scholarly journals Induction of antigen-specific tolerance to bone marrow allografts with CD4+CD25+ T lymphocytes

Blood ◽  
2004 ◽  
Vol 103 (11) ◽  
pp. 4216-4221 ◽  
Author(s):  
Olivier Joffre ◽  
Nathalie Gorsse ◽  
Paola Romagnoli ◽  
Denis Hudrisier ◽  
Joost P. M. van Meerwijk
Keyword(s):  
Bone Marrow ◽  
T Cells ◽  
T Lymphocytes ◽  
Regulatory T Cells ◽  
Third Party ◽  
Antigen Specificity ◽  
Minor Histocompatibility Antigens ◽  
Regulatory T Lymphocytes

Abstract Thymus-derived regulatory T lymphocytes of CD4+CD25+ phenotype regulate a large variety of beneficial and deleterious immune responses and can inhibit lethal graft-versus-host disease in rodents. In vitro, CD4+CD25+ T cells require specific major histocompatibility complex (MHC)/peptide ligands for their activation, but once activated they act in an antigen-nonspecific manner. In vivo, regulatory T cells are also activated in an antigen-specific fashion, but nothing is known about antigen specificity of their suppressor-effector function. Here we show that CD4+CD25+ regulatory T lymphocytes isolated from naive mice and activated in vitro with allogeneic antigen-presenting cells (APCs) induced specific long-term tolerance to bone marrow grafts disparate for major and minor histocompatibility antigens; whereas “target” bone marrow was protected, third-party bone marrow was rejected. Importantly, in mice injected with a mix of target and third-party bone marrows, protection and rejection processes took place simultaneously. These results indicate that CD4+CD25+ regulatory T cells can act in an antigen-specific manner in vivo. Our results suggest that CD4+CD25+ regulatory T cells could in the future be used in clinical settings to induce specific immunosuppression. (Blood. 2004;103:4216-4221)

scholarly journals CD8α+ plasmacytoid precursor DCs induce antigen-specific regulatory T cells that enhance HSC engraftment in vivo

Blood ◽  
2011 ◽  
Vol 117 (8) ◽  
pp. 2494-2505 ◽  
Author(s):  
Yiming Huang ◽  
Larry D. Bozulic ◽  
Thomas Miller ◽  
Hong Xu ◽  
Lala-Rukh Hussain ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Regulatory T Cells ◽  
Foxp3 Expression ◽  
Cell Receptor ◽  
Third Party ◽  
Antigen Specificity ◽  
Clinical Potential

Abstract CD8-positive/T-cell receptor–negative (CD8+/TCR−) graft facilitating cells (FCs) are a novel cell population in bone marrow that potently enhance engraftment of hemopoietic stem cells (HSCs). Previously, we showed that the CD11c+/B220+/CD11b− plasmacytoid-precursor dendritic cell (p-preDC) FC subpopulation plays a critical but nonredundant role in facilitation. In the present study, we investigated the mechanism of FC function. We report that FCs induce antigen-specific CD4+/CD25+/FoxP3+ regulatory T cells (Tregs) in vivo. The majority of chimeric Tregs were recipient derived. Chimeric Tregs harvested at ≥ 4 weeks after transplantation significantly enhanced engraftment of donor- and recipient-derived HSCs, but not third-party HSCs, in conditioned secondary recipients, demonstrating antigen specificity. Although Tregs were present 2 and 3 weeks after transplantation, they did not enhance engraftment. In contrast, week 5 and greater Tregs potently enhanced engraftment. The function of chimeric Tregs was directly correlated with the development of FoxP3 expression. Chimeric Tregs also induced significantly stronger suppression of T-cell proliferation to donor antigen in vitro. Removal of p-preDC FCs resulted in impaired engraftment of allogeneic HSCs and failure to produce chimeric Tregs, suggesting that the CD8α+ p-preDC subpopulation is critical in the mechanism of facilitation. These data suggest that FCs induce the production of antigen-specific Tregs in vivo, which potently enhance engraftment of allogeneic HSCs. FCs hold clinical potential because of their ability to remain tolerogenic in vivo.

Suppression of NK Cell-Mediated Bone Marrow Cell Rejection by CD4+CD25+ Regulatory T Cells: Linkage of Adaptive to Innate Responses.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 2195-2195
Author(s):  
William J. Murphy ◽  
Isabel Bareo ◽  
Alan M. Hanash ◽  
Lisbeth A. Welniak ◽  
Kai Sun ◽  
...  
Keyword(s):  
Bone Marrow ◽  
T Cells ◽  
Regulatory T Cells ◽  
Cell Function ◽  
Nk Cell ◽  
Marrow Cell ◽  
Poly I:C ◽  
Hybrid Resistance

Abstract While a link between the innate to adaptive immune system has been established, studies demonstrating direct effects of T cells in regulating Natural Killer (NK) cell function have been lacking. Naturally occurring CD4+CD25+ regulatory T cells (Tregs) have been shown to potently inhibit adaptive responses by T cells. We therefore investigated whether Tregs could affect NK cell function in vivo. Using a bone marrow transplantation (BMT) model of hybrid resistance, in which parental (H2d) marrow grafts are rejected by the NK cells of the F1 recipients (H2bxd), we demonstrate that the in vivo removal of host Tregs significantly enhances NK-cell mediated BM rejection. This heightened rejection was mediated by the specific NK cell Ly-49+ subset previously demonstrated to reject the BMC in this donor/host pairing. The depletion of Tregs could also further increase rejection already enhanced by treating recipients with the NK cell activator, poly I:C. Although splenic NK cell numbers were not significantly altered, increased splenic NK in vitro cytotoxic activity was observed from the recovered cells. The regulatory role of Tregs was confirmed in adoptive transfer studies in which transferred CD4+CD25+ Tregs resulted in abrogation of NK cell-mediated hybrid resistance. Thus, Tregs can potently inhibit NK cell function in vivo and their depletion may have therapeutic ramifications with NK cell function in BMT and cancer therapy.

Safely Improving the in Vivo Survival of Tumor Specific Cytotoxic T Lymphocytes by Co-Transfer of IL7 Receptor Alpha Chain and icaspase9

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 3534-3534
Author(s):  
Juan F Vera ◽  
Valentina Hoyos ◽  
Barbara Savoldo ◽  
Concetta Quintarelli ◽  
Greta A Giordano ◽  
...  
Keyword(s):  
T Cells ◽  
T Lymphocytes ◽  
Cytotoxic T Lymphocytes ◽  
Adoptive Transfer ◽  
Fold Increase ◽  
Suicide Gene ◽  
Antigen Specificity ◽  
Anti Tumor Activity

Abstract Providing a proliferative and survival advantage to tumor-specific cytotoxic T lymphocytes (CTLs) remains a challenge in the adoptive therapy of cancer patients. It is now evident that the in vivo expansion of T cells after adoptive transfer is best accomplished in the lymphodepleted host due to the increased production of endogenous IL15 and IL7, which help restore lymphopoiesis. We have found that antigen activated cytotoxic T lymphocytes (CTLs) directed to tumor associated epitopes (for example derived from EBV, or from cancer testis antigens such as PRAME) down regulate a chain of IL7R, a common γ chain cytokine receptor, impairing their capacity to respond to IL7. We hypothesized that despite receptor downregulation, the signal transduction pathway for IL7R would remain intact in the CTLs so that forced expression of IL7Rα would restore IL7 responsiveness and improve in vivo expansion and survival of CTLs. We used EBV-specific CTLs as our model, and showed in vitro that a functional IL-7Ra molecule can be expressed in CTLs using retroviral gene transfer so that the percentage of receptor + cells increased from 2.4%±0.5% to 50%±20. This modification restored the in vitro proliferation of genetically modified CTLs in response to IL7 so that cell numbers increased from 1×106 cells to 0.1×109 (range, 0.6×108 to 0.3×109)] comparable with the effects of IL2 [from 1×106 cells to 0.7×109 (range, 0.7×107 to 1.6×109)] In contrast, control EBV-CTL with IL7 progressively declined in number (p<0.001) These effects were accomplished without alteration of antigen specificity or responsiveness to other common γ chain cytokines, and cell survival remained antigen dependent. In a xenogeneic mouse model, CTLs expressing IL7Ra significantly expanded in vivo in response to EBV-tumor antigen and the administration of IL7. By day 15, both control CTLs and IL7Ra+ CTLs had modestly proliferated in response to IL-2 (2.3 fold, range 1.1–5.1 for control CTLs, and 2.67 fold, range 0.6 to 8.15 for IL7Ra+ CTLs). In contrast, only IL7Ra+ CTLs significantly expanded in the presence of IL7, showing a 6.09 fold increase (range 0.7 to 25.2) compared to mice that received control CTLs and IL7 (0.9 fold, range 0.5–1.7) (p<0.0001). Modified CTLs also provided enhanced anti-tumor activity. SCID mice engrafted i.p with 3×106 tumor cells marked with Firefly luciferase, showed a rapid increase in signal in the absence of CTLs (Fold increase in luminance = 29.8 median, range 4.4 to 103) by day 14 after tumor engraftment. Similar tumor growth was observed in mice receiving IL7Ra+ CTLs without cytokines (luminance increase14.4 fold, range 1 to 90). In contrast, mice receiving IL7Ra+ CTLs and either IL2 or IL7, had a decline in tumor luminance (fold expansion 0.7, range 0.08 to 2.9, and 0.8, range 0.004 to 3.5, respectively p<0.0001). Although growth of the transgenic T cells remained antigen dependent, as a further safety measure, we incorporated an inducible suicide gene based on icaspase9 that can be activated by exposure to a small chemical inducer of dimerization (CID) (AP20187). Incorporation of this suicide gene did not affect the in vitro or in vivo anti-tumor activity of the CTL’s but allowed them to be rapidly eliminated. So that after a single dose of CID (50 nM) the transgenic population were decreased by >98.5% We conclude that forced expression of the IL-7Ra by CTLs can be used to recapitulate the response of these cells to this cytokine and thereby promote their in vivo anti-tumor activity after adoptive transfer either in a lymphodepleted host or after the administration of the recombinant protein.

scholarly journals Induction of tolerance to bone marrow allografts by donor-derived host nonreactive ex vivo–induced central memory CD8 T cells

Blood ◽  
2010 ◽  
Vol 115 (10) ◽  
pp. 2095-2104 ◽  
Author(s):  
Eran Ophir ◽  
Yaki Eidelstein ◽  
Ran Afik ◽  
Esther Bachar-Lustig ◽  
Yair Reisner
Keyword(s):  
Bone Marrow ◽  
T Cells ◽  
T Cell ◽  
Ex Vivo ◽  
Memory Cell ◽  
Central Memory ◽  
Third Party ◽  
High Survival

Abstract Enabling engraftment of allogeneic T cell–depleted bone marrow (TDBM) under reduced-intensity conditioning represents a major challenge in bone marrow transplantation (BMT). Anti–third-party cytotoxic T lymphocytes (CTLs) were previously shown to be endowed with marked ability to delete host antidonor T cells in vitro, but were found to be less effective in vivo. This could result from diminished lymph node (LN) homing caused by the prolonged activation, which induces a CD44+CD62L− effector phenotype, and thereby prevents effective colocalization with, and neutralization of, alloreactive host T cells (HTCs). In the present study, LN homing, determined by imaging, was enhanced upon culture conditions that favor the acquisition of CD44+CD62L+ central memory cell (Tcm) phenotype by anti–third-party CD8+ cells. These Tcm-like cells displayed strong proliferation and prolonged persistence in BM transplant recipients. Importantly, adoptively transferred HTCs bearing a transgenic T-cell receptor (TCR) with antidonor specificity were efficiently deleted only by donor-type Tcms. All these attributes were found to be associated with improved efficacy in overcoming T cell–mediated rejection of TDBM, thereby enabling high survival rate and long-term donor chimerism, without causing graft-versus-host disease. In conclusion, anti–third-party Tcms, which home to recipient LNs and effectively delete antidonor T cells, could provide an effective and novel tool for overcoming rejection of BM allografts.

scholarly journals IL-27 confers a protumorigenic activity of regulatory T cells via CD39

2019 ◽  
Vol 116 (8) ◽  
pp. 3106-3111 ◽  
Author(s):  
Young-Jun Park ◽  
Heeju Ryu ◽  
Garam Choi ◽  
Byung-Seok Kim ◽  
Eun Sook Hwang ◽  
...  
Keyword(s):  
Bone Marrow ◽  
T Cells ◽  
Regulatory T Cells ◽  
Antitumor Immunity ◽  
Suppressive Activity ◽  
Independent Manner ◽  
Bone Marrow Chimera ◽  
Cell Responses

Expression of ectonucleotidase CD39 contributes to the suppressive activity of Foxp3+ regulatory T cells (Tregs) by hydrolyzing immunogenic ATP into AMP. The molecular mechanism that drives CD39 expression on Tregs remains elusive. We found that tumor-infiltrating Tregs (Ti-Tregs) failed to up-regulate CD39 in mice lacking EBI3 subunit of IL-27 or IL-27Ra. Mixed bone marrow chimera and in vitro studies showed that IL-27 signaling in Tregs directly drives CD39 expression on Ti-Tregs in a STAT1-dependent, but STAT3- and T-bet–independent, manner. Tregs stimulated with IL-27 showed enhanced suppressive activities against CD8+ T cell responses in vitro. Moreover, IL-27Ra–deficient Tregs and STAT1-deficient Tregs were less efficient than WT Tregs in suppressing antitumor immunity in vivo. CD39 inhibition significantly abolished IL-27–induced suppressive activities of Tregs. Thus, IL-27 signaling in Tregs critically contributes to protumorigenic properties of Tregs via up-regulation of CD39.

scholarly journals Pre-Clinical Trial to Evaluate the Efficacy of Delayed Administration of Ixazomib in the Prophylaxis of Chronic Graft-Versus-Host Disease

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 4521-4521 ◽  
Author(s):  
Teresa Ramos ◽  
Alfonso Rodríguez-Gil ◽  
Melanie Nufer ◽  
María Victoria Barbado ◽  
Estefania Garcia Guerrero ◽  
...  
Keyword(s):  
Bone Marrow ◽  
T Cells ◽  
T Lymphocytes ◽  
Graft Versus Host Disease ◽  
Activation Markers ◽  
Graft Versus Host ◽  
Post Transplant ◽  
Activated T Lymphocytes

Abstract Introduction: Although survival rates have improved over the years, chronic graft-versus-host disease (cGvHD) remains the most frequent and severe complication in the long term after allogeneic hematopoietic stem cell transplantation (allo-HSCT). All the strategies developed to reduce its incidence are based on procedures aimed to decrease the risk of acute GvHD that, consequently, can also reduce the risk of cGvHD, mainly using immunosuppression in the early post-transplant period. These strategies induce apoptosis of donor T lymphocytes, responsible for a/cGvHD but also for graft-versus-tumor response. In the present project, we have evaluated a new strategy aimed to reduce the risk of cGvHD by manipulating the immune response not in the early post-transplant period, but in later phases. cGvHD develops via a complex cellular and molecular network involving thymus damage and unusual antigen presentation leading to aberrant T- and B-cell reactions characterized by Th17/Tc17 differentiation, macrophage sequestration in tissue, alloantibody formation, and fibrosis. Most of these cell populations are dependent on NF-kB for their activation. Ixazomib is a second-generation proteasome inhibitor for oral administration, representing an ideal candidate for prophylaxis in GvHD. Objective: We propose to develop a murine model of cGVHD and progressive onset cGvHD to test the efficacy of delayed administration of ixazomib as a novel strategy to decrease the risk of cGvHD. Methods: For in vitro studies, peripheral blood mononuclear cells from healthy donors were stimulated in the presence of different concentrations of ixazomib. After 48h and 120h, apoptosis was analyzed, and activation markers were evaluated. For in vivo studies, a murine model of progressive cGvHD (pcGvHD - allowing the recipient to survive to mild aGvHD, thus favoring autoreactive T cells to expand and cause cGvHD) and a scleroderma model of cGvHD were used. Ixazomib at 0.75mg/Kg/twice weekly, 2X from day +21 in the pcGvHD, with or without cyclosporine A (CyA) at 5mg/Kg/day from day 0 until the end of the study. For scleroderma cGvHD group, 3mg/Kg/2X from day +30 was used up to 120 days post-transplant. Flow cytometry was used to evaluate the different lymphocyte populations in the different target organs of the GvHD. Results: In vitro results showed that activated T lymphocytes are sensitive to the proapoptotic effect of ixazomib (>100nM), while high concentrations of the drug are necessary to cause apoptosis in the non-activated cells (5000nM). Through CD27 and CD45 expression, we verified that ixazomib has a proapoptotic effect mainly on naïve and effector cells. We also verified a decrease in the activation markers of CD3+CD25+INF-γ+ cells (p˂0.01, 1000nM, n = 4). In the animal trial, the survival of the pcGvHD model mice treated with ixazomib was significantly higher as compared to untreated mice, with a significant decrease in the signs of pcGvHD (Figure 1A). In addition, the combination of CyA and ixazomib improved survival as compared to those mice receiving CyA or ixazomib alone as well as untreated controls (Figure 1B). In the scleroderma cGvHD model, we observed that the animals treated with ixazomib showed significantly less signs of GvHD (p˂0.0001) (Figure 2). Multiparametric flow cytometer analyses showed that in the pcGvHD model, mice treated with ixazomib had a decrease of effector CD4 T-cells in bone marrow (p=0.06) and spleen (p=0.015) when compared to untreated mice. Also an increase of CD19+ cells in the colon (p=0.04), liver (p=0.035), lymph nodes (LN) (p=0.037) and lungs (p=0.03) was observed. Regarding the regulatory T cells (Foxp3+), the treated mice had a significant increase in LN (p=0.02), Peyer patches (p=0.015) and thymus (p=0.028) as compared to untreated mice. Conclusion: In vitro studies show that ixazomib induces apoptosis on activated T lymphocytes and decreases the expression of activation markers. Our in vivo model indicates that the combination of CyA and ixazomib for the prevention of pcGvHD and cGVHD after allogeneic HSCT is promising and merits further investigation in clinical trials. Figure 1. Survival of pcGvHD. Figure 2. Score graphic of scleroderma cGvHD. BM - Bone marrow, mice that were transplanted with BM from BALBc mice (Syngeneic). Disclosures Ramos: Takeda Oncology: Research Funding.

Fucosylated Antigen-Specific Cytotoxic T Lymphocytes Demonstrate Enhanced Killing of Leukemia Targets

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 1888-1888
Author(s):  
Gheath Alatrash ◽  
Mao Zhang ◽  
Na Qiao ◽  
Pariya Sukhumalchandra ◽  
Madhushree Zope ◽  
...  
Keyword(s):  
Bone Marrow ◽  
T Cells ◽  
T Lymphocytes ◽  
Cell Surface ◽  
Adhesion Molecules ◽  
Cytotoxic T Lymphocytes ◽  
Cell Surface Expression ◽  
Target Cells

Abstract Introduction Immunotherapy using cytotoxic T lymphocytes (CTL) has shown efficacy in the management of leukemia. However the efficacy of CTL, whether they are engineered and adoptively transferred or administered as part of allogeneic stem cell transplantation, must be balanced by their off-target toxicities, which at times can be lethal. Fucosylation, which is mediated by fucosyl transferases, is a process by which fucose sugar groups are added to cell surface receptors. Fucosylated T cells have been shown to preferentially home to inflamed tissues, including bone marrow. In view of recent data showing that fucosylation with fucosyltransferase (FT)-VI facilitates homing of regulatory T cells (T-regs) to inflamed tissues and cord blood engraftment into the bone marrow, we hypothesized that fucosylation could enhance the efficacy of CTL that target leukemia antigens. In this study, we tested whether ex vivo fucosylation of CTL that target the HLA-A2 restricted leukemia peptides, CG1 (derived from cathepsin G) and PR1 (derived from neutrophil elastase and proteinase 3), with the novel enzyme FT-VII enhances their migration and anti-leukemia functions. Experimental design CG1- and PR1-CTL were generated using standard methodologies. Fucosylation was achieved by incubating T cells with FTVII enzyme and GDP fucose (Targazyme). To study migration, fucosylated and non-fucosylated CTL were passed through chambers coated with a HUVEC barrier and migrated CTL were detected using cell fluorescence. To examine CTL surface markers, cells were stained for standard co-stimulatory and adhesion molecules and were analyzed using flow cytometry. Calcein AM cytotoxicity assays were used to determine the effects of fucosylation on CTL killing of target cells. In vitro effects of fucosylation on leukemia-CTL specificity was accomplished using standard CFU assays. For in vivo assessment of fucosylation on activity of CTL, NSG mice were engrafted with U937-A2 human acute myeloid leukemia (AML) cells or primary AML and were treated with intravenous injections of 5.0 x 105 fucosylated or non-fucosylated CTL. Mice were followed twice weekly and were sacrificed for bone marrow and tissue analysis at prespecified time points or when they became moribund. Results Fucosylated CG1-CTL and PR1-CTL showed approximately 2-fold higher migration through the HUVEC cell barrier compared to non-fucosylated CTL. Analysis of T cell surface expression of chemokine/adhesion molecules showed an approximately a 5-fold increase in CD49d and CD195, and a 50% increase in CXCR1 and CXCR3 following fucosylation. Fucosylation enhanced the cytotoxicity of leukemia specific-CTL against primary HLA-A2+ leukemia and HLA-A2+ U937 cells at increasing effector to target ratios. For primary patient AML, we show enhanced leukemia killing by fucosylated-PR1-CTL in comparison with non-fucosylated-PR1-CTL at the 20:1 effector to target (E:T) ratio (25-fold higher killing ) and the 10:1 E:T ratio (4-fold higher killing). Similar results were seen using the U937-A2 AML cell line favoring fucosylated-CG1-CTL: 20-fold higher killing at 20:1 E:T ratio and a 9-fold higher killing at the 10:1 E:T ratio. In vitro CFU assays using HLA-A2+ healthy donor bone marrow showed no change in the specificity of the antigen specific CTL following fucosylation. Specifically we show 283 and 295 colonies in the fucosylated and non-fucosylated CG1-CTL groups, respectively (P >0.05). These were also compared to irrelevant peptide HIV-CTL, which demonstrated 286 and 269 CFUs in the fucosylated and non-fucosylated HIV-CTL groups, respectively (P >0.05). In vivo experiments using CG1-CTL against primary AML showed 5-fold higher killing of AML by fucosylated CTL vs. non-fucosylated CTL. Similar results were also seen using U937-A2 AML targets. Conclusion Fucosylation with FT-VII enhances the efficacy of leukemia-targeting CTL against primary human AML and AML cell lines. These data demonstrate a novel approach to enhance the efficacy of antigen specific CTL that could be used in adoptive cellular immunotherapy approaches for leukemia. Disclosures No relevant conflicts of interest to declare.

scholarly journals Interleukin 10 Gene-Modified Bone Marrow-Derived Dendritic Cells Attenuate Liver Fibrosis in Mice by Inducing Regulatory T Cells and Inhibiting the TGF-β/Smad Signaling Pathway

2019 ◽  
Vol 2019 ◽  
pp. 1-15 ◽  
Author(s):  
Yejin Xu ◽  
Xinyue Tang ◽  
Min Yang ◽  
Shengguo Zhang ◽  
Shanshan Li ◽  
...  
Keyword(s):  
Bone Marrow ◽  
T Cells ◽  
Dendritic Cells ◽  
Liver Fibrosis ◽  
Regulatory T Cells ◽  
Interleukin 10 ◽  
Smad Signaling Pathway ◽  
Culture Supernatants

Aim. To explore the therapeutic effects and mechanisms of interleukin 10 gene-modified bone marrow-derived dendritic cells (DC-IL10) on liver fibrosis. Methods. In vitro, BMDCs were transfected with lentiviral-interleukin 10-GFP (LV-IL10-GFP) at the MOI of 1 : 40. Then, the phenotype (MHCII, CD80, and CD86) and allo-stimulatory ability of DC-IL10 were identified by flow cytometry, and the levels of IL-10 and IL-12 (p70) secreted into the culture supernatants were quantified by ELISA. In vivo, DC-IL10 was injected into mice with CCl4-induced liver fibrosis through the tail vein. Lymphocytes were isolated to investigate the differentiation of T cells, and serum and liver tissue were collected for biochemical, cytokine, histopathologic, immune-histochemical, and Western blot analyzes. Results. In vitro, the expressions of MHCII, CD80, and CD86 in DC-IL10 were significantly suppressed, allogeneic CD4+T cells incubated with DC-IL10 showed a lower proliferative response, and the levels of IL-10 and IL-12 (p70) secreted into the DC-IL10 culture supernatants were significantly increased and decreased, respectively. In vivo, regulatory T cells (Tregs) were significantly increased, while ALT, AST, and inflammatory cytokines were significantly reduced in the DC-IL10 treatment group, and the degree of hepatic fibrosis was obviously reversed. The TGF-β/smad pathway was inhibited following DC-IL10 treatment compared to the liver fibrosis group. Conclusion. IL-10 genetic modification of BMDCs may maintain DC in the state of tolerance and allow DC to induce T cell hyporesponsiveness or tolerance. DC-IL10 suppressed liver fibrosis by inducing Treg production and inhibiting the TGF-β/smad signaling pathway.

CD8+/TCR− Facilitating Cells Induce Generation of Regulatory T cells in Vivo: Chimeric Regulatory T Cells are Antigen-Specific and Enhance Engraftment of Allogeneic HSC in Non-Obese Diabetic (NOD) Mice.

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 2318-2318
Author(s):  
Yiming Huang ◽  
Larry D Bozulic ◽  
Thomas Miller ◽  
Hong Xu ◽  
Lala-Rukh Hussain ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Cell Proliferation ◽  
T Cells ◽  
Regulatory T Cells ◽  
Nod Mice ◽  
Strong Suppression ◽  
Hematopoietic Stem ◽  
Cell Based Therapy

Abstract CD8+/TCR− graft facilitating cells (FC) are a novel tolerogenic cell population in bone marrow that potently enhance engraftment of hematopoietic stem cells (HSC) in allogeneic and syngeneic recipients. The CD11c+/CD11b−/B220+ plasmacytoid precursor dendritic cell (p-preDC) subpopulation of FC (p-preDC FC) comprises over 60% of FC total and plays a critical and nonredundant role in facilitation. FC prevent graft-versus-host disease and remain tolerogenic after in vivo infusion. Regulatory T cells (Treg) are immunomodulatory cells that maintain tolerance in vivo. They can be generated in vitro via co-culture with p-preDC. There is great interest regarding the use of Treg as a cell-based therapy to induce graft/host tolerance in vivo. However, a major challenge to the clinical use of Treg has been to obtain sufficient numbers of cells for in vivo use and maintain their tolerogenic properties in vivo after in vitro expansion. Here, we evaluated whether FC function by inducing the production of Tregin vivo and examined the function of these chimeric Tregin vivo and in vitro. HSC (c-Kit+Sca-1+Lin−; KSL) were sorted from donor B6 and NOD mice. 10,000 B6 HSC and 1,000 NOD HSC were transplanted by tail-vein injection into recipient NOD mice conditioned with 950 cGy of total body irradiation (TBI). Spleen, thymus, and bone marrow were harvested from recipient NOD mice 5 weeks after transplantation. CD4+CD25+Foxp3+ Treg were analyzed by flow cytometry. FC induced the generation of both donor and recipient CD4+CD25+Foxp3+ Tregin vivo; the majority of Treg were recipient-derived (89% to 97%). To test the function of Treg from HSC + FC chimeras (chimeric Treg), CD8− CD4+CD25+ Treg were sorted from the spleen of chimeras 5 weeks after transplantation. 50,000 chimeric Treg plus 10,000 B6 HSC were transplanted into NOD recipients conditioned with 950 cGy TBI. Recipients of 50,000 Treg from naïve B6 spleens (B6 Treg) + HSC or HSC alone served as controls. Five of 26 recipients of HSC alone engrafted and survived up to 100 days. Only 2 of 5 recipients of HSC plus 50,000 B6 Treg engrafted and none of the recipients exhibited durable engraftment beyond 100 days. In striking contrast, 100% (4 of 4) recipients of HSC + 50,000 chimeric Treg engrafted durably, with survival ≥ 100 days. Chimeric Treg function was confirmed in vitro by MLR suppressor assays, as evidenced by strong suppression of T cell proliferation. Sorted chimeric Treg demonstrated an 87.2% suppression of cell proliferation when plated in a 1:1 ratio with naïve NOD responder cells and B6 stimulator cells. Moreover, when plated at a 1:4 and 1:8 ratio with naïve NOD responders, Treg suppressive function titrated to 62.7% and 43.3%, respectively. In contrast, sorted Treg from naïve B6 animals showed 75.8%, 35.4, and 29.4% suppression when plated in ratios of 1:1, 1:4, and 1:8, respectively. Taken together, these data suggest that FC induce the production of antigen-specific Tregin vivo and chimeric Treg are superior to naïve Treg in suppressing the proliferation of effector T cells and potently enhance engraftment of allogeneic HSC.

scholarly journals In Vitro–expanded Antigen-specific Regulatory T Cells Suppress Autoimmune Diabetes

2004 ◽  
Vol 199 (11) ◽  
pp. 1455-1465 ◽  
Author(s):  
Qizhi Tang ◽  
Kammi J. Henriksen ◽  
Mingying Bi ◽  
Erik B. Finger ◽  
Greg Szot ◽  
...  
Keyword(s):  
T Cells ◽  
Regulatory T Cells ◽  
Autoimmune Diabetes ◽  
Interleukin 2 ◽  
Disease Onset ◽  
Cellular Immunotherapy ◽  
Antigen Specificity ◽  
Cell Functions

The low number of CD4+ CD25+ regulatory T cells (Tregs), their anergic phenotype, and diverse antigen specificity present major challenges to harnessing this potent tolerogenic population to treat autoimmunity and transplant rejection. In this study, we describe a robust method to expand antigen-specific Tregs from autoimmune-prone nonobese diabetic mice. Purified CD4+ CD25+ Tregs were expanded up to 200-fold in less than 2 wk in vitro using a combination of anti-CD3, anti-CD28, and interleukin 2. The expanded Tregs express a classical cell surface phenotype and function both in vitro and in vivo to suppress effector T cell functions. Most significantly, small numbers of antigen-specific Tregs can reverse diabetes after disease onset, suggesting a novel approach to cellular immunotherapy for autoimmunity.

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