Selective inhibition of hepatic collagen accumulation in experimental liver fibrosis in rats by a new prolyl 4-hydroxylase inhibitor

In order to detect and characterize allergen-specific T cells in the airways of atopic asthmatics, we measured proliferation and cytokine production by bronchoalveolar lavage (BAL) T cells isolated from Dermatophagoides pteronyssinus (Der p)-sensitive asthmatics and nonatopic control subjects, and compared the results with those generated using peripheral blood (PB) T cells. BAL and PB mononuclear cells were collected 24 h after segmental allergen challenge by fibreoptic bronchoscopy and venepuncture, respectively. T cells purified from BAL and PB were stimulated with autologous, irradiated antigen-presenting cells and D. pteronyssinus extract or a control, nonallergen antigen (M. tuberculosis purified protein derivative [PPD]). IL-5 and IFN-gamma concentrations were measured in culture supernatants by ELISA, and T-cell proliferation by 3H-thymidine uptake. D. pteronyssinus-induced proliferation of T cells derived from both BAL and PB was elevated in asthmatics when compared with control subjects (p < 0.05), whereas PPD-induced proliferation was equivalent in both compartments. In the asthmatics, D. pteronyssinus-induced proliferative responses of equivalent numbers of BAL and PB T cells obtained after allergen challenge were statistically equivalent. Nevertheless, BAL T cells stimulated with D. pteronyssinus produced significantly greater amounts of IL-5 than did PB T cells (p < 0.05). Allergen-induced proliferation and IL-5 production by BAL T cells in the asthmatics after segmental allergen challenge correlated with the percentages of eosinophils in the BAL fluid (p < 0.01). Further, BAL T cells from asthmatic patients produced significantly higher amounts of IL-5 than did the same number of cells from nonatopic control subjects (p < 0.05). We conclude that, in D. pteronyssinus-sensitive asthmatics, allergen-specific T cells can be detected in the bronchial lumen after allergen challenge and that allergen-induced proliferation and IL-5 production by these cells correlates with local eosinophil influx. Although bronchial luminal T cells show an equivalent proliferative response to allergen stimulation as compared with PB T cells, they do produce more IL-5, consistent with the hypothesis that local differentiation or priming of these cells within the bronchial mucosal environment results in upregulation of allergen-induced IL-5 secretion.     

The importance of CD54 and CD86 costimulation in T cells stimulated with Candida albicans and Dermatophagoides farinae antigens in patients with atopic dermatitis

A number of studies have demonstrated an increased frequency of allergen-specific T cells producing increased amounts of interleukin-4 (IL-4) and IL-5, but little interferon-y in both the peripheral blood and skin lesions of patients with atopic dermatitis (AD). In this study, to further clarify the characteristics of T cells obtained from AD patients, we examined the dependency of the antigen-specific proliferation of peripheral blood mononuclear cells (PBMC) from AD patients on costimulatory molecules. The antigens used were Candida albicans and Dermatophagoides farinae, for which AD patients show increased levels of IgE antibodies. PBMC from control healthy donors stimulated with these antigens incorporated [3H]-thymidine much more than PBMC from AD patients. The addition of anti-CD54, -CD40, -CD80 and -CD86 monoclonal antibodies to the cultures showed that the PBMC required only CD54 and CD86 for stimulation with C. albicans, but required CD54, CD80 and CD86 for stimulation with D. farinae. Among these monoclonal antibodies, the anti-CD54 antibody suppressed the proliferative responses of most PBMC, most effectively followed by the anti-CD86 antibody. However, there were no significant differences in the requirement for costimulatory molecules of PBMC proliferation stimulated with C. albicans or D. farinae between AD patients and healthy donors. Since many studies have suggested that T-helper type 1 and T-helper type 2 immune responses are different in their dependency on CD80 or CD86 costimulation, our present results suggest that the allergen-specific T cells of AD patients are not completely shifted to a T-helper type 2 subset.     

Allergen-induced cytokine production in atopic disease and its relationship to disease severity

The Th2 cytokines, interleukin (IL)-4 and IL-5, have an important role in atopic disease. CD30 is a transmembrane molecule that may be expressed on a proportion of activated T-lymphocytes and has been reported to be a marker for Th2 phenotype. Our objective was to compare the in vitro cytokine responses and CD30 expression of peripheral blood mononuclear cells (PBMCs) to stimulation with house dust mite antigen (Dermatophagoides pteronyssinus) in atopic asthmatics, atopic nonasthmatics, and normal subjects, and to see if atopic asthmatic cytokine production correlated with symptomatic disease activity and whether cytokine production was allergen-specific. Eighteen atopic asthmatics (all were allocated a symptomatic disease score), 6 atopic nonasthmatics, and 7 healthy nonatopic individuals were studied. Resting serum IL-4 levels were measured, then PBMCs were separated using Lymphoprep density centrifugation and cultured in modified RPMI 1640 medium. PBMCs were stimulated with IL-2 alone or with D. pteronyssinus (1,000 subcutaneous units/ml) with IL-2 and harvested after 5 and 10 d. Using monoclonal antibodies and flow cytometry we obtained the percentage of CD4+ T cells expressing CD30 and the intensity of CD30 staining. Culture supernatants were analyzed for IL-4 and interferon gamma (IFN-gamma) using an enzyme-linked immunosorbent assay. In 9 atopic asthmatics PBMCs were also stimulated nonspecifically using phytohemagglutinin (PHA). IL-4 was detectable in the serum of atopic subjects but not in normal subjects. Stimulation of PBMCs with D. pteronyssinus produced significant amounts of IL-4 in atopic asthmatics and atopic nonasthmatics, but minimal quantities in normal subjects. Much lower levels of IFN-gamma were produced by atopic asthmatics in response to D. pteronyssinus compared to atopic nonasthmatics. IFN-gamma levels had an inverse correlation with asthmatic symptom score. CD4+ T-cell expression of CD30 also correlated inversely with IFN-gamma production and IFN-gamma:IL-4 ratio. PHA produced minimal levels of IL-4 compared to specific allergen stimulation. It is concluded that different groups of atopic patients exhibit different patterns of allergen-induced cytokine production. In vitro allergen-induced cytokine production in atopic asthmatics correlated with symptomatic disease activity, and is allergen-specific.     

Requirement of CD28-CD86 co-stimulation in the interaction between antigen-primed T helper type 2 and B cells

The interaction between CD28 and its ligands, CD80 and CD86, is crucial for an optimal activation of antigen-specific T cells. However, the requirement of CD80 or CD86 co-stimulation in Th2 cell differentiation and activation is controversial. Freshly isolated murine CD4+ and CD8+ T cells were incubated with P815 transfectants expressing a similar level of either CD80 or CD86 in the presence of anti-CD3 mAb. Both CD80 and CD86 co-stimulated the proliferation of CD4+ and CD8+ T cells at comparable time-kinetics and magnitude, but CD86 alone was able to co-stimulate IL-4 and especially IL-10 production in CD4+ T cells. In typical Th2-dependent immune responses elicited by Nippostrongylus brasillensis infection, the anti-CD86 mAb treatment but not the anti-CD80 mAb treatment efficiently inhibited antigen-specific IgE and IgG1 production, which was accompanied with the reduced IL-4 production. Our results suggest that CD86 co-stimulation plays a dominant role not only in the primary activation of Th2 cells but also in the secondary interaction between antigen-primed Th2 cells and B cells.     

Costimulation through B7-2 (CD86) is required for the induction of a lung mucosal T helper cell 2 (TH2) immune response and altered airway responsiveness

The recruitment of eosinophils into the airways after allergen exposure is dependent on interleukin (IL) 5 secreted from antigen-specific CD4+ T cells of the T helper cell (Th) 2 subset. However, while it is established that costimulation through CD28 is required for TCR-mediated activation and IL-2 production, the importance of this mechanism for the induction of a Th2 immune response is less clear. In the present study, we administered the fusion protein CTLA-4 immunoglobulin (Ig) into the lungs before allergen provocation to determine whether CD28/CTLA-4 ligands are required for allergen-induced eosinophil accumulation and the production of Th2 cytokines. Administration of CTLA-4 Ig inhibited the recruitment of eosinophils into the lungs by 75% and suppressed IgE in the bronchoalveolar lavage fluid. CTLA-4 Ig also inhibited the production of IL-4, IL-5, and IL-10 by 70-80% and enhanced interferon-gamma production from CD3-T cell receptor-activated lung Thy1.2+ cells. Allergen exposure upregulated expression of B7-2, but not B7-1, on B cells from the lung within 24 h. Moreover, airway administration of an anti-B7-2 monoclonal antibody (mAb) inhibited eosinophil infiltration, IgE production, and Th2 cytokine secretion comparable in magnitude to that observed with CTLA-4 Ig. Treatment with an anti-B7-1 mAb had a small, but significant effect on eosinophil accumulation, although was less effective in inhibiting Th2 cytokine production. The anti-B7-2, but not anti-B7-1, mAb also inhibited antigen-induced airway hyperresponsiveness in vivo. In all of the parameters assessed, the combination of both the anti-B7-1 and anti-B7-2 mAb was no more effective than anti-B7-2 mAb treatment alone. We propose that strategies aimed at inhibition of CD28 interactions with B7-2 molecules may represent a novel therapeutic target for the treatment of lung mucosal allergic inflammation.     

Involvement of CD40 ligand-CD40 and CTLA4-B7 pathways in murine acute graft-versus-host disease induced by allogeneic T cells lacking CD28

The blockade of B7, using B7 antagonists such as anti-CD80 and/or -CD86 mAbs or CTLA4Ig in vivo, has been shown to induce an efficient suppression of T cell-mediated immune responses in allograft, allergy, and autoimmune models. However, this treatment does not result in complete tolerance. In this study, we examined CD28-B7-independent activation pathways in the pathogenesis of graft-vs-host disease (GVHD) using allogeneic T cells from CD28-deficient mice. Acute GVHD was induced in the absence of CD28 on donor T cells and its manifestations were obvious in the lymphoid tissues. The CD28-independent GVHD was significantly improved by treatment with anti-CD40 ligand (CD40L) mAb. In contrast, treatment with anti-CD80 plus anti-CD86 mAbs exacerbated the clinical manifestations of GVHD and increased the T cell response against host alloantigen, resulting in the expression of CTLA4, CD40L, and CD25 on splenic T cells. These data suggested that the CD40L-CD40 pathway significantly contributed to the CD28-independent pathogenesis of acute GVHD, whereas the CTLA4-B7 pathway acted protectively in the development of GVHD. These results imply that selectively blockading CD28, instead of disrupting both CD28 and CTLA4, would be a better therapeutic strategy for GVHD. Additionally, the simultaneous use of CD40 antagonists may be advantageous.     

CD40 expressed on thymic epithelial cells provides costimulation for proliferation but not for apoptosis of human thymocytes

Human thymic epithelial cells express CD40, so we examined the possible role of CD40 in activation of thymocytes. We observed that both CD4+CD8- and CD4-CD8+ thymocytes proliferate after stimulation by anti-CD3 mAb in the presence of cultured thymic epithelial cells. Costimulation of CD4+ thymocytes by thymic epithelial cells is partly inhibited by an anti-CD40 mAb, but this mAb has no effect on costimulation of CD8+ thymocytes. The selective costimulatory ability of CD40 for CD4+ thymocytes was confirmed in experiments in which thymocytes were stimulated with anti-CD3 in the presence of murine P815 cells transfected with CD40 cDNA. The level of costimulation induced by P815-CD40 was comparable with that induced by P815 cells expressing CD80 (B7.1). Treatment of thymocytes with the Ca2+ ionophore ionomycin and the phorbol ester PMA or with anti-CD3 mAb resulted in up-regulation of the CD40 ligand, suggesting that this molecule is involved in CD40-mediated costimulation of human thymocytes. Costimulation of thymocytes by CD80 strongly increased anti-CD3-induced death of fetal thymocytes. In contrast, costimulation by CD40 did not increase anti-CD3-mediated apoptosis of these thymocytes. To confirm that CD40 does not affect anti-CD3-induced cell death, we established a variant of the Jurkat T leukemic cell line that constitutively expresses CD40L and analyzed the sensitivity of this cell line for activation-induced apoptosis. In contrast to CD80, CD40 failed to increase anti-CD3-mediated apoptosis in CD40L+ Jurkat cells, whereas both CD40 and CD80 strongly increased IL-2 production induced by anti-CD3. These findings suggest that costimulation by CD40 is involved in clonal expansion of CD4+ thymocytes but not in activation-induced cell death.     

Selective inhibition of T cell proliferation but not expression of effector function by human alveolar macrophages

BACKGROUND: Alveolar macrophages are thought to play an important part in regulating lung immune responses. While it is clear that human alveolar macrophages suppress T cell proliferation in vitro, the mechanisms by which this is achieved are not clear, nor is it known whether alveolar macrophages also inhibit other aspects of T cell function. METHODS: Peripheral blood mononuclear cells were stimulated with phytohaemagglutinin or house dust mite allergen, and cultured with variable numbers of autologous alveolar macrophages obtained by bronchoalveolar lavage from 20 normal subjects. RESULTS: Alveolar macrophages induced a reversible inhibition of T cell proliferation in response to both mitogen and allergen stimulation, with the latter being considerably more susceptible to inhibition. This was achieved via heterogenous mechanisms, involving both soluble factors derived from alveolar macrophages and cell-cell contact. Despite inhibiting proliferation, alveolar macrophages had little or no effect on T cell calcium flux, the characteristic changes in CD3, CD2, CD28 and interleukin-2 (IL-2) receptor expression which accompany normal T cell activation, and IL-2 and interferon gamma secretion. In contrast, alveolar macrophages inhibited the tyrosine phosphorylation of proteins which may be involved in IL-2 receptor-associated signal transduction. CONCLUSIONS: The immunoregulatory properties of alveolar macrophages are relatively selective, allowing T cell activation and cytokine secretion while inhibiting T cell proliferation within the lung.     

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BACKGROUND: Multiple sensory neuropeptides are present in human airways and may contribute to diseases such as asthma. This study quantified and characterised substance P (SP), neurokinin A (NKA), and calcitonin gene related peptide (CGRP) immunoreactivity in bronchoalveolar lavage fluid in asthmatic and normal subjects. METHODS: Using specific radioimmunoassay (RIA), SP, NKA and CGRP were measured in bronchoalveolar lavage fluid from asthmatic subjects (n = 5), normal subjects (n = 5), atopic non-asthmatic subjects (n = 6), and asthmatic subjects four hours after allergen challenge (n = 12). Peptide immunoreactivity was characterised using high performance liquid chromatography (HPLC) and RIA. RESULTS: No SP or CGRP immunoreactivity was detected in any of the fractions from samples after extraction, HPLC, and RIA. Non-specific binding resulted in spurious SP immunoreactivity being detected in bronchoalveolar lavage fluid when no extraction process was employed. NKA was detected in significant amounts in asthmatic (median 550, range 425-625 pg/ml) and normal subjects (median 725, range 350-1425 pg/ml). The level of NKA was significantly higher in the asthmatic subjects after allergen challenge (median 750, range 350-1250 pg/ml) than in unchallenged asthmatic subjects (median 600, range 425-600 pg/ml, p < 0.01). CONCLUSIONS: Extraction and characterisation of peptides from bronchoalveolar lavage fluid must be performed to ensure that the measured immunoreactivity represents target peptide. NKA is present in bronchoalveolar lavage fluid in high concentrations and is the predominant tachykinin. The concentrations of NKA are similar in normal subjects and subjects with mild asthma.     

Local expansion of allergen-specific CD30+Th2-type gamma delta T cells in bronchial asthma

BACKGROUND: T lymphocytes infiltrating airways during the allergic immune response play a fundamental role in recruiting other specialized cells, such as eosinophils, by secreting interleukin 5 (IL-5), and promoting local and systemic IgE synthesis by producing IL-4. Whether these presumed allergen-specific T cells are of mucosal or systemic origin is still a matter of conjecture. MATERIALS AND METHODS: Immunophenotype, IL-4 production, and in vitro proliferative response to specific or unrelated allergens were analyzed in the bronchoalveolar lavage (BAL) fluid lymphocyte suspensions obtained from untreated patients with allergic asthma. Healthy subjects and patients affected by pulmonary sarcoidosis, a granulomatous lung disease characterized by infiltrating Th1 CD4+ lymphocytes, served as controls. RESULTS: The proportions of gamma delta T lymphocytes, mostly CD4+ or CD4- (-)CD8-, was higher in asthmatic subjects than in controls (p < 0.05). Most BAL gamma delta CD4+ lymphocytes of asthmatic patients displayed the T cell receptor (TCR)-gamma delta V delta 1 chain. While CD30 antigen coexpression on the surface of BAL alpha beta(+) T lymphocytes was low (ranging from 5 to 12%), about half of pulmonary gamma delta T cells coexpressed it. These cells produced IL-4 and negligible amounts of interferon-gamma (IFN gamma), and proliferated in vitro in response to purified specific but not unrelated allergens. In contrast, control or sarcoidosis gamma delta T cells never displayed the CD30 surface molecule or produced significant quantities of IL-4. CONCLUSIONS: These findings not only confirm our previous hypothesis that the allergen-specific Th2-type lymphocytes found in the lungs of asthmatic patients are gamma delta T cells belonging to airway mucosal immunocytes, but also strongly support the notion that asthma is a local rather than a systemic disease.     

Organization of the human RH50A gene (RHAG) and evolution of base composition of the RH gene family

The alpha4 chain (CD49d), which constitutes one of the chains of alpha4beta1 (very late activating antigen-4 [VLA-4]) and alpha4beta7 integrins, mediates migration of T cells to extravascular spaces. The interaction between VLA-4 and vascular cell adhesion molecule-1 (VCAM-1) has been shown to be the critical pathway for the selective accumulation of eosinophils and basophils at sites of allergic inflammation. T lymphocytes are also specifically recruited into allergic sites, including the allergic asthmatic airway. Increased numbers of activated CD4+ cells expressing the DR antigen subset of the human leukocyte antigens (HLA-DR) appear in the allergic lung 48 h after allergen inhalation. The mechanisms by which these cells localize into the lung are still unknown. We report that stimulation of allergen-specific T cells with allergen in vitro resulted in enhanced expression of alpha4 chain (CD49d) as measured by receptor density on allergen-specific T-cell lines and T-cell clones. Kinetic studies showed that CD49d density was enhanced over a 24- to 48-h period in a time-dependent fashion, and was coordinately upregulated with HLA-DR expression. We also demonstrated that increased expression of CD49d on T-cell lines 24 h and 48 h after stimulation correlated with increased adhesion to the CS-1 fragment of fibronectin. In contrast, lymphocyte function-associated antigen-1b (LFA-1b) (CD11b), LFA-3 (CD58), and intercellular adhesion molecule-1 (ICAM-1) (CD54) expression did not change with allergen stimulation. We also showed that CD49d receptor density on T cells obtained by bronchoalveolar lavage (BAL) of allergic patients before and 48 h after allergen challenge was significantly higher than that on T cells taken from BAL of normal subjects and from controls with other inflammatory lung diseases. Taken together, these findings indicate that allergen stimulation activates allergen-specific T cells and coordinately induces increased CD49d receptor expression and binding to counterligands. We postulate that allergen-driven upregulation of CD49d, which together with the beta1 chain constitutes VLA-4 integrin, may be responsible for the selective accumulation of T cells in the allergic asthmatic lung.     

T cell proliferation-augmenting activities of the gene 3 protein derived from a phage library clone with CD80-binding activity

We have isolated a phage clone, F2, by panning a phage library with a CTLA4-conformation recognizing mAb (anti-CTLA4 mAb). The unique sequence of 15 amino acids with an internal disulfide bond was inserted in the gene 3 proteins of F2 phage clone (F2-g3p). We show here that 1) F2-g3p was recognized with anti-CTLA4 mAb but not with anti-CD28 mAb, and 2) F2-g3p bound to CD80 but not to CD86. The surface plasmon resonance analysis showed that F2-g3p strongly bound CD80. F2-g3p inhibited the binding of CTLA4 to CD80 but not to CD86. In contrast, F2-g3p weakly inhibited the binding of CD28 with CD80. When hen egg lysozyme (HEL)-primed lymph node cells were stimulated with HEL in the presence of F2-g3p in vitro, cell proliferation was highly potentiated. In the absence of antigenic stimulation, F2-g3p induced no T cell proliferation, indicating the costimulatory nature of F2-g3p. The T cell-augmenting activity of the F2 clone was eliminated when the F2 clone was preincubated with CD80-Ig before the addition to the cultures, indicating the involvement of CD80-binding in the F2-g3p-mediated immunopotentiation. Thus, the F2 motif conferred CD80-binding activity and an immunoregulatory function to the g3p.     

Human effector memory T cells express CD86: a functional role in naive T cell priming

The glycoprotein CD86 expressed on APCs provides a costimulatory signal necessary for an efficient activation of naive T cells. In contrast, there is controversy about the condition of expression and the function of CD86 on T cells. In this study, we have analyzed the phenotype and the biological activity of CD86+ T cells generated from human PBMC. Results show that CD86 expression on T cells is induced by long term stimulation via CD3 and IL-2R and is down-regulated as the cells become quiescent. The CD86-expressing cells are memory effector T cells: 1) they express CD45RO and high levels of the activation markers CD25, CD54, and HLA-Dr; 2) they selectively express CD30, CD40-ligand, and CD70; and 3) in response to stimulation, most of them produce IFN-gamma before dying by apoptosis. We then analyzed whether CD86 expressed on T cells is functional. Results show that paraformaldehyde-fixed CD86+ T cells enhance the proliferation and production of IFN-gamma by anti-CD3 mAb-stimulated naive T cells and induce proliferation of resting allogenic T cells. All these effects are prevented by neutralizing anti-CD86 mAbs. In contrast, we report no autocrine effect of CD86 in CD86+ T cell activation. In conclusion, these data show that human memory effector T cells express a functional form of CD86 that can costimulate naive T cell responses.     

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Eosinophil differentiation is thought to occur by the action of interleukin (IL)-5 on CD34(+) progenitor cells. The allergen-induced increase in eosinophil numbers in isolated airway preparations in vitro, and detection of increased numbers of circulating CD34(+) cells in atopic subjects, led us to the hypothesis that the eosinophil infiltration of the airway in asthma may result from local mucosal differentiation, in addition to recruitment from the bone marrow. We examined CD34(+) cell numbers by immunohistochemistry and IL-5 receptor alpha (IL-5Ralpha) messenger RNA (mRNA) expression by in situ hybridization in bronchial biopsies from atopic asthmatic patients, and from atopic and nonatopic control subjects. CD34(+) cell numbers were increased in the airway in atopic asthmatic and atopic nonasthmatic subjects. In contrast, CD34(+)/ IL-5Ralpha mRNA+ cells were increased in asthmatic subjects when compared with both atopic and nonatopic control subjects. Airway numbers of CD34(+)/IL-5Ralpha mRNA+ cells were correlated to airway caliber in asthmatic subjects and to eosinophil numbers. These findings support the concept that eosinophils may differentiate locally in the airway in asthma.     

CD4+ T cells can induce airway hyperresponsiveness to allergen challenge in the brown norway rat

Airway hyperresponsiveness to inhalational challenge with methacholine (MCh) develops by 32 h after allergen challenge of actively sensitized BN rats. To test the hypothesis that CD4+ T cells mediate allergen-induced hyperresponsiveness independent of IgE-mediated mechanisms, we administered CD4+ T cells, CD8+ T cells, and a mixture of CD4+ and CD8+ T cells (total T cells) isolated from the cervical lymph nodes of rats sensitized with ovalbumin (OA) to naive BN rats that underwent aerosol challenge with either OA or bovine serum albumin (BSA) 2 d later. Responsiveness to MCh was measured 2 d before transfer of T cells and 32 h after challenge with OA or BSA. Airway responsiveness increased significantly in recipients of CD4+ T cells after OA challenge, but not in any other of the treatment groups. Analysis of bronchoalveolar lavage (BAL) cells for major basic protein expression by immunostaining showed eosinophilia in OA-challenged CD4+ and total T-cell recipients. Cells retrieved by bronchoalveolar lavage showed increased expression of IL-5 mRNA (in situ hybridization) in CD4+ T cell recipients after OA challenge compared with other groups. Interferon-gamma mRNA was expressed to the greatest extent in CD8+ recipients, but it was elevated in both OA- and BSA-challenged animals. We conclude that CD4+ T cells can induce airway hyperresponsiveness after inhalational challenge with allergen and this is associated with IL-5 production and eosinophilia. CD8+ T cells may have a negative regulatory effect on responsiveness, possibly mediated by interferon-gamma.     

CD9-mediated costimulation of TCR-triggered naive T cells leads to activation followed by apoptosis

The induction of full activation or death in TCR-triggered T cells depends largely on whether appropriate costimulatory signals are provided. In this study, we show that the costimulation of CD9 on naive T cells during TCR stimulation results in transient, albeit potent, activation followed by apoptosis, rather than full activation. Anti-CD9 mAb synergized with suboptimal doses of anti-CD3 mAb in inducing T cell activation. [3H]TdR incorporation determined 2 days after CD9 costimulation was as potent as that induced by CD28 costimulation. In contrast to progressive T cell proliferation induced by CD28 costimulation, CD9 costimulation led to the induction of apoptosis of once-activated T cells. Although IL-2R expression was induced significantly earlier and to a greater degree after CD9 costimulation than after CD28 costimulation, CD9 costimulation only transiently produced a small amount of IL-2 and induced apparently low levels of bcl-xL compared with those observed in CD28 costimulation. Addition of rIL-2 to cultures of CD9 costimulation induced strikingly enhanced expression of bcl proteins, especially of bcl-xL, and protected TCR-stimulated T cells from apoptosis. These data indicate that CD9-mediated costimulation of TCR-triggered naive T cells leads to activation followed by apoptosis as the result of failure to generate a positive signal for sufficient levels of IL-2 production.     

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CD80 and CD86 (B7-1 and B7-2) are the ligands on antigen-presenting cells (APCs) which bind CD28 and deliver the costimulatory signals necessary for T cell activation. The reasons for the existence of two CD28 binding molecules are not well understood. We created a mutant version of CTLA4-Ig that could selectively bind CD80 and block CD28-CD80 interaction but leave CD28-CD86 binding intact. CD80 blockade prevented antigen-induced accumulation of eosinophils and lymphocytes in the lung of immunized mice, but did not block antigen induced systemic blood eosinophilia or IgE antibody production. No preferential expression of CD80 could be demonstrated on a population of lung APC consisting mainly of macrophages. These results indicate that CD80 costimulation is not necessary for the induction of Th2 immune responses but rather for the maintenance or amplification of lung inflammatory responses.