4+ cells, IFN-γ and IL-4.
Results:  Natural killer T cells have been detected in small numbers in the majority of AE specimens as well as in atopy patch test (APT) reactions, allergic contact dermatitis (ACD) and irritant contact dermatitis (ICD). In AE, the proportion of NKT cells among CD3+ cells was approximately 5%. NKT cells expressed both IFN-γ and IL-4 in AE, APT and ACD but predominantly IFN-γ in ICD.
Conclusion:  Natural killer T cells are part of the inflammatory infiltrate of AE as well as APT, ACD and ICD, suggesting a pathogenic role of NKT cells in eczematous skin disorders. The pattern of IFN-γ and IL-4 cytokine expression by NKT cells varied depending on the type of eczematous disease.     

Cyclosporin A (CyA) reduces sCD30 serum levels in atopic dermatitis: a possible new immune intervention

Atopic dermatitis (AD) is a chronic inflammatory skin disease frequently associated with asthma, rhinitis, and food allergy. Lymphocytes producing Th2-type cytokines (such as interleukin [IL]-3, IL-4, and IL-5) have been thought to have a key role in the pathogenesis of the disease. We have recently demonstrated that elevated serum levels of the soluble form of CD30 (sCD30), an activation marker of Th2-cell clones, correlates with disease activity in pediatric patients suffering from AD. Clinical trials have demonstrated that cyclosporin A (CyA) treatment resulted in significant improvement of clinical symptoms in patients affected with AD. In this study, we evaluated the role of CyA in modulating sCD30 release in a group of adult patients affected by severe AD treated with CyA at the dosage of 3.5 mg/kg body weight for 12 weeks. Our results demonstrated, in parallel with an improvement of clinical symptoms, a significant reduction of serum levels of both IL-4 and sCD30, thus suggesting that CyA can prevent the activation of Th2 cells observed in AD.     

Cutaneous CD30+ cells in children with atopic dermatitis

BACKGROUND: CD30 expression can be considered a marker of Th2 cells. We investigated the presence of CD30+ cells in the lesional skin of children with atopic dermatitis (AD). We also analyzed the possible relationship between CD30+ cells and serum soluble CD 30 (sCD30) levels, and IgE, soluble interleukin-2 (IL-2) receptor (sIL-2R) or soluble CD23 (sCD23) levels in the blood, and clinical score. METHODS: Ten eczematous children (4 males, 6 females; median age: 4 years and 5 months; range: 11 months to 14 years), 9 sex- and age-matched control children and an adult control group were studied. A clinical score (SCORAD index), was given to eczematous lesions. Blood was taken for the determination of IgE, sCD30, sIL-2R and sCD23 levels. Punch biopsies of lesional skin were stained with hematoxylin and eosin or incubated with anti-CD30 monoclonal antibodies. Skin prick tests (SPTs) were also performed. RESULTS: In the biopsy specimens, CD30 expression was observed in high proportions of infiltrating cells. In children with AD, total serum IgE, sCD30, sIL-2R, sCD23 and eosinophils were significantly elevated compared to controls. CD30+ cells were not associated with serum IgE, sCD30, sIL-2R, sCD23, or SPT results, score of inflammatory cells in lesional skin or clinical score. Children with AD who had high total IgE and specific IgE antibodies did not differ from those with normal total IgE and negative specific IgE in respect of age, clinical score, number of CD30+ cells, sCD30, sIL-2R and sCD23 levels, score of inflammatory cells in skin or clinical score. CONCLUSION: Our results showed remarkable numbers of CD30+ cells in the lesional skin and high sCD30 in the serum of children with AD. CD30+ cells did not correlate with systemic markers of IgE reaction.     

Soluble CD30 is more relevant to disease activity of atopic dermatitis than soluble CD26

It is suggested that CD30 and CD26 are surface molecules expressed on activated Th2 and Th1 cells, respectively. We examined plasma levels of soluble CD26 (sCD26) and sCD30 in patients with atopic dermatitis (AD) when their eruptions were aggravated and in non-atopic healthy controls, and then analysed the possible correlation between these values and the levels of several clinical markers. The plasma levels of both sCD30 and sCD26 were significantly higher in AD patients than in controls, both in exacerbation status and after conventional treatment. Multiple regression analyses showed that plasma sCD30 was a much better predictor of the levels of serum IgE, serum LDH and plasma sCD25, and the area and the score of AD eruption than sCD26, although elevated levels of both sCD30 and sCD26 are associated with these clinical predictors of AD. Importantly, sCD30 plasma levels decreased significantly in AD patients after conventional treatment, while no significant transition was noted in the concentration of sCD26. Moreover, a significant reduction of sCD30 levels was observed in the group of patients whose eruption score was reduced > 50%, whereas it was not in those < 50%. These findings provide evidence that the successful treatment of AD is associated with down-activation of Th2.     

Soluble CD30 in pediatric patients with atopic dermatitis

Atopic dermatitis (AD) is a chronic, inflammatory skin disease in which a pathogenetic role of Th2 cells has been supposed. This study investigated the presence of soluble CD30 (sCD30), an activation marker of T-cell clones able to produce Th2-type cytokines, in sera from pediatric patients affected by AD ( n =25) with no symptoms of asthma or rhinitis. The severity of the disease was graded by both the SCORAD and Costa et al. clinical scoring systems. Serum levels of sCD30 were significantly higher in patients with AD in respect to both normal donors ( n =20) and urticaria patients ( n = 10), and a positive correlation between serum sCD30 and clinical score was found ( r =0.508; P =0.01) when AD patients were evaluated by Costa et al.'s method. Furthermore, a significant association ( r -=0.443; P =0.027) between sCD30 and serum levels of the soluble interleukin (IL)-2 receptor (sIL-2R) was observed in AD. The presence of high amounts of sCD30 in atopic patients seems to confirm the role of this molecule as an activation marker useful for in vivo evaluation of a Th2 immune response, and the correlation observed with both clinical score and sIL-2R levels indicates the role of sCD30 as an additional marker of disease activity in pediatric patients with AD.     

CD30 expression on circulating memory CD4+ T cells as a Th2-dominated situation in patients with atopic dermatitis

Th2 clones have been reported to express CD30 preferentially, but whether T cells producing Th2-type cytokines may favor CD30 expression in the in vivo state remains unknown. We investigated the expression of CD30 on circulating T cells in atopic dermatitis (AD) as a Th2-dominated disorder. Peripheral blood mononuclear cells were prepared from 51 AD patients and 14 nonatopic controls, and their phenotypes were analyzed with flow cytometry. Cytokine production by stimulated CD4+ T cells was also assessed by the single-cell-staining method. Flow cytometric analysis clearly revealed that CD30+ T cells were identifiable in the blood of AD patients with greater frequency compared to controls. The important finding was that CD30 expression was restricted to a small but substantial population of memory (CD45RO+) CD4+ T cells, but not CD8+ ones. In AD patients, it was demonstrated that the percentages of CD30+ cells within CD45RO+ CD4+ T cells correlated well with the disease severity, serum IgE levels, peripheral eosinophil counts, and tendency toward Th2-dominant cytokine pattern as determined by the ratio of interleukin-4 to interferon-gamma production. This study suggests that CD30 expression in circulating T cells might serve as an in vivo marker for the Th2-dominated condition.     

Soluble CD30 and cyclosporine in severe atopic dermatitis

BACKGROUND: Allergen-reactive Th2-like cells expressing membrane CD30 are present in the circulation of atopic dermatitis (AD) patients during seasonal allergen exposure. Moreover, CD30+ T cells are present in the lesional skin of AD patients and high levels of soluble CD30 (sCD30) are found in the serum of the same atopic patients. To investigate the immunosuppressive capacity of cyclosporin A (CsA) in AD patients, the sCD30 serum level was determined before and after CsA treatment (5 mg/kg/day) in 10 patients with severe, refractory AD. The sCD30 serum levels before and after CsA therapy together with other serum parameters were correlated with disease activity. METHODS: sCD30 serum levels were detected using a commercial sandwich ELISA; serum eosinophil cationic protein (ECP) levels were determined using a radioimmunoassay (RIA). RESULTS: In all AD patients sCD30 serum levels were increased ranging from 36 to 300 U/ml, with a mean value equal to 135.7 U/ml. After 6 weeks of CsA treatment, not only was there a significant difference between serum sCD30 levels before (mean 135.7) and after (mean 96.2) treatment but even the serum ECP levels before (mean 57.78) and after (mean 18.69) therapy showed an important reduction. Moreover, no significant difference was found between the mean of serum IgE levels before and after treatment, although the values showed a correlation (p = 0.0003). No significant correlations could be demonstrated between sCD30 levels and serum IgE or between sCD30 and ECP serum levels nor between sCD30 levels and blood eosinophil count after CsA treatment. Moreover, a positive correlation (p = 0.001) was instead documented between sCD30 and the severity of the disease. CONCLUSIONS: In this study, CsA therapy results in clinical improvement together with a statistically significant reduction in sCD30 and ECP serum levels in AD patients.     

Circulating T Cells of Patients with Active Psoriasis Respond to Streptococcal M-Peptides Sharing Sequences with Human Epidermal Keratins

Psoriasis is a T-cell mediated inflammatory skin disease which has been associated with group A, β-haemolytic streptococcal infections. Four 20 a.a. long M6-peptides sharing 5–6 a.a. sequences with human epidermal keratins were identified. To investigate the role of potentially cross-reactive T cells in the pathogenesis of psoriasis, interferon-γ (IFN-γ) and interleukin-4 (IL-4) responses of circulating T cells to these peptides were analysed by ELISPOT and RT-PCR in 14 psoriatic patients, 12 healthy individuals and six patients with atopic dermatitis (AD). Untreated psoriatic patients' responses were significantly higher to these peptides than healthy and AD controls, while responses to a control M6-peptide, not sharing sequences with keratin, were negligible in all groups. No difference was found in response to streptokinase/streptodornase (SK/SD). M6-protein and peptides exclusively elicited IFN-γ production, with little IL-4 production, even in AD patients. Interferon-γ responses to all the M6-peptides were abolished after successful treatment of psoriatic patients, but responses to SK/SD were unaffected. The results indicate that active psoriasis is associated with Th1-like cells responding to streptococcal M6-peptides sharing sequences with human epidermal keratin. This is consistent with the hypothesis that psoriasis may be induced and exacerbated in susceptible individuals by M-protein specific Th1-like cells that cross-react with human epidermal keratin.     

Rheumatoid Inflammatory T-Cell Clones express mostly Th1 but also Th2 and Mixed (Th0-Like) Cytokine Patterns

This study was performed in order to characterize whether T cells from rheumatoid synovial inflammation belong to the Th1- or Th2-like functional subsets. Cytokine production was studied in 26 CD4⁺αβ⁺ and 2 CD8⁺αβ T-cell clones from the synovial fluid, the synovial membrane and peripheral blood of 5 patients. Fifteen of the CD4⁺ clones were raised against various mycobacterial antigens and 11 CD4⁺ clones and 2 CD8⁺ clones were raised unspecifically using PHA and/or IL-2. The specificities of these clones are not known. In the mycobacterial antigen-specific group, all CD4⁺'αβ T-cell clones produced IFN-γ at high levels, while the production of IL-4 was generally absent or low (< 1 ng/ml), consistent with a Thl-like profile. Some of these clones, however, also produced various amounts of IL-10 which has been regarded as a Th2 product but can be produced also in lower amounts by Thi cells. One HSP-65-specific clone produced levels of IL-4 and IL-10 in the same order as that of IFN-γ, thus appearing to be Th0-like. Among the 11 unspecific CD4⁺ clones, 7 showed a Thl-like pattern but with lower levels of IFN-γ than the antigen-specific clones. However, three clones did not produce any IFN-γ activity but produced IL-4 and one of them also produced distinct amounts of IL-10, compatible with a Th2-like pattern. In addition, one of the clones also showed an almost equally strong IFN-γ and IL-4 production, thus most likely representing a Th0-like clone.     

Elevated serum levels of soluble CD30 in patients with atopic dermatitis (AD)

The immunopathology of AD is still unclear, but evidence for an immune response polarized towards Th2 activity has been provided. The CD30 molecule belongs to the tumour necrosis factor (TNF) receptor family and is expressed on activated T cells with a sustained expression in Th2 cells. This molecule also exists in a soluble form (sCD30). Elevated serum levels of sCD30 have been found in patients with Hodgkin's disease, chronic hepatitis B infection and HIV infection. Studies were undertaken to compare the serum levels of sCD30 in patients with AD (n=49) and healthy non-atopic controls (n=94). The presence of sCD30 was analysed with ELISA. A significantly higher concentration of sCD30 was noted in AD patients, median sCD30 level 29 U/ml (range 1–708 U/ml), compared with healthy non-atopic controls (P< 0.001), where the median level was 11 U/ml with a range of 1–1042 U/ml. No correlation was found between sCD30 levels and total serum IgE, or between the AD patients' SCORAD values and concentration of sCD30. sCD30 levels were also analysed in 20 AD patients, which during ketoconazole treatment had improved their clinical scores and reduced their serum IgE and eosinophil cationic protein levels. However, no significant decrease in sCD30 levels was noted after treatment. The results show that patients with AD have elevated levels of sCD30, but without correlation to total serum IgE or disease activity.     

Progressive polarization towards a T helper/cytotoxic type-1 cytokine pattern during age-dependent maturation of the immune response inversely correlates with CD30 cell expression and serum concentration.

In order to investigate the T cell cytokine profile during age-dependent maturation of the immune response, we evaluated the cytokine expression of CD4+ and CD8+ circulating cells by flow cytometric single-cell analysis after non-specific stimulation in vitro in different age groups of normal individuals, from cord blood to adulthood. Moreover, we correlated these lymphocyte cytokine patterns with the expression/release of CD30, a member of the tumour necrosis factor (TNF) receptor superfamily, which has been suggested to be related to the T helper/cytotoxic (Th(c))2-type immune responses, in order to verify this association in vivo, in non-pathological conditions. The results showed a progressive increase of circulating Th(c)1-type, interferon-gamma (IFN-gamma)- and/or IL-2-producing T cells along with ageing and, conversely, a stable number, although higher than in cord blood samples, of CD4+/IL-4+ T cells in the post-natal groups. In addition, serum levels of soluble CD30 (sCD30) and numbers of circulating CD4+/CD30+ and CD8+/CD30+ T cells were significantly higher in children aged < 5 years in comparison with those found either in cord blood or in blood from both older children and adults. These data support the concept of a progressive polarization of the Th(c) cell cytokine profile towards the Th(c)1 pattern during age-dependent maturation of the immune response. Moreover, the peak of CD30 expression/release in early infancy before the Th(c)1 shifting occurs, although not associated with a significant increase of circulating IL-4+ T cells, raises the question of the possible relationship in vivo between CD30 and Th(c)2-type immune responses.     

Acquisition of Interleukin-5 Secretion by Human Naive T-Helper Cells is Regulated by Distinct Signals from both the T-Cell Receptor/CD3 Complex and CD2

We developed a human naive T-helper (Th) cell differentiation model with anti-CD3 monoclonal antibody (MoAb), using a B-cell line as source of costimulation. In this system, we examined the contribution of the T-cell receptor (TCR)/CD3-derived signals and that of lymphocyte function-associated antigen-1 (LFA-1) and CD2 in regulating Th-cell subset differentiation. We found that lowering the level of anti-CD3 MoAb decreased tumour necrosis factor-α (TNF-α) production, while increasing secretion of the Th2 cytokines, interleukin-5 (IL-5) and interleukin-13 (IL-13). Secretion of interferon-γ (IFN-γ) was not influenced by the strength of the anti-CD3 signal. Under conditions where Th0 cells are generated, co-culture with anti-CD2 F(ab')₂ MoAb led to the generation of Th cells that secreted 30–35% less IL-5, while not affecting secretion of IFN-γ or granulocyte–macrophage colony-stimulating factor (GM-CSF). By contrast, anti-CD18 MoAb F(ab')₂ inhibited IFN-γ and GM-CSF levels only in the primary stimulation, but did not affect cytokine levels after restimulation. Neither anti-CD2 nor anti-CD18 F(ab')₂ MoAbs could alter cytokine secretion profiles of peripheral blood-derived memory/effector Th cells. Our results indicate that acquisition of IL-5 secretion capability by Th cells is regulated mainly by signals transduced by the TCR/CD3 complex and by the presence of interleukin-4 (IL-4), while the CD2/LFA-3 pathway plays an additional, but minor, role. These regulatory CD2-derived signals, however, are distinct from those generated by the TCR/CD3 complex.     

Thymus and activation-regulated chemokine in atopic dermatitis: Serum thymus and activation-regulated chemokine level is closely related with disease activity

BACKGROUND: Atopic dermatitis (AD) is a chronic and relapsing inflammatory skin disease characterized by the predominant infiltration of TH2-type cells in lesional skin. Thymus and activation-regulated chemokine (TARC/CCL17) is a chemokine that attracts CC chemokine receptor 4-positive (CCR4+) or CCR8+ cells. OBJECTIVE: The purpose of this study was to investigate the participation of TARC in AD. METHODS: We measured serum TARC levels in 40 patients with AD, 20 healthy control subjects, and 20 patients with psoriasis. We also examined disease activity by using SCORAD score; serum soluble E-selectin, soluble IL-2 receptor, IgE, and GM-CSF levels; and eosinophil numbers in peripheral blood, as well as correlations between TARC levels and these factors. The positivity of CCR4 of CD4+CD45RO+ cells in PBMCs was examined by using FACS analysis. Immunohistochemical staining of TARC and GM-CSF was performed in the lesional skin of patients with AD. RESULTS: The serum TARC levels of patients with AD were significantly higher than those of healthy control subjects and patients with psoriasis. The serum TARC levels significantly correlated with eosinophil number (r = 0.61), SCORAD score (r = 0.60), and serum soluble E-selectin levels (r = 0.58) and weakly correlated with serum soluble IL-2 receptor levels (r = 0.34) in patients with AD. The TARC levels of patients with AD decreased after the treatment in accordance with the improvement of clinical symptoms. The CCR4 positivity of CD4+CD45RO+ cells in PBMCs of patients with AD was also higher than that of healthy control subjects. Immunohistochemical staining revealed that TARC was positive in keratinocytes in the epidermis and in vascular endothelial cells, T cells, and dendritic cells in the dermis. CONCLUSION: Serum TARC levels are associated with disease activity of AD, and TARC may play an important role in the pathogenesis of AD.     

Peripheral Immune Response in Pulmonary Tuberculosis

Cellular immune response and delayed-type hypersensitivity reactions are considered to play a major role in the immunopathogenesis of pulmonary tuberculosis (PTB). But the exact mechanism is still to be clarified. Th1 cells are mainly involved in cellular immune responses in PTB and provide a normal healing process with minimal or no sequela whereas Th2 cell and CD8⁺ T lymphocyte responses may lead to more severe type of disease. In this study, we investigated the peripheral blood immune responses in PTB. The study group consisted of acid fast positive young male soldiers with PTB and a negative HIV serology. The control group included healthy young volunteer male soldiers without a history of PTB. Intracytoplasmic cytokine content of CD8⁺ T cells and lymphocytes, including IL-2, IL-4, IL-5, IL-10 and IFN-γ were determined by flow cytometry, and IL-2, IL-4, IL-5, IL-10, IFN-γ and TNF-α serum levels were measured by cytometric bead array (CBA). No difference was observed between the percentages of T, B, NK cells and HLA-DR expression in both groups, however, the number of CD3⁺HLA-DR⁺ activated T cell percentages was higher in PTB group as compared to healthy subjects. IL-2, IL-4, IL-5, IL-10 contents of lymphocytes and IFN-γ⁺CD8⁺ T cells were found to be significantly lower in PTB patients when compared with healthy subjects, and in parallel, serum IL-2, IL-4, IL-5 and TNF-α levels were also significantly lower in PTB patients. In conclusion we suggest that, CD8⁺ T cells producing both Th1 and Th2 type cytokines, may play important role in the peripheral immune response to mycobacteria.     

Characterization of chemokine receptor expression and cytokine production in circulating CD4+ T cells from patients with atopic dermatitis: up-regulation of C-C chemokine receptor 4 in atopic dermatitis

BACKGROUND: Th2 and Th1 cells have been suggested to express CCR3/CCR4 and CCR5/CXCR3, respectively. OBJECTIVE: We examined CCR3, CCR4, CCR5 and CXCR3 expression and cytokine production in peripheral blood CD4+ T cells from patients with atopic dermatitis (AD), which has been postulated to be a Th2-type cell-mediated disease, and then analysed the possible correlation between these values and the levels of several clinical parameters. METHODS: Intracellular cytokine production and chemokine receptor expression in peripheral blood CD4+ T cells from 40 AD patients and 20 sex- and age-matched healthy control subjects were studied by flow cytometry. RESULTS: The frequencies of IL-4- and IL-13-producing CD4+ T cells from patients with AD were significantly higher than those from healthy control subjects (IL-4:3.9 +/- 2.1% vs. 1.6 +/- 0.7%, P = 0.0005, IL-13:4.0 +/- 2.1% vs. 1.8 +/- 0.8%, P = 0.0023), whereas the frequencies of IL-2- and IFN-gamma-producing CD4+ T cells were significantly decreased in AD patients (IL-2:38.1 +/- 10.3% vs. 51.3 +/- 6.3%, P = 0.0003, IFN-gamma: 9.9 +/- 3.5% vs. 26.4 +/- 4.6%, P < 0.0001). The percentage of CCR4+ cells in CD4+ CD45RO+ T cells in AD patients was significantly higher than that in healthy control subjects (24.4 +/- 8.0% vs. 10.9 +/- 2.3%, P < 0.0001) and was correlated positively with the total serum IgE, serum lactic dehydrogenase (LDH) level, eosinophil number, eruption score, and IL-4 and IL-13 secretion in CD4+ T cells, and inversely with IL-2 and IFN-gamma secretion in CD4+ T cells. In contrast, CCR3 was not detected on circulating CD4+ T cells even in AD patients. On the other hand, the percentage of CCR5+ or CXCR3+ cells in CD4+ CD45RO+ T cells in AD patients was significantly decreased (CCR5:23.2 +/- 7.0% vs. 28.4 +/- 5.4%, P = 0.023, CXCR3:29.9 +/- 11.4% vs. 38.5 +/- 6.7%, P = 0.028) and was positively correlated with eruption score (P < 0.05). Multiple regression analyses showed that the percentage of CCR4 expression highly correlated with serum IgE, LDH, eosinophil number and eruption in AD patients. CONCLUSION: CCR4+ cells might be involved in the aetiopathogenesis of AD.     

Interleukin-4 and Interferon-Gamma Production by Leishmania Stimulated Peripheral Blood Mononuclear Cells from Nonexposed Individuals

Interferon-gamma (IFN-γ) and interleukin-4 (IL-4) production by Leishmania reactive peripheral blood mononuclear cells (PBMC) from non-exposed individuals was investigated. IFN-γ was measured in culture supernatants after antigen stimulation. For the measurement of IL-4, antigen stimulated cells were pulsed with PMA and ionomycin before IL-4 release was measured. L. donovani and L. major antigens induced IL-4 production (105–1748pg/ml) in 13 and seven cultures, and IFN-γ production (1.7- > 66IU/ml) in 14 and 11 of 20 cultures, respectively. IL-4 production rose steeply after 6 days of antigen stimulation suggesting a response due to antigen recognition. Both IL-4 and IFN-γ production was abrogated by depletion of CD2⁺ or CD4⁺ but not CD8⁺ cells. CD2⁺ or CD4⁺ but not CD8⁺ enriched cultures produced cytokines as unseparated PBMC. Thus, in non-exposed individuals circulating Leishmania reactive CD4⁺ T cells could be demonstrated. The cells from different individuals showed different patterns of IFN-γ and/or IL-4 production upon antigenic stimulation. In experimental leishmaniasis the early balance between IFN-γ and IL-4 is important for the clinical outcome. Our findings call for studies of the importance of cytokine production by cross-reactive T cells for the outcome of L. donovani infections in humans and show that the method for IL-4 detection is useful for this purpose.     

Suppression of Experimental Autoimmune Myasthenia Gravis After CD8 Depletion is Associated with Decreased IFN-γ and IL-4

CDS⁺ T cells can perform both Th1 - and Th2-like functions by producing cytokines such as interferonγ (IFN-γ) and interleukin-4 (IL-4), as well as the immune response down-regulating transforming growth factor-β (TGF-β), which are all involved in the development of experimental autoimmune myasthenia gravis (EAMG), a model for human MG. We have reported that depletion of CD8⁺ T cells results in the suppression of EAMG accompanied by the down-regulation of AChR-specific B cell responses and AChR-reactive IFN-γ secreting Th1-like cells. To identify the involvement of IFN-γ, IL-4 and TGF-β in the development of EAMG after CD8⁺ T cell depletion, the expression of mRNA for these cytokines was studied in mononuclear cells from popliteal, inguinal and mesenteric lymph nodes, spleen and thymus by adopting in situ hybridization with complementary DNA oligonucleotide probes. Depletion of CD8⁺ T cells resulted in decreased levels of IFN-7 and IL-4 mRNA expressing cells in different lymphoid organs except thymus, but no change in the numbers of TGF-β mRNA expressing cells. The results imply that the suppression of EAMG after depletion of CD8⁺ T cells is caused by decreasing the effector factors but not by increasing the suppressor factor(s).     

The effects of CD40- and interleukin (IL-4)-activated CD23+ cells on the production of IL-10 by mononuclear cells in Graves' disease: the role of CD8+ cells

The possible roles of CD8+ cells in the abnormal T cell-dependent B-cell activation in Graves' disease were investigated by analysing lymphocyte subsets in peripheral blood mononuclear cells (PBMC) and their production of soluble factors and cytokines such as IL-10 in patients with Graves' disease, Hashimoto's thyroiditis and normal controls. The PBMC were separated into CD8+ and CD8-depleted cells by magnetic separation columns, and cultured for 7 days with or without anti-CD40 monoclonal antibodies and IL-4. The culture supernatant was assayed for sCD23 and IL-10 using EIA, and the remaining cells were analysed by flow cytometry. Stimulation with anti-CD40 antibody together with IL-4 increased sCD23 levels and the number of CD23+ cells. The latter was further augmented by depletion of CD8+ cells. This combination of B cell stimulants increased production of IL-10 by PBMC from patients with Graves' disease. The CD40- and IL-4-activated production of IL-10 was decreased by CD8+ cell depletion. In contrast, constitutive production of IL-10 was increased after CD8+ cell depletion in a group of patients with low basal secretion levels (<35 ng/ml). It was, however, decreased in a group with higher basal production levels, but such a relationship was not found in the normal control group. Thus, T cell-dependent B-cell activation via a CD40 pathway activates CD23+ cells, leading to over-production of IL-10 and a shift of the Th1/Th2 balance to Th2 dominance, while CD8+ cells may suppress this activation to counteract the Th2 deviation in Graves' disease.     

CD4(+)IL-13(+) cells in peripheral blood well correlates with the severity of atopic dermatitis in children

BACKGROUND: In atopic dermatitis (AD) a Th1/Th2 imbalance has been reported, and interleukin (IL)-13 seems to play a pivotal role in the inflammatory network. We tried to assess the correlation between the immunological marker CD4(+)IL-13(+) and the clinical phase of extrinsic AD in children. METHODS: Twenty children with AD were studied. Assessed parameters were: clinical severity (SCORAD index), total serum immunoglobulin E (IgE), blood eosinophil count, and percentage of CD4(+)IFNgamma(+), CD4(+)IL-4(+), CD4(+)IL-13(+) T cells. Determinations were carried out in the acute phase and after clinical remission were achieved. Ten nonatopic-matched children served as controls. RESULTS: At baseline, AD was mild in 25%, moderate in 50% and severe in 25% of children. In the acute phase a significant relationship between the eosinophil count and the SCORAD index was found (P = 0.0001). Blood CD4(+)IL-4(+) were significantly higher in the AD group (median 17.0, range: 13.7-21.4) than in controls (12.6, 6.4-17.2, P < 0.0001). CD4(+)IL-13(+) cells in the AD group well correlated (P = 0.0007) with SCORAD index. At remission, a significant correlation between SCORAD index and eosinophil count was found (P < 0.03) and the percentage of CD4(+)IL-13(+) cells globally decreased (P < 0.0001), while no difference was found among SCORAD classes. CONCLUSION: This study confirms the Th2 profile predominance in the peripheral blood of children with AD, and evidences close relationship between the number of CD4(+)IL-13(+) T cells and the disease's severity.     

Preferential expression of T(h)2-type chemokine and its receptor in atopic dermatitis

Lesional skin of patients with atopic dermatitis (AD) is histologically characterized by hypertrophy of the skin, and the infiltration of a large number of eosinophils and T cells into the dermis. Recent studies have indicated that Th2 cells play a crucial role in the pathogenesis of AD skin. Chemokines and their receptors are implicated in the development of symptoms of various skin diseases such as AD and psoriasis vulgaris (psoriasis). We have examined the in situ expression of a typical Th2-type chemokine, thymus- and activation-regulated chemokine (TARC), and its receptor (CCR4) using immunohistochemical techniques. TARC was found to be highly expressed in the basal epidermis of the lesional skin of AD patients and only slightly in the non-lesional skin. On the other hand, no positive cells were seen in the lesional skin of psoriasis. Consistently, CCR4+ cells were present predominantly in the lesional skin of AD patients, but not in the non-lesional skin. In contrast, in the lesional skin of psoriasis patients, cells positive for CCR5, which is expressed on Th1 cells, were abundantly present. Interestingly, psoralen plus ultraviolet A therapy reduced the number of CCR4+ cells in the AD skin lesions. These results suggest that Th2-type cytokines such as TARC are involved in the pathogenesis of skin lesions in AD patients through the preferential recruitment of Th2 cells.