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.     

Assessment of tissue viability using near-infrared spectroscopy

Both anti-CD40 antibodies and anti-immunoglobulin (Ig) coupled to Sepharose induced proliferation of resting B cells and suppressed lipopolysaccharide (LPS)-induced B-cell differentiation to immunoglobulin secretion at comparable levels determined with the plaque-forming assay and Ig RNA steady state levels. Anti-CD40 antibodies also increased the proliferation of B cells stimulated by T helper cells in vitro while suppressing their differentiation to Ig secretion. Further, B cells preactivated by anti-Ig, anti-CD40 or a combination of the two mitogens could be restimulated by anti-CD40 but not by anti-Ig antibodies. Phenotypic divergence of Ig and CD40 signals regarding surface expression of activation markers was observed. Restimulation of anti-Ig- or anti-CD40-prestimulated cells with anti-Ig induced apoptosis whereas apoptosis could be inhibited when cells were recultivated with anti-CD40.     

Perspective evaluation of the post-graduate courses of the Nursing Department at UFRN (1980-1992)

Superantigens have been used to study peripheral tolerance in CD4+ T cells. The superantigen SEB induces T cell anergy by promoting the differentiation of SEB-activated virgin T cells into anergic memory T cells. Memory T cells from SEB or antigen-primed mice do not proliferate when they are cultured with SEB. The present studies were performed to determine whether memory T cells fail to interact with SEB antigen-presenting cells or whether SEB promotes incomplete or negative signals in memory T cells. When murine virgin and memory T cells were separated on the basis of CD45RB expression and cultured with SEB-pulsed B cells, SEB induced the expression of CD25, which then mediated proliferation when IL-2 was added to the cultures. In addition, SEB promoted the expression of the CD40L, which is required for T helper cell function. Finally, PMA induced a costimulatory signal leading to the proliferation of these cells. Surprisingly, the agents, i.e., IL-2 and PMA, which induced TM cell proliferation in conjunction with SEB failed to induce lymphokine secretion. However, in the presence of IL-4 plus IL-5, the T memory cells induced the SEB-pulsed B cells to secrete IgM and IgG. These results suggest that memory T cells are not simply unresponsive to SEB but are actively anergized.     

Diminished expression of CD40 ligand may contribute to the defective humoral immunity in patients with MHC class II deficiency

Major histocompatibility complex (MHC) class II deficiency (bare lymphocyte syndrome, BLS) is a rare primary immunodeficiency classified as a subgroup of severe combined immunodeficiency. We studied T and B lymphocyte function by examining the CD40 ligand/CD40 system in three BLS patients from two unrelated families. CD40 ligand expression by maximally activated BLS T cells was diminished. This abnormality may represent immunological na?veté rather than a general T cell defect, since expression of activation marker CD69 and proliferative responses to PHA or anti-CD3 were normal, and BLS T cells primed and restimulated in vitro expressed normal amounts of CD40 ligand. BLS B cells proliferated and produced IgE if stimulated with anti-CD40 or soluble CD40 ligand and IL-4. Activation of BLS B cells with soluble CD40 ligand and IL-4 induced normal expression of activation markers, although MHC class II expression remained absent. Depressed antibody titers, lack of amplification and failure to undergo isotype switching in response to immunization with bacteriophage phi x 174 demonstrated defective T cell help. We conclude that BLS B cells are functionally normal if appropriately stimulated, and that the defective humoral immunity observed may be related to diminished expression of CD40 ligand on BLS T cells.     

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.     

CD28-induced cytokine production and proliferation by thymocytes are differentially regulated by the p59fyn tyrosine kinase

CD28 is a 44-kDa homodimeric receptor that is expressed on the majority of T cells. Engagement of the CD28 receptor by soluble anti-CD28 mAb in conjunction with phorbol ester (PMA) induces the production of cytokines and the proliferation of resting T cells via signal transduction pathways independent of the TCR. Evidence is provided herein that CD28 signals leading to cytokine production do not require the p59fyn (Fyn) tyrosine kinase, whereas CD28-mediated proliferation is dependent on the presence of the Fyn kinase in thymic, but not lymph node, cells. The defect in proliferation is not due to failure of IL-2R signaling, since addition of high concentrations of exogenous IL-2 can overcome the proliferative defect. Analysis of CD28-directed induction of the IL-2R alpha (CD25)-chain, which confers high affinity binding to IL-2, showed that Fyn-deficient thymocytes, but not lymph node cells, failed to up-regulate CD25 expression following anti-CD28 and PMA stimulation. Thus, the Fyn tyrosine kinase is critically required for thymic CD28-mediated CD25 expression and proliferation but not for CD28-mediated cytokine production.     

Partial activation of naive CD4 T cells and tolerance induction in response to peptide presented by resting B cells

Tolerance is thought to occur when Ag is presented to T cells in the absence of costimulatory interactions from APC accessory molecules. Of the professional APC, the resting B cell may be the main tolerizing cell in vivo. We have analyzed several aspects of activation of naive transgenic CD4 cells stimulated with resting or activated B cells presenting peptide Ag. Similar results were obtained with stimulation from peptide presenting fibroblast APC lacking or expressing B7-1 with intracellular adhesion molecule-1. TCR ligation with little or no accessory molecule coreceptor engagement induced efficient blastogenesis; up-regulation of CD25, CD44, CD69, CD95 and CD71; and down-regulation of CD62L over a 48-h period. Accessory molecule help enhanced the expression of CD25, CD44, CD69, and CD71, but to very modest degrees. Only two molecules, CD40 ligand and IL-2, were found to be extremely dependent on accessory molecule help, with little or no expression evident with peptide presented on resting B cells or class II-positive fibroblasts. T cells induced on resting B cells expanded minimally over 3 days, and this was followed by extensive cell death and hyporesponsiveness of the resulting cells. These studies suggest that under tolerizing conditions, such as Ag presentation by resting B cells, much of the naive CD4 response is induced efficiently. Partial activation, however, may be the overall result due to the lack of CD40 ligand expression, which may regulate costimulatory activity in APC and, in turn, may contribute to limiting the production of IL-2 required for T cell expansion and survival.     

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.     

Specific regulation of VLA-4 and alpha 4 beta 7 integrin expression on human activated T lymphocytes

Modulation of VLA integrins was studied in several human T cell clones upon specific and nonspecific cellular activation. Human activated T lymphocytes down-regulated both alpha 4 beta 1 and alpha 4 beta 7 integrins upon specific recognition of alloantigens (cytotoxic T cells) or in the presence of Staphylococcus enterotoxin B (superantigen recognizing noncytotoxic T cells). In contrast, the expression of other membrane integrins, such as VLA-1 and VLA-5 integrins, was not modified. Down-regulation of alpha 4 beta 1 and alpha 4 beta 7 integrins was observed as early as 3 h after stimulation, lasted later than 72 h and was partially inhibited by cytochalasin D. Interestingly, neither target cells nor NK cells modulated CD49d expression after interaction with T cells of K562, respectively, suggesting that CD49d expression was linked to specific T cell activation. The down-regulation of the CD49d chain in T cell clones stimulated with immobilized anti-CD3 mAbs confirmed the role of TCR-mediated activation in CD49d regulation. However, the CD3-independent cellular aggregation induced by soluble anti-CD43 mAb was also able to strongly down-regulate alpha 4 beta 1 and alpha 4 beta 7. The present work shows the first evidence that CD49d subunit-bearing integrin expression is distinctly regulated from other integrins after Ag or superantigen recognition by human activated T cells. CD49d modulation may be relevant for the traffic and tissue localization of locally activated T cells during immune responses.     

Young people in transition: the relationship between homesickness and self-disclosure

To assess cell-mediated immunity to Toxoplasma gondii, we evaluated the expression of the activation antigens CD69, CD71, and CD25 on T lymphocytes by flow cytometry after specific in vitro stimulation of whole blood from 127 T. gondii-positive and 63 T. gondii-negative patients. T lymphocytes from many seropositive individuals did not express CD69 at 24 h after T. gondii antigen stimulation, but CD71 and CD25 were easily detectable on T cells from seropositive individuals 7 days after specific activation. CD25 was mainly expressed by stimulated CD4(+) T cells, and its detection on total T cells was both a sensitive (98%) and a specific (97%) indicator of prior T. gondii infection. These results make flow cytometric detection of CD25 an excellent candidate for screening cell-mediated immunity to T. gondii in vitro and an interesting tool for the diagnosis of congenital infection.     

Induction of IL-12 p40 messenger RNA expression and IL-12 production of macrophages via CD40-CD40 ligand interaction

The mechanism of IL-12 production has been studied by stimulating macrophages or B cell lines with LPS, Staphylococcus aureus, or phorbol diester. However, since IL-12 plays an important role in the activation of T cells interacting with APC, it is important to study the mechanism of IL-12 production induced by T helper cell-APC interaction. We and others have demonstrated that IL-12 is produced in cultures where Th1 cells are stimulated with Ag or APC. In the present experiments, we studied a role of CD40-CD40 ligand (CD40L) interaction in IL-12 production and obtained the following results: 1) incubation of normal Th1 clone with APC in the presence of Ag induced IL-12 p40 and p35 mRNA accumulation and IL-12 production, and the addition of anti-CD40L blocked the p40 mRNA accumulation and IL-12 production but not p35 mRNA accumulation; 2) when Th1 clone from a CD40L-deficient mouse was used in the incubation, p35 mRNA accumulation was induced, but neither p40 mRNA accumulation nor IL-12 production was induced; 3) CD40L+ Th1 clone, or insect cell membrane expressing mouse CD40L, induced p40 mRNA accumulation and IL-12 production but not p35 mRNA accumulation. These results indicate that the CD40-CD40L interaction plays a critical role in IL-12 p40 mRNA accumulation and bioactive IL-12 production and that p35 mRNA accumulation was regulated via a different mechanism than CD40-CD40L interaction. Most of the cells producing IL-12 were Mac-1+ macrophages.     

CD40 activation boosts T cell immunity in vivo by enhancing T cell clonal expansion and delaying peripheral T cell deletion

In this report we show that activation of APC with an agonist anti-CD40 mAb profoundly alters the behavior of CD4 T cells in vivo. Stimulation of mice with anti-CD40 2 days before, but not 1 day after, administration of superantigen (SAg) enhanced CD4 and CD8 T cell clonal expansion by approximately threefold. Further, CD40 activation also delayed peripheral T cell deletion after activation. Dying, activated T cells were quantitated by detecting extracellular phosphatidylserine with concomitant staining for SAg-reactive T cells using a TCR Vbeta-specific mAb. Upon close examination, it was shown that CD40 activation delayed the death of the activated T cells. Additionally, it was found that enhanced survival of CD4 T cells was equally dependent on APC expression of B7-1 and B7-2. This is in contrast to CD8 T cells, which did not depend as much on B7-1 as B7-2. Thus, CD40 activation indirectly promotes T cell growth and delays the death of SAg-stimulated CD4 T cells in vivo. These data suggest that one way CD40 activation promotes a more robust immune response is by indirectly increasing the production of effector T cells and by keeping them alive for longer periods of time.     

Blockade of CD40-CD40 ligand pathway induces tolerance in murine contact hypersensitivity

Interactions between CD40 on antigen-presenting cells and its ligand (CD40L) on T cells has been implicated in T cell-mediated immune responses. Previously, we have shown that contact hypersensitivity (CHS), a cell-mediated cutaneous immune response in reaction to haptens, could be subclassified based on whether the hapten primed for Th1 or Th2 cytokines in cells isolated from draining lymph nodes. We also found that tolerance to a Th2-priming hapten could be induced only by simultane blockade of the CD40-CD40L and B7-CD28 at the time of sensitization. Here we demonstrate that blockade of CD40-CD40L signaling alone induces long-lasting unresponsiveness to the Th1 hapten 2,4-dinitrofluorobenzene (DNFB), and inhibits antigen-specific T cell proliferation in vitro. We find that CD40-CD40L signaling is required in the sensitization but not elicitation phase of DNFB-induced CHS, as treatment of mice with anti-CD40L monoclonal antibody (mAb) does not affect the response to hapten challenge in previously sensitized and untreated animals. Examination of cytokine production shows that anti-CD40L mAb decreases interferon-gamma production by draining lymph node cells from DNFB-sensitized mice, and reciprocally increases interleukin (IL)-4 production. Consistent with this Th1 to Th2 immune deviation, anti-CD40L mAb prevents the induction of IL-12 mRNA in regional lymph nodes, an event which is normally seen within 12 h following hapten sensitization. In contrast, suppression of CHS by CTLA4Ig decreased the production of all cytokines by draining lymph node cells. Together, these data show that blockade of the CD40-CD40L pathway by itself is sufficient to induce tolerance to DNFB-induced CHS, and that this is associated with blockade of IL-12 induction and Th1 to Th2 immune deviation.     

Properties of mouse CD40: the role of homotypic adhesion in the activation of B cells via CD40

Stimulation of human B cells via CD40 is known to induce their homotypic aggregation. We show here that anti-mouse CD40 monoclonal antibodies (mAb) also induce B cells to form large, spherical, extremely stable clusters. This clustering is markedly enhanced by co-stimulation with either interleukin-4 (IL-4) or anti-immunoglobulin (Ig). The aggregation is slow in onset, and is largely (but not completely) abrogated by anti-LFA-1 mAb, but not by mAb directed against other potentially important adhesion molecules on B cells. Anti-LFA-1 mAb also partially suppressed DNA synthesis induced by anti-CD40, but not by other B cell mitogens, suggesting that clustering is an important component of B cell activation via CD40. This concept is supported by analyses of the phenotype of clustered B cells: the cells within clusters express higher levels of various activation markers, and also more of them are in cell cycle than non-clustered cells. These results therefore suggest that CD40 stimulation may either induce B cells to secrete soluble factors which act in an autocrine way to promote B cell activation, or that clustering generates cell contact-mediated signals which are important in the activation cascade.     

Induction and inhibition of CD40-CD40 ligand interactions: a new strategy underlying host-virus relationships

Interaction between CD40 and the CD40 ligand (CD40L) is required for mouse mammary tumor virus (MMTV) propagation. We found that Fas was expressed on B cells and CD40L on a small subset of viral superantigen-cognate T cells 12 h after MMTV(SW) infection. CD40L and Fas were down-regulated after 24 h. All CD4 T cells then became resistant to anti-CD3-induced CD40L induction in vitro for 2 wk. Initiation of CD40L expression and its rapid shut-off was associated with IL-12 production and was controlled by IFN-gamma and shedding of soluble CD40. These results suggest that a rapid, transient CD40-CD40L interaction involving a small number of cells is sufficient for MMTV propagation. Modulation of CD40L expression may be a major mechanism regulating the balance between viral propagation and host defenses, allowing mutual survival.     

Activation of peripheral blood T cells by interaction and migration through endothelium: role of lymphocyte function antigen-1/intercellular adhesion molecule-1 and interleukin-15

Cell adhesion molecules have a key role in the migration of T cells to inflammatory foci. However, the effect of the endothelial-lymphocyte interaction on the activation of the latter cells remains unresolved. We have studied the effect of resting and stimulated endothelial cells (ECs) on the activation of peripheral blood T cells (PBTLs), as assessed by the expression of CD69 and CD25 activation antigens. The incubation of PBTLs with tumor necrosis factor-alpha-activated EC monolayers, either alive or fixed, induced the expression of CD69 but not CD25, preferentially in the CD8(+) CD45RO+ cell subset. Furthermore, it induced the production of cytokines such as IFN-gamma, but not that of interleukin-2 (IL-2) and IL-4. EC treated with other stimuli such as IL-1beta, IFN-gamma, or lipopolysaccharide also showed the same proactivatory effect on T cells. Lymphocyte activation was almost completely inhibited by blocking anti-CD18 and anti-intercellular adhesion molecule-1 (anti-ICAM-1) monoclonal antibodies (MoAbs), but only slightly affected by MoAbs against CD49d, vascular cell adhesion molecule-1, and anti-IL-15. In addition, the interaction of PBTL with immobilized ICAM-1 induced CD69 expression in the same memory T-cell subset. IL-15 induced T-cell activation with expression of CD69 and CD25, and production of IFN-gamma, and its effect was additive with that triggered by cell adhesion to either EC or immobilized ICAM-1. The transmigration of PBTLs through either confluent EC monolayers or ICAM-1-coated membranes also induced efficiently the expression of CD69. When IL-15 was used as chemoattractant in these assays, a further enhancement in CD69 expression was observed in migrated cells. Together these results indicate that stimulated endothelium may have an important role in T-cell activation, through the lymphocyte function antigen-1/ICAM-1 pathway, and that IL-15 efficiently cooperates in this phenomenon. These observations could account for the abundance of CD69(+) cells in the lymphocytic infiltrates of several chronic inflammatory diseases.     

The real and imagined harmful effects of rewards: implications for clinical practice

The actions of a humanised therapeutic CD4 mAb YHB.46 on T cell activation were investigated in vitro. Soluble YHB.46 IgG or YHB.46-derived F(ab')2 fragments caused inhibitions of up to 100% of the proliferation of purified CD4+ T cells activated with immobilised CD3 mAb. The inhibitory effects of the CD4 mAb were equally potent in both CD45RA+ and CD45RO+ T cell subset proliferation assays. Inhibitory effects on DNA synthesis were nto explicable by increased T cell apoptosis. YHB.46 was inhibitory even when added 70 h after exposure of cells to immobilised CD3 mAb, but it had little effect on IL-2 receptor-driven proliferation signals. The CD4 mAb inhibited the CD3-induced expression of the CD25 and CD69 activation markers on the T cell surface and suppressed CD40 ligand expression, but not that of CD25 and CD69, when their expression was induced by phorbol ester plus ionomycin. YHB.46 also exerted a profound inhibitory effect on the production of IL-2, IL-4, and IL-10, irrespective of whether T cells were activated with CD3 mAb or with phorbol ester plus ionomycin. The inhibitory effects of YHB.46 on CD4+ T cell proliferation were partially prevented by the addition of exogenous IL-2 or autologous monocytes and were completely prevented by activating T cells with a novel CD3-CD28 bivalent F(ab')2 reagent. However, the inhibitory effects of YHB.46 on T cell proliferation were equipotent in the presence or the absence of CTLA-4Ig, showing that the CD4 mAb was not acting on CD28-induced activation signals per se. Our results show that the inhibitory effects of YHB.46 on T cell activation do not involve CD28 or IL-2 receptor signalling, but are directed at the TCR-mediated G0-G1 transition. These findings in vitro predict that YHB.46 may act as a potent immunosuppressant in the clinical context.     

CD40 ligand expressed on B cells in the BXSB mouse model of systemic lupus erythematosus

Male BXSB mice, unlike female BXSB, develop a severe early onset lupus-like disease that has been linked to an intrinsic B cell defect. In investigating this B cell defect the present study showed that male, but not female, BXSB contained a higher percentage of large, activated splenic B cells that were more responsive to anti-CD40 mAb-induced proliferation. The hyperactivity of the large B cells from the male mice was also observed in the absence of anti-CD40 mAb or any other stimuli. In examining the mechanism of the B cell hyperactivity, it was found that 20% of unstimulated large B cells from male mice, unlike large B cells from female mice, expressed CD40 ligand (CD40L), a molecule normally expressed on activated CD4+ cells. The percentage of large B cells from the male BXSB that expressed CD40L was increased to 43% by stimulation with LPS. A functional role for CD40L expression on B cells was confirmed by showing that CD40-Ig blocked the spontaneous proliferation of the large B cells from male mice. In addition, the stimulatory capacity of the large B cells from the male mice was demonstrated by their ability to induce DNA synthesis in small B cells in a CD40L-dependent manner. These results demonstrated that large B cells from male BXSB expressed functionally active CD40L. It is likely that the B cell CD40L expression and increased susceptibility to CD40 signaling due to an intrinsic B cell hyperactivity promotes autoimmune disease in BXSB mice.     

Normal growth in high-risk hyperlipidemic children and adolescents with dietary intervention

We have analyzed the effect of complete T cell activation (anti-CD3 plus anti-CD28) on the activation of NF-kappaB in CD45RA+ (naive) and CD45RO+ (memory/effector) T cells. Long exposure (24 h) induced stronger NF-kappaB DNA binding in CD45RA+ cells than in CD45RO+ cells. Analysis of the nuclear c-Rel protein indicated that after anti-CD3+anti-CD28 stimulation the level of c-Rel was higher in CD45RA+ cells. Analysis of the cytoplasmic inhibitor IkappaBalpha indicated that anti-CD3+anti-CD28 stimulation induced a long-lasting degradation in CD45RA+ cells but in CD45RO+ cells the degradation process was more rapid. Because the CD28 costimulus is known to induce the production of reactive oxygen intermediates (ROIs), the intracellular ROI levels in CD45RA+ and CD45RO+ cells were compared by flow cytometry. ROIs were produced in both cell types, but more strongly in CD45RA+ cells. The data presented in this study further emphasize the differences between CD45RA+ and CD45RO+ T lymphocytes in ROI-dependent signaling pathways.     

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.