Cytokine profile of protective anti-Trichinella spiralis CD4+ OX22− and non-protective CD4+ OX22+ thoracic duct cells in rats: secretion of IL-4 alone does not determine protective capacity

We analysed the cytokine profile of a T cell subset (CD4⁺ CD45 RC⁻) that confers protection against Trichinella spiralis infection in rats. These CD4⁺ cells are generated in the gut and appear in the thoracic duct lymph within 72 h after infection. Cytokine mRNA levels for IL-2, IL-3, IL-4, IL-5, IL-10 and IFN-γ and functional cytokine secretion for IL-4, IL-5, IFN-γ, TNF-α and mast cell differentiation activity were tested in vitro following stimulation with T. spiralis antigens. Compared to a non-protective T cell population (CD4⁺ CD45 RC⁺ or CD8⁺), also isolated from the same thoracic lymph, no significant differences were observed in the levels of mRNA for IL-2, IL-3, IL-4, IL-5, IL-10 or IFN-γ in the protective CD4⁺ CD45 RC⁻ cells. However, analysis of the cytokine activities in culture supernatant of these T cell subsets following 24 h stimulation in vitro with T. spiralis antigens showed that significant IL-4 and IL-5 activity but little IFN-γ or TNF-α was secreted by the protective CD4⁺ CD45 RC⁻ cells. Whereas the non-protective CD4⁺ CD45 RC⁺ cells secreted significant levels of IL-4, IFN-γ, mast cell differentiating activity and TNF-α but little IL-5 activity. Nonprotective CD8⁺ cells were found to secrete IL-4 but not IL-5. Production of IL-4 was essentially equal for both protective and non-protective T cell subsets. These findings suggest that the presence or absence of IFN-γ secretion, rather than IL-4 alone, determines whether a T cell subset has protective activity against T. spiralis infection in rats.     

A potential immunopathogenic role for reduced IL-35 expression in allergic asthma

Objective: Allergic asthma is a chronic airway inflammation resulting from an imbalance of T helper (Th) cell responses to allergens. Interleukin (IL)-35 has been shown to have potent immunoregulatory properties. Whether IL-35 participates in the immunopathogenesis of allergic asthma patients is still unknown. Methods: CD4⁺ T cells and CD4⁺CD25^? T cells were obtained from peripheral blood mononuclear cells (PBMCs) using magnetic separation. The concentration of IL-35 in plasma was measured by enzyme-linked immunosorbent assay (ELISA). The mRNA expression levels of the IL-35 subunits, EBI3 and IL-12p35, were detected by quantitative real-time PCR (qPCR). The proliferative responses of CFSE-labeled CD4⁺CD25^? T cells in the presence or absence of rhIL-35 were evaluated by flow cytometry. Cytokine production of activated CD4⁺CD25^? T cells was examined by flow cytometry and ELISA. Results: IL-35 protein and mRNA levels were decreased in allergic asthmatics. The frequencies of CD4⁺CD25⁺Foxp3⁺ Tregs and CD4⁺IL-12p35⁺ T cells in allergic asthma patients were lower than in healthy controls. Moreover, the addition of rhIL-35 suppressed CFSE⁺CD4⁺CD25^? T cell proliferation in vitro in a dose-dependent manner, and the suppression induced by rhIL-35 was associated with decreases in IL-4 but not IFN-γ and IL-17 production of activated CD4⁺CD25^? T cells. The increased level of Th1/Th2 was observed in allergic asthmatics in the presence of rhIL-35. Conclusions: Our data suggest that IL-35 can effectively suppress the proliferation and IL-4 production of activated CD4⁺CD25^? T cells in allergic asthma, and that IL-35 may be a new immunotherapy for asthma patients.     

Increased IL-4- and IL-17-producing CD8+ cells are related to decreased CD39+CD4+Foxp3+ cells in allergic asthma

Objective: In allergic asthma, regulatory T cell (Treg) number and function are decreased. Antigen-primed CD8⁺ T cells play an indispensable role in the full development of airway inflammation and airway hyper-responsiveness (AHR) occurring in asthma. In this study, we investigated the relationship between subpopulations of CD8⁺ T cells and CD39⁺ Tregs. Methods: Female C57BL/6 mice were used to develop the model of allergic asthma. Experimental mice were immunized with ovalbumin (OVA) by intra-peritoneal (i.p) injection and then challenged with OVA by intra-tracheal administration. Control mice were immunized with vehicle by i.p injection and challenged with OVA. Airway inflammation was determined by histology and AHR was measured by an invasive method. Levels of interferon (IFN)-γ, IL-4, and IL-17 in bronchoalveolar lavage fluid (BALF) were determined by enzyme-linked immunosorbent assay. The frequencies of CD8⁺IFN-γ⁺ cells (Tc1), CD8⁺IL-4⁺ cells (Tc2), CD8⁺IL-17⁺cells (Tc17), and CD39⁺Tregs were measured by flow cytometry. The correlation between CD39⁺Tregs and Tc subsets was analyzed by Pearson’s test. Results: Experimental mice displayed phenotypes of allergic asthma, including inflammatory cell infiltration into the lungs, goblet cell hyperplasia, increased airway resistance, and increased IL-4 and IL-17 in BALF. Compared to control mice, experimental mice displayed lower CD39⁺Tregs and Tc1 but higher Tc2 and Tc17. There was a negative correlation between CD39⁺Tregs and Tc2 or Tc17. Conclusion: In allergic asthma, increased Tc2 and Tc17 are possibly related to insufficient CD39⁺Tregs.     

Cell-contact-dependent activation of CD4+ T cells by adhesion molecules on synovial fibroblasts

Objective: To determine how cell–cell contact with synovial fibroblasts (SF) influence on the proliferation and cytokine production of CD4^+?T cells.

Methods: Naïve CD4^+?T cells were cultured with SF from rheumatoid arthritis patients, stimulated by anti-CD3/28 antibody, and CD4^+?T cell proliferation and IFN-γ/IL-17 production were analyzed. To study the role of adhesion molecules, cell contact was blocked by transwell plate or anti-intracellular adhesion molecule-1 (ICAM-1)/vascular cell adhesion molecule-1(VCAM-1) antibody. To study the direct role of adhesion molecules for CD4^+?T cells, CD161^+?or CD161^- naïve CD4^+?T cells were stimulated on plastic plates coated by recombinant ICAM-1 or VCAM-1, and the source of IFN-γ/IL-17 were analyzed.

Results: SF enhanced naïve CD4^+?T cell proliferation and IFN-γ/IL-17 production in cell-contact and in part ICAM-1-/VCAM-1-dependent manner. Plate-coated ICAM-1 and VCAM-1 enhanced naïve CD4^+?T cell proliferation and IFN-γ production, while VCAM-1 efficiently promoting IL-17 production. CD161^+?naïve T cells upregulating LFA-1 and VLA-4 were the major source of IFN-γ/IL-17 upon interaction with ICAM-1/VCAM-1.

Conclusion: CD4^+?T cells rapidly expand and secrete IFN-γ/IL-17 upon cell-contact with SF via adhesion molecules. Interfering with ICAM-1-/VCAM-1 may be beneficial for inhibiting RA synovitis.     

The immune-modulatory effects of a mixed herbal formula on dendritic cells and CD4+ T lymphocytes in the treatment of dust mite allergy asthma and perennial allergic rhinitis

Objective: Asthma and allergic rhinitis are chronic inflammatory diseases of the conducting nasal airway. Traditional Chinese medicine has long been used for supplemental therapy of allergic diseases, especially asthma and allergic rhinitis. We previously reported the effects of a mixed herbal formula in patients with allergic rhinitis. However, the immune-modulatory mechanism underlying the effects of herbal medicine for the treatment of allergic diseases remains unclear. Methods: We investigated the physiologic changes in dendritic cell (DC) and CD4⁺ T cell activity in patients with asthma and allergic rhinitis who were treated with a mixed herbal formula composed of Xin-yi-san?+?Xiao-qing-long-tang?+?Xiang-sha-liu- jun-zi-tang. Specifically, we set up in vitro autologous or heterologous co-culture experiments between DCs and CD4⁺ T cells, and used flow cytometry and ELISA to analyze the expression of surface molecules on DCs and the release of cytokines by CD4⁺ T cells. Results: Expression of HLA-DR on DCs was suppressed following treatment with the mixed herbal formula. Surface expression of CD40, CD54 and CD86 on DCs was also attenuated after treatment. In autologous co-cultures, CD4⁺ T cells increased their IL-10 production while decreasing TNF-α production. In heterologous co-cultures, IL-10 secretion by T cells was enhanced, while IL-12, IL-4, IL-5 and TNF-α secretion were reduced. Conclusion: Our mixed herbal formula attenuated the allergic reaction by modifying the physiologic function of the DC–CD4⁺ T cell interaction. Further investigations are necessary to understand the mechanism of immune modification mediated by the mixed herbal formula.     

IFN-γ stimulation of dental follicle mesenchymal stem cells modulates immune response of CD4+ T lymphocytes in Der p1+ asthmatic patients in vitro

BackgroundHouse dust mite (Dermataphagoides pteronyssinus) is a widespread risk factor in the development of asthma. CD4⁺ T lymphocytes have an important role in the pathogenesis of allergic asthma by polarizing to Th2 cells.ObjectiveWe aimed to evaluate the immunoregulatory effects of dental follicle mesenchymal stem cells with and without IFN-γ stimulation on peripheral blood mononuclear cells of house dust mite sensitive asthmatic patients, and compared those with Dexamethasone as a systemic steroid.Material and methodsPBMC of asthmatic patients and healthy individuals separately cultured with or without DF-MSCs in the presence and absence of IFN-γ or Der p1 or Dexamethasone for 72 h. CD4⁺ T proliferation, cell viability, CD4⁺CD25⁺FoxP3⁺ Treg cell frequency and cytokine profiles of PBMC were evaluated via flow cytometry.ResultsDF-MSCs suppressed proliferation of CD4⁺ T lymphocytes (p_CDmix < 0.01, p_Derp1 < 0.01, p_IFN < 0.005) by increasing the number of FoxP3 expressing CD4⁺CD25⁺ T regulatory cells (p_CDmix < 0.005, p_Derp1 < 0.01, p_IFN < 0.001) and suppressed lymphocyte apoptosis (p_CDmix < 0.05, p_Derp1 < 0.05, p_IFN < 0.05), while Dexamethasone increased the apoptosis and decreased Treg cell frequency in asthmatic patients. IFN-γ stimulation increased the suppressive effect of DF-MSCs and also enhanced the frequency of FoxP3 expressing CD4⁺CD25⁺ T regulatory cells. The cytokine levels were regulated by DF-MSCs by reducing IL-4 cytokine levels (p_CDmix < 0.01, p_Derp1 < 0.05, p_IFN < 0.05) and upregulating IFN-γ levels (p_CDmix < 0.01, p_Derp1 < 0.05, p_IFN < 0.005) in asthmatic patients.ConclusionIFN-γ stimulated DF-MSCs were found to have a high modulatory effect on CD4⁺ T cell responses, while Dexamethasone had an apoptotic effect on CD4⁺ T cells in asthmatic patients. DF-MSCs may be a new cell-based therapy option for allergic diseases including asthma.     

Demonstration of Low-Regulatory CD25High+CD4+ and High-Pro-inflammatory CD28−CD4+ T-Cell Subsets in Patients with Ulcerative Colitis: Modified by Selective Granulocyte and Monocyte Adsorption Apheresis

Low-CD25^High+CD4⁺, a subset of regulatory CD25⁺CD4⁺ T cells and high-inflammatory CD28⁻CD4⁺ T cells can exacerbate ulcerative colitis (UC). This study sought to investigate the frequency of CD25^High+CD4⁺ and CD28⁻CD4⁺ T cells in patients with UC and the changes in these cells during Adacolumn granulocyte and monocyte adsorption apheresis (GMA). Subjects were 12 patients with active UC, 11 with quiescent UC, and 14 healthy volunteers (HVs). The mean clinical activity index was 15.7 ± 2.2 in active UC and 4.5 ± 1.1 in quiescent UC. Peripheral blood samples were stained with CD4, CD25, and CD28 antibodies for flow cytometry. Patients with active UC received GMA and blood samples were examined before and after the first GMA session. Patients with active UC (P < 0.04) or quiescent UC (P < 0.02) had a higher percentage of CD28⁻D4⁺T cells compared with HVs, while the percentage of CD28⁺CD4⁺ T cells was lower in both UC groups compared with HVs (P = 0.03 and P < 0.02). Patients with active UC had a lower percentage of CD25^High+CD4⁺T cells compared with quiescent UC patients (P < 0.001). A significant increase in CD25^High+CD4⁺ T cells was associated with GMA (P < 0.03). Low CD25^High+CD4⁺ and high CD28⁻CD4⁺ are prominent features in UC. The increase in CD25^High+CD4⁺ T cells induced by GMA should contribute to improved immune function. Additional studies are warranted, since a low frequency of CD25^High+CD4⁺ ₋ and a high frequency of CD28⁻CD4⁺ ₋ expressing T cells might be a predictor of clinical response to GMA.     

Interleukin-12 gene adjuvant increases the immunogenicity of virus-like particles of human papillomavirus type 16 regional variant strain

ObjectivesTo analyze the immunogenicity of virus-like particles (VLP) of human papillomavirus type 16 (HPV16) isolated in East China and the adjuvant potential of interleukin-12 (IL-12).MethodsThe variant HPV16 L1VLP expressed in sf9 insect cells were purified with cesium chloride gradient centrifugation. BALB/c mice were vaccinated with VLP (L1N), VLP with Freund's adjuvant (L1A) or VLP with IL-12 recombinant plasmid (L1P). HPV16 VLP specific IgG and IFN-γ level in the serum were detected by ELISA, and the percentage of CD4⁺ and CD8⁺ in spleen cells was detected with flow cytometry.ResultsThe titers of serum IgG antibodies in vaccinated groups were higher than in negative control and the serum antibodies mainly recognized conformation-dependent HPV16 VLP epitopes. Splenic CD4⁺ and CD8⁺ T cell subsets increased after vaccination in every experimental group, and CD8⁺ increased obviously in L1P group. The ratio of CD4⁺/CD8⁺ decreased in L1P group and increased in the other two groups, compared to control group. Vaccination induced specific secretion of IFN-γ in the serum of vaccinated group (p < 0.05), especially in the L1P group.ConclusionsVLP of HPV16 variant strain isolated in East China could induce humoral immunity and cellular immunity in mice, and IL-12 recombinant plasmid can enhance cellular immunity.     

CD4+Foxp3+ T cells,interleukin-35 (IL-35) and IL-10 in systemic lupus erythematosus patients: Relation to disease activity

Aim of the workTo evaluate three subtypes of CD4⁺Foxp3⁺ T cells, interleukin-35 (IL-35) and IL-10 in systemic lupus eryhtematosus (SLE) patients and study their relation to disease activity.Patients and methodsFifty SLE patients were included and divided according to the SLE disease activity index (SLEDAI) into 2 equal groups with activity or in remission. Twenty healthy subjects were included as controls. All subjects underwent flow cytometric analysis of CD4, CD25, CXCR5 and Foxp3 expression on T cells. Serum IL-35 and IL-10 levels were measured by ELISA.ResultsPatients were 46 females and 4 males with a mean age of 38.0 ± 10.0 years, disease duration of 9.2 ± 6.0 years. The mean SLEDAI was 6.8 ± 3.7 in active ones. SLE patients especially those with activity had significantly reduced percents of CD4⁺CD25⁺Foxp3⁺ and CD4⁺CXCR5⁺Foxp3⁺ T cells, but increased percents of CD4⁺CD25⁻Foxp3⁺ T cells. This was accompanied by significant higher levels of serum IL-35 and IL-10 (p < 0.0001). The SLEDAI in active patients significantly correlated with CD4⁺CD25⁻Foxp3⁺ T cell percent, serum IL-35 and IL-10 levels (p < 0.05) and inversely with the CD4⁺CD25⁺Foxp3⁺ and CD4⁺CXCR5⁺Foxp3⁺ T cell percents (p < 0.05). At cut-off values of 3.29% for CD4⁺CD25⁺Foxp3⁺ T cell, 7.62% for CD4⁺CD25⁻Foxp3⁺ T cell, 1.77% for CD4⁺CXCR5⁺Foxp3⁺ T cell, 22.04 pg/ml for IL-35 level and 30.51 pg/ml for serum IL-10 level were found to be highly sensitive and specific for detecting lupus activity.ConclusionCD4⁺Foxp3⁺ T cells, IL-35 and IL-10 showed high sensitivity and specificity for detecting SLE activity and may be considered as potentially promising therapeutic targets.     

Serum Levels of Interleukins (IL)-4, IL-5, IL-13, and Interferon-γ in Acute Asthma

T-cell activation and alteration of cytokine levels are involved in the pathogenesis of bronchial asthma. However, the profile of circulating T-lymphocyte subsets and related cytokines during acute asthmatic attacks is still unclear. We hypothesized that serum levels of interleukin (IL)-4, IL-5, and IL-13 would be increased, whereas IFN-γ would be decreased in acute asthma. The subjects enrolled in this study included 58 acute asthmatics, 22 asymptomatic asthmatics, and 10 healthy controls. Serum levels of IL-4, IL-5, IL-13, and IFN-γ were measured using a sandwich enzyme-linked immunosorbent assay. We correlated serum levels of IL-4, IL-5, IL-13, and IFN-γ with initial forced expiratory volume in 1 sec (FEV₁). Compared with control subjects, acute asthmatics had significantly increased levels of circulating IL-4 (p < 0.001), IL-5 (p < 0.001), and IL-13 (p < 0.001), although the differences were of borderline significance in serum IFN-γ (p = 0.069). There were also significant differences in the circulating levels of IL-4, IL-5, and IL-13 between acute asthmatics and asymptomatic asthmatics. There was no significant association between initial FEV₁ and serum levels of IL-4 or IL-13, however, among acute asthmatics, a lower initial FEV₁ was associated with higher IL-5 and/or lower IFN-γ levels. Our results suggest that serum levels of IL-4, IL-5, and IL-13 may be elevated in acute asthma, and that higher levels of IL-5 and/or lower levels of IFN-γ are associated with severe airway obstruction.     

Therapeutic Effect of Intratracheal Administration of Murine IL-4 Receptor Antagonist on Asthmatic Airway Inflammation

Objective. It is well known that IL-4 and IL-13 play critical roles in the pathogenesis of asthma. In this study, by overexpressing murine IL-4 receptor antagonist (mIL-4RA), a competitive antagonist for both IL-4 and IL-13, we investigated the therapeutic effects of mIL-4RA on mouse asthmatic airway inflammation. Material and methods. BALB/c mice were randomly divided into four groups: healthy control mice; ovalbumin (OVA) sensitized/challenged mice; OVA sensitized/challenged mice intratracheally administered with mIL-4RA plasmid (mIL-4RA group); and OVA sensitized/challenged mice intratracheally administered with control plasmid (control plasmid group). The airway inflammation was determined by histopathological examinations. Cytokines were measured by enzyme-linked immunosorbent assay (ELISA). Flow cytometry was used to analyze CD4 and CD8 T-lymphocyte subsets. Results. Compared to the control plasmid-treated mice, intratracheal administration of mIL-4RA expressing plasmid on the sensitization phase protected the mice from the subsequent induction of asthmatic airway inflammation. The eosinophilic infiltration in bronchoalveolar lavage fluid (BALF) was significantly reduced compared to that of the control (p < 0.01). Interestingly, intratracheal administration of mIL-4RA regulated the Th1/Th2 cytokine imbalance in local airway with increased IL-13 levels and decreased IFN-γ levels compared to the control plasmid group. However, although we did see the decreased level of IL-4 and IL-13 in serum, the serum level of IFN-γ is not changed in the mIL-4RA group, suggesting that mIL-4RA could not correct the imbalance of Th1/Th2 cytokines in serum. In addition, intratracheal administration of mIL-4RA had no effect on the ratio of CD4/CD8 T-lymphocyte subsets in the peripheral blood, lung, or spleen. Conclusions. This study demonstrated that intratracheal administration of mIL-4RA attenuated the asthmatic inflammation and regulated the Th1/Th2 cytokine imbalance in local airway with minimal systemic effects. This method may serve as a potential therapeutic option for treating asthma.     

Detection of type 2-like T-helper cells in hepatitis C virus infection: Implications for hepatitis C virus chronicity

One striking clinical feature of hepatitis C virus (HCV) infection is that more than 50% of patients with acute hepatitis C will develop chronic infection. To investigate its possible mechanisms, we examined the activation of type 2-like T-helper (Th2-like) cells relating to the development of chronicity. Peripheral blood CD4⁺ T-cell proliferation and cytokine secretion in response to a panel of recombinant HCV antigens including core (C22), envelope 1 (E1), E2, nonstructural (NS) protein 4 (C100), fusion protein of NS3 and NS4 (C200), and NS5 were assayed in 17 patients with acute hepatitis C. All six patients with self-limited disease had a significant CD4⁺ T-cell proliferation to C22, E1, C100, C200, and NS5, running parallel with the antigen-stimulated secretion of interleukin (IL)-2 and interferon γ (IFN-γ), but not with interleukin (IL)-4 and IL-10, indicating predominant Th1 responses. Among the remaining 11 patients who developed chronicity, 6, 2, and 9 cases showed a specific CD4⁺ T-cell response to C22, C100, and C200, respectively, and the responses were significantly lower than those of cases with recovery in terms of stimulation index (SI) (P < .05) and of antigen-stimulated IL-2 and IFN-γ production. Importantly, IL-4 and IL-10 (Th2 responses) were detectable, and C22-specific Th2-like T-cell clones could be generated from patients with chronicity. The data suggested that activation of Th2 responses in acute hepatitis C patients may play a role in the development of chronicity.(Hepatology 1997 Feb;25(2):449-58)     

Augmented human-tumor-cytolytic activity of peripheral blood lymphocytes and cells from a mixed lymphocyte/tumor culture activated by interleukin-12 plus interleukin-2, and the phenotypic characterization of the cells in patients with advanced carcinoma

In this study, we evaluated the ability of combination regimens of interleukin-12 (IL-12) and interleukin-2 (IL-2) to induce effective killer cells against human tumors in vitro, in peripheral blood lymphocytes (PBL) from 15 cancer patients and mixed lymphocyte/tumor culture (MLTC) cells from 16 cancer patients, and carried out a phenotypic analysis of the cells responsible for the lysis of the human tumors. The freshly prepared PBL were cultivated with IL-2 alone or IL-12/IL-2 for 10 days [lymphokine-activated killer (LAK) cell generation system]. The MLTC cells (PBL cultured with mitomycin-C-treated allogeneic G-415 tumor cells for 3 days) were further cultivated with IL-2 or IL-12/IL-2 for 7 days [cytotoxic T lymphocytes (CTL) generation system]. The cytolytic activities of the lymphoid cells cultivated with IL-12/IL-2 were significantly augmented in both the LAK and CTL generation systems, as compared with those of cells treated with IL-2 alone. In the LAK generation system, the cytolytic activities of the cells cultivated with IL-12/IL-2 were significantly decreased by the method of negative selection of CD11b⁺ or CD56⁺ cells using immunomagnetic beads. The CD8⁺-depleted cells showed a slight decrease of activity. The killer cell activities of the CD4⁺-depleted cells remained unchanged. In the CTL generation system, the activity was markedly reduced by the elimination of the CD8⁺ or CD11b⁺ or CD56⁺ cells. The combined data suggested that IL-12/IL-2-induced killer effector cells in the LAK generation system were mainly of the natural killer (NK) type, comprising CD8^–CD11b⁺, CD8^– CD16b^–, CD3^–CD56⁺, and partly possible CD8⁺ CD11b^–T cells. CD8⁺ CD11b^–T cells mixed with cells of the NK type, comprising CD8^–CD11b⁺, CD8^– CD16b^– and CD3^–CD56⁺ cells, were the population of killer effector cells induced by IL-12/IL-2 in the CTL generation system.Abbreviations IL interleukin - LAK lymphokine-activated killer - CTL cytotoxic T lymphocytes - PBL peripheral blood lymphocytes - NK natural killer - MLTC mixed lymphocyte/tumor cell culture - TIL tumor-infiltrating lymphocytes     

The role of Tα1 on the infective patients after hematopoietic stem cell transplantation

The present study was designed to investigate the effect of thymosin α1 (Tα1) administration in infective recipients of hematopoietic stem cell transplantation (HSCT) for hematologic malignancies. Eight patients were enrolled in our study, including seven allo-HSCT patients and one auto-HSCT patient. These patients were allocated randomly into the treatment group (four cases) and control group (four cases). Tα1 was used in the treatment group to test its effectiveness in infection control. The concentrations of cytokines IFN-γ, IL-2, IL-4, IL-10, and IL-12 were observed, and the levels of CD3⁺, CD4⁺, and CD8⁺ T cells, as well as of CD4⁺/CD8⁺ and CD4⁺/CD25⁺ regulatory T cell (Treg) were measured. When Tα1 was administered for 2 weeks, the concentrations of these cytokines were increased after 1 month in the treatment group. Interestingly, the levels of IFN-γ, IL-2, IL-10, and IL-12 were increased in the treatment group more than those in the control group, whereas there were no significant differences between the treatment and control group in the levels of CD3⁺, CD4⁺, and CD8⁺ T cells, or in CD4⁺/CD8⁺ or CD4⁺/CD25⁺ Treg cells. Notably, Tα1 administration did not cause acute or chronic graft versus host disease (GVHD). We conclude that Tα1 administration is safe and may impact favorably on immune function, and that it may improve resistance to infection and induce immunotolerance without GVHD.     

Endothelial Dysfunction in Patients with Asthma: The Role of Polymorphisms of ACE and Endothelial NOS Genes

Both atopy and asthma are claimed to be associated with a Th-2 cytokine pattern. We sought to determine the contribution of atopy and asthma to the observed Th-2/Th-1 imbalance in these conditions. Of 60 children aged 6–16 years that were included in the study, 13 were nonatopic nonasthmatic, 15 atopic nonasthmatic, 14 nonatopic asthmatic, and 18 atopic asthmatic. Atopic children had positive skin prick tests to grass pollens only. All children were studied after an asymptomatic and drug-free period of at least three months. Total IgE was measured in serum. Peripheral blood mononuclear cells were cultured and stimulated in vitro with phytohemagglutinin and interferongamma (IFN-γ) and interleukin-4 (IL-4) measured in the supernatants. Total IgE was significantly higher in atopic asthmatics compared to nonatopic asthmatics (p = 0.004), and nonatopic nonasthmatics (p = 0.001), but was not different from atopic nonasthmatics (p > 0.05). On the other hand, IL-4 was significantly elevated in atopic asthmatics and in nonatopic asthmatics compared to nonatopic nonasthmatics (p = 0.037 and p = 0.009, respectively). Although atopic asthmatics had lower IFN-γ values than nonatopic asthmatics, the difference did not reach statistical significance. No correlation was detected between any two parameters. Our results suggest that both atopy and asthma contribute to the increased levels of IL-4 and that, whereas nonatopic asthma is associated with increases in both IL-4 and IFN-γ release by mononuclear cells, only atopic asthma is characterized by a Th-2 type cytokine dominance.     

Functional analysis of synovial fluid and peripheral blood T cells from patients with rheumatoid arthritis

Rheumatoid arthritis is a T cell-mediated autoimmune disease. The lack of knowledge of the involved target antigens severely hampers research on relevant T cells in patients. Here we describe the functional analysis of freshly isolated T cells from the peripheral blood and the site of the lesion (synovial fluid or synovial membrane) of patients with rheumatoid arthritis. Healthy donors and osteoarthritis patients served as controls. Using various polyclonal stimuli, we analyzed CD4⁺ T cells with respect to proliferation and their ability to produce lymphokines. Our data show that lesion-derived CD4⁺ T cells of patients with rheumatoid arthritis are severely defective in proliferation and lymphokine (interleukin-2, interleukin-4, tumor necrosis factor-, interferon-) production. This activation defect was most pronounced at lower cell densities and was present in both synovial fluid derived and synovial membrane derived CD4⁺ T cells of all patients tested. No difference was found between responses of synovial fluid derived CD4⁺ T cells from osteoarthritis patients and those observed with peripheral blood derived T cells from all groups. The observed defect in lesion-derived CD4⁺ T cells from rheumatoid arthritis patients was not due to the effect of inflammatory factors in the synovial fluid because preincubation with synovial fluid could not induce a similar defect in control T cells. Together, our data show a rheumatoid arthritis specific, general defect in the activation of lesion-derived CD4⁺ T cells.Abbreviations IFN Interferon - IL Interleukin - MNC Mononuclear cells - OA Osteoarthritis - PB Peripheral blood - PSA Phosphate-buffered saline+bovin serum albumin+NaN₃ - PIAlb Phosphate-buffered saline+0.38% trisodiumcitrate+10% human albumin - RA Rheumatoid arthritis - SF Synovial fluid - SM Synovial membrane - TNF Tumor necrosis factor     

IFN-γ and TNF-α secretion by CD4+ and CD8+ TCRαβ+ T-cell clones derived early after allogeneic bone marrow transplantation

Abstract: Secretion of the potentially antileukaemic cytokines IFN-γ and TNF-α was investigated for CD4 ⁺ and CD8 ⁺ TCRαβ⁺ T-cell clones derived from 4 leukaemia patients 3–6 weeks after allogeneic BMT. We investigated cytokine secretion in response to the activation signal accessory cells + phytohaemagglutinin + Interleukin 2. All clones derived after BMT were capable of IFN-γ and TNF-α secretion, and both for CD4⁺ (n = 96) and CD8⁺ (n = 8) T cells quantities of IFN-γ and TNF-α were significantly correlated with one another. When comparing the overall results for posttransplant and normal T-cell clones derived from 2 bone marrow donors (n = 65), both CD4⁺ and CD8⁺ TCRαβ⁺ T-cell clones showed increased IFN-γ production, and CD4⁺ but not CD8⁺ clones showed a decreased TNF-α secretion. The results suggest that noncytotoxic T cells derived after allogeneic BMT can produce IFN-γ and TNF-α and may thus be capable of mediating antileukaemic effects.     

Increased surface receptor Fas (CD95) levels on CD4+ lymphocytes in patients with primary intestinal lymphangiectasia

Objective. Exudative enteropathy secondary to primary intestinal lymphangiectasia (PIL) is characterized by lymphopenia, hypogammaglobulinemia and hypoalbuminemia resulting from leakage of lymph fluid into the intestinal tract. The objective of this study was to better characterize the lymphopenia of PIL-confirmed patients. Material and methods. T-cell markers and T-cell proliferation/capacities (differentiation, activation and death) were analyzed for phenotype in 9 patients (6 F, 3 M, aged from 18 to 72 years). Results. Mean counts of CD4 and CD8 subsets were significantly decreased, 174±123/µl and 134±77/µl compared with controls, 858±260/µl and 482±164/µl, respectively (p<0.0001). Significant depletion of naïve (CD45RA⁺ CD62L⁺) CD4⁺ T cells was noted, with a mean expression of 7±4% compared with controls, 45±1% (p<0.0001). Both CD4⁺ and CD8⁺ T-cell mean subsets were activated as assessed by their proportion expressing the late activation markers HLA-DR, 18±7% and 19±9% compared with controls, 6±3% and 10±6%, respectively (p<0.0001 and p<0.01). The mean expression of CD95/Fas on CD4⁺ T cells was significantly higher in patients than in controls, 83±16% versus 45±13% (p<0.0001). No major abnormality of T-cell proliferation/capacities was observed. Conclusions. Our results suggest that the T-cell loss in PIL patients is probably due to various mechanisms including enteric lymphocytes loss and activation of residual T cells, leading to death. Moreover, this loss is not compensated by a sufficient increase in T-cell thymic production.     

The Decreased CD4+CD25+ FoxP3+ T Cells in Nonstimulated Allergic Rhinitis Patients Sensitized to House Dust Mites

Objective. Regulatory (CD4⁺CD25⁺) T cells have been shown to play an important role in the development of allergic diseases. This study aims to investigate CD4⁺CD25⁺ T cells, Forkhead box P3 (FoxP3⁺ cells), and T-helper 1/T-helper 2 (Th1/Th2) cytokines in newly diagnosed allergic rhinitis (AR) patients. Methods. Altogether, 10 subjects with AR and 12 age-matched nonallergic healthy subjects were included in this study. CD4⁺CD25⁺ T cells, FoxP3⁺ T cells in peripheral blood mononuclear cells (PBMCs) were evaluated by flow cytometry, and the Th1/Th2 cytokine levels were determined by cytometric bead array immunoassay in both PBMC supernatants and nasal lavage fluids. Results. The percentage of CD4⁺CD25⁺ T cells were significantly higher, whereas the percentage of FoxP3⁺ cells were lower in AR patients compared with healthy subjects. In PBMC culture supernatants, interleukin-10 (IL-10) levels were significantly lower (p = .012), whereas IL-4, IL-5, and tumor necrosis factor-α (TNF-α) levels in nasal lavage fluids were higher in AR patients compared with healthy subjects (p = .026, p = .015, p = .03, respectively). Conclusions. Our findings indicate that decrease in CD4⁺CD25⁺FoxP3⁺ T cell fraction and diminished levels of IL-10 are noteworthy without allergen stimulation in house dust mite AR patients.