Chang-Qing Xia

IL-6 produced by dendritic cells from lupus-prone mice inhibits CD4+CD25+ T cell regulatory functions

The B6.Sle1.Sle2.Sle3 triple congenic mouse (B6.TC) is a model of lupus coexpressing the three major NZM2410-derived susceptibility loci on a C57BL/6 background. B6.TC mice produce high titers of antinuclear nephrogenic autoantibodies and a highly penetrant glomerulonephritis. Previous studies have shown the Sle1 locus is associated with a reduced number of regulatory T cells (Treg) and that Sle3 results in intrinsic defects of myeloid cells that hyperactivate T cells. In this report, we show that B6.TC dendritic cells (DCs) accumulate in lymphoid organs and present a defective maturation process, in which bone marrow-derived, plasmacytoid, and myeloid DCs express a significantly lower level of CD80, CD86, and MHC class II. B6.TC DCs also induce a higher level of proliferation in CD4+ T cells than B6 DCs, and B6.TC DCs block the suppressive activity of Treg. B6.TC DCs overproduce IL-6, which is necessary for the blockade of Treg activity, as shown by the effect of anti-IL-6 neutralizing Ab in the suppression assays. The overproduction of IL-6 by DCs and the blockade of Treg activity maps to Sle1, which therefore not only confers a reduced number of Treg but also blocks their ability to regulate autoreactive T cells. Taken together, these results provide a genetic and mechanistic evidence for systemic autoimmunity resulting from an impaired regulatory T cell compartment in both number and function and for Sle1-expressing DCs playing a major role in the latter defect though their production of IL-6.

Dexamethasone Induces IL-10-Producing Monocyte- Derived Dendritic Cells with Durable Immaturity

It is highly desirable that immature dendritic cells (DC) used for tolerance induction maintain steady immature state with predominant interleukin (IL)‐10 production. In this study, we attempted to develop DC with durable immaturity and other tolerogenic features by using dexamethasone (Dex). We found DC derived from human monocytes in the presence of 10−7 m Dex were negative for CD1a. Compared with control transduced DC (Ctrl‐DC), Dex‐DC expressed lower CD40, CD80 and CD86 but equivalent human leucocyte antigen‐DR. Both immature Dex‐ and Ctrl‐DC did not express CD83. Nevertheless, upon stimulation of lipopolysaccharide (LPS) or CD40 ligand, the expression of CD40, CD80, CD83 and CD86 was upregulated on Ctrl‐DC but not on Dex‐DC. The immaturity of Dex‐DC was durable following Dex removal. Interestingly, Dex‐DC maintained production of large amount of IL‐10 and little IL‐12 five days after Dex removed. Further study indicated that high‐level IL‐10 production by Dex‐DC was associated with high‐level phosphorylation of extracellular signal‐regulated kinase (ERK) as blockade of this enzyme markedly attenuated IL‐10 production. Furthermore, Dex‐DC sustained the capability of high phosphorylation of ERK and IL‐10 production 5 days after Dex removal. In addition, Dex‐DC had significantly lower activity in stimulating T‐cell proliferation. Neutralization of IL‐10, to some extent, promoted DC maturation activated by LPS, as well as T‐cell stimulatory activity of Dex‐DC. The above findings suggest that IL‐10‐producing Dex‐DC with durable immaturity are potentially useful for induction of immune tolerance.

Transfusion of Apoptotic β-Cells Induces Immune Tolerance to β-Cell Antigens and Prevents Type 1 Diabetes in NOD Mice

In vivo induction of β-cell apoptosis has been demonstrated to be effective in preventing type 1 diabetes in NOD mice. Based on the notion that steady-state cell apoptosis is associated with self-tolerance and the need for developing a more practical approach using apoptotic β-cells to prevent type 1 diabetes, the current study was designed to investigate apoptotic β-cells induced ex vivo in preventing type 1 diabetes. The NIT-1 cell line serves as a source of β-cells. Apoptotic NIT-1 cells were prepared by ultraviolet B (UVB) irradiation. Three weekly transfusions of UVB-irradiated NIT-1 cells (1 × 105/mouse) or PBS were used to determine whether transfusions of UVB-irradiated NIT-1 cells induce immune tolerance to β-cell antigens in vivo and prevent type 1 diabetes. The suppression of anti–β-cell antibodies, polarization of T-helper (Th) cells, and induction of regulatory T-cells by UVB-irradiated NIT-1 cell treatment were investigated. The transfusions of apoptotic NIT-1 cells suppress anti–β-cell antibody development and induce Th2 responses and interleukin-10–producing regulatory type 1 cells. Importantly, this treatment significantly delays and prevents the onset of diabetes when 10-week-old NOD mice are treated. Adoptive transfer of splenocytes from UVB-irradiated NIT-1 cell–treated mice prevents diabetes caused by simultaneously injected diabetogenic splenocytes in NOD-Rag−/− mice. Moreover, the proliferation of adoptively transferred carboxyfluorescein diacetate succinimidyl ester–labeled β-cell antigen–specific T-cell receptor–transgenic T-cells in UVB-irradiated NIT-1–cell treated mice is markedly suppressed. The transfusion of apoptotic β-cells effectively protects against type 1 diabetes in NOD mice by inducing immune tolerance to β-cell antigens. This approach has great potential for immune intervention for human type 1 diabetes.

Heparin Induces Differentiation of CD1a+ Dendritic Cells from Monocytes: Phenotypic and Functional Characterization

Dendritic cells (DCs) play important roles in initiation and regulation of immune responses. DCs derived from human monocytes can be classified according to presence of CD1a molecules. Although CD1a+ DCs can be prepared from monocytes in media containing GM-CSF, IL-4, and FCS, it has been reported that CD1a+ DCs could not be easily obtained from monocytes using media containing human serum or plasma. In this study, we demonstrate for the first time that heparin can reliably induce differentiation of CD1a+ DCs from monocytes with or without autologous serum or plasma. The development of CD1a+ DCs is heparin concentration dependent (0–50 U/ml). Comparing with CD1a− DCs developed without heparin, CD1a+ DCs express higher CD40 and CD80 and lower CD86. Both CD1a+ and CD1a− DCs express similar levels of HLA-DR. CD80, CD86, HLA-DR, and CD40 are proportionally up-regulated when both types of DCs are stimulated with LPS or LPS plus IFN-γ. The effect of heparin is neutralized by heparin-binding proteins, such as protamine sulfate, platelet factor-4, and β-thromboglobulin. Functionally, heparin-treated DCs respond to LPS or LPS plus IFN-γ with higher IL-10 and less IL-12 production than heparin-untreated DCs. Heparin-treated DCs are more potent in priming allogeneic and autologous CD4+ T cells to proliferate and to produce both type 1 and type 2 cytokines. The results of our study show that CD1a+ DCs can be prepared from monocytes ex vivo without using xenogeneic serum and may be used for immunotherapy.

Extracorporeal photopheresis-induced immune tolerance: a focus on modulation of antigen-presenting cells and induction of regulatory T cells by apoptotic cells.

Purpose of reviewThis review is intended to introduce recent advances in the research surrounding extracorporeal photopheresis (ECP) with a focus on how apoptotic cells modulate antigen-presenting cells and induce regulatory T cells, given that ECP therapy induces apoptosis of leukocytes collected through leukapheresis. Recent findingsIt has been suggested that ECP therapy, unlike other immunosuppressive regimens, does not cause global immunosuppression, but induces immune tolerance. Recent clinical and animal studies demonstrate that ECP therapy induces antigen-specific regulatory T cells, including CD4+CD25+FoxP3+ T cells and IL-10-producing Tr1 cells, that may arise secondarily to the induction of tolerogenic antigen-presenting cells (APCs) by infusion of apoptotic cells. It has also been suggested that ECP therapy may induce IL-10-producing regulatory B cells and regulatory CD8+ T cells. Finally, several recent studies, which examined the cellular elements involved in the uptake of apoptotic cells, demonstrated that apoptotic cells modulate APCs through binding to specific receptors, particularly TAM receptors that provide inhibitory signals that block APC activation. SummaryECP therapy induces immune tolerance through modulation of antigen-presenting cells as well as induction of regulatory T cells. ECP therapy has great potential in the management of allogeneic transplantation and autoimmune diseases.

MicroRNA-17-92 controls T-cell responses in graft-versus-host disease and leukemia relapse in mice

MicroRNAs (miRs) play important roles in orchestrating many aspects of the immune response. The miR-17-92 cluster, which encodes 6 miRs including 17, 18a, 19a, 20a, 19b-1, and 92-1, is among the best characterized of these miRs. The miR-17-92 cluster has been shown to regulate a variety of immune responses including infection, tumor, and autoimmunity, but the role of this cluster in T-cell response to alloantigens has not been previously explored. By using major histocompatibility complex (MHC)-matched, -mismatched, and haploidentical murine models of allogeneic bone marrow transplantation (allo-BMT), we demonstrate that the expression of miR-17-92 on donor T cells is essential for the induction of graft-versus-host disease (GVHD), but dispensable for the graft-versus-leukemia (GVL) effect. The miR-17-92 plays a major role in promoting CD4 T-cell activation, proliferation, survival, and Th1 differentiation, while inhibiting Th2 and iTreg differentiation. Alternatively, miR-17-92 may promote migration of CD8 T cells to GVHD target organs, but has minimal impact on CD8 T-cell proliferation, survival, or cytolytic function, which could contribute to the preserved GVL effect mediated by T cells deficient for miR-17-92. Furthermore, we evaluated a translational approach and found that systemic administration of antagomir to block miR-17 or miR-19b in this cluster significantly inhibited alloreactive T-cell expansion and interferon-γ (IFNγ) production, and prolonged the survival in recipients afflicted with GVHD while preserving the GVL effect. Taken together, the current work provides a strong rationale and demonstrates the feasibility to target miR-17-92 for the control of GVHD while preserving GVL activity after allo-BMT.

Peptide‐Pulsed Immature Dendritic Cells Reduce Response to β Cell Target Antigens and Protect NOD Recipients from Type I Diabetes

Abstract:  Our previous work demonstrated peptide‐pulsed mature myeloid dendritic cells (DC) presenting β cell antigens induce tolerance. Here we determine whether immature DC (iDC) presenting dominant (insulin β9–23 chain, proinsulin C19–A3) or ignored (glutamic acid decarboxylase 6578‐97) antigen determinants promote tolerance. Nonobese diabetic (NOD) mice were given injections of either unpulsed or peptide‐pulsed myeloid iDC beginning at 9 weeks of age for 3 consecutive weeks. Diabetes incidence in recipients of unpulsed iDC was comparable to unmanipulated animals (∼80%), whereas GAD6578–97 pulsed iDC recipients were protected from the disease (P= 0.05). We also analyzed splenic T cell proliferation responses to the panel of studied peptides in diabetic and nondiabetic recipients. When stimulated with insulin or proinsulin peptide, nondiabetic mice receiving the peptide‐pulsed iDC had a 21‐ to 31‐fold or 3.9‐ to 9.0‐fold reduction in T cell response, respectively, as compared to the response of diabetic unpulsed recipients. However, only a 2.6‐ to 3.1‐fold reduction in response to β chain peptide, and a 1.5‐ to 3.4‐fold reduction in proinsulin response were observed in diabetic mice receiving peptide‐pulsed iDC. The reduction was not specific to the immunizing peptide, as reduced proliferation was observed to other diabetes‐target peptides. We conclude that protective iDC‐based therapies require target antigen presentation, and ignored determinants may be preferable perhaps due to an available naïve T cell repertoire. In addition, iDC presenting peptides induce a nonspecific reduction in T cell responses to β cell antigens, possibly through the induction of regulatory T cells.

LBH589 enhances T cell activation in vivo and accelerates graft-versus-host disease in mice.

Histone deacetylase inhibitors (HDACis) are a new class of compounds that induce acetylation of histone lysine tails in chromatin and modify gene expression. The Food & Drug Administration approved HDACi, Vorinostat, or suberoylanilide hydroxamic acid (SAHA), has been shown to inhibit tumor cell growth and the production of proinflammatory cytokines. In preclinical allogeneic transplant models, SAHA induces graft-versus-host disease (GVHD) amelioration in treated mice without impairing graft-versus-leukemia. LBH589 (Panobinostat), a structurally novel cinnamic hydroxamic acid class, is an HDACi more potent than SAHA. In the current work, we tested the hypothesis that LBH589 would be highly effective in the prevention of GVHD. Using mouse model of allogeneic bone marrow transplant (BMT), we unexpectedly found that treatment with LBH589 accelerated GVHD, in contrast to the treatment with SAHA that alleviated GVHD. Accelerated GVHD in the recipients treated with LBH589 was associated with elevated Th1 cytokines in recipient serum, enhanced CXCR3 expression on donor T cells, and T cell infiltration in the liver. The current study highlights the distinct effects of pan HDACi on allogeneic BMT and alerts that LBH589 (Panobinostat) could have an adverse effect on GVHD, and possibly on other inflammatory diseases.

Rabbit Polyclonal Mouse Antithymocyte Globulin Administration Alters Dendritic Cell Profile and Function in NOD Mice to Suppress Diabetogenic Responses

Mouse antithymocyte globulin (mATG) prevents, as well as reverses, type 1 diabetes in NOD mice, through mechanisms involving modulation of the immunoregulatory activities of T lymphocytes. Dendritic cells (DC) play a pivotal role in the generation of T cell responses, including those relevant to the autoreactive T cells enabling type 1 diabetes. As Abs against DC are likely generated during production of mATG, we examined the impact of this preparation on the phenotype and function of DC to elucidate novel mechanisms underlying its beneficial activities. In vivo, mATG treatment transiently induced the trafficking of mature CD8− predominant DC into the pancreatic lymph node of NOD mice. Splenic DC from mATG-treated mice also exhibited a more mature phenotype characterized by reduced CD8 expression and increased IL-10 production. The resultant DC possessed a potent capacity to induce Th2 responses when cultured ex vivo with diabetogenic CD4+ T cells obtained from BDC2.5 TCR transgenic mice. Cotransfer of these Th2-deviated CD4+ T cells with splenic cells from newly diabetic NOD mice into NOD.RAG−/− mice significantly delayed the onset of diabetes. These studies suggest the alteration of DC profile and function by mATG may skew the Th1/Th2 balance in vivo and through such actions, represent an additional novel mechanism by which this agent provides its beneficial activities.

Anti-thymocyte globulin (ATG) differentially depletes naïve and memory T cells and permits memory-type regulatory T cells in nonobese diabetic mice

BackgroundATG has been employed to deplete T cells in several immune-mediated conditions. However, whether ATG administration affects naïve and memory T cell differently is largely unknown.The context and purpose of the studyIn this study, we assessed how murine ATG therapy affected T cell subsets in NOD mice, based on their regulatory and naïve or memory phenotype, as well as its influence on antigen-specific immune responses.ResultsPeripheral blood CD4+ and CD8+ T cells post-ATG therapy declined to their lowest levels at day 3, while CD4+ T cells returned to normal levels more rapidly than CD8+ T cells. ATG therapy failed to eliminate antigen-primed T cells. CD4+ T cell responses post-ATG therapy skewed to T helper type 2 (Th2) and possibly IL-10-producing T regulatory type 1 (Tr1) cells. Intriguingly, Foxp3+ regulatory T cells (Tregs) were less sensitive to ATG depletion and remained at higher levels following in vivo recovery compared to controls. Of note, the frequency of Foxp3+ Tregs with memory T cell phenotype was significantly increased in ATG-treated animals.ConclusionATG therapy may modulate antigen-specific immune responses through inducing memory-like regulatory T cells as well as other protective T cells such as Th2 and IL-10-producing Tr1 cells.