Role of the autologous rosette-forming T cells in the concanavalin A-induced suppressor cell function

It was recently reported that the human autologous rosette-forming T cells (Tar cells) are devoid of Fc receptors for IgM and IgG but that they give rise in vitro to Tμ and Tγ cells and that these cells participate actively in feedback inhibition. We now investigated whether Tar cells participate in the concanavalin A-induced suppressor cell function, using two indicator systems, namely, mitogen- or alloantigen-induced DNA synthesis and mitogen-driven polyclonal immunoglobulin synthesis. When Tar cells were removed from peripheral blood T cells (T-Tar) there was no generation of suppression determined on both DNA synthesis and immunoglobulin production. When Tar cells were readded to T-Tar cells suppressor activity was restored. When purified Tar cells were activated by concanavalin A they showed the highest degree of suppression as compared to that exerted by total T cells or T-Tar cells. Tar cells lose their capacity to generate suppression when treated with mitomycin C before their activation; however, they are no longer sensitive to mitomycin C treatment once they have completed the period of activation. These results together with our previous findings showing that Tar cells give rise in vitro to Tμ and Tγ cells after their activation by Con-A suggest that Tar cells participate in the Con-A-induced suppressor cell function as precursors of the suppressor effector T cells.     

Suppression of interleukin-2 production by human concanavalin A-induced suppressor cells

When PHA-activated normal responder cells (R cells) were cocultured with mononuclear cells (MN cells) which had been preincubated for 48 hr in medium alone (C cells) an enhanced proliferative response was observed. This enhancement was only obtained when the R cells were cultured with allogeneic C cells or when PHA was in the cocultures for the entire culture period. This effect was due to greater production of interleukin 2 (IL-2) by irradiated C cells in the presence of allogeneic or mitogenic stimulation. Con A-treated mononuclear cells (S cells) cultured with PHA-activated allogeneic or autologous responder cells showed reduced [3H]thymidine incorporation and IL-2 production as compared to activated R cells alone. Glutaraldehyde-treated S cells (which retained the ability to absorb IL-2) did not affect the proliferative response or IL-2 production by the R cells, indicating that passive absorption of IL-2 was not entirely responsible for suppression induced by S cells. S cells, pretreated with IL-2, still inhibited R-cell activity. These results show that Con A-treated MN cells suppressed or prevented [3H]thymidine incorporation by actively inhibiting IL-2 production.     

Human suppressor T cells induced by concanavalin A: suppressor T cells belong to distinctive T cell subclasses.

Human peripheral blood lymphocytes were stimulated by concanavalin A (Con A) and then evaluated by their suppressive activity for thymus-derived (T) cell- and bone marrow-derived (B) cell-proliferative responses to mitogen and allogeneic cells. Con A-activated T cells markedly suppressed these responses, but Con A-activated B cells failed to demonstrate suppressor activity. Discontinuous bovine serum albumin (BSA) density gradient separation of T cells which had been activated by Con A demonstrated that a fraction containing blast cells as well as fractions containing unproliferated cells manifest the same degree of suppressor capabilities. However, when density gradient separation of T cells followed by subsequent incubation with Con A was performed, fractions of proliferating cells of low density exhibited no suppression; a fraction containing high density T cells produced marked suppression, but this fraction incorporated only little thymidine in response to Con A. Thus, these studies indicate that Con A-induced suppressor T cells belong to a distinctive subpopulation which has already been programmed to express this function before exposure to Con A and that cell proliferation may not be a prerequisite for the development of such suppressor T cells.     

In vitro effects of 4-hydroperoxycyclophosphamide on concanavalin A-induced human suppressor T cells

Summary Treatment in vitro of human peripheral blood lymphocytes (PBL) with ConA induced the generation of suppressor cells which inhibited T cell blastogenic response to ConA and of allogeneic response in the mixed lymphocyte reaction (MLR). Treatment of PBL with 4-hydroperoxycyclophosphamide (4-HPCy) before incubation with ConA markedly decreased the generation of suppressor cells by ConA. The effect of 4-HPCy on generation of suppressor cells was more pronounced in the test of ConA stimulation than in the MLR. Treatment with 4-HPCy had no effect on suppressor cells already induced as shown by incubation of PBL with 4-HPCy after incubation with ConA.     

Effects of concanavalin A-induced cells on the proliferative response of T cells. Concanavalin A-induced suppressor and amplifier cells to the proliferative response of human T cells to trinitrophenyl-modified autologous lymphocytes.

Effects of Con A-induced human mononuclear cells on the proliferative response of peripheral T cells were examined by using TNP-modified autologous lymphocytes as stimulator cells. Cells induced by incubation with Con A contained both suppressor cells and amplifier cells. The former were induced from nylon wool-nonadherent T cells and these precursor cells were sensitive to mitomycin treatment. On the other hand, amplifier precursor cells were nylon wool-nonadherent T cells and were resistant to mitomycin treatment. Cell proliferation was required for the induction of suppressor cells but not for the induction of amplifier cells. Con A-induced suppressor effector cells were both nylon wool-adherent and nonadherent cells, on the contrary, Con A-induced amplifier effector cells were nonadherent cells. A small number of macrophages enhanced the suppressive activity of nonadherent T cells when added at the induction phase of suppressor T cells.     

Cell-based immunotherapy with suppressor CD8+ T cells in rheumatoid arthritis

The chronic persistence of rheumatoid synovitis, an inflammation driven by activated T cells, macrophages, and fibroblasts causing irreversible joint damage, suggests a failure in physiologic mechanisms that down-regulate and terminate chronic immune responses. In vitro CD8(+)CD28(-)CD56(+) T cells tolerize APCs, prevent the priming of naive CD4(+) T cells, and suppress memory CD4(+) T cell responses. Therefore, we generated CD8(+)CD28(-)CD56(+) T cell clones from synovial tissues, expanded them in vitro, and adoptively transferred them into NOD-SCID mice engrafted with synovial tissues from patients with rheumatoid arthritis. Adoptively transferred CD8(+)CD28(-)CD56(+) T cells displayed strong anti-inflammatory activity. They inhibited production of IFN-gamma, TNF-alpha, and chemokines in autologous and HLA class I-matched heterologous synovitis. Down-regulation of costimulatory ligands CD80 and CD86 on synovial fibroblasts was identified as one mechanism of immunosuppression. We propose that rheumatoid synovitis can be suppressed by cell-based immunotherapy with immunoregulatory CD8(+) T cells.     

The protective role of myeloid-derived suppressor cells in concanavalin A-induced hepatic injury

The mechanism underlying T cell-mediated fulminant hepatitis is not fully understood. In this study, we investigated whether myeloid derived suppressor cells (MDSCs) could prevent the concanavalin A (ConA)- induced hepatitis through suppressing T cell proliferation. We observed an increase in the frequencies of MDSCs in mouse spleen and liver at early stage of ConA treatment, implicating that the MDSCs might be involved in the initial resistance of mice against ConAmediated inflammation. Subpopulation analysis showed that the MDSCs in liver of ConA-induced mice were mainly granulocytic MDSCs. Adoptive transfer of the bone marrow-derived MDSCs into ConA-treated mice showed that the MDSCs migrated into the liver and spleen where they suppressed T cell proliferation through ROS pathway. In addition, the frequencies of MDSCs in mice were also significantly increased by the treatment with immune suppressor glucocorticoids. Transfer of MDSCs into the regulatory T cell (Treg)- depleted mice showed that the protective effect of MDSCs on ConA-induced hepatitis is Treg-independent. In conclusion, our results demonstrate that MDSCs possess a direct protective role in T cell-mediated hepatitis, and increasing the frequency of MDSCs by either adoptive transfer or glucocorticoid treatment represents a potential cell-based therapeutic strategy for the acute inflammatory disease.     

Apolipoprotein A-II suppressed concanavalin A-induced hepatitis via the inhibition of CD4 T cell function

Con A-induced hepatitis has been used as a model of human autoimmune or viral hepatitis. During the process of identifying immunologically bioactive proteins in human plasma, we found that apolipoprotein A-II (ApoA-II), the second major apolipoprotein of high-density lipoprotein, inhibited the production of IFN-γ by Con A-stimulated mouse and human CD4 T cells. Con A-induced hepatitis was attenuated by the administration of ApoA-II. The beneficial effect of ApoA-II was associated with reduced leukocyte infiltration and decreased production of T cell-related cytokines and chemokines in the liver. ApoA-II inhibited the Con A-induced activation of ERK-MAPK and nuclear translocation of NFAT in CD4 T cells. Interestingly, exacerbated hepatitis was observed in ApoA-II-deficient mice, indicating that ApoA-II plays a suppressive role in Con A-induced hepatitis under physiological conditions. Moreover, the administration of ApoA-II after the onset of Con A-induced hepatitis was sufficient to suppress disease. Thus, the therapeutic effect of ApoA-II could be useful for patients with CD4 T cell-related autoimmune and viral hepatitis.     

Effects of atomic bomb radiation on the differentiation of B lymphocytes and on the function of concanavalin A-induced suppressor T lymphocytes

The differentiation of peripheral blood B lymphocytes into immunoglobulin-producing cells (Ig-PC) by pokeweed mitogen (PWM) and the function of concanavalin A (Con A)-induced suppressor T lymphocytes were examined to elucidate the late effects of atomic bomb radiation. A total of 140 individuals, 70 with an exposure dose of 100 rad or more and an equal number with an exposure dose of 0 rad matched by sex and age, were selected from the Nagasaki Adult Health Study (AHS) sample. Both the differentiation of peripheral blood B lymphocytes into Ig-PC by PWM and the function of Con A-induced suppressor T lymphocytes tended to be more depressed in the exposed group than in the control group, but a statistically significant difference could not be observed between the two groups. The function of Con A-induced suppressor T lymphocytes tended to decrease with age, but a statistical significance was detected only for percentage suppression against IgM-PC.     

In vitro immune response of human peripheral lymphocytes. II. Properties and functions of concanavalin A-induced suppressor T cells.

Con A-stimulation of human peripheral T lymphocytes induced both suppressor and helper T cells. ConA-generated suppressor T cells inhibited PWM-induced IgG and IgM production in PBL. Lower concentrations of Con A (0.5 micrograms/ml) or shorter incubation periods (6 to 24 hr) induced mainly helper T cells, while higher concentrations of Con A (10 micrograms/ml) or longer incubation periods (at least 48 hr) induced suppressor T cells. Con A-generated suppressor T cells were sensitive to mitomycin treatment and exerted their suppressor function on the early phase of differentiation and/or proliferation of B cells but not on the final differentiation of B cells to Ig-producing cells. The identity of the MHC was not required for the expression of suppressor function. Suppressor T cells competed with helper T cells in PWM-induced Ig-production of PBL. This experimental system can be applied to estimate the regulatory function of T cells in several disease states.     

Activity of concanavalin A-induced suppressor cells in human MLR: differential effects on primary MLR, secondary MLR, and memory cell precursors

The relative sensitivity of the MLR responses of freshly isolated human lymphocytes to Con A-induced suppressor cells (SC) was compared to that of lymphocytes that had been primed previously in vitro. Fresh and primed cells were suppressed 72 and 17%, respectively, when cultured under similar conditions (p less than 0.0005). Titration of SC indicated that equivalent suppression in the two populations would require greater than a 50-fold excess of SC in the primed cells. Neither preincubation of the primed cells for 24 or 72 hr with the SC before restimulation, addition of fresh autologous cells to the primed cells, nor preincubation of the SC with fresh cells before addition to the primed cells increased suppression of primed cells. SC added at the beginning of a primary MLC were, however, very suppressive for both the primary response (67% suppression) and the subsequent secondary response (81% suppression). These data indicate that although the human primary MLR and the precursor of the memory cell are both sensitive to the suppressors induced by Con A, the memory cell itself possess an intrinsic resistance to such suppressors that is not related to simple kinetic phenomena nor to the loss in vitro of an intermediate regulatory cell. Cell depletion experiments suggest that resistance to nonspecific suppression may occur at the level of the helper cell.     

Altered memory T cell differentiation in patients with early rheumatoid arthritis.

The chronic immune response in rheumatoid arthritis (RA) might be driven by activated Th1 cells without sufficient Th2 cell differentiation to down-modulate inflammation. To test whether disordered memory T cell differentiation contributes to the typical Th1-dominated chronic inflammation in RA we investigated differentiation of resting CD4+ memory T cells in patients with early (6 wk to 12 mo) untreated RA and in age- and sex-matched healthy controls in vitro. No difference in cytokine secretion profiles of freshly isolated memory T cells was detected between patients and controls. A cell culture system was then employed that permitted the differentiation of Th effectors from resting memory T cells by short term priming. Marked differences were found in response to priming. Th2 cells could be induced in all healthy controls by priming with anti-CD28 in the absence of TCR ligation. By contrast, priming under those conditions resulted in Th2 differentiation in only 9 of 24 RA patients. Exogenous IL-4 could overcome the apparent Th2 differentiation defect in seven patients but was without effect in the remaining eight patients. In all patients a marked decrease in IL-2-producing cells and a significant increase in well-differentiated Th1 cells that produced IFN-gamma but not IL-2 were evident after priming with anti-CD3 and anti-CD28. The data suggest that CD4+ memory T cells from patients with early untreated RA manifest an intrinsic abnormality in their ability to differentiate into specific cytokine-producing effector cells that might contribute to the characteristic Th1-dominated chronic (auto)immune inflammation in RA.     

Cell-cell interactions in synovitis: antigen presenting cells and T cell interaction in rheumatoid arthritis

The synovial tissue in rheumatoid arthritis (RA) patients is enriched with mature antigen presenting cells (APCs) and many T lymphocytes. Interactions between APCs and T cells are essential for the initiation and amplification of T-cell-dependent immune responses, and may therefore play an important role in the chronic inflammatory processes in the synovium. The nature of the antigen(s) involved in RA still remains elusive. However, interactions and signaling through the costimulatory molecules CD28-CD80/86 and CD40-CD40L are critical during APC-T cell interaction for optimal cell activation. This review discusses how such costimulatory signals can be involved in the initiation and amplification of the inflammatory reactions in the synovium. Blocking of the signaling pathways involved in APC-T cell interactions might provide a specific immuno-therapeutic approach for the treatment of RA.     

Cell-cell interactions in synovitis: Antigen presenting cells and T cell interaction in rheumatoid arthritis

The synovial tissue in rheumatoid arthritis (RA) patients is enriched with mature antigen presenting cells (APCs) and many T lymphocytes. Interactions between APCs and T cells are essential for the initiation and amplification of T-cell-dependent immune responses, and may therefore play an important role in the chronic inflammatory processes in the synovium. The nature of the antigen(s) involved in RA still remains elusive. However, interactions and signaling through the costimulatory molecules CD28-CD80/86 and CD40-CD40L are critical during APC–T cell interaction for optimal cell activation. This review discusses how such costimulatory signals can be involved in the initiation and amplification of the inflammatory reactions in the synovium. Blocking of the signaling pathways involved in APC–T cell interactions might provide a specific immuno-therapeutic approach for the treatment of RA.     

Impaired beta cell function is present in nondiabetic rheumatoid arthritis patients

Introduction

To investigate how markers of β-cell secretion (proinsulin-processing metabolites) are expressed in rheumatoid arthritis (RA) patients and their potential relation with the insulin resistance (IR) observed in these patients.

Methods

The 101 RA patients and 99 nondiabetic sex- and age-matched controls were included. IR by homeostatic model assessment (HOMA2), and β-cell secretion, as measured by insulin, split and intact proinsulin, and C-peptide levels were determined for both groups. Multiple regression analysis was performed to compare IR between groups and to explore the interrelations between RA features, proinsulin metabolites, and IR. Data were adjusted for glucocorticoids intake and for IR classic risk factors.

Results

Compared with controls, RA patients showed higher HOMA-IR (β coef., 0.40 (95% CI, 0.20 to 0.59); P = 0.00). When data were adjusted for glucocorticoids intake, noncorticosteroid patients maintained a higher IR index (β, 0.14 (0.05 to 0.24); P = 0.00). Impaired insulin processing in RA patients was detected by the onset of elevated split proinsulin levels (β, 0.70 pmol/L (0.38 to 1.02); P = 0.00). These data remained significant also when adjusted for prednisone intake (β, 0.19 (0.00 to 0.36) pmol/L; P = 0.04). Split proinsulin-to-C-peptide ratios were higher in RA patients undergoing corticosteroid therapy (β, 0.25 (0.12 to 0.38); P = 0.03) and were nearly significant in comparison between noncorticosteroids patients and controls (β, 0.16 (-0.02 to 0.34); P = 0.08). Interestingly, the impact of HOMA-IR on the ratio of intact proinsulin to C-peptide was higher in controls compared with patients (β, 6.23 (1.41 to 11.06) versus 0.43 (-0.86 to 1.71); P = 0.03).

Conclusions

β-Cell function is impaired in nondiabetic and in RA patients not taking corticoids by a mechanism that seems to be, at least in part, independent of IR.     

MAP kinase p38 and its relation to T cell anergy and suppressor function of regulatory T cells

Diverse regulatory T cell populations (Treg) are important for the control of self tolerance and immune homeostasis. These include naturally occurring CD4+CD25+ Treg (nTreg) and induced Treg (iTreg). Tolerogenic dendritic cells, modulated by IL-10, are able to convert peripheral T cells into iTreg. These are anergic and characterized by a G1 cell cycle arrest, dependent on elevated levels of the cdk inhibitor p27Kip1. Novel data revealed a distinct pattern of MAP kinase activation in iTreg different from clonal T cell anergy, with enhanced activation of the p38-MAPKAP-K2/3 pathway. p38 is involved in cell cycle control and its activity is a prerequisite for the induction and maintenance of the anergic state in iTreg. Inhibition of p38 leads to down regulation of p27Kip1, cell cycle progress and loss of regulatory T cell function. Here, we discuss these data in light of the role of p38 and p27Kip1 in T cell activation, anergy induction and cell cycle control.     

Atorvastatin upregulates regulatory T cells and reduces clinical disease activity in patients with rheumatoid arthritis

In this study, we investigated the hypothesis that regulatory T cells (T(reg)) are involved in the immunomodulatory effects of statins on rheumatoid arthritis (RA) patients. The 12-week study cohort consisted of 55 RA patients and 42 control subjects allocated to either a group treated with atorvastatin (AT) (20 mg/day) or a non-AT group. T(reg) numbers, suppressive function, serum inflammatory markers, and disease activity were evaluated before and after the therapy. Furthermore, the effects of AT on the frequency and suppressive function of T(reg) were determined in vitro. Our data revealed that the suppressive function of T(reg) from RA patients significantly decreased compared with that of control subjects. AT significantly reduced erythrosedimentation, C-reactive protein, and disease activity. Concomitantly, T(reg) numbers and suppressive functions were significantly improved by AT. Consistent with the in vivo experiments, AT promoted the generation of T(reg) from primary T cells and enhanced preexisting T(reg) function in vitro. Moreover, we showed that PI3K-Akt-mTOR and ERK signal pathways were involved in the induction of T(reg) by AT. In conclusion, AT significantly increased T(reg) numbers and restored their suppressive function in the RA patients, and this may be relevant in the modulation of uncontrolled inflammation in this disorder.     

Mechanical interactions between dendritic cells and T cells correlate with T cell responsiveness

Ag recognition is achieved through the communication across intercellular contacts between T cells and APCs such as dendritic cells (DC). Despite remarkable progress in delineating detailed molecular components at the intercellular contacts, little is known about the functional roles of physical cross-junctional adhesion between T and DC in shaping T cell responses. In addition, the mechanisms underlying sensitivity and specificity of Ag discrimination by T cells at intercellular contacts remain to be elucidated. In this study, we use single-cell force spectroscopy to probe the mechanical interactions between DC and T cells in response to stimulation with a panel of altered peptide ligands. The results show that intercellular interactions of DC-T cell conjugates exhibited different ranges of interaction forces in peptide-dependent manners that match the ability of the peptides to activate T cells. Elevated calcium mobilization and IL-2 secretion by T cells were only promoted in response to antigenic peptides that induce strong interaction forces, suggesting that mechanically stable DC-T cell contacts are crucial for driving T cell activation. Strong interactions were not solely dependent on cell-surface molecules such as TCRs and the adhesion molecule LFA-1, but were also controlled by cytoskeletal dynamics and the integrity of membrane lipid rafts. These data provide novel mechanical insights into the effect of Ag affinity on intercellular contacts that align with T cell responsiveness.