Regulation of murine chronic colitis by CD4+CD25- programmed death-1+ T cells

Naturally arising CD4(+)CD25(+) regulatory T (T(R)) cells are engaged in the maintenance of self tolerance and prevention of autoimmune diseases. However, accumulating evidence suggests that a fraction of peripheral CD4(+)CD25(-) T cells also possesses regulatory activity. Programmed death-1 (PD-1) is a new member of the CD28/CTLA-4 family, which has been implicated in the maintenance of peripheral self tolerance. Here, we identified a subpopulation of CD4(+)CD25(-)PD-1(+) T cells in the spleen of naive mice that constitutively expressed CTLA-4 and FoxP3 and was hypoproliferative in response to anti-CD3 antibody stimulation in vitro. However, the CD4(+)CD25(-)PD-1(+) T cells uniquely produced large amounts of IL-4 and IL-10 in response to anti-CD3 and anti-CD28 mAb stimulation, unlike the CD4(+)CD25(+) T(R) cells. The CD4(+)CD25(-)PD-1(+) T cells exhibited a suppressor activity against the proliferation of anti-CD3 antibody-stimulated CD4(+)CD25(-)PD-1(-) T cells in vitro, which was partially abrogated by anti-CTLA-4 mAb, but not by anti-IL-10 or anti-PD-1 mAb. Remarkably, the CD4(+)CD25(-)PD-1(+) T cells inhibited the development of colitis induced by adoptive transfer of CD4(+)CD45RB(high) T cells into C.B17-scid/scid mice, albeit to a lesser extent than CD4(+)CD25(+) T(R) cells, in a CTLA-4-dependent manner. These results indicate that the CD4(+)CD25(-)PD-1(+) T cells contain substantial amounts of T(R) cells that are involved in the maintenance of peripheral tolerance.     

Anti-CD45RB antibody deters xenograft rejection by modulating T cell priming and homing

Pancreatic islet xenotransplantation has been advocated as a way of overcoming the shortage of human donor tissue for the treatment of type 1 diabetes. However, the potent immune response against xenografts is a major barrier to their use. We show that a short course of the anti-CD45RB antibody, MB23G2, prolongs survival of fetal pig pancreas grafts in mice. To investigate this effect further we used an i.p. xenograft model in which both donor pig cells and host inflammatory cells can be expediently recovered and analyzed. Graft prolongation was associated with reduced T cell and macrophage infiltration, and reduced production of both T(h)1 and T(h)2 cytokines at the graft site. Graft survival was further increased and T cell infiltration further reduced by combining anti-CD45RB antibody with co-stimulation blockade. The primary effect of anti-CD45RB antibody may be on CD4 T cells, in keeping with the marked reduction in T cell cytokine production in both spleen and graft sites. This concurs with previous studies in allogeneic models that indicate that this antibody perturbs T cell responses by modifying signaling via the TCR. In addition, anti-CD45RB treatment led to reduced expression of LFA-1 and CD62 ligand (CD62L) on CD4 T cells, independent of antigenic challenge. LFA-1 may enhance co-stimulation, and both LFA-1 and CD62L are involved in T cell trafficking. Their reduced expression provides an explanation why the T cell pool is reduced in lymph nodes. We conclude that modulation of inflammation against xenografts by anti-CD45RB antibody is due to effects on both T cell priming and trafficking.     

B淋巴细胞在抗CD45RB抗体诱导的移植免疫耐受中的作用

 目的: 探讨B淋巴细胞在抗CD45RB抗体诱导的移植免疫耐受中的作用。方法: 抗CD45RB抗体对BALB/c裸鼠进行预处理后制备脾脏单细胞悬液,与BALB/c小鼠T淋巴细胞和C57BL/6小鼠脾细胞混合培养,流式细胞术分析Th1、Th2、Treg和Tm淋巴细胞。以B6.μMT^-/-小鼠为受体、BALB/c小鼠为供体建立皮肤移植模型,移植后向受体鼠腹腔注射抗CD45RB单抗,监测脾淋巴细胞CD3⁺CD45RB^hi细胞比例。在混合淋巴培养过程中加入抗CD45RB单抗,分离B细胞,建立以BALB/c小鼠为供体、B6.μMT^-/-小鼠为受体的心脏移植模型,通过尾静脉注射B细胞给B6.μMT^-/-小鼠,观察受体鼠生存期和B细胞分布。结果: 在裸鼠体内用抗CD45RB抗体处理过的B淋巴细胞,与T淋巴细胞混合培养时,可使Treg和Th2淋巴细胞比例明显升高,Th1淋巴细胞的比例明显下降,Tm细胞无明显变化。在体内B淋巴细胞缺失的情况下,抗CD45RB抗体依然能够降低T细胞表面CD45RB的表达,与对照组B淋巴细胞存在组相比,抗CD45RB抗体对T淋巴细胞表面CD45RB下调更为快速,但最终CD3⁺CD45RB^hi T细胞比例无明显变化。体外抗CD45RB抗体处理过的B淋巴细胞可以延长受体鼠的生存时间。B6.μMT^-/-鼠在接受抗CD45RB抗体处理的B细胞并进行同种异体心脏移植后,B细胞可向胸腺迁移。结论: 在抗CD45RB抗体诱导的免疫耐受中,B淋巴细胞可能通过介导各T淋巴细胞亚群比例发挥着重要作用,且在中枢耐受中也起到一定作用,但是仅靠B淋巴细胞无法形成完全耐受。     

Targeting CD45RB alters T cell migration and delays viral clearance

CD45 is a receptor tyrosine phosphatase essential for TCR signaling. One isoform, CD45RB, is down-regulated in memory cells and targeting CD45RB with a specific antibody has been shown to inhibit graft rejection. Its role in immunity to infection, however, has not been tested. Here, we report the effect of anti-CD45RB antibody treatment on the induction of anti-influenza CD8+ T cells and viral clearance. Anti-CD45RB-treated mice had delayed pulmonary viral clearance compared with untreated mice whose infection was completely cleared by day 8 post-infection. In anti-CD45RB-treated mice, the total CD4+ and CD8+ T cell numbers in both the lungs and mediastinal nodes were substantially reduced at days 5 and 8; this effect was less marked for the spleen. CD8+ T cells specific for influenza virus were also reduced compared with the control group in all three organs at day 8. By day 11, when both treated and control groups showed no virus remaining in the lungs, specific CD8+ T cell numbers were at similar low levels. Homing to lymph nodes and lung of dye-labeled T cells was greatly inhibited (by >80%) by anti-CD45RB treatment. This reduced homing corresponded with reduced CD62L and beta1-integrin expression in both uninfected and infected mice. Since CD62L plays a critical role in homing lymphocytes to lymph nodes, and high levels of CD62L and alpha4beta1-integrin are expressed by lymphocytes that home to bronchus-associated lymphoid tissue, we suggest that reduced expression of these molecules is a key explanation for the delay in immune responses.     

CD4~+CD25~+CD127~-T细胞在抗CD45RB抗体诱导免疫耐受中的作用

目的:研究CD4+CD25+CD127-T细胞在抗CD45RB抗体诱导的免疫耐受中所发挥的作用,从而阐明抗CD45RB抗体诱导免疫耐受作用的机制。方法:体内实验建立小鼠异位心脏移植模型,观察抗CD45RB抗体对移植物生存期的影响。体外实验观察抗CD45RB抗体对T细胞增殖抑制能力和对CD4+CD25+CD127-T细胞生成的影响。流式细胞术检测外周血和混合细胞中CD4+CD25+CD127-T细胞百分率,ELISA法检测血清和培养液中IL-2和IL-10含量,Real-TimePCR法检测脾脏和混合细胞中Foxp3基因的表达,移植心脏病理学观察。结果:抗CD45RB抗体显著延长移植物存活时间(P0.01),对ConA刺激引起的T淋巴细胞增殖具有明显的抑制能力(P0.05)。与对照组相比,实验组CD4+CD25+CD127-T细胞百分率和Foxp3 mRNA表达量均明显增加(P0.05),实验组IL-2水平较对照组明显降低(P0.05),但IL-10含量较对照组明显升高(P0.05)。病理结果显示对照组移植心脏出现典型细胞免疫性损伤病理改变,而实验组中几乎无炎性细胞浸润现象。结论:抗CD45RB抗体能显著延长移植物存活时间,其诱导免疫耐受机制与上调CD4+CD25+CD127-T细胞百分率和增加Foxp3 mRNA表达量有关。     

Non-responsiveness of antigen-experienced CD4 T cells reflects more stringent co-stimulatory requirements.

We recently reported that previously activated T cells, irrespective of the nature of the first stimulus they encountered, are unable to respond to Staphylococcal enterotoxin B (SEB), nor to soluble anti-CD3 monoclonal antibody (mAb) presented by splenic antigen-presenting cells (APC). Such previously activated T cells are, however, fully capable of responding to plate-bound anti-CD3 plus splenic APC. These data suggest differential integration of the T-cell receptor (TCR) and co-stimulatory signalling pathways in naive versus antigen-experienced T cells. Consistent with this hypothesis, anti-CD28 mAb restores the proliferative capacity of resting ex vivo CD45RBlo CD4+ T cells (representing previously activated T cells) to both soluble anti-CD3 mAb and SEB. Interestingly, mAb-mediated engagement of cytotoxic T-lymphocyte antigen-4 (CTLA-4) completely negates the rescue effects mediated by anti-CD28 mAb in CD45RBlo cells. Nevertheless, the non-responsiveness of CD45RBlo CD4+ T cells cannot be reversed by anti-CTLA-4 Fab fragments, indicating that it is not related to negative regulatory effects of CTLA-4 engagement itself. Interestingly, the addition of interleukin-2 (IL-2) restores the proliferative capacity of CD45RBlo CD4+ T cells to SEB and soluble anti-CD3 mAb. Moreover, when rescued by IL-2, the cells are less susceptible to the negative regulatory effects of CTLA-4 engagement. Together, these findings suggest that the non-responsiveness of CD45RBlo CD4+ T cells to certain stimuli may be related to inadequate TCR signalling, primarily affecting IL-2 production.     

IL-2协同抗CD45RB抗体调节Treg/Th17细胞的比例并延长小鼠移植皮肤存活时间

目的:研究IL-2在抗CD45RB抗体诱导免疫耐受中对Treg/Th17细胞分化的影响,进一步阐明抗CD45RB抗体诱导免疫耐受的机制。方法:用免疫磁珠分选C57BL/6小鼠脾脏中的CD4+T细胞,在抗CD45RB抗体与IL-2的作用下培养72小时后,流式检测Treg/Th17细胞的变化。以BALB/c小鼠为供体,C57BL/6小鼠为受体建立同种异基因皮肤移植模型,分别给予抗CD45RB抗体及IL-2等治疗,术后1、3、5、7、9天取受体鼠脾细胞,动态检测Treg/Th17细胞的变化;术后第9天取移植皮肤HE染色观察炎性细胞的浸润情况;观察并记录移植皮肤的存活时间。结果:CD4+T细胞在IL-2联合抗CD45RB抗体的作用下培养72小时后,Treg比例升高,Th17细胞比例下降;IL-2联合抗CD45RB抗体治疗后明显延长小鼠移植皮肤的存活时间。结论:IL-2可以明显增强抗CD45RB抗体诱导免疫耐受的形成,使Treg细胞上调,下调Th17细胞,有利于免疫耐受的形成。     

CD45 isoform RB as a molecular target to oppose lipopolysaccharide-induced microglial activation in mice

CD45 is a membrane-bound protein tyrosine phosphatase expressed on all hemopoietic cells with multiple splice variants, including RA, RB, RC and RO. Our previous studies have shown that cross-linking of CD45 with an anti-CD45 antibody markedly inhibits LPS-induced microglia activation. In order to determine which of the CD45 isoforms may be responsible for these effects, we have investigated the expression of CD45 isoforms on cultured microglial cells using flow cytometric analysis. Data reveal that CD45RB is the predominant isoform expressed in murine primary cultured microglial cells. Furthermore, incubation of these cultured cells with anti-CD45RB antibody results in a reduction of microglial activation induced by LPS as evidenced by TNF-alpha production. As a validation of these findings in vivo, brain homogenates from anti-CD45RB antibody (MG23G2)-injected animals that had been treated with LPS demonstrate a significant decrease in TNF-alpha levels compared to control mice treated with LPS plus vehicle. Taken together, these findings suggest that therapeutic agents that specifically stimulate the microglial CD45RB signaling pathway may be effective in suppressing microglial activation associated with several neurodegenerative disorders.     

CD28/CTLA-4/B7 and CD40/CD40L costimulation and activation of regulatory T cells

Costimulatory signals are crucial for T cell activation. Attempts to block costimulatory pathways have been effective in preventing unwanted immune reactions. In particular, blocking the CD28/cytotoxic T lymphocyte antigen(CTLA)-4/B7 interaction(using CTLA-4Ig) and the CD40/CD40 L interaction(using anti-CD40 L antibodies) prevents T cell mediated autoimmune diseases, transplant rejection and graft vs host disease in experimental models. Moreover, CTLA-4Ig is in clinical use to treat rheumatoid arthritis(abatacept) and to prevent rejection of renal transplants(belatacept). Under certain experimental conditions, this treatment can even result in tolerance. Surprisingly, the underlying mechanisms of immune modulation are still not completely understood. We here discuss the evidence that costimulation blockade differentially affects effector T cells(Teff) and regulatory T cells(Treg). The latter are required to control inappropriate and unwanted immune responses, and their activity often contributes to tolerance induction and maintenance. Unfortunately, our knowledge on the costimulatory requirements of Treg cells is very limited. We therefore summarize the current understanding ofthe costimulatory requirements of Treg cells, and elaborate on the effect of anti-CD40 L antibody and CTLA-4Ig treatment on Treg cell activity. In this context, we point out that the outcome of a treatment aiming at blocking the CD28/CTLA-4/B7 costimulatory interaction can vary with dosing, timing and underlying immunopathology.     

On the role of CD26 in CD4 memory T cells

We studied an in vivo mouse model to evaluate the relationships between CD26--a glycoprotein with dipeptidyl peptidase IV (DPP-IV) activity implicated in the regulation of immune functions--and T cells expressing the effector/memory phenotype CD45RB. We report that CD26 does not define a differentiation stage of CD4 T cells because the density and frequency of CD26 on CD4 T cells from the spleen, inguinal and mesenteric lymph node was similar within the CD45RB+ (na?ve) and CD45RB- (antigen primed) subsets. This observation was confirmed using CD4 T cells from a T-cell receptor transgenic (tg) model. CD4 tg T cells specific for ovalbumin (OVA) were adoptively transferred and challenged in vivo with antigen. CD26 expression was the same on naive and antigen-stimulated CD4 T cells. Depleting CD4 T cells with an anti-CD4 antibody preferentially depleted the CD45RB+ subset. In CD4 depleted animals CD26 expression was not altered on the CD45RB- subset but the density of CD26 was marginally increased on the remaining CD45RB+ CD4 T cells. The results suggest that, unlike the human, CD26 in the mouse was not directly linked with T cell activation.     

Decidual and peripheral blood CD4+CD25+ regulatory T cells in early pregnancy subjects and spontaneous abortion cases

Human pregnancy represents a situation of semiallograft to maternal host. Therefore, it has been reported that tolerance to the fetal allograft represents a mechanism for maintaining a pregnancy. CD4(+)CD25(bright) regulatory T cells are known to play an important role in the development and maintenance of tolerance in peripheral tissues. However, the potential role of CD4(+)CD25(bright) T cells in maintaining human pregnancy has not been reported. In this study, we show that early human pregnancy decidua contains an abundance of CD4(+)CD25(bright) T cells, which express CD152(CTLA-4) at a high level. CD4(+)CD25(bright) T cells mediate potent inhibition of autologous T-cell proliferation by anti-CD3 stimulation. Furthermore, these cells inhibit the proliferation of autologous CD4(+)CD25(-) T cells in a dose-dependent fashion. This suppressive function of decidual CD4(+)CD25(+) T cells required cell-to-cell contact. The proportion of decidual CD4(+)CD25(bright) T cells was significantly lower in specimens from spontaneous abortion compared to those from specimens from induced abortions. These results suggest that decidual CD4(+)CD25(bright) T cells contribute to the mechanisms mediating maternal immune tolerance of conceptus antigens and therefore might contribute to the maintenance of pregnancy.     

Induction of human CD4+ regulatory T cells by mycophenolic acid-treated dendritic cells

Depending on their degree of maturation, costimulatory molecule expression, and cytokine secretion, dendritic cells (DC) can induce immunity or tolerance. DC treated with mycophenolic acid during their maturation (MPA-DC) have a regulatory phenotype and may therefore provide a new approach to induce allograft tolerance. Purified CD4(+) T cells stimulated in a human in vitro model of mixed culture by allogeneic MPA-DC displayed much weaker proliferation than T cells activated by mature DC and were anergic. This hyporesponsiveness was alloantigen-specific. Interestingly, T cells stimulated by MPA-DC during long-term coculture in four 7-day cycles displayed potent, suppressive activity, as revealed by marked inhibition of the proliferation of naive and preactivated control T cells. These regulatory T cells (Tregs) appeared to have antigen specificity and were contact-dependent. Tregs induced by MPA-DC were CD25(+)glucocorticoid-induced TNFR(+)CTLA-4(+)CD95(+), secreted IL-5 and large amounts of IL-10 and TGF-beta, and displayed enhanced forkhead box p3 expression. These results obtained in vitro demonstrate that human MPA-DC can induce allospecific Tregs that may be exploited in cell therapy to induce allograft tolerance.     

Biased activation of human T lymphocytes due to low extracellular pH is antagonized by B7/CD28 costimulation

As T cell response to tumor-associated antigens may be impaired by the acidic microenvironment typical of solid tumors, we assessed the effect of extracellular pH (pH(e)) on the activation and proliferation of human T lymphocytes and generation of the cytotoxic response. T lymphocytes stimulated with anti-CD3 mAb or PHA at low pH(e) were unable to secrete IL-2 and IFN-gamma and their ability to progress through the cell cycle was impaired. T lymphocytes also displayed up-regulation of IFN-gammaR2 chain and CTLA-4 expression, rendering them sensitive to negative regulatory signals. Agonistic mAb against CD28, but not against CD2, completely restored cytokine production and cell cycle progression, but down-regulated IFN-gammaR2 and CTLA-4 expression. The anti-CD28mAb rescued the CTL response of allogeneic anti-tumor cultures generated at low pH(e). Following anti-CD28 mAb treatment, T cells synthesized cyclooxygenase-2 (Cox-2) protein, which is involved in the early phases of T cell activation. This rescue of T cell activation was independent of the inducible 6-phosphofructo-2-kinase (iPFK-2) pathway, which stimulates proliferation in hypoxic and acidic conditions. The restoration of proliferative and cytotoxic T cell responses by CD28-triggering provides insight into the mechanisms by which B7 enhances the T cell anti-tumor response in vivo.     

A role for anti-CD45RB monoclonal antibody treatment upon dendritic cells

Selective interference with CD45RB isoform by monoclonal antibody (anti-CD45RBmAb) reliably induces donor-specific tolerance. Dendritic cells (DCs) are the most potent antigen-presenting cells that are capable of activating na?ve T cells. The purposes of the present study were to investigate the roles of anti-CD45RBmAb on the phenotypes and functioning of DCs and to further illustrate the mechanism of anti-CD45RBmAb-inducing immunologic tolerance. DCs from C57BL/6 mice were cultured and treated with various doses of anti-CD45RB monoclonal antibody. Cell phenotype, cycle and phagocytic ability were detected by flow cytometry. The production of IL-10 and IL-12 in the supernatants of mature DCs was measured with ELISA. Exosomes (Dex) were recovered from the supernatant of DCs cultured for 6 days in depleted medium, and effects of DCs and Dex on the ability of T-cell proliferation were detected by mixed lymphocyte culture. Anti-CD45RBmAb could inhibit DCs maturation in a dose-dependent manner, and the effects of exosomes (Dex) on DCs enhance or inhibition proliferation of T cells were also in a dose-dependent manner. Anti-CD45RBmAb could profoundly inhibit the maturation and functioning of DCs and generate tolerogenic dendritic cells (tDCs) as well as Dex, suggesting mechanistic contributions to tolerance development from the DCs through interactions with T cells.     

Molecular associations on the T cell surface correlate with immunological memory

Different isoforms of CD45 are expressed on naive and memory CD4 T cells in the mouse, as revealed by an antibody to a set of isoforms of CD45 that utilize exon B, called CD45RB. Cloned TH1 and TH2 lines also differ for expression of isoforms detected by this antibody. Differential expression of CD45 isoforms correlates with different behavior of cell surface molecules involved in transmembrane signal transduction. On naive T cells, CD4, CD45 and the CD3/T cell receptor complex behave as independent entities. On memory T cells, these three molecules are stably associated on the T cell surface. Furthermore, on TH2 cells, which express intermediate levels of CD45RB, CD4 is stably associated with CD45 isoforms other than CD45RB, but this complex is not associated with the CD3/T cell receptor. These results lead us to propose that immunological memory in CD4 T cells consists of an altered structure of the T cell's specific signal transduction apparatus controlled by low-molecular weight CD45 isoforms. This altered receptor structure would allow the more sensitive triggering of the T cell characteristic of memory cells. The organization of multimolecular signal transduction systems may be a general means by which cells alter their physiological behavior, allowing the acquisition of new phenotypic characteristics.     

Blocking CD40 - CD154 and CD80/CD86 - CD28 interactions during primary allogeneic stimulation results in T cell anergy and high IL-10 production.

Although CD28 triggering provides an important co-stimulatory signal to T cells, blocking the CD80/CD86 - CD28 interaction with CTLA-4lg fusion protein is not sufficient for tolerance induction in vivo or in vitro. According to more recent data, interruption of the CD40 - CD154 interaction might complement the effect of CTLA-4lg and induce graft acceptance. We studied the effects of a blocking anti-CD40 monoclonal antibody (mAb) and/or blocking anti-CD80/anti-CD86 mAb in cultures of human peripheral blood mononuclear cells (PBMC) stimulated with allogeneic PBMC. T cells activated by alloantigens in the presence of anti-CD80, anti-CD86 and anti-CD40 entered a state of alloantigen-specific non-responsiveness as evidenced upon restimulation by lack of proliferation, cytotoxic activity, and IL-2, IL-5 and IL-13 production. IFN-gamma production during restimulation was less than in the control cultures, while the production of IL-10 was enhanced. Addition of recombinant IL-2 during the restimulation rescued alloantigen-specific activity. We conclude that the simultaneous blocking of the CD40 - CD154 and CD80/CD86 - CD28 interaction during allogeneic T cell activation induces T cell anergy. Since anergic cells induced by this treatment still produce high levels of IL-10, the latter could contribute to modulation of antigen-presenting cell activity and to bystander suppression of residual reactive T cells.     

Reassessment of the role of CD8+ T cells in the induction of allograft tolerance by donor-specific blood transfusion.

Donor-specific allograft tolerance can be induced in adult rats by pregraft donor-specific blood transfusion (DST). We have previously shown that DST elicits in recipients the expansion of a donor-specific CD8+ T cell clone displaying the Vbeta18-Dbeta1-Jbeta2.7 TCR rearrangement, which rapidly infiltrates allografts after transplantation, suggesting a regulatory function for this clone in DST-induced tolerance. In this study, recipients were pretreated before grafting, using an anti-CD8 monoclonal antibody to deplete CD8+ T cells. CD8 depletion before DST and transplantation abrogated allograft tolerance, and the CD8+ T cell clone was absent from allografts. These effects were not observed when CD8 depletion was performed after DST but before transplantation. We conclude that CD8+ T cells play a role in the induction of allograft tolerance by DST.     

Clinical amelioration of murine lupus by a peptide based on the complementarity determining region-1 of an autoantibody and by cyclophosphamide: similarities and differences in the mechanisms of action

Systemic lupus erythematosus (SLE) is an autoimmune disease characterized by autoantibodies and systemic clinical manifestations. In this study we investigated the beneficial effects on murine lupus accomplished by a peptide based on the sequence of the complementarity-determining region 1 of an anti-DNA autoantibody (hCDR1) when given alone or in combination with cyclophosphamide (CYC), and determined the mechanisms underlying those effects. SLE-afflicted (NZB x NZW) F(1) mice were treated for 12 weeks with injections of hCDR1, CYC or a combination of both drugs. We found that hCDR1 and CYC ameliorated serological and renal manifestations of the diseased mice, down-regulated interferon-gamma and interleukin-10, and up-regulated transforming growth factor-beta. These effects were associated with an increment of naive CD4(+) cells at the expense of the number of CD4(+) cells with the memory/activated phenotype. Further, the number of CD8(+) cells in the diseased mice was increased by the two drugs, resulting in a significant decrease in the CD4 : CD8 ratio. However, whereas the frequency and activity of CD4(+) CD25(+) CD45RB(low) regulatory T cells and the expression of cytotoxic T-lymphocyte antigen 4 (CTLA-4) in CD4(+) cells were up-regulated by hCDR1 treatment, they were minimally affected following treatment with CYC. CTLA-4 played an important role in the activity of the hCDR1-induced CD4(+) CD25(+) cells as manifested by down-regulation of CD28 expression, decrease of activation-induced apoptosis, and modulation of the cytokine profile in CD4(+) CD25(-) cells derived from SLE-afflicted mice. Thus, although the two drugs have similar ameliorative effects, hCDR1 but not CYC elicits regulatory pathways that are of importance for tolerance induction in SLE.     

The phosphatidylinositol phosphatase PTEN is under control of costimulation and regulates proliferation in human T cells

The phosphatidylinositol phosphatase gene PTEN is a dual specific phosphatase acting on phospho amino acids but also on three phosphorylated inositol phospholipids. Present results demonstrate that PTEN is inducible by costimulatory signals in human CD4(+) T cells. PTEN expression was up-regulated on RNA and protein level in freshly isolated human CD4(+) T cells following stimulation with CD28 or CD2. In contrast, PTEN expression was high but remained CD28 and CD2 unresponsive in lymphoma cells. Intracellular staining revealed PTEN expression in CD4(+) T cell populations stimulated with anti-CD28 or anti-CD28 / anti-CD3. Stimulation with anti-CD3 alone did not induce PTEN expression. Inhibition of PTEN expression by antisense oligonucleotides in CD4(+) T cells stimulated with non-mitogenic anti-CD28 resulted in massively increased proliferation, which was sensitive to the phosphatidylinositol 3-kinase (PI3 K) inhibitor wortmannin. Although CD28 and CD2 induce PI3 K signal transduction, wortmannin did not block PTEN up-regulation by CD28 or CD2 indicating that PTEN gene expression is PI3 K independent. These results demonstrate that PTEN negatively controls costimulatory signals by antagonizing PI3 K activity in the absence of TCR engagement.