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.     

Increase of chemokine interferon-inducible protein-10 (IP-10) in the serum of patients with autoimmune liver diseases and increase of its mRNA expression in hepatocytes

To clarify the role of IP-10 in autoimmune liver diseases, we studied the serum levels of IP-10 in 14 patients with autoimmune hepatitis (AIH), 23 patients with primary biliary cirrhosis (PBC), and 65 patients with chronic viral hepatitis (20 type B and 45 type C). The hepatic expression of IP-10 mRNA and the correlation between the serum levels of IP-10 and clinical parameters were also evaluated. In addition to 20 healthy controls, 16 rheumatoid arthritis (RA) patients were included as an extrahepatic inflammatory disease. The serum level of IP-10 was significantly (P < 0.02) higher in patients with AIH, PBC, and chronic hepatitis B and C than in healthy controls, and it was significantly correlated (P < 0.05) with the serum levels of aspartate aminotransferase and alanine aminotransferase in patients with AIH, PBC, and chronic hepatitis B and C. The serum level of IP-10 was not elevated in RA patients. After successful treatment of AIH and chronic hepatitis C, the serum level of IP-10 decreased to the same level as in healthy volunteers. As we previously showed in cases with chronic hepatitis B or C, in situ hybridization in both AIH and PBC cases demonstrated the expression of IP-10 mRNA in hepatocytes around focal or lobular necrosis surrounded by infiltrating mononuclear cells, whereas IP-10 mRNA was not expressed in areas around the damaged bile ducts in PBC cases. The present results suggest that IP-10 is specifically produced by hepatocytes in inflammatory areas irrespective of the aetiology of hepatitis, and that IP-10 may help to recruit T cells to the hepatic lesions in autoimmune liver diseases as well as in chronic viral hepatitis.     

Allergen immunotherapy induces a suppressive memory response mediated by IL-10 in a mouse asthma model

BACKGROUND: Human studies have demonstrated that allergen immunotherapy induces memory suppressive responses and IL-10 production by allergen-specific T cells. Previously, we established a mouse model in which allergen immunotherapy was effective in the suppression of allergen-induced asthma manifestations. OBJECTIVE: In this study, we examined whether immunotherapy induces a long-lasting effect and investigated the role of IL-10 in successful immunotherapy. METHODS: Ovalbumin-sensitized BALB/c mice were treated with 3 injections of ovalbumin (1 mg, subcutaneous) on alternate days. After a short interval (1 week) and after a long interval (5 weeks), mice were challenged by ovalbumin inhalation, and subsequently, airway reactivity, airway eosinophilia, ovalbumin-specific IgE, and T(H)2 cytokine profile were measured. Flow cytometry and blocking of IL-10 receptors in vivo were used to gain insight in the role of IL-10 in the beneficial effects of allergen immunotherapy. RESULTS: After a long interval between ovalbumin immunotherapy and ovalbumin challenge, the development of airway eosinophilia and hyperresponsiveness to methacholine were as strongly suppressed as after a short interval. These suppressive effects coincided with significantly reduced serum ovalbumin-specific IgE levels and T(H)2 cytokine production. On immunotherapy, the IL-5:IL-10 ratio in the bronchoalveolar lavage fluid shifted toward IL-10. In ovalbumin-restimulated lung cell and thoracic lymph node cultures from these mice, IL-5 levels dramatically decreased, whereas the percentage of IL-10(+)CD4(+) T cells was not affected. Finally, in mice treated with mAb against IL-10 receptors, the beneficial effects of immunotherapy were largely abrogated. CONCLUSION: These data demonstrate that allergen immunotherapy induces a memory suppressive effect in which IL-10 is essential.     

Chronic exposure to superantigen induces regulatory CD4(+) T cells with IL-10-mediated suppressive activity

The repeated injection of bacterial superantigens (SAg), such as staphylococcus enterotoxin (SE) A or B, has been shown in mice to induce a state of unresponsiveness characterized by the lack of secretion of Th1 lymphokines, such as IL-2 and IFN-gamma, following subsequent SAg challenge. We made the observation, in vivo as well as in vitro, that unresponsiveness to SAg could be transferred from SEA- to SEB-reactive T cells (and reversibly from SEB- to SEA-specific T cells) in C57BL/6 mice but not in BALB/c mice. Since C57BL/6 mice, unlike BALB/c mice, possess TCR V(beta)3+ and V(beta)11+ T cells able to react with both SEA and SEB, we hypothesized that SAg-unresponsive V(beta)3(+) and V(beta)11+ T cells could mediate linked suppression of other SAg-reactive T cells. To analyze further this possibility, spleen cells from BALB/c mice made unresponsive to SEB were tested for their capacity to suppress the response of normal BALB/c cells to SEB. The production of both IFN-gamma and IL-2 following SEB stimulation was greatly impaired in co-cultures containing CD4(+) T cells, but not CD8(+) T cells, isolated from unresponsive animals. In vivo, the production of both IFN-gamma and IL-2 responses to SEB was dramatically reduced in animals adoptively transferred with unresponsive spleen cells. This suppression was abrogated in recipients injected with neutralizing anti-IL-10 antibodies. Moreover, in animals made unresponsive to SEB, SAg-reactive CD4(+) T cells were found to express high levels of CTLA-4, a molecule recently described to play an essential role in the suppressive function of regulatory T cells. Taken together these results demonstrate that the repetitive injection of SAg induces the differentiation of regulatory CD4(+) T cells capable of suppressing SAg-reactive naive T cells.     

The role of different subsets of T regulatory cells in controlling autoimmunity

T regulatory cells-in addition to clonal deletion and anergy-are essential for the downregulation of T cell responses to both foreign and self antigens, and for the prevention of autoimmunity. Recent progress has been made in characterising the different subsets of T regulatory cells, the factors that drive their differentiation, and their mode of action. The resolution of these mechanisms will make it possible to use T regulatory cells therapeutically in human autoimmune diseases.     

HIV-induced IL-6/IL-10 dysregulation of CD4 cells is associated with defective B cell help and autoantibody formation against CD4 cells

To analyse CD4 cell cytokine secretion and helper/suppressor function at a clonal level we established 446 CD4⁺ T cell clones (TCC) in four healthy controls, three HIV^? haemophilia patients, four CDC II,III and four CDC IV patients. Spontaneous TCC secretion of Th1 cytokines (IL-2, interferon-gamma (IFN-γ)) and Th2 cytokines (IL-4, IL-6, IL-10) was determined by ELISA. TCC helper and suppressor functions were tested in a pokeweed mitogen (PWM)-stimulated allogeneic co-culture system using a reverse haemolytic plaque assay for assessment of B cell responses. There was a significant association of TCC surface marker expression (Leu-8, CD45RA) with TCC IL-6 secretion in healthy controls (P < 0.01), HIV^? patients (P 0.001) and CDC II,III patients (P 0.01) but not in CDC IV patients. Likewise, TCC expression of Leu-8 and CD45RA was significantly associated with TCC suppressor function in healthy controls (P 0.0005) but not in HIV-infected patients. A reduced TCC helper frequency (10% of TCC) and an enhanced TCC suppressor frequency (> 80% of TCC) were detected only in those HIV-infected patients who showed an excessively increased TCC IL-6 secretion (> 70% of TCC) together with a significantly diminished TCC IL-10 secretion (10% of TCC). CD4 cell autoantibodies also were found only in patients with this type of cytokine dysregulation. These data indicate that CD4 cell surface markers lose their functional relevance in HIV-infected patients. HIV-induced IL-6/IL-10 dysregulation of CD4⁺ T cells, i.e. the up-regulation of spontaneous IL-6 and down-regulation of spontaneous IL-10 secretion, appears to be involved in inducing CD4 helper defects and may promote autoantibody formation against CD4 cells.     

Influence of breast-feeding on the production of cytokines

PROBLEM: Recently, we reported increases in the production of interferon-gamma (IFN-gamma), interleukin-2 (IL-2), and IL-4 during the postpartum period. The present study was undertaken to investigate whether these increases might be explained by increased prolactin while breast-feeding. METHOD: Whole blood from 41 women who were breast-feeding, 13 women not breast-feeding, and 31 healthy non-pregnant women was stimulated with phorbol 12-myristate 13-acetate and ionomycin, and the levels of cytokines in the supernatant were measured by enzyme-linked immunosorbent assay. Their serum levels of prolactin were measured by enzyme immunoassay. RESULTS: Increases in IFN-gamma, IL-2, IL-4, and IL-10 production were observed in women who were breast-feeding but not in women who were not breast-feeding. Serum levels of prolactin correlated with the levels of IFN-gamma in culture supernatant. CONCLUSIONS: These results suggest that breast-feeding induces production of cytokines and that IFN-gamma production is enhanced by physiological concentrations of prolactin.     

IL-10 stimulates production of platelet-activating factor by monocytes of patients with active systemic lupus erythematosus (SLE)

IL-10 displays modulatory properties on the synthesis of platelet-activating factor (PAF), a potent inflammatory mediator of vascular injury. Despite the fact that IL-10 is considered to be an anti-inflammatory cytokine, IL-10 levels correlate with disease activity in SLE. Moreover, in SLE IL-10 is unable to exert its immunosuppressive and anti-inflammatory effects. We have investigated the ability of IL-10 to stimulate PAF production from monocytes of SLE patients. Spontaneous and IL-10-stimulated PAF production by peripheral blood monocytes was measured in active (n = 13) and inactive (n = 14) SLE patients and in 15 normal control subjects. We observed that monocytes derived from patients with active SLE, but not from controls or inactive SLE, spontaneously produced significant amounts of PAF. Moreover, IL-10 enhanced the synthesis of PAF from monocytes of active SLE patients only. IL-10-induced PAF production correlated with the severity of the disease and with the extent of proteinuria. These results indicate that IL-10 only stimulates the synthesis of PAF from monocytes of SLE patients when immunologically active, suggesting that IL-10 may possess a paradoxical proinflammatory effect in SLE by promoting the production of PAF, a secondary mediator of inflammation.