Urocanic acid enhances IL-10 production in activated CD4+ T cells

The immunosuppressive effects of UV radiation have been well documented. This suppression has been attributed to the action of the cis form of urocanic acid (UCA), a photoproduct of trans-UCA, a natural constituent of the skin. Here, we show that mouse spleen cells preincubated with cis-UCA have a diminished proliferative response to allogeneic cells in MLC and to stimulation with anti-CD3 mAb. Cells preincubated with cis-UCA also had a decreased ability to serve as APC and to stimulate the proliferation of allogeneic lymphocytes in MLC. Simultaneously, the production of IL-2 and IFN-gamma by cells preincubated with cis-UCA was decreased. However, IL-10 gene expression and IL-10 protein secretion by spleen cells stimulated in the presence of cis-UCA were significantly enhanced. The principal cell population displaying the cis-UCA-induced elevated production of IL-10 was CD4+ T cells, which were shown to be a direct target of cis-UCA action. This was also supported by the observation that production of IL-10 by stimulated splenic non-T cells or by macrophages was not altered by cis-UCA. The enhanced production of IL-10 by activated CD4+ T cells may represent a novel pathway of UVB radiation-induced, cis-UCA-mediated immunosuppression. We suggest that the elevated production of IL-10 by activated CD4+ T cells may account for the suppressor T cell phenomena described in UV-irradiated recipients.     

IL-10 impacts autoimmune diabetes via a CD8+ T cell pathway circumventing the requirement for CD4+ T and B lymphocytes

IL-10 is essential for an early phase of diabetes in nonobese diabetic (NOD) mice, but later becomes protective against its development. The mechanism by which IL-10 mediates the pathway to diabetes in these mice is unknown. Herein, we dissected the cellular and costimulation requirements for diabetes in transgenic (tg) NOD mice that expressed IL-10 in their pancreatic islets (IL-10-NOD mice). We found that IL-10 alone did not cause diabetes because the offspring (IL-10-NOD-scid mice) from back-crosses of IL-10-NOD mice with NOD-scid mice had no diabetes. Moreover, these IL-10-NOD-scid mice were free of lymphocytic infiltration. Treatment of IL-10-NOD mice with depleting anti-CD4 mAb or control mAb had no effect on diabetes. Surprisingly, depletion of CD8+ T cells by treatment with the corresponding mAb inhibited diabetes without attenuating insulitis, demonstrating a critical role for CD8+ T cells in the disease process. Interestingly, B cell-deficient IL-10-NOD mice readily developed diabetes with kinetics and incidence similar to those observed in wild-type mice, demonstrating that B lymphocytes as APCs were not required in the disease process. Administration of anti-CD40 ligand (CD40L) mAb did not prevent disease, indicating that CD40/CD40L costimulation is not required for diabetes in IL-10-NOD mice. Immunization of IL-10-NOD mice with CFA or heat-shock protein 65, known to block diabetes in NOD mice, had no effect on their diabetes. We demonstrate that IL-10 contributes early to the pathology of diabetes via a CD8+ T cell pathway, eliminating the requirement for B lymphocytes and CD40-CD40L costimulation. Our findings provide a mechanism for the participation of IL-10 in the early development of diabetes.     

Reciprocal regulation of CD30 expression on CD4+ T cells by IL-4 and IFN-gamma

Prior studies have implicated CD30 as a marker for Th2 cells, but the mechanism that underlies this correlation was unknown. We show here that CD30 was expressed on activated CD4+ T cells in the presence of IL-4. In the absence of endogenously produced IL-4, however, even Th2 lineage cells lost CD30 expression. Thus, CD30 is not an intrinsic marker of Th2 cells, but is inducible by IL-4. CD30 was also found to be down-regulated by IFN-gamma. Committed Th1 effector cells do not express CD30, although differentiating Th1 lineage cells temporarily express CD30. The transient expression of CD30 on differentiating Th1 lineage cells was mainly the result of endogenously produced IL-4 induced by IL-12. Culture of IL-12-primed cells under conditions that reverse the phenotype (Ag plus IL-4) resulted in two cell populations based upon their ability to express CD30. One population responded to IL-4 upon restimulation and became a CD30-positive, Th0-like cell population, while the other remained CD30 negative and synthesized only IFN-gamma. Thus, CD30 expressed on CD4+ T cells reflected the ability of CD4+ T cells to respond to IL-4.     

Differential capacities of CD4+, CD8+, and CD4-CD8- T cell subsets to express IL-18 receptor and produce IFN-gamma in response to IL-18

IL-12 and IL-18 have the capacity to stimulate IFN-gamma production by T cells. Using a T cell clone, we reported that IL-18 responsiveness is generated only after exposure to IL-12. Here, we investigated the induction of IL-18 responsiveness in resting CD8+, CD4+, and CD4-CD8- T cells. Resting T cells respond to neither IL-12 nor IL-18. After stimulation with anti-CD3 plus anti-CD28 mAbs, CD8+, CD4+, and CD4-CD8- T cells expressed IL-12R, but not IL-18R, and produced IFN-gamma in response to IL-12. Cultures of T cells with anti-CD3/anti-CD28 in the presence of rIL-12 induced IL-18R expression and IL-18-stimulated IFN-gamma production, which reached higher levels than that induced by IL-12 stimulation. However, there was a substantial difference in the expression of IL-18R and IL-18-stimulated IFN-gamma production among T cell subsets. CD4+ cells expressed marginal levels of IL-18R and produced small amounts of IFN-gamma, whereas CD8+ cells expressed higher levels of IL-18R and produced more IFN-gamma than CD4+ cells. Moreover, CD4-CD8- cells expressed levels of IL-18R comparable to those for CD8+ cells but produced IFN-gamma one order higher than did CD8+ cells. These results indicate that the induction of IL-18R and IL-18 responsiveness by IL-12 represents a mechanism underlying enhanced IFN-gamma production by resting T cells, but the operation of this mechanism differs depending on the T cell subset stimulated.     

Age-related increase in the fraction of CD27-CD4+ T cells and IL-4 production as a feature of CD4+ T cell differentiation in vivo

The influence of ageing on phenotype and function of CD4+ T cells was studied by comparing young (19-28 years of age) and aged (75-84 years of age) donors that were selected using the SENIEUR protocol to exclude underlying disease. An age-related increase was observed in the relative number of memory cells, not only on the basis of a decreased CD45RA and increased CD45RO expression, but also on the basis of a decrease in the fraction of CD27+CD4+ T cells. Our observation that the absolute number of CD45RO+CD4+ T cells was increased, while absolute numbers of CD27-CD4+ T cells remained unchanged in aged donors, indicates that the latter subset does not merely reflect the size of the CD45RO+CD4+ T cell pool. The increased fraction of memory cells in the aged was functionally reflected in an increased IL-4 production and T cell proliferation, when cells were activated with the combination of anti-CD2 and anti-CD28, whereas IL-2 production was comparable between both groups. No differences were observed with respect to proliferative T cell responses or IL-2 production using plate-bound anti-CD3 or phytohaemagglutinin (PHA). The observation that IL-4 production correlated with the fraction of memory cells in young donors but not in aged donors suggests different functional characteristics of this subset in aged donors.     

Interleukin-10 dose-dependent regulation of CD4+ and CD8+ T cell-mediated graft-versus-host disease

BACKGROUND: Endogenous interleukin (IL)-10 production has been associated with the lack of graft-versus-host disease (GVHD) in human recipients of MHC-disparate donor grafts. Paradoxically, we have shown that the exogenous administration of high doses (30 microg/dose) of IL-10 to murine recipients of MHC-disparate grafts accelerates GVHD lethality. METHODS: The effects of IL-10 on GVHD mediated by either CD4+ or CD8+ T cells was examined in studies involving exogenous IL-10 administration or the infusion of T cells from IL-10-deficient (-/-) donor mice. The role of interferon (IFN)-gamma on IL-10-induced GVHD acceleration was studied using IFN-gamma-deficient (-/-) donor mice or neutralizing monoclonal antibody. RESULTS: IL-10 was found to have a dose-dependent effect on the GVHD lethality mediated by either CD4+ or CD8+ T cells. High doses of exogenous IL-10 accelerated GVHD lethality. IFN-gamma release was not responsible for the IL-10 facilitation of GVHD lethality. Paradoxically, low doses of IL-10 protected mice against GVHD lethality. The GVHD protective effect of the bioavailability of small amounts of IL-10 was confirmed by demonstrating that the infusion of T cells from IL-10 -/- donors accelerated GVHD lethality. CONCLUSIONS: The results suggest that IL-10 has a dose-dependent effect on the GVHD lethality mediated by CD4+ or CD8+ T cells, such that high doses accelerate lethality, while low amounts of bioavailable IL-10 are protective.     

Inhibitory and stimulatory effects of IL-10 on human CD8+ T cells

IL-10 is a well-documented immunosuppressant that inhibits macrophage-dependent Ag presentation and CD4+ T cell proliferation in vitro. We report that IL-10 inhibits alloantigen-specific proliferative responses and induces a long lasting anergic state in human purified CD8+ T cells when added concomitantly with the Ag in the presence of APC. Moreover, the generation of allospecific cytotoxic activity is inhibited by IL-10. These effects are indirect and are mediated through inhibition of the costimulatory functions of APC. In contrast, IL-10 has no direct inhibitory effects on the proliferation of purified CD8+ T cells activated by anti-CD3 mAb and promotes the growth of activated CD8+ T cells in combination with low doses of IL-2. Taken together, these results indicate that IL-10 has differential effects on CD8+ T cells depending on their state of activation, which may explain both the enhancing and inhibitory effects observed after IL-10 treatment in different in vivo experimental models.     

Regulation of IL-17, IFN-gamma and IL-10 in human CD8(+) T cells by cyclic AMP-dependent signal transduction pathway

In the present study, the expression of interleukin 17 (IL-17) by human CD8(+) T lymphocytes and its regulation following PKA activation was determined and compared with that of interferon gamma (IFN-gamma) and IL-10. IL-17 mRNA was highly expressed in human CD8(+) T lymphocytes at least at the same level than in CD4(+) T cells that were isolated from peripheral blood mononuclear cells (PBMC). Expression of IL-17 mRNA in CD8(+) T cell was induced by prior activation of PBMC for 18 h with Ca2+ ionophore and phorbol myristate acetate (PMA). Furthermore, our results clearly showed that CD8(+) T cells are sensitive to elevation of cAMP and PKA activation pathway. Data demonstrated a significant inhibition of IL-17 as well as of IFN-gamma mRNA expression in CD8(+) T cells isolated from activated PBMC cultured in the presence of either dibutyryl cAMP (db-cAMP) or PGE2. In contrast, IL-10 mRNA expression was strongly enhanced in the same experimental conditions. The differential expression of IL-10 and IFN-gamma production in CD8(+) T cells was also observed at the protein level as it was measured by a double immunofluorescence technique and flow cytometry analysis. Taken together, these results provide evidence that human CD8(+) T cells are also the source of massive expression of IL-17, and that PKA plays a prominent role in the switch of CD8(+) T cells to a Th2 like profile and an inhibition of IL-17 expression, thus suggesting that the activation of cAMP signal transduction pathway may have consequences for the relative role of CD8(+) T cells in the immune and inflammatory process.     

LPS-stimulated SJL macrophages produce IL-12 and IL-18 that inhibit IgE production in vitro by induction of IFN-gamma production from CD3intIL-2R beta+ T cells

SJL mice are known for their poor IgE production upon helminth infection. In this study, we have demonstrated that SJL standard B cells (85% IgM+ or B220+), prepared by complement-mediated T cell lysis, failed to proliferate and to produce IgE and IgG1 in response to LPS plus IL-4 in vitro. This diminished IgE production was restored by anti-IL-12 and enhanced by additional treatment with anti-IL-18, suggesting active suppression by the cells that produce IL-12 and IL-18. Indeed, SJL standard B cells were contaminated with Mac-1+ cells. Therefore, we removed macrophages by passing standard B cells through a Sephadex G-10 column (G10). Resultant cells (95% IgM+), designated as G10-B cells, responded to LPS and IL-4 by their proliferation and differentiation. G-10 treatment markedly diminished the proportion of B220- cells and Mac-1+ cells in SJL standard B cells. Furthermore, addition of SJL B220- cells dose dependently and MHC independently inhibited LPS plus IL-4-induced B cell growth and IgE production in SJL and BALB/c B cells. B220- cells in SJL standard B cells contained Mac-1+ cells (51%) and Fas ligand+ CD4-CD8- double-negative CD3intIL-2R beta+ T cells (26%). Thus, IL-12 and IL-18 produced by LPS-stimulated Mac-1+ cells stimulate this unique subpopulation of T cells to produce IFN-gamma, which in combination with Fas ligand, inhibits IgE production from the B cells. Our present results indicate that Mac-1+ cells and double-negative CD3intIL-2R beta+ T cells, uniquely abundant in the spleens of SJL mice, inhibit IgE production, indicating their new role in IgE response.     

IL-9 production of naive CD4+ T cells depends on IL-2, is synergistically enhanced by a combination of TGF-beta and IL-4, and is inhibited by IFN-gamma

Dense CD4+ T cells isolated from naive mice produce only trace amounts of IL-9 when stimulated by immobilized anti-CD3 in combination with anti-CD28 Abs. In this situation, IL-9 production is significantly stimulated by TGF-beta and further enhanced by the addition of IL-4, which, by itself, has only a minimal influence. IFN-gamma was found to inhibit the enhancing effect of IL-4. However, increasing amounts of IL-4 in the presence of a constant concentration of IFN-gamma could overcome the inhibitory activity of IFN-gamma. The application of CD4+ T cells isolated from IL-2 knockout mice unequivocally revealed that IL-2 is essential for the production of IL-9 by T cells. In addition, the use of T cells from IL-4 knockout mice elucidated that the basic (IL-2 + TGF-beta) mediated IL-9 production is independent of IL-4. Therefore, our results demonstrate that optimal IL-9 production of naive dense CD4+ T cells is positively regulated at different levels: 1) by IL-2, which is essential for IL-9 secretion; 2) followed by TGF-beta, which promotes a considerable increase in IL-9 production above the level induced by IL-2; and 3) finally, by IL-4, which requires the presence of IL-2 and TGF-beta to strongly enhance the production of IL-9. IFN-gamma inhibits the production of IL-9 mainly at the level of IL-4 by neutralizing the effect of this cytokine.     

CD4+ type 1 and CD8+ type 2 T cell subsets in human leishmaniasis have distinct T cell receptor repertoires

The mechanism of protective immunity and immunologic resistance against intracellular pathogens is believed to involve the activation of Ag-specific T cells. The T cells involved in protection/resistance to Leishmania can be studied using localized American cutaneous leishmaniasis (LCL) as a model, because the disease is often self-healing. Our study was undertaken to identify specific T cell populations that had accumulated in LCL lesions on the basis of TCR V beta gene usage. RNA was derived from skin lesions and blood of eight LCL patients, as well as from purified CD4+ and CD8+ subsets from the lesions and blood of three patients. After synthesis of cDNA, V beta gene usage was assessed by polymerase chain reaction. In all eight patients, several V beta gene families were overrepresented in lesions compared to blood. More importantly, the TCR V beta repertoires of both lesional CD4+ and CD8+ subsets were skewed compared to the repertoire of the respective subsets in the blood of the same donor. The overrepresented V beta s in the CD4+ and CD8+ subsets from lesions were in most instances disparate, particularly with the V beta 6 TCR skewed in the lesional CD8+ subset. Not only were the TCR repertoires of the overrepresented V beta in the lesional CD4+ and CD8+ subsets generally distinct, but the cytokine mRNA expressed by these subsets were also discrete. Strikingly, the CD4+ subset was characterized by IFN-gamma mRNA expression and the CD8+ subset by IL-4 and IL-10 mRNA expression. These data indicate that the pathogenesis of human leishmaniasis may be explained by the balance of CD4+ type 1 and CD8+ type 2 T cells, which probably recognize distinct sets of Ag.     

The relation between diabetes mellitus and IFN-gamma, IL-12 and IL-10 productions by CD4+ alpha beta T cells and monocytes in patients with pulmonary tuberculosis

Membrane glycoproteins (gps) play an important role in cell-cell interactions during epidermal maturation, and we have previously shown an up-regulation of PNA-binding gps in cultured human keratinocytes treated with interferon gamma (IFN-gamma). The protein kinase C (PKC) pathway is known to play a key role in the regulation of proliferation and differentiation of keratinocytes and is also reported to be involved in some IFN-gamma-mediated effects. In order to evaluate the cellular mechanisms and whether PNA-binding gp expression is related to the differentiative activity of the lymphokine, we studied the effects of PKC agonists and antagonists and the role of retinoic acid (RA), in the induction of these gps in cultured human keratinocytes stimulated with IFN-gamma and processed for protein analysis. The expression of PNA-binding gps was revealed by incubation of SDS-polyacrylamide gels with 125I-PNA. The PKC antagonists (H7, sphingosine) as well as RA downregulated the IFN-gamma-induced PNA-reactive gps, whereas staurosporine and TPA upregulated their expression. These results provide evidence that PNA-reactive gps are late highly IFN-gamma-sensitive markers of keratinocyte differentiation, drastically modulated through selective isoforms of PKC.     

Interleukin-12 primes human CD4 and CD8 T cell clones for high production of both interferon-gamma and interleukin-10

Interleukin-12 (IL-12) induces differentiation of T helper 1 (Th1) cells, primarily through its ability to prime T cells for high interferon-gamma (IFN-gamma) production. We now report that the presence of IL-12 during the first several days of in vitro clonal expansion in limiting dilution cultures of polyclonally stimulated human peripheral blood CD4+ and CD8+ T cells also induces stable priming for high IL-10 production. This effect was demonstrated with T cells from both healthy donors and HIV+ patients. Priming for IL-4 production, which requires IL-4, was maximum in cultures containing both IL-12 and IL-4. IL-4 modestly inhibited the IL-12-induced priming for IFN-gamma, but almost completely suppressed the priming for IL-10 production. A proportion of the clones generated from memory CD45RO+ cells, but not those generated from naive CD45RO- CD4+ T cells, produced some combinations of IFN-gamma, IL-10, and IL-4 even in the absence of IL-12 and IL-4, suggesting in vivo cytokine priming; virtually all CD4+ clones generated from either CD45RO(-) or (+) cells, however, produced high levels of both IFN-gamma and IL-10 when IL-12 was present during expansion. These results indicate that each Th1-type (IFN-gamma) and Th2-type (IL-4 and IL-10) cytokine gene is independently regulated in human T cells and that the dichotomy between T cells with the cytokine production pattern of Th1 and Th2 cells is not due to a direct differentiation-inducing effect of immunoregulatory cytokines, but rather to secondary selective mechanisms. Particular combinations of cytokines induce a predominant generation of T cell clones with anomalous patterns of cytokine production (e.g., IFN-gamma and IL-4 or IFN-gamma and IL-10) that can also be found in a proportion of fresh peripheral blood T cells with "memory" phenotype or clones generated from them and that may identify novel Th subsets with immunoregulatory functions.     

Processing and release of IL-16 from CD4+ but not CD8+ T cells is activation dependent

IL-16 is synthesized as a precursor molecule of 68 kDa (pro-IL-16) that is processed by caspase-3, a member of the IL-1 converting enzyme (ICE) family. This cleavage results in a 13-kDa carboxy terminal peptide, which constitutes the bioactive secreted form of IL-16. We have previously reported constitutive IL-16 mRNA expression and pro-IL-16 protein in CD4+ and CD8+ T cells. Although bioactive IL-16 protein is present in unstimulated CD8+ T cells, there is no bioactive IL-16 present in CD4+ T cells. Along these lines, unstimulated CD8+ T cells contain active caspase-3. In the current studies we investigated the regulation of IL-16 protein and mRNA expression in CD4+ T cells and determined the kinetics of secretion following stimulation of the TCR. CD4+ T cells release IL-16 protein following antigenic stimulation, and this release is accelerated in time by costimulation via CD28. However, CD3/CD28 costimulation did not alter IL-16 mRNA appearance or stability in either CD4+ or CD8+ T cells. The secretion of bioactive IL-16 from CD4+ T cells correlated with the appearance of cleavage of pro-caspase-3 into its 20-kDa active form. Thus, resting CD8+ T cells contain active caspase-3 that is capable of cleaving pro-IL-16, whereas CD4+ T cells require activation for the appearance of active caspase-3. The mechanism of release or secretion of bioactive IL-16 is currently unknown, but does not correlate with cellular apoptosis.     

IL-15 promotes IL-12 production by human monocytes via T cell-dependent contact and may contribute to IL-12-mediated IFN-gamma secretion by CD4+ T cells in the absence of TCR ligation

At inflammatory sites, the number of activated bystander T cells exceeds that of Ag-activated T cells. We investigated whether IL-15, a monocyte-derived cytokine that shares several biologic activities with IL-2, may contribute to bystander T cell activation in the absence of IL-2 and triggering Ag. The addition of IL-15 to cocultures of monocytes and T cells stimulates CD4+ but not CD8+ T cells to produce IFN-gamma. IFN-gamma production requires endogenous IL-12, the production of which in turn is dependent upon CD40/CD154 interactions between CD4+ T cells and monocytes. Indeed, non-TCR-activated CD4+ but not CD8+ T cells express significant levels of CD154. IL-15 may enhance IFN-gamma in this system by up-regulating CD40 expression on monocytes and IL-12Rbeta1 expression on CD4+ T cells. Conversely, using neutralizing anti-IL-15 mAb, we show that the ability of IL-12 to augment IFN-gamma secretion is partly mediated by endogenous IL-15. Finally, in the absence of monocytes, a synergistic effect between exogenous IL-12 and IL-15 is necessary to induce IFN-gamma production by purified CD4+ T cells, while IL-15 alone induces T cell proliferation. It is proposed that this codependence between IL-12 and IL-15 for the activation of inflammatory T cells may be involved in chronic inflammatory disorders that are dominated by a Th1 response. In such a response, a self-perpetuating cycle of inflammation is set forth, because IL-15-stimulated CD4+ T cells may activate monocytes to release IL-12 that synergizes with IL-15 to induce IL-12 response and IFN-gamma production.     

Protective CD4+ and CD8+ T cells against influenza virus induced by vaccination with nucleoprotein DNA

DNA vaccination is an effective means of eliciting both humoral and cellular immunity, including cytotoxic T lymphocytes (CTL). Using an influenza virus model, we previously demonstrated that injection of DNA encoding influenza virus nucleoprotein (NP) induced major histocompatibility complex class I-restricted CTL and cross-strain protection from lethal virus challenge in mice (J. B. Ulmer et al., Science 259:1745-1749, 1993). In the present study, we have characterized in more detail the cellular immune responses induced by NP DNA, which included robust lymphoproliferation and Th1-type cytokine secretion (high levels of gamma interferon and interleukin-2 [IL-2], with little IL-4 or IL-10) in response to antigen-specific restimulation of splenocytes in vitro. These responses were mediated by CD4+ T cells, as shown by in vitro depletion of T-cell subsets. Taken together, these results indicate that immunization with NP DNA primes both cytolytic CD8+ T cells and cytokine-secreting CD4+ T cells. Further, we demonstrate by adoptive transfer and in vivo depletion of T-cell subsets that both of these types of T cells act as effectors in protective immunity against influenza virus challenge conferred by NP DNA.     

Development of CD4 and CD8 single positive T cells in human thymus organ culture: IL-7 promotes human T cell production by supporting immature T cells

This paper describes novel model systems to study the development of human T cells. Fragments of neonatal human thymus (HUNT) can be cultured in vitro; the initial majority population of CD4, CD8 double-positive (DP) thymocytes is not maintained in organ culture. These cells are rapidly replaced by populations of CD4 or CD8 single-positive (SP) T cells. In addition, allogeneic thymic chimeras can be established by the addition of human cord blood (HUCB) mononuclear cells as a source of T progenitor cells to the organ cultures. Culture results in the acquisition of a mature SP T cell phenotype by the donor cells similar to that found when HUCB is allowed to develop in xenogeneic murine scid/scid fetal thymus organ culture. The number of immature and mature T cells produced by organ cultures can be differentially increased by the addition of exogenous IL-7, stem cell growth factor, IL-1, or GM-CSF. Anti-IL-7 antibody inhibits T cell production. Taken together, the results suggest that human T cell development occurs in these in vitro systems in a similar manner, regardless of the species origin of the thymic stromal cells in the culture, and that exogenous cytokines can be used to expand subpopulations of developing T cells.     

CD4+ T cells orchestrate both amplification and deletion of CD8+ T cells

This review focuses on the role of CD4+ T cells in regulating immune responses, orchestrating both the amplification and deletion of immune cells, particularly CD8+ T cells. These two functions, which represent only an apparent contradiction, appear to be two faces of the same process of regulation. In fact, because the immune response, once activated, needs to be carefully controlled or switched off when the antigenic stimulus is eliminated, the immune system has developed several strategies either to regulate clonal amplification or to avoid useless expansion of activated cells. In particular, we have reported many data demonstrating that CD4+ T cells may be indicated as the regulatory element in the activation as well as the deletion of CD8+ T cells. New data are also reported on the ability of anergic CD4+ T cells to suppress CD8+ T-cell activation through induction of apoptosis, and on the need for CD8+ T cells for antigen recognition in inducing cell death in CD4+ T cells. Moreover, the central role of CD4+ T cells in the maintenance of peripheral tolerance has been widely described.     

Enhanced murine CD4+ T cell responses induced by the CD2 ligand CD48

The physiological functions of murine CD2 and its ligand CD48 are uncertain. We have examined the role of the CD2-CD48 interaction in murine T cell activation using a series of Chinese hamster ovary (CHO) cell transfectants. CHO cells expressing I-Ad together with CD48 induced more potent activation of OVA-specific, I-Ad-restricted DO11.10-transgenic T cells than CHO cells expressing I-Ad alone. CD48 augmented proliferation and IL-2 production in response to antigen. The enhancing effect of CD48 was of the same magnitude as that seen for CD80 (B7-1). Conjugate assays revealed the ability of CD48 to increase adhesion between T cells and CHO transfectants. The enhancing effects of CD48 on T cell-antigen-presenting cell adhesion and T cell activation were inhibited by anti-CD2 monoclonal antibody. This report provides the first evidence that the CD2 ligand CD48 contributes to the interactions of murine CD4+ T cells with antigen-presenting cells.     

Sequential production of IL-2, IFN-gamma and IL-10 by individual staphylococcal enterotoxin B-activated T helper lymphocytes

Upon primary activation, T helper (Th) cell populations express different cytokines transiently and with different kinetics. Stimulation of naive murine splenic Th cells with the bacterial superantigen Staphylococcus aureus enterotoxin B (SEB) in vitro results in expression of IL-2, IFN-gamma and IL-10 with fast, intermediate and slow kinetics, respectively. This first report of a functional analysis of cells separated alive according to cytokine expression shows that these cytokines are not produced by different Th cell subpopulations, but can be expressed sequentially by individual Th cells. Th cells, activated with SEB for 1 day and isolated according to expression of IL-2, using the cellular affinity matrix technology, upon continued stimulation with SEB later secrete most of the IFN-gamma and IL-10. Likewise, after 2 days of SEB culture, cells expressing IFN-gamma, separated according to specific surface-associated IFN-gamma as detected by magnetofluorescent liposomes, 1 day later secrete IL-10. Thus, individual Th1 cells can contribute to the control of their own IFN-gamma expression by sequential expression of first IL-2, supporting their proliferation, and later IL-10, down-regulating the production of IFN-gamma-inducing monokines and limiting the pro-inflammatory effects of IFN-gamma.