Xiaochun Wan

Modulation of T-cell responses to alloantigens by TR6/DcR3

TR6 (DcR3) is a new member of the TNF receptor (TNFR) family that lacks a transmembrane domain in its sequence, indicating that it is a secreted molecule. TR6 can bind to FasL and prevent FasL-induced apoptosis; it can also associate with LIGHT, another TNF family member. The role of TR6 in immune responses was investigated in this study. According to flow cytometry, recombinant human TR6-Fc binds to human LIGHT expressed on 293 cells or on activated human T cells and competes with the LIGHT receptor TR2 for the binding to LIGHT on these cells. Human TR6 could cross-react with mouse LIGHT in immunoprecipitation. TR6-Fc also downregulates cytotoxic T lymphocyte activity in vitro and graft-versus-host responses in mice. Moreover, TR6-Fc modulates lymphokine production by alloantigen-stimulated mouse T cells. TR6-Fc ameliorated rejection response to mouse heart allograft. These results indicate that TR6 can dampen T-cell responses to alloantigens. Such regulatory effects of TR6 probably occur via interference with interaction between pairs of related TNF and TNFR family members, LIGHT/TR2 being one of the possible candidate pairs.

A TNF Family Member LIGHT Transduces Costimulatory Signals into Human T Cells

DcR3/TR6 is a secreted protein belonging to the TNFR family. It binds to Fas ligand, LIGHT, and TL1A, all of which are TNF family members. LIGHT is expressed on activated T cells. Its known receptors are TR2 and LTβR on the cell surface, and TR6 in solution. In the present study, we report soluble TR6-Fc or solid-phase TR6-Fc costimulated proliferation, lymphokine production, and cytotoxicity of human T cells in the presence of TCR ligation. These costimulating effects were blocked by soluble LIGHT but not by soluble Fas-Fc. TR6-Fc could also effectively costimulate gld/gld mouse T cells. We further demonstrated that TR6 bound to both Th1 and Th2 cells, according to flow cytometry, and that the association was inhibited by soluble LIGHT. Cross-linking Th1 and Th2 cells with solid-phase TR6-Fc along with a suboptimal concentration of anti-CD3 enhanced proliferation of both Th1 and Th2 cells, and augmented Th1 but not Th2 lymphokine production. These data suggest that TR6 delivers costimulation through its ligand(s) on the T cell surface, and at least the major part of such costimulation is via LIGHT.

A proteasome inhibitor effectively prevents mouse heart allograft rejection

BACKGROUND We have previously demonstrated in vitro that proteasome inhibitors could suppress proliferation and induce apoptosis of activated T cells. This finding suggests that such inhibitors could be used as a novel category of immunosuppressants in blocking allograft rejection. METHODS The proteasome inhibitor dipeptide boronic acid (DPBA) was tested in vitro for its inhibitory effect on mouse T-cell proliferation and lymphokine secretion. DPBA was also used in vivo to treat mouse heterotopic heart allograft rejection. Possible side effects of this compound were examined according to blood chemistry of mice treated with DPBA. RESULTS DPBA suppressed the T-cell proliferation and potently inhibited interleukin (IL)-2, IL-6, IL-10, IL-13, and IFN-gamma produced by anti-CD3-activated T cells. Given i.p. starting 1 day after transplantation at 0.66 mg/kg per day for 16 days, or at 1 mg/kg per day for 4 days followed by 0.5 mg/kg per day for 12 days, DPBA could prolong heart allograft survival to 35.5 days (mean survival time, MST) and to 36.2 days, respectively. The control group had MST of 7.3 days. When administrated 72 hr post operation at 1 mg/kg per day for 4 days, DPBA could prolong the graft survival to 19.8 days. During the course of these effective dosages, DPBA had no apparent toxicity in the liver, kidney, pancreas, or heart, according to analysis of blood chemistry. CONCLUSIONS The proteasome inhibitor could repress allograft rejection in mice without apparent side-effects at the effective dosages. This finding has opened a new dimension in development of novel immunosuppressants for organ transplantation.

Cross-Linking of EphB6 Resulting in Signal Transduction and Apoptosis in Jurkat Cells

Eph kinases are the largest family of receptor tyrosine kinases (RTK), and their ligands are cell surface molecules. The known functions of Eph kinases are mainly pattern formation in the CNS. Although several Eph kinases are expressed at high levels in hemopoietic cells and in the thymus, we have no knowledge of the functions of any Eph kinase in the immune system. In this study, we have demonstrated that an Eph kinase, EphB6, was expressed at high levels in Jurkat leukemic T cells. Co-cross-linking of EphB6 and CD3 led to an altered profile of lymphokine secretion along with proliferation inhibition of Jurkat cells. The cells subsequently underwent Fas-mediated apoptosis. Although EphB6 has no intrinsic kinase activity, its cross-linking triggered general protein tyrosine phosphorylation in Jurkat cells. EphB6 was found to associate with a number of molecules in the signaling pathways, notably Cbl. EphB6 cross-linking resulted in Cbl dephosphorylation and dissociation from Src homology 2 domain-containing tyrosine phosphatase-1 (SHP-1). Our results show that EphB6 has important functions in T cells, and it can transduce signals into the cells via proteins it associates with.

MicroRNA-21 regulates T-cell apoptosis by directly targeting the tumor suppressor gene Tipe2

MicroRNAs (MiRs) are short noncoding RNAs that can regulate gene expression. It has been reported that miR-21 suppresses apoptosis in activated T cells, but the molecular mechanism remains undefined. Tumor suppressor Tipe2 (or tumor necrosis factor-α-induced protein 8 (TNFAIP8)-like 2 (TNFAIP8L2)) is a newly identified anti-inflammatory protein of the TNFAIP8 family that is essential for maintaining immune homeostasis. We report here that miR-21 is a direct target of nuclear factor-κB and could regulate Tipe2 expression in a Tipe2 coding region-dependent manner. In activated T cells and macrophages, Tipe2 expression was markedly downregulated, whereas miR-21 expression was upregulated. Importantly, Tipe2-deficient T cells were significantly less sensitive to apoptosis. Conversely, overexpression of Tipe2 in EL-4 T cells increased their susceptibility to activation-induced apoptosis. Therefore, Tipe2 provides a molecular bridge between miR-21 and cell apoptosis; miR-21 suppresses apoptosis in activated T cells at least in part through directly targeting tumor suppressor gene Tipe2.

Compromised kidney graft rejection response in Vervet monkeys after withdrawal of immunosuppressants tacrolimus and sirolimus.

BACKGROUND In nonprimates, organ allografts are often not rejected after withdrawal of immunosuppression. In this study, we examined whether such a phenomenon also occurs in primates. METHODS Vervet monkeys were transplanted with renal allografts and treated for 60 days with tacrolimus, or tacrolimus plus sirolimus. The drugs were totally withdrawn on day 61. The survival of the monkeys was monitored, and their response to donor- or third party-derived alloantigens was examined in vivo and in vitro. RESULTS The majority (80-100%) of the grafts survived for at least additional 30 days with no signs of acute rejection. The compromised rejection is donor-specific, because recipient monkeys failed to reject a donor-derived skin graft, but a third-party skin graft was rejected. In vitro mixed lymphocyte reaction and interleukin-2 production in the mixed lymphocyte reaction between the recipients and their donors or between the recipients and a third party had no discernable patterns, and thus did not reflect the in vivo status of the immune system. Although the recipients could not reject the graft acutely after drug withdrawal, the kidney grafts and the donor-derived skin grafts had pathological findings of chronic rejection. CONCLUSIONS The rejection response of the monkeys to an established graft after withdrawal of immunosuppression is compromised. The compromised rejection is specific and is not due to a permanent alteration of the immune system by the initial drug treatment. The allografts are not inert but have low levels of interaction with the recipient immune system.

DcR3/TR6 modulates immune cell interactions

DcR3/TR6, a secreted protein, is a member of TNF receptor family. Its ligands include FasL, LIGHT, and TL1A, all TNF family members. TR6 can interfere with FasL‐ or LTβR‐mediated apoptosis; it can also inhibit T‐cell costimulation by blocking the two‐way signaling between TR2 and LIGHT, and the one‐way signaling from TL1A to DR3. In this study, we discovered that TR6 was secreted by peripheral blood mononuclear cells (PBMC) stimulated by T‐cell mitogens. It inhibited actin polymerization of T cells upon mitogen stimulation, and repress T‐cell pseudopodium formation, which is known to be important for cell–cell interaction. As a consequence, T‐cell aggregation stimulated by alloantigens, anti‐CD3 or PHA was suppressed by either soluble or solid phase TR6‐Fc. This result suggests that TR6 might regulate T‐cell interaction with other cells such as antigen‐presenting cells (APC) or their fellow T cells by preventing them from forming inseparable cell clusters, which are undesirable for the progression of immune responses.

NK Cells Alleviate Lung Inflammation by Negatively Regulating Group 2 Innate Lymphoid Cells

Group 2 innate lymphoid cells (ILC2s) play an important role in orchestrating type II immune responses. However, the cellular mechanisms of group 2 innate lymphoid cell regulation remain poorly understood. In this study, we found that activated NK cells inhibited the proliferation of, as well as IL-5 and IL-13 production by, ILC2s in vitro via IFN-γ. In addition, in a murine model of ILC2 expansion in the liver, polyinosinic-polycytidylic acid, an NK cell–activating agent, inhibited ILC2 proliferation, IL-5 and IL-13 production, and eosinophil recruitment. Such effects of polyinosinic-polycytidylic acid were abrogated in NK cell–depleted mice and in IFN-γ–deficient mice. Adoptively transferring wild-type NK cells into NK cell–depleted mice resulted in fewer ILC2s induced by IL-33 compared with the transfer of IFN-γ–deficient NK cells. Importantly, during the early stage of papain- or bleomycin-induced lung inflammation, depletion of NK cells resulted in increased ILC2 numbers and enhanced cytokine production by ILC2s, as well as aggravated eosinophilia and goblet cell hyperplasia. Collectively, these data show that NK cells negatively regulate ILC2s during the early stage of lung inflammation, which represents the novel cellular interaction between two family members of ILCs.

The MicroRNA-21 in Autoimmune Diseases.

MicroRNA-21 (miR-21) is an oncomiR and significantly upregulated in a wide range of cancers. It is strongly involved in apoptosis and oncogenesis, since most of its reported targets are tumor suppressors. Recently, miR-21 was found to be correlated with the pathogenesis of autoimmune diseases and may play an essential role in regulating autoimmune responses. In particular, miR-21 promotes Th17 cell differentiation, which mediates the development of multiple autoimmune diseases. In this article, we review the current research on the mechanisms that regulate miR-21 expression, the potential of miR-21 as a diagnostic biomarker for autoimmune disease and the mechanisms by which miR-21 promotes the development of autoimmune disease. We also discussed the therapeutic potential of targeting miR-21 in treating patients with autoimmune disease.

Treating psoriasis by targeting its susceptibility gene Rel

Psoriasis is a chronic inflammatory disorder of the skin. Accumulating evidence indicates that the Rel gene, a member of the NF-κB family, is a risk factor for the disease. We sought to investigate whether psoriasis can be prevented by directly targeting the Rel gene transcript, i.e., the c-Rel mRNA. Using chemically-modified c-Rel specific siRNA (siRel) and poly(ethylene glycol)-b-poly(l-lysine)-b-poly(l-leucine) (PEG-PLL-PLLeu) micelles, we successfully knocked down the expression of c-Rel, and showed that the expression of cytokine IL-23, a direct target of c-Rel that can drive the development of IL-17-producing T cells, was markedly inhibited. More importantly, treating mice with siRel not only prevented but also ameliorated imiquimod (IMQ)-induced psoriasis. Mechanistic studies showed that siRel treatment down-regulated the expression of multiple inflammatory cytokines. Taken together, these results indicate that the susceptibility gene Rel can be targeted to treat and prevent psoriasis.