Apoptosis Of Th1 Cells Research Articles

TNFR25SF AND GALECTIN-9 COMBINATION THERAPY REDUCES HERPES SIMPLEX VIRUS-1 INDUCED STROMAL KERATITIS BY EXPANDING REGULATORY T CELLS AND REDUCING PATHOGENIC TH1 CELLS (170.17)

Abstract Ocular infection with HSV-1 results in a chronic immunoinflamammtory reaction in the cornea, which is primarily orchestrated by CD4 T cells. Hence, targeting proinflammatory CD4 T cells or increasing the representation of cells that regulate their function are relevant therapeutic strategies. In the present report, we use an agonistic mAb (4C12) to TNF receptor 25 (TNFR25) that selectively expands numbers of regulatory T cells (Tregs). Treatment early after ocular HSV infection with 4C12 caused a three fold increase in the Treg numbers in the cornea along with a consequent reduction in SK lesion severity. However, 4C12 treatment was less effective if given 6 days after infection since it expanded proinflammatory T effectors that also express TNFR25. To suppress the latter population, galectin-9 (Gal-9) was used which causes apoptosis of Th1 cells. When therapy with both 4C12 and gal-9 was used in combination, the recipients showed significantly reduced angiogenesis and SK lesions over single treatment controls. The beneficial outcome of the combination therapy was attributed to the expansion of the Treg population that expressed CD103 needed for trafficking to inflammatory sites along with a marked reduction in the infiltration of pathogenic CD4 T cells. Levels of several proinflammatory cytokines and chemokines were also reduced. Our results demonstrate that combination therapy may be a promising approach to control HSV induced SK lesions, a common cause of human blindness.

Foxp3+ regulatory T cells exert asymmetric control over murine helper responses by inducing Th2 cell apoptosis

AbstractFoxp3+ regulatory T cells play a pivotal role in maintaining self-tolerance and immune homeostasis. In the absence of regulatory T cells, generalized immune activation and multiorgan T cell–driven pathology occurs. Although the phenomenon of immunologic control by Foxp3+ regulatory T cells is well recognized, the comparative effect over different arms of the immune system has not been thoroughly investigated. Here, we generated a cohort of mice with a continuum of regulatory T-cell frequencies ranging from physiologic levels to complete deficiency. This titration of regulatory T-cell depletion was used to determine how different effector subsets are controlled. We found that in vivo Foxp3+ regulatory T-cell frequency had a proportionate relationship with generalized T-cell activation and Th1 magnitude, but it had a surprising disproportionate relationship with Th2 magnitude. The asymmetric regulation was associated with efficient suppression of Th2 cells through additional regulations on the apoptosis rate in Th2 cells and not Th1 cells and could be replicated by CTLA4-Ig or anti–IL-2 Ab. These results indicate that the Th2 arm of the immune system is under tighter control by regulatory T cells than the Th1 arm, suggesting that Th2-driven diseases may be more responsive to regulatory T-cell manipulation.

Reciprocal Regulation of The Survival and Apoptosis of Th17 and Th1 Cells in The Colon

The immediate early response gene X-1 (IEX-1) is a stress-inducible gene involved in the regulation of cell growth, apoptosis and inflammation. Acute colitis was induced by treatment of IEX-1 knockout (KO) and wild type (WT) control mice with dextran sulfate sodium (DSS), whereas chronic colitis was induced in Rag-/- mice by adoptive transfer of CD4(+) CD45RB(hi) T cells isolated from the two strains of mice. The diseases and responses of lamina propria lymphocytes were analyzed in the mice. IEX-1 KO mice produced IL-17 in the colon significantly greater than WT control mice following DSS treatment owing to better survival and differentiation of both IL-17-secreting γδ T cells and Th17 cells. The altered level of IL-17 production contributed critically to the reduced colon inflammation in IEX-1 KO mice, and administration of neutralizing anti-IL-17 antibody increased susceptibility of the animal to the disease. Strikingly, in contrast to the better survival of T cells producing IL-17, lack of IEX-1 enhanced apoptosis in proinflammatory T cells producing interferon gamma (IFN-γ). Enhanced apoptosis in Th1 cells and better survival of Th17 cells may both result in a delayed onset of colitis in Rag-/- mice receiving pathogenic CD4(+) CD45RB(hi) T cells isolated from IEX-1 KO animals compared to those mice transferred with WT counterparts The present study demonstrates the refractoriness of IEX-1 knockout (KO) mice to DSS-induced colitis and diminished pathogenesis of IEX-1-deficient CD4(+) CD45RB(hi) T cells. These data demonstrate that IEX-1 reciprocally regulates T-cell survival and apoptosis in a subset-dependent fashion. Inhibition of IEX-1 may thus offer novel strategies for colitis treatment by simultaneous induction of apoptosis in proinflammatory Th1 cells while promoting the survival and differentiation of a protective T-cell subset.

Effects of deoxycycline induced lentivirus encoding FasL gene on apoptosis of Th1 cells

Fas/Fas ligand (FasL)-mediated apoptosis plays a critical role in deletion of activated T cells. This study aimed to construct the lentivirus encoding FasL gene induced by deoxycycline and evaluate its effects on apoptosis of Th1 cells. A plasmid expression system encoding FasL was constructed through utilizing the controlling gene and target gene of the Tet-on system controlled by the PBI plasmid containing the bidirectional promoter. The lentiviral vector that consisted of the vector plasmid, the packaging plasmid and the envelop plasmid was isolated and purified. 293 T cells were cotransfected with three plasmids using Lipofectamine 2000. 48 h post transfection, the viral supernatant was collected to determine the virus titre. The FasL protein expression in 293T cells was detected by Western blotting. Spleen lymphocytes of rats were transfected with the lentiviral vector system encoding FasL gene and 5 days later were induced by doxycycline for 24 h, followed by detection of FasL mRNA. The apoptosis index of Th1 cells was measured through both Annexin V-FITC flow cytometry and TUNEL. Additionally flow cytometry was adopted to determine changes of the quantity of Th1 cells. The FasL protein was expressed in 293T cells transfected by the expressing FasL lentiviral vector following deoxycycline induction, while it was not expressed in 293T cells without deoxycycline induction. The virus titre was 4 × 108 u/l for 293T cells. The deoxycycline induced lentiviral vector system encoding FasL gene was successfully transfected into spleen lymphocytes of rats. The FasL RNA expression significantly increased in the deoxycycline plus lentivirus group, compared to that in the control group and the deoxycycline group. A higher apoptosis index and an obviously smaller quantity of Th1 cells were obtained in the deoxycycline plus lentivirus group than in the other two groups. The lentivirus system encoding FasL induced by deoxycycline promotes apoptosis of Th1 cells. Key words : FasL, lentiviral vector, Th1 cell, apoptosis.

Galectin‐9 ameliorates acute GVH disease through the induction of T‐cell apoptosis

Galectins comprise a family of animal lectins that differ in their affinity for β-galactosides. Galectin-9 (Gal-9) is a tandem-repeat-type galectin that was recently shown to function as a ligand for T-cell immunoglobin domain and mucin domain-3 (Tim-3) expressed on terminally differentiated CD4(+) Th1 cells. Gal-9 modulates immune reactions, including the induction of apoptosis in Th1 cells. In this study, we investigated the effects of Gal-9 in murine models of acute GVH disease (aGVHD). First, we demonstrated that recombinant human Gal-9 inhibit MLR in a dose-dependent manner, involving both Ca(2+) influx and apoptosis in T cells. Next, we revealed that recombinant human Gal-9 significantly inhibit the progression of aGVHD in murine BM transplantation models. In conclusion, Gal-9 ameliorates aGVHD, possibly by inducing T-cell apoptosis, suggesting that gal-9 may be an attractive candidate for the treatment of aGVHD.

OP06 Tim3 down-regulation renders CD4 effector cells less susceptible to T-reg control in patients with autoimmune hepatitis

IntroductionIn autoimmune hepatitis (AIH), the extent of CD4 effector immune responses is associated with defective CD4posCD25pos regulatory T-cells (T-regs). Engagement of T-cell-immunoglobulin-and-mucin-domain-3 (Tim3) on Th1-cells by galectin-9, expressed by T-regs,...

TIM-3 is a receptor for phosphatidylserine and allelic variants differentially mediate uptake of apoptotic cells (130.41)

Abstract TIM-3 is a member of the T cell/transmembrane, Ig and mucin (TIM) gene family. TIM-3 is expressed on Th1 cells and dendritic cells and generates an inhibitory signal resulting in apoptosis of Th1 cells. We found that TIM-3 is receptor for phosphatidylserine (PtdSer) exposed on apoptotic cells. A co-crystal structure showed a similar binding mode to TIM-4/PtdSer with a deep binding pocket coordinating calcium ion with PtdSer and flanked by two hydrophobic loops that can extend into a membrane. Fibroblasts transfected with BALB/c TIM-3 more effectively bound to and phagocytosed apoptotic cells than HBA TIM-3 transfected cells. Liposomes containing equal amounts of PtdSer and phosphatidylcholine reduced phagocytosis in a concentration dependent-manner. This suggests that functional differences in TIM-3 alleles may contribute to the Th1/Th2 differences between the strains. T or B cells expressing TIM-3 formed conjugates with but failed to phagocytose apoptotic cells, suggesting a role in signaling in lymphocytes without engulfment. These findings indicate that TIM-3 expressing cells can respond to apoptotic cells through TIM-3/PtdSer binding, but the consequence of TIM-3 engagement of PtdSer depends on the polymorphic variants of and type of cell expressing TIM-3. These findings establish a new paradigm for TIMs as PtdSer receptors and unify the function of the TIM gene family, which has been associated with ashma and autoimmunity and shown to modulate peripheral tolerance.

Regulation of TRAIL Expression in T helper cells (85.17)

Abstract TNF-related apoptosis-inducing ligand (TRAIL/Apo2 ligand), along with FasL and TNFα, is one of three members of the TNF cytokine family that induce apoptosis. TRAIL can bind to several different receptors including those containing a cytoplasmic death domain and others that lack such a domain, termed decoy receptors. Here we found that TRAIL was preferentially expressed in Th2 cells, and, though exogenous TRAIL did not cause death in these cells, it did induce apoptosis of Th1 cells. Since it differentially induced apoptosis in Th1 and Th2 cells, TRAIL was likely to modulate the balance between them in determining the type of immune response. However, the regulation mechanism of TRAIL expression in T lymphocytes is not fully understood. We showed that A1.1 T hybridoma cells constitutively expressed TRAIL at a low level that could be boosted by adding IFN-γ. When Th2 cells were treated with high dosage of IFN-γ upon restimulation, TRAIL expression was boosted as well, whereas when Th1 cells were treated with high dosage of IL-4 upon restimulation, TRAIL was surprisingly detected at a comparable level to that in Th2 cells without any cytokine treatment. Interestingly, although Th1 cells were the main producers of IFNγ, Th2 cells expressed IFNγ receptors. All these results suggest that IFN-γ secreted by Th1 cells and IL-4 secreted by Th2 cells may be potent inducers of TRAIL under certain circumstances and therefore can provide some insight into clinical applications.

Activation-Induced T Helper Cell Death Contributes to Th1/Th2 Polarization following MurineSchistosoma japonicumInfection

In chronic infectious diseases, such as schistosomiasis, pathogen growth and immunopathology are affected by the induction of a proper balanced Th1/Th2 response to the pathogen and by antigen-triggered activation-induced T cell death. Here, by using S. japonicum infection or schistosome antigens-immunized mouse model, or antigens in vitro stimulation, we report that during the early stage of S. japonicum infection, nonegg antigens trigger Th2 cell apoptosis via the granzyme B signal pathway, contributing to Th1 polarization, which is thought to be associated with worm clearance and severe schistosomiasis. Meanwhile, after the adult worms lay their eggs, the egg antigens trigger Th1 cell apoptosis via the caspase pathway, contributing to Th2 polarization, which is associated with mild pathology and enhanced survival of both worms and their hosts. Thus, our study suggests that S. japonicum antigen-induced Th1 and Th2 cell apoptosis involves the Th1/Th2 shift and favorites both hosts and parasites.

Galectin-9 Ameriorates Acute GVHD by Inducing T-Cell Apoptosis and Increasing Regulatory T Cell.

Abstract 1338Poster Board I-360Galectins are a family of soluble animal lectins that differ in their affinity for b-galactosides. Fifteen members of the human galectin family have been described to date. Galectin-9 (Gal-9) is a tandem-repeat-type galectin that was recently found to serve as a ligand for T cell immunoglobulin and mucin domain-3 (Tim-3), which is a Th1-specific type 1 membrane protein. Gal-9 modulates immune reactions, such as induction of apoptosis in Th1 cells. We herein investigated the effects of Gal-9 treatment on acute graft-versus-host disease (aGVHD) in a murine model. First, we assessed whether recombinant human (rh) Gal-9 can inhibit the mixed lymphocyte reaction (MLR) by culturing irradiated (30 Gy) splenic cells from 7- to 8-week-old female BDF1 mice with splenic cells from 7- to 8-week-old female C57BL/6J mice in the presence of various concentrations of Gal-9 for 10 days. rhGal-9 inhibited MLR in a dose-dependent manner. We then studied whether this effect was mediated by rhGal-9-induced apoptosis by culturing splenic cells from BDF1 mice with plate-bound anti-CD3 monoclonal antibody and Gal-9. Flow cytometric analysis revealed that rhGal-9 increased the number of Annexin-V+ cells in a dose-dependent manner (Figure. 1A). Thus, rhGal-9 inhibited MLR by inducing splenic cell apoptosis. This suppressive effect of Gal-9 on MLR was inhibited by lactose but not by sucrose (Figure. 1B). Taken together, Gal-9 induces T cell apoptosis through Ca2+ influx induced by binding to b-galactoside, resulting in the suppression of MLR. We then assessed whether rhGal-9 treatment altered aGVHD. Recipient B6D2F1 mice received a myeloablative dose (9 Gy) of total body irradiation from an X-ray irradiator. Six to eight hours later, each recipient mouse was injected i.v. with 4 × 106 TCD-BM cells and 5 × 106 mononuclear splenocytes. aGVHD clinical scores were evaluated once or twice a week. After aGVHD developed, recipient mice were treated with rhGal-9 (30 mg/mouse) or vehicle by i.p. injection for 14 consecutive days. The administration of rhGal-9 significantly attenuated aGVHD as compared to vehicle-treated mice (Figure. 2). Histological analyses also confirmed that aGVHD was declined by rhGal-9 treatment. Additionally, we investigated the effects of Gal-9 treatment on different T ell subsets. To analyze the effect of Gal-9 on donor lymphocytes, splenic mononuclear cells from GFP Tg mice were used for the induction of aGVHD. The gating parameter was first set to isolate the lymphocyte population of peripheral blood leukocytes, and then set for GFP+ cells. Gal-9 treatment decreased the frequency of CD4+/Tim-3+ cells and CD8+/intracellular IFN-g+ cells, whereas CD4+/CD25+ and CD25+/Foxp3+ Treg cells were increased by rhGal-9 treatment. These results suggest that Gal-9 regulates aGVHD by increasing regulatory T cells. In conclusion, Gal-9 ameliorates aGVHD by inducing T-cell apoptosis and also by increasing regulatory T cells. [Display omitted] [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Galectin-9 in Allergic Airway Inflammation and Hyper-Responsiveness in Mice

Background: Galectin-9 (Gal-9) is a member of the growing family of β-galactoside-binding lectins. Gal-9 is an eosinophil chemoattractant and inducer of Th1 cell apoptosis. These effects suggest its potential role in the pathogenesis of asthma. Our aim was to study the expression of Gal-9 in an ovalbumin (OVA)-induced mouse model of allergic asthma. Methods: To investigate the significance of Gal-9 in allergic inflammation and airway hyperresponsiveness (AHR), a group of BALB/c mice was sensitized and challenged with OVA (G_OVA). Another group of animals was allergized with OVA and also treated with dexamethasone (DEX) (G_OVA+DEX). The control group (G_PBS) received phosphate-buffered saline instead of OVA as placebo. Airway reactivity to intravenous methacholine was assessed. Results: The percentage of Gal-9-positive cells and their intracellular Gal-9 content and Th1/Th2 cytokine levels in the bronchoalveolar lavage (BAL) were determined by flow cytometry. Gal-9 mRNA expression and protein level were measured in the lung tissue by real-time RT-PCR and Western blot. In G_OVAmice, airway inflammation and mucus hypersecretion developed. DEX treatment inhibited the main features of experimental asthma. The number of Gal-9-positive lymphocytes, eosinophil and neutrophil granulocytes and the levels of Th2 cytokines were higher in the BAL of G_OVA compared to G_PBS or G_OVA+DEXmice. Moreover, Gal-9 protein level was elevated in the lungs of G_OVA mice. Conclusions: These results suggest that Gal-9 plays a role as a mediator contributing to the development of allergic airway inflammation. Gal-9 may serve as a recruiter of eosinophil granulocytes and promoter of Th2 dominance.

Novel Function of CRTH2 in Preventing Apoptosis of Human Th2 Cells through Activation of the Phosphatidylinositol 3-Kinase Pathway

It is now well established that interaction of PGD(2) with chemoattractant receptor- homologous molecule expressed on Th2 cells (CRTH2) promotes chemotaxis and proinflammatory cytokine production by Th2 lymphocytes. In this study we show a novel function of CRTH2 in mediating an inhibitory effect of PGD(2) on the apoptosis of human Th2 cells induced by cytokine deprivation. This effect was mimicked by the selective CRTH2 agonist 13,14-dihydro-15-keto-PGD(2), inhibited by the CRTH2 antagonists ramatroban and TM30089, and not observed in CRTH2-negative T cells. D prostanoid receptor 1 (DP(1)) or the thromboxane-like prostanoid (TP) receptor did not play a role in mediating the effects of PGD(2) on the apoptosis of Th2 cells because neither the DP(1) antagonist BW868C nor the TP antagonist SQ29548 had any effect on the antiapoptotic effect of PGD(2). Apoptosis of Th2 cells induced by Fas ligation was not suppressed by treatment with PGD(2), illustrating that activation of CRTH2 only inhibits apoptosis induced by cytokine deprivation. Treatment with PGD(2) induced phosphorylation of Akt and BAD, prevented release of cytochrome c from mitochondria, and suppressed cleavage of caspase-3 and poly(ADP-ribose) polymerase in Th2 cells deprived of IL-2. The PI3K inhibitor LY294002 blocked the effect of PGD(2) both on the signaling events and on the apoptotic death of Th2 cells. These data suggest that in addition to promoting the recruitment and activation of Th2 cells, PGD(2) may also impede the resolution of allergic inflammation through inhibiting apoptosis of Th2 cells.

Failure of galectin-9 expression by uterine endometrial epithelial cells as a cause for preeclampsia

Preeclampsia is a severe pregnancy complication that originates in the placenta and is characterized by shallow trophoblast invasion into the spiral arteries. Immunological imbalances associated with abnormal uterine spiral arteries remodeling during pregnancy have been identified to contribute to the onset and progression of preeclampsia. Interferon (IFN)-γ has a bilateral role in mediating uterine spiral artery remodeling and may lead to preeclampsia under abnormal circumstances. Until recently, the mechanism that regulates the balance between IFN-γ-mediated artery remodeling and IFN-γ-induced Th1 cell activation is ambiguous; but recent studies suggest an important part for galectin-9 in the immune regulation. Therefore, we hypothesize that the galectin-9 expression by uterine endometrial epithelial cells plays a key role in the regulation of the dual function of IFN-γ. Engaging galectin-9 with its receptor on activated Th1 cells causes an inhibitory signal, resulting in apoptosis of Th1 cells and negatively regulates Th1 type immunity. We further hypothesize that failure of galectin-9 expression by endometrial epithelial cells may dampen the endovascular remodeling process and thus result in preeclampsia. This hypothesis proposes a new mechanism in the immunological balance at the uteroplacental interface. Also this hypothesis will help to find out new cause for preeclamspsia and provide new strategy for disease treatment.

New insights into mechanisms of immunoregulation in 2007

Substantial progress in understanding the mechanisms of immune regulation in allergic diseases and asthma has been made during the last year. In asthma, rhinitis, and atopic dermatitis the immune system is activated by allergens, autoantigens, and components of superimposed infectious agents. Immune regulation in the lymphatic organs and in the tissue has an important role in the control and suppression of allergic disease in all stages of the inflammatory process, such as cell migration to tissues, cells gaining an inflammatory and tissue-destructive phenotype in the tissues, and their interaction with resident tissue cells to augment the inflammation. After the discovery of regulatory T cells, the importance of their unique suppressive capacity was strongly emphasized for the suppression of effector T-cell responses. However, it seems that all 3 subsets of effector T(H)1, T(H)2, and T(H)17 cells, as well as regulatory T cells, regulate each other at the level of transcription, major cytokines, and surface molecules. This review highlights key advances in immune regulation that were published in the Journal of Allergy and Clinical Immunology.

Delayed maturation of an IL-12–producing dendritic cell subset explains the early Th2 bias in neonatal immunity

Primary neonatal T cell responses comprise both T helper (Th) cell subsets, but Th1 cells express high levels of interleukin 13 receptor α1 (IL-13Rα1), which heterodimerizes with IL-4Rα. During secondary antigen challenge, Th2-produced IL-4 triggers the apoptosis of Th1 cells via IL-4Rα/IL-13Rα1, thus explaining the Th2 bias in neonates. We show that neonates acquire the ability to overcome the Th2 bias and generate Th1 responses starting 6 d after birth. This transition was caused by the developmental maturation of CD8α+CD4− dendritic cells (DCs), which were minimal in number during the first few days of birth and produced low levels of IL-12. This lack of IL-12 sustained the expression of IL-13Rα1 on Th1 cells. By day 6 after birth, however, a significant number of CD8α+CD4− DCs accumulated in the spleen and produced IL-12, which triggered the down-regulation of IL-13Rα1 expression on Th1 cells, thus protecting them against IL-4–driven apoptosis.

The immunosuppressive drug thalidomide induces T helper cell type 2 (Th2) and concomitantly inhibits Th1 cytokine production in mitogen- and antigen-stimulated human peripheral blood mononuclear cell cultures.

Thalidomide is an effective immunomodulatory drug in man, but its mechanism of action remains unclear. We hypothesized that, in addition to its reported inhibitory effects on production of monocyte-derived tumour necrosis factor-alpha (TNF-alpha), thalidomide might be effective at the level of Th immunoregulation. In a comparative study with the immunosuppressant cyclosporin A, we have demonstrated a potent and specific effect of thalidomide on cytokine production relating to the distinct Th1 and Th2 subsets. It induced and enhanced the production of IL-4 and IL-5 and, at the same dose (1000 ng/ml), significantly inhibited interferon-gamma (IFN-gamma) production in phytohaemagglutinin (PHA)-stimulated human peripheral blood mononuclear cell (PBMC) cultures. Stimulation of PBMC with recall antigen (streptokinase:streptodornase (SKSD)) at 144 h in the absence of thalidomide resulted in a predominantly Th1 response, with the production of IFN-gamma and IL-2. Thalidomide switched this response from a Th1 to a Th2 type. The effect was most pronounced at 1000 ng/ml thalidomide, where inhibition of IFN-gamma and enhancement of IL-4 production was maximal. In unstimulated cultures thalidomide alone induced IL-4 production. Cyclosporin A, in contrast, inhibited both Th1 and Th2 cytokine production by PHA-stimulated PBMC. Time course data from thalidomide-treated cultures revealed that the augmented IL-4 production diminished as the culture time increased, whereas IFN-gamma production was significantly increased. This response might be due to activation-induced apoptosis of Th2 cells or the induction of Th2 cell anergy, in the continued presence of stimulating agents, with the emergence of IFN-gamma-secreting Th1 cells when Th2 antagonism declines. The effects of thalidomide and related compounds may enhance our understanding of the mechanisms of T helper cell selection, offer the possibility of controlled therapeutic switching between Th1 and Th2 responses, and may lead to a rational approach for the treatment of some T cell-mediated immunological disorders.

Role of GADD45β in the regulation of synovial fluid T cell apoptosis in rheumatoid arthritis

Rheumatoid arthritis (RA) is characterized by persistent Th1 cell infiltration and production of inflammatory cytokines in the location of joint lesion. It is known that infiltrated Th1 cells in the synovial fluid (SF) of RA patients are resistant to apoptosis. Here we demonstrate that Th1 cells accumulated in patient SF expressed a high level of GADD45β ( Growth Arrest and DNA Damage-inducible 45β) which further inhibited Th1 cell apoptosis. Interestingly, in vitro culture of T cells with SF from RA patients increased GADD45β expression in Th1 cells and inhibited their apoptosis. Silencing of GADD45β by RNAi abolished the anti-apoptotic effect of RA SF, which was accompanied by down-regulation of Bcl-2 and up-regulation of Bax. Further analysis showed that TNF-α and IL-12 in RA SF could stimulate GADD45β expression in Th1 cells and inhibit their apoptosis. Taken together, our results suggest a novel mechanism by which specific cytokines in the RA SF elevate GADD45β expression in local Th1 cells and subsequently leading to the enhanced T cell survival.

Tim‐3 Signaling in T cells and Dendritic Cells

The T‐Cell Immunoglobulin and Mucin 3 Protein (Tim‐3) is a type I transmembrane protein that plays a role in regulating T helper responses and promoting tolerance. The extracellular portion of Tim‐3 consists of an immunoglobulin‐V and a mucin‐like domain followed by a transmembrane domain and a cytoplasmic tail that has 6 tyrosine residues. While previously thought to be specifically expressed by certain T cell subsets, we have also found Tim‐3 expressed on the surface of dendritic cells. Cross‐linking of Tim‐3 induces apoptosis in TH1 cells but activates dendritic cells. Therefore, our hypothesis is that the signaling pathway of Tim‐3 differs in lymphoid and myeloid cells. Our preliminary data indicate that AE7 (mouse TH1 clone) and D2SC/1 (mouse dendritic cell line) cells stimulated with anti‐Tim‐3 antibody display different tyrosine phosphorylation patterns. We are currently trying to identify specific phospho‐proteins present only in the lysates of Tim‐3 activated AE7 cells but not D2SC/1 cells. In addition, we have generated truncation and point mutants of Tim‐3 to determine which, if any, of the 6 tyrosine residues are required for the signaling capabilities of Tim‐3 in T cells and DC's. With these studies we aim to identify proximal signaling events downstream of Tim‐3 in lymphoid versus myeloid cells.

A small subset of dendritic cells controls the Th2 bias of neonatal immunity through an interplay between IL‐12 cytokine and IL‐13Rα1 expression

It is known that neonates are susceptible to microbial infection and allergic reactions. This is likely due to the lack of Th1 but excess of Th2 cells. The mechanism underlying this Th1/Th2 unbalance has not been clearly elucidated. When exposed to antigen (Ag) on the day of birth, both Th1 and Th2 cells develop in the primary response but Th1 cells up‐regulate IL‐13Rα1 chain which associates with IL‐4Rα to form an IL‐4Rα/IL‐13Rα1 heteroreceptor. During rechallenge with Ag, IL‐4 from Th2 cells utilizes this heteroreceptor to drive apoptosis of Th1 cells, hence creating a secondary Th2 immunity bias. Herein, it is shown that day 6 after birth represents a point at which the neonate acquires the ability to develop Th1 responses and overcome the Th2 bias. This transition lies on delayed developmental maturation of the CD8α+CD4‐ dendritic cell (DC) subset. Indeed, during the first 6 days of birth the neonates had minimal CD8α+CD4‐ DCs and were unable to produce significant IL‐12 which led to up‐regulation of IL‐13Rα1 expression on primary Th1 cells and formation of IL‐4Rα/IL‐13Rα1 heteroreceptor. By day 6 accumulation of CD8α+CD4‐DCs reaches optimal level leading to significant IL‐12 that opposes IL‐13Rα1 up‐regulation on Th1 cells and minimizes IL‐4Rα/IL‐13Rα1 expression.This research was supported by grants 2RO1AI48541 and R21AI06279 from NIH. CMH is supported by training grant GM08396‐16A1 from NIGMS.

Activation of Tim-3–Galectin-9 pathway improves survival of fully allogeneic skin grafts

T cell immunoglobulin and mucin domain (Tim)-3 is a molecule expressed on terminally differentiated murine Th1 cells but not on Th2 cells. Identification of Galectin-9 as a ligand for Tim-3 has now firmly established the Tim-3–Galectin-9 pathway as an important regulator of Th1 immunity, which results in apoptosis of Th1 cells. Here, we demonstrate that engagement of Tim-3 by mouse recombinant Galectin-9 remarkably suppresses allograft rejection and improves survival of allogeneic skin grafts. Furthermore, administration of recombinant Galectin-9 decreases Tim-3 positive cells in draining lymph node and selectively inhibits production of IFN-γ after skin transplantation. At last, even low dose of Galectin-9 (1 μg/ml) can obviously inhibit TCR crosslinking-induced primary T cell proliferation in vitro. These observations suggest that Tim-3–Galectin-9 pathway plays an important role in the termination of productive Th1-immune response and could lead to developing novel therapies in transplant medicine.