RESUMEN
Myeloid-derived suppressor cells (MDSCs) are markedly increased in cancer patients and tumor-bearing mice and promote tumor growth and survival by inhibiting host innate and adaptive immunity. In this study, we generated and characterized MDSCs from murine-induced pluripotent stem cells (iPSCs). The iPSCs were co-cultured with OP9 cells, stimulated with GM-CSF, and became morphologically heterologous under co-culturing with hepatic stellate cells. Allogeneic and OVA-specific antigen stimulation demonstrated that iPS-MDSCs have a T-cell regulatory function. Furthermore, a popliteal lymph node assay and autoimmune hepatitis model showed that iPS-MDSCs also regulate immune responsiveness in vivo and have a therapeutic effect against hepatitis. Taken together, our results demonstrated a method of generating functional MDSCs from iPSCs and highlighted the potential of iPS-MDSCs as a key cell therapy resource for transplantation and autoimmune diseases.
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Linfocitos T CD8-positivos/metabolismo , Células Madre Pluripotentes Inducidas/metabolismo , Células Supresoras de Origen Mieloide/metabolismo , Animales , Diferenciación Celular , Línea Celular , Proliferación Celular , Humanos , Ratones , Ratones Endogámicos C57BLRESUMEN
BACKGROUND: Islet transplantation is a promising therapeutic approach to restore the physical response to blood glucose in type 1 diabetes. Current chronic use of immunosuppressive reagents for preventing islet allograft rejection is associated with severe complications. In addition, many of the immunosuppressive drugs are diabetogenic. The induction of transplant tolerance to eliminate the dependency on immunosuppression is ideal, but remains challenging. METHODS: Addition of hepatic stellate cells allowed generation of myeloid-derived suppressor cells (MDSC) from precursors in mouse bone marrow. Migration of MDSC was examined in an islet allograft transplant model by tracking the systemic administered MDSC from CD45.1 congenic mice. RESULTS: The generated MDSC were expressed C-C chemokine receptor type 2 (CCR2), which was enhanced by exposure to interferon-γ. A single systemic administration of MDSC markedly prolonged survival of islet allografts without requirement of immunosuppression. Tracking the administered MDSC showed that they promptly migrated to the islet graft sites, at which point they exerted potent immune suppressive activity by inhibiting CD8 T cells, enhancing regulatory T cell activity. MDSC generated from CCR2 mice failed to be mobilized and lost tolerogenic activity in vivo, but sustained suppressive activity in vitro. CONCLUSIONS: MDSC migration was dependent on expression of CCR2, whereas CCR2 does not directly participate in immune suppression. Expression of CCR2 needs to be closely monitored for quality control purpose when MDSC are generated in vitro for immune therapy.
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Rechazo de Injerto/inmunología , Tolerancia Inmunológica , Trasplante de Islotes Pancreáticos/inmunología , Células Supresoras de Origen Mieloide/inmunología , Receptores CCR2/biosíntesis , Tolerancia al Trasplante/inmunología , Animales , Diferenciación Celular , Modelos Animales de Enfermedad , Regulación de la Expresión Génica , Rechazo de Injerto/metabolismo , Rechazo de Injerto/prevención & control , Masculino , Ratones , Ratones Endogámicos C57BL , Células Supresoras de Origen Mieloide/metabolismo , Reacción en Cadena de la Polimerasa , ARN/genética , Receptores CCR2/genética , Trasplante HomólogoRESUMEN
PURPOSE OF REVIEW: To highlight the results of the ongoing research on the mechanisms of liver-induced tolerance focusing on results from the last year. RECENT FINDINGS: The liver is exposed to a massive antigenic burden of dietary and commensal products from the gastrointestinal tract via portal vein, most of which are necessary for survival. To prevent the immune system from destroying these foreign yet beneficial elements, the liver has developed unique mechanisms to suppress immune responses. It is thought that these mechanisms of acquired tolerance may also underlie the spontaneous acceptance of liver allografts observed after transplantation in many species. The fact that isolated hepatocyte transplants are acutely rejected, suggests that nonparenchymal liver cells play a critical role in spontaneous liver allograft acceptance. IFN-γ, a key inflammatory cytokine produced by T effector (Tef) cells, is paradoxically compulsory for spontaneous liver allograft acceptance. Analysis of IFN-γ signaling points to liver mesenchymal nonparenchymal liver cell that eliminate infiltrating Tef cells via expression of B7-H1, IL-10, and tumor growth factor-ß, as well as the enhancement of Tregs and MDSCs. Thus, liver mesenchymal cells are thought to promote tolerance by eliminating alloreactive Tef cells and enhancing suppressor cells (T and B). SUMMARY: The research during last year offered some key insights into the mechanisms of liver-induced tolerance. Through interactions with activated T cells and B cells via IFN-γ/B7-H1 pathways, liver mesenchymal cells have been shown to be critical components of liver-specific tolerance induction.
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Tolerancia Inmunológica/inmunología , Trasplante de Hígado/métodos , Hígado/inmunología , Humanos , Hígado/patologíaRESUMEN
We demonstrated previously that mouse hepatic stellate cells (HSCs) suppress T cells via programmed death-ligand 1 (PD-L1), but it remains unknown whether they exert any effects on B cells, the other component of the adaptive immune system. In this study, we found that mouse HSCs directly inhibited B cells and that PD-L1 was also integrally involved. We found that HSCs inhibited the upregulation of activation markers on activated B cells, as well as the proliferation of activated B cells and their cytokine/Ig production in vitro, and that pharmaceutically or genetically blocking the interaction of PD-L1 with programmed cell death protein 1 impaired the ability of HSCs to inhibit B cells. To test the newly discovered B cell-inhibitory activity of HSCs in vivo, we developed a protocol of intrasplenic artery injection to directly deliver HSCs into the spleen. We found that local delivery of wild-type HSCs into the spleens of mice that had been immunized with 4-hydroxy-3-nitrophenylacetyl-Ficoll, a T cell-independent Ag, significantly suppressed Ag-specific IgM and IgG production in vivo, whereas splenic artery delivery of PD-L1-deficient HSCs failed to do so. In conclusion, in addition to inhibiting T cells, mouse HSCs concurrently inhibit B cells via PD-L1. This direct B cell-inhibitory activity of HSCs should contribute to the mechanism by which HSCs maintain the liver's immune homeostasis.
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Linfocitos B/inmunología , Antígeno B7-H1/inmunología , Antígeno B7-H1/metabolismo , Células Estrelladas Hepáticas/inmunología , Animales , Ficoll/análogos & derivados , Ficoll/inmunología , Homeostasis , Inmunoglobulina G/sangre , Inmunoglobulina M/sangre , Hígado/inmunología , Activación de Linfocitos , Ratones Endogámicos C57BL , Ratones Noqueados , Receptor de Muerte Celular Programada 1/genética , Bazo/inmunologíaRESUMEN
The liver, which is a metabolic organ, plays a pivotal role in tolerance induction. Hepatic stellate cells (HpSCs), which are unique non-parenchymal cells, exert potent immunoregulatory activity during cotransplantation with allogeneic islets effectively protecting the islet allografts from rejection. Multiple mechanisms participate in the immune tolerance induced by HpSCs, including the marked expansion of myeloid-derived suppressor cells (MDSCs), attenuation of effector T cell functions and augmentation of regulatory T cells. HpSC conditioned MDSC-based immunotherapy has been conducted in mice with autoimmune disease and the results show that this technique may be promising. This article demonstrates how HpSCs orchestrate both innate immunity and adaptive immunity to build a negative network that leads to immune tolerance.
Asunto(s)
Células Estrelladas Hepáticas/inmunología , Tolerancia Inmunológica , Hígado/inmunología , Inmunidad Adaptativa , Traslado Adoptivo/métodos , Animales , Comunicación Celular , Rechazo de Injerto/inmunología , Rechazo de Injerto/prevención & control , Supervivencia de Injerto , Células Estrelladas Hepáticas/metabolismo , Células Estrelladas Hepáticas/trasplante , Humanos , Inmunidad Innata , Inmunosupresores/uso terapéutico , Hígado/metabolismo , Trasplante de Órganos/efectos adversos , Linfocitos T/inmunologíaRESUMEN
Hepatic stellate cells (HSCs) inhibit T cells, a process that could help the liver to maintain its immunoprivileged status. HSCs secrete latent TGF-ß1, but the detailed mechanisms by which latent TGF-ß1 is activated and whether it plays any role in HSC-mediated T cell suppression remain unclear. Glycoprotein A repetitions predominant (GARP) is a surface marker of activated regulatory T cells. GARP binds latent TGF-ß1 for its activation, which is critical for regulatory T cells to suppress effector T cells; however, it is still unclear whether GARP is present on HSCs and whether it has any impact on HSC function. In this study, we found that TGF-ß1(+/-) HSCs, which produce reduced levels of TGF-ß1, showed decreased potency in inhibiting T cells. We also found that pharmaceutical or genetic inhibition of the TGF-ß1 signaling pathway reduced the T cell-inhibiting activity of HSCs. Additionally, using isolated primary HSCs, we demonstrated that GARP was constitutively expressed on HSCs. Blocking GARP function or knocking down GARP expression significantly impaired the potency of HSCs to suppress the proliferation of and IFN-γ production from activated T cells, suggesting that GARP is important for HSCs to inhibit T cells. These results demonstrate the unexpected presence of GARP on HSCs and its significance in regard to the ability of HSCs to activate latent TGF-ß1 and thereby inhibit T cells. Our study reveals a new mechanism for HSC-mediated immune regulation and potentially for other conditions, such as liver fibrosis, that involve HSC-secreted TGF-ß1.
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Células Estrelladas Hepáticas/inmunología , Proteínas de la Membrana/metabolismo , Linfocitos T Reguladores/inmunología , Factor de Crecimiento Transformador beta1/metabolismo , Animales , Diferenciación Celular/inmunología , Proliferación Celular , Células Cultivadas , Activación Enzimática , Humanos , Inflamación/inmunología , Interferón gamma/biosíntesis , Hígado/inmunología , Cirrosis Hepática/inmunología , Cirrosis Hepática/patología , Activación de Linfocitos/inmunología , Proteínas de la Membrana/biosíntesis , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Unión Proteica/inmunología , Interferencia de ARN , ARN Interferente Pequeño , Transducción de Señal/inmunologíaRESUMEN
UNLABELLED: Liver tolerance was initially recognized by the spontaneous acceptance of liver allografts in many species. The underlying mechanisms are not completely understood. However, liver transplant (LT) tolerance absolutely requires interferon (IFN)-γ, a rejection-associated inflammatory cytokine. In this study, we investigated the rejection of liver allografts deficient in the IFN-γ receptor and reveal that the liver graft is equipped with machineries capable of counterattacking the host immune response through a mesenchyme-mediated immune control (MMIC) mechanism. MMIC is triggered by T effector (Tef) cell-derived IFN-γ that drives expression of B7-H1 on graft mesenchymal cells leading to Tef cell apoptosis. We describe the negative feedback loop between graft mesenchymal and Tef cells that ultimately results in LT tolerance. Comparable elevations of T-regulatory cells and myeloid-derived suppressor cells were observed in both rejection and tolerance groups and were not dependent on IFN-γ stimulation, suggesting a critical role of Tef cell elimination in tolerance induction. We identify potent MMIC activity in hepatic stellate cells and liver sinusoidal endothelial cells. MMIC is unlikely exclusive to the liver, given that spontaneous acceptance of kidney allografts has been reported, although less commonly, probably reflecting variance in MMIC activity. CONCLUSION: MMIC may represent an important homeostatic mechanism that supports peripheral tolerance and could be a target for the prevention and treatment of transplant rejection. This study highlights that the graft is an active participant in the equipoise between tolerance and rejection and warrants more attention in the search for tolerance biomarkers.
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Tolerancia Inmunológica/inmunología , Mesodermo/inmunología , Receptores de Interferón/inmunología , Inmunología del Trasplante/fisiología , Animales , Citocinas/inmunología , Modelos Animales de Enfermedad , Rechazo de Injerto/inmunología , Supervivencia de Injerto/inmunología , Factores Inmunológicos/metabolismo , Trasplante de Hígado/efectos adversos , Masculino , Mesodermo/metabolismo , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Transgénicos , Receptores de Interferón/metabolismo , Sensibilidad y Especificidad , Linfocitos T Reguladores/inmunología , Linfocitos T Reguladores/metabolismo , Trasplante Homólogo , Receptor de Interferón gammaRESUMEN
A major complication of factor VIII (F.VIII) infusion therapies for the treatment of hemophilia A is the formation of antibodies (inhibitors) against F.VIII, a T-cell-dependent, B-cell-mediated process. To date, attempts to inhibit formation of the inhibitors have been limited in success. We have shown that hepatic stellate cells (HSCs) promote the development of myeloid-derived suppressor cells (MDSCs). The HSC-induced MDSCs are potent regulators of T-cell and B-cell responses. Here we show that MDSCs can be propagated from hemophilia A mouse bone marrow cells in coculture with HSCs. These cells exhibit a suppressive phenotype and display a marked ability to inhibit T-cell proliferation induced by dendritic cells in response to F.VIII. MDSCs can also inhibit proliferation and activation of B cells stimulated by immunoglobulin M and interleukin 4. Administration of HSC-induced MDSCs induces CD4(+) T cell and B220(+) B-cell hyporesponsiveness to F.VIII and reduces inhibitor formation in hemophilia A mice. These results suggest that MDSCs could serve as a form of immunotherapy for preventing inhibitor formation via induction of immune tolerance.
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Autoanticuerpos/biosíntesis , Factor VIII/inmunología , Hemofilia A/inmunología , Células Estrelladas Hepáticas/citología , Células Mieloides/citología , Animales , Secuencia de Bases , Cartilla de ADN , Ratones , Ratones Endogámicos C57BL , Reacción en Cadena de la PolimerasaRESUMEN
We recently demonstrated that hepatic stellate cells induce the differentiation of myeloid-derived suppressor cells (MDSCs) from myeloid progenitors. In this study, we found that adoptive transfer of these MDSCs effectively reversed disease progression in experimental autoimmune myasthenia gravis (EAMG), a T cell-dependent and B cell-mediated model for myasthenia gravis. In addition to ameliorated disease severity, MDSC-treated EAMG mice showed suppressed acetylcholine receptor (AChR)-specific T cell responses, decreased levels of serum anti-AChR IgGs, and reduced complement activation at the neuromuscular junctions. Incubating MDSCs with B cells activated by anti-IgM or anti-CD40 Abs inhibited the proliferation of these in vitro-activated B cells. Administering MDSCs into mice immunized with a T cell-independent Ag inhibited the Ag-specific Ab production in vivo. MDSCs directly inhibit B cells through multiple mechanisms, including PGE2, inducible NO synthase, and arginase. Interestingly, MDSC treatment in EAMG mice does not appear to significantly inhibit their immune response to a nonrelevant Ag, OVA. These results demonstrated that hepatic stellate cell-induced MDSCs concurrently suppress both T and B cell autoimmunity, leading to effective treatment of established EAMG, and that the MDSCs inhibit AChR-specific immune responses at least partially in an Ag-specific manner. These data suggest that MDSCs could be further developed as a novel approach to treating myasthenia gravis and, even more broadly, other diseases in which T and B cells are involved in pathogenesis.
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Linfocitos B/inmunología , Miastenia Gravis Autoinmune Experimental , Células Mieloides , Linfocitos T/inmunología , Animales , Autoanticuerpos/inmunología , Linfocitos B/patología , Dinoprostona/inmunología , Células Estrelladas Hepáticas/inmunología , Células Estrelladas Hepáticas/patología , Inmunoglobulina G/inmunología , Inmunoglobulina M/inmunología , Ratones , Miastenia Gravis Autoinmune Experimental/inmunología , Miastenia Gravis Autoinmune Experimental/patología , Miastenia Gravis Autoinmune Experimental/terapia , Células Mieloides/inmunología , Células Mieloides/patología , Células Mieloides/trasplante , Receptores Nicotínicos/inmunología , Linfocitos T/patologíaRESUMEN
Hepatic stellate cells (HSC) are a major source of the immunoregulatory metabolite all-trans retinoic acid (ATRA), which may contribute to the generation of tolerogenic dendritic cells (DCs) in the liver. The present study seeks to clarify the mechanism(s) through which ATRA promotes the development of tolerogenic DCs. Although bone marrow-derived ATRA-treated DCs (RA-DCs) and conventional DCs had comparable surface phenotype, RA-DCs had diminished stimulatory capacity and could directly inhibit the expansion of DC/OVA-stimulated OT-II T cells. Arginase-1 (Arg-1) was found promote suppression because 1) ATRA was a potent inducer of Arg-1 protein and activity, 2) the Arg-1 inhibitor N(w)-hydroxy nor-l-arginine partially reversed suppression, and 3) the suppressive function of RA-DCs was partially compromised using OT-II T cells from GCN2(-/-) mice, which are insensitive to Arg-1. Inducible NO synthase (iNOS), however, was found to be a more significant contributor to RA-DC function because 1) ATRA potentiated the expression of IFN-γ-induced iNOS, 2) suppressive function in RA-DCs was blocked by the iNOS inhibitor N(G)-monomethyl-l-arginine, monoacetate salt, and 3) RA-DCs derived from iNOS(-/-) mice exhibited near complete loss of tolerogenic function, despite sustained Arg-1 activity. The expression of iNOS and the suppressive function of RA-DCs were dependent on both IFN-γ and ATRA. Furthermore, the in vivo behavior of RA-DCs proved to be consistent with their in vitro behavior. Thus, we conclude that ATRA enhances both Arg-1 and iNOS expression in IFN-γ-treated DCs, resulting in a tolerogenic phenotype. These findings elucidate mechanisms through which ATRA may contribute to liver immune tolerance.
Asunto(s)
Antineoplásicos/farmacología , Arginasa/inmunología , Células de la Médula Ósea/inmunología , Células Dendríticas/inmunología , Tolerancia Inmunológica/efectos de los fármacos , Óxido Nítrico Sintasa de Tipo II/inmunología , Tretinoina/farmacología , Animales , Arginasa/genética , Regulación Enzimológica de la Expresión Génica/efectos de los fármacos , Regulación Enzimológica de la Expresión Génica/genética , Regulación Enzimológica de la Expresión Génica/inmunología , Tolerancia Inmunológica/genética , Tolerancia Inmunológica/inmunología , Interferón gamma/genética , Interferón gamma/inmunología , Ratones , Ratones Noqueados , Óxido Nítrico Sintasa de Tipo II/genética , Linfocitos T/inmunologíaRESUMEN
BACKGROUND: Islet transplantation is an alternative to pancreas transplantation to cure type 1 diabetes, but both require chronic immunosuppression, which is often accompanied by deleterious side effects. The purpose of this study was to explore prolongation of islet allograft survival by cotransplantation with myeloid-derived suppressor cells (MDSCs) without requirement of immunosuppression and determine the role of inducible nitric oxide synthase (iNOS) produced by MDSCs in immune regulation. METHODS: Bone marrow cells were isolated from wild-type (WT) or iNOS mice and cultured in the presence of granulocyte-macrophage colony-stimulating factor and hepatic stellate cells (HSCs), resulting in the generation of MDSCs. WT or iNOS MDSCs were cotransplanted with islet allografts under the renal capsule of diabetic recipient mice. RESULTS: Addition of HSCs into DC culture promoted generation of MDSCs (instead of DCs). MDSCs had elevated expression of iNOS upon exposure to IFN-γ and inhibited T-cell responses in an MLR culture. Cotransplantation with WT MDSCs markedly prolonged survival of islet allografts, which was associated with reduced infiltration of CD8 T cells resulting from inhibited proliferative response. These effects were significantly attenuated when MDSCs were deficient in iNOS. Furthermore, iNOS MDSCs largely lost their ability to protect islet allografts. CONCLUSIONS: Cotransplantation with HSC-induced MDSCs significantly extends islet allograft survival through iNOS-mediated T-cell inhibition. The results demonstrate the potential use of in vitro generated MDSCs as a novel adjunctive immunotherapy for islet transplantation.
Asunto(s)
Trasplante de Islotes Pancreáticos , Células Mieloides/trasplante , Óxido Nítrico Sintasa de Tipo II/fisiología , Aloinjertos , Animales , Rechazo de Injerto , Supervivencia de Injerto , Inmunoterapia , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Linfocitos T/inmunologíaRESUMEN
Hepatic stellate cells (HSCs) interact with immune cells to actively participate in regulating immune response in the liver which is mediated by the effector molecules, including B7-H1. We demonstrated here that expression of B7-H1 on HSCs was markedly enhanced by interferon-(IFN-) γ stimulation. IFN- γ stimulated HSCs inhibited T-cell proliferation via induction of T-cell apoptosis (22.1% ± 1.6%). This immunosuppressive effect was inhibited by preincubation with an anti-B7-H1 antibody, or inhibitor of the MEK/ERK pathway inhibited IFN- γ mediated expression of B7-H1. Thus, regulation of B7-H1 expression on HSCs by IFN- γ represents an important mechanism that regulates immune responses in the liver favoring tolerogenicity rather than immunogenicity. Involvement of MEK/ERK pathway provides a novel target for therapeutic approaches.
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Células Estrelladas Hepáticas/efectos de los fármacos , Interferón gamma/farmacología , Proteína Quinasa 1 Activada por Mitógenos/inmunología , Proteína Quinasa 3 Activada por Mitógenos/inmunología , Transducción de Señal/efectos de los fármacos , Linfocitos T/efectos de los fármacos , Animales , Antracenos/farmacología , Anticuerpos Neutralizantes/farmacología , Apoptosis , Antígeno B7-H1/antagonistas & inhibidores , Antígeno B7-H1/genética , Antígeno B7-H1/inmunología , Proliferación Celular , Cromonas/farmacología , Regulación de la Expresión Génica/efectos de los fármacos , Células Estrelladas Hepáticas/citología , Células Estrelladas Hepáticas/inmunología , Tolerancia Inmunológica , Prueba de Cultivo Mixto de Linfocitos , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Proteína Quinasa 1 Activada por Mitógenos/antagonistas & inhibidores , Proteína Quinasa 1 Activada por Mitógenos/genética , Proteína Quinasa 3 Activada por Mitógenos/antagonistas & inhibidores , Proteína Quinasa 3 Activada por Mitógenos/genética , Morfolinas/farmacología , Transducción de Señal/inmunología , Linfocitos T/citología , Linfocitos T/inmunologíaRESUMEN
BACKGROUND: The liver is an immunologic privileged organ; liver allografts are accepted across major histocompatibility complex barriers in many species. However, hepatocyte transplants are acutely rejected, suggesting a role for liver nonparenchymal cells in regulating the immunoresponse. We have shown potent immunoregulatory activity of hepatic stellate cells (HSCs) in mice. The aim of this study was to examine the immunoregulatory activity of human HSCs. METHODS: HSCs were isolated from normal human livers for analyses of their impact on T-cell response. RESULTS: HSCs expressed low HLA-DR and costimulatory molecules CD40 and CD80 but constitutively expressed high levels of CD54. Interferon-γ stimulated HSCs to express B7-H1 in a dose-dependent manner and produce the suppressive cytokines interleukin-6, interleukin-10, and transforming growth factor-ß but did not affect expression of HLA-DR, CD40, and CD80. Human HSCs did not stimulate allogeneic T-cell proliferative response, indicating that they are not professional antigen-presenting cells. HSCs markedly inhibited T-cell response elicited by either allogeneic antigen-presenting cells or CD3/CD28 beads, which was associated with increases in activated CD4 and CD8 T-cell apoptosis. Addition of anti-B7-H1 blocking antibody significantly reversed the inhibitory effect. CONCLUSIONS: Human HSCs demonstrate potent immunoregulatory activity via B7-H1-mediated induction of apoptosis in activated T cells. Understanding of the involved mechanisms may lead to development of novel therapeutic approaches for treatment of liver diseases.
Asunto(s)
Antígeno B7-H1/inmunología , Rechazo de Injerto/inmunología , Células Estrelladas Hepáticas/inmunología , Trasplante de Hígado/inmunología , Transducción de Señal/inmunología , Linfocitos T/inmunología , Anticuerpos Monoclonales/inmunología , Anticuerpos Monoclonales/farmacología , Apoptosis/inmunología , Antígeno B7-1/inmunología , Antígeno B7-1/metabolismo , Antígeno B7-H1/metabolismo , Antígenos CD40/inmunología , Antígenos CD40/metabolismo , Comunicación Celular/inmunología , Células Cultivadas , Rechazo de Injerto/metabolismo , Células Estrelladas Hepáticas/citología , Células Estrelladas Hepáticas/metabolismo , Humanos , Tolerancia Inmunológica/inmunología , Inmunofenotipificación , Molécula 1 de Adhesión Intercelular/inmunología , Molécula 1 de Adhesión Intercelular/metabolismo , Hígado/citología , Hígado/inmunología , Hígado/metabolismo , Trasplante HomólogoRESUMEN
BACKGROUND: Despite extensive research, the mechanism of immature dendritic cells (DCs) induced immune hyporesponsiveness remains incompletely understood. METHODS: Recipient DCs from C3H mouse bone marrow cells were incubated with donor antigen from splenic lymphocytes of C57BL/6 mouse; these DCs were transfected with CD80/86 specific siRNA using lentiviral vectors. Flow cytometry was used to evaluate expression of CD80/86 on the antigen-pulsed recipient DCs. Immune regulatory activity was examined by mixed lymphocyte reaction, in which irradiated DCs were cultured with C3H spleen T cells. After the reaction, interleukin (IL)-2, IL-4, IL-10, and interferon (INF)-γ levels of mixed lymphocyte reaction culture supernatant were measured by enzyme-linked immunosorbent assay. The apoptotic T lymphocytes were identified by Annexin V and CD3 staining. RESULTS: There was a significant inhibition of CD80/86 expression in DCs transfected with CD80/86 lentiviral vectors compared with the control groups (P < 0.05), indicating the specificity of RNA interference. Enzyme-linked immunosorbent assay results showed a significant reduction of INF-γ, IL-2 and IL-10 in the CD80/86 lentivirus transfected group compared to the control groups (P < 0.05). There was no significant difference in IL-4 levels between the groups (P > 0.05). We also showed that CD80/86 low DCs loaded with alloantigen (1) stimulated low T cell proliferative responses via the indirect recognition pathway and (2) enhanced apoptotic activity (P < 0.05) in co-cultured T cells. CONCLUSIONS: Lentiviral vector transfection can effectively and specifically knock down target genes in DCs. The CD80/86 low DCs may show tolerogenic activity via induction of T-cell apoptosis, thereby modulating the activity of recipient-derived DCs. The use of this approach may potentially be clinically applicable.
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Apoptosis , Antígeno B7-1/fisiología , Antígeno B7-2/fisiología , Células Dendríticas/inmunología , Interferencia de ARN , Linfocitos T/inmunología , Animales , Antígeno B7-1/genética , Antígeno B7-2/genética , Lentivirus/genética , Activación de Linfocitos , Ratones , Ratones Endogámicos C3H , Ratones Endogámicos C57BL , Linfocitos T/citologíaRESUMEN
Myeloid-derived suppressor cells (MDSCs) play an important role in the regulation of the immune response. MDSC expansion occurs in many circumstances, including cancer, inflammation, stresses, and transplant tolerance. Liver transplants in mice are spontaneously accepted, but hepatocyte transplants are acutely rejected, suggesting the immunoregulatory activities of liver nonparenchymal cells. We have reported that hepatic stellate cells (HpSCs), the stromal cells in the liver, are immensely immunosuppressive and can effectively protect islet transplants via induction of MDSCs. The present study shows that the addition of HpSCs into dendritic cell (DC) culture promoted development of MDSCs, instead of DCs, which was highly dependent on complement component 3 (C3) from HpSCs. The C3(-/-) HpSCs lost their ability to induce MDSCs and, consequently, failed to protect the cotransplanted islet allografts. HpSCs produced complement activation factor B and factor D which then enhanced C3 cleavage to activation products iC3b and C3d. Addition of exogenous iC3b, but not C3d, into the DC culture led to the differentiation of MDSCs with potent immune-inhibitory function. These findings provide novel mechanistic insights into the differentiation of myeloid cells mediated by local tissue cells, and may assist in the development of MDSC-based therapy in clinical settings.
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Diferenciación Celular/inmunología , Complemento C3/fisiología , Células Dendríticas/inmunología , Células Estrelladas Hepáticas/inmunología , Trasplante de Islotes Pancreáticos/inmunología , Células Mieloides/inmunología , Animales , Células Presentadoras de Antígenos/inmunología , Western Blotting , Células Dendríticas/citología , Diabetes Mellitus Tipo 1/inmunología , Diabetes Mellitus Tipo 1/patología , Diabetes Mellitus Tipo 1/prevención & control , Citometría de Flujo , Supervivencia de Injerto , Células Estrelladas Hepáticas/patología , Células Estrelladas Hepáticas/trasplante , Técnicas para Inmunoenzimas , Terapia de Inmunosupresión , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Noqueados , Células Mieloides/citología , ARN Mensajero/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Linfocitos T Citotóxicos/inmunología , Linfocitos T Reguladores/inmunología , Trasplante HomólogoRESUMEN
Interleukin (IL)-13 is an immunoregulatory cytokine secreted by numerous immune cells. Its functions are similar to those of IL-4 and they share a common receptor. This cytokine has been included in recent studies on human tumors and malignant diseases, evoking a scientific interest to investigate the role of IL-13 and its receptors as novel biomarkers and targets for therapy. Colorectal cancer is one of the most common human malignancies, its prognosis is not promising and the efficacy of molecular-targeted therapy has not been established. This review summarizes the currently available data on the role of IL-13 and its receptors in colorectal cancer, including the signaling pathways involved in mediating the effects of IL-13, the role of IL-13 and/or its receptors in the prediction of cancer and several drugs targeting IL-13 or its receptors that are currently under evaluation.
RESUMEN
Allogeneic islet transplantation is a promising approach for restoring normoglycemia in type 1 diabetic patients. Current use of immunosuppressive therapies for management of islet transplant recipients can be counterintuitive to islet function and can lead to complications in the long term. The induction of donor-specific tolerance eliminates the dependency on immunosuppression and allows recipients to retain responses to foreign antigens. The mechanisms by which tolerance is achieved involve the deletion of donor-reactive T cells, induction of T-cell anergy, immune deviation, and generation of regulatory T cells. This review will outline the various methods used for inducing donor-specific tolerance in islet transplantation and will highlight the previously unforeseen potential of tissue stromal cells in promoting islet engraftment.
RESUMEN
BACKGROUND: The CXCR3 receptor and its three interferon-inducible ligands (CXCL9, CXCL10 and CXCL11) have been implicated as playing a central role in directing a Th1 inflammatory response. Recent studies strongly support that the CXCR3 receptor is a very attractive therapeutic target for treating autoimmune diseases, such as rheumatoid arthritis, multiple sclerosis and psoriasis, and to prevent transplant rejection. We describe here the in vitro and in vivo pharmacological characterizations of a novel and potent small molecule CXCR3 antagonist, SCH 546738. RESULTS: In this study, we evaluated in vitro pharmacological properties of SCH 546738 by radioligand receptor binding and human activated T cell chemotaxis assays. In vivo efficacy of SCH 546738 was determined by mouse collagen-induced arthritis, rat and mouse experimental autoimmune encephalomyelitis, and rat cardiac transplantation models. We show that SCH 546738 binds to human CXCR3 with a high affinity of 0.4 nM. In addition, SCH 546738 displaces radiolabeled CXCL10 and CXCL11 from human CXCR3 with IC50 ranging from 0.8 to 2.2 nM in a non-competitive manner. SCH 546738 potently and specifically inhibits CXCR3-mediated chemotaxis in human activated T cells with IC90 about 10 nM. SCH 546738 attenuates the disease development in mouse collagen-induced arthritis model. SCH 546738 also significantly reduces disease severity in rat and mouse experimental autoimmune encephalomyelitis models. Furthermore, SCH 546738 alone achieves dose-dependent prolongation of rat cardiac allograft survival. Most significantly, SCH 546738 in combination with CsA supports permanent engraftment. CONCLUSIONS: SCH 546738 is a novel, potent and non-competitive small molecule CXCR3 antagonist. It is efficacious in multiple preclinical disease models. These results demonstrate that therapy with CXCR3 antagonists may serve as a new strategy for treatment of autoimmune diseases, including rheumatoid arthritis and multiple sclerosis, and to prevent transplant rejection.
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Enfermedades Autoinmunes/tratamiento farmacológico , Quimiotaxis de Leucocito/efectos de los fármacos , Rechazo de Injerto/prevención & control , Supervivencia de Injerto/efectos de los fármacos , Trasplante de Corazón , Piperazinas/farmacología , Pirazinamida/análogos & derivados , Receptores CXCR3/antagonistas & inhibidores , Linfocitos T/efectos de los fármacos , Animales , Artritis Experimental/tratamiento farmacológico , Artritis Experimental/inmunología , Artritis Reumatoide/tratamiento farmacológico , Artritis Reumatoide/inmunología , Enfermedades Autoinmunes/inmunología , Quimiotaxis de Leucocito/inmunología , Modelos Animales de Enfermedad , Encefalomielitis Autoinmune Experimental/tratamiento farmacológico , Encefalomielitis Autoinmune Experimental/inmunología , Rechazo de Injerto/inmunología , Humanos , Técnicas In Vitro , Ratones , Esclerosis Múltiple/tratamiento farmacológico , Esclerosis Múltiple/inmunología , Unión Proteica , Pirazinamida/farmacología , Ensayo de Unión Radioligante , Ratas , Linfocitos T/inmunologíaRESUMEN
PURPOSE: To test whether retinal pigment epithelial (RPE) cells are able to induce myeloid-derived suppressor cell (MDSC) differentiation from bone marrow (BM) progenitors. METHODS: BM cells were cocultured with or without RPE cells in the presence of GM-CSF and IL-4. Numbers of resultant MDSCs were assessed by flow cytometry after 6 days of incubation. The ability of the RPE cell-induced MDSCs to inhibit T cells was evaluated by a CFSE-based T-cell proliferation assay. To explore the mechanism by which RPE cells induce MDSC differentiation, PD-L1-deficient RPE cells and blocking antibodies against TGF-ß, CTLA-2α, and IL-6 were used. RPE cell-induced MDSCs were adoptively transferred into mice immunized with interphotoreceptor retinoid-binding protein in complete Freund's adjuvant to test their efficacy in suppressing autoreactive T-cell responses in experimental autoimmune uveitis (EAU). RESULTS: RPE cells induced the differentiation of MDSCs. These RPE cell-induced MDSCs significantly inhibited T-cell proliferation in a dose-dependent manner. PD-L1-deficient RPE cells induced MDSC differentiation as efficiently as wild-type RPE cells, and neutralizing TGF-ß or CTLA-2α did not alter the numbers of induced MDSCs. However, blocking IL-6 reduced the efficacy of RPE cell-induced MDSC differentiation. Finally, adoptive transfer of RPE cell-induced MDSCs suppressed IRBP-specific T-cell responses that led to EAU. CONCLUSIONS: RPE cells induce the differentiation of MDSCs from bone marrow progenitors. Both cell surface molecules and soluble factors are important in inducing MDSC differentiation. PD-L1, TGF-ß, and CTLA-2α were not measurably involved in RPE cell-induced MDSC differentiation, whereas IL-6 was important in the process. The induction of MDSCs could be another mechanism by which RPE cells control immune reactions in the retina, and RPE cell-induced MDSCs should be further investigated as a potential approach to therapy for autoimmune posterior uveitis.
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Enfermedades Autoinmunes/inmunología , Autoinmunidad , Activación de Linfocitos/inmunología , Epitelio Pigmentado de la Retina/inmunología , Linfocitos T/inmunología , Uveítis/inmunología , Animales , Enfermedades Autoinmunes/patología , Diferenciación Celular , Células Cultivadas , Modelos Animales de Enfermedad , Citometría de Flujo , Ratones , Ratones Endogámicos C57BL , Epitelio Pigmentado de la Retina/patología , Uveítis/patologíaRESUMEN
BACKGROUND: Side effects of lifetime immunosuppression for cell transplants often outweigh the benefits; therefore, induction of transplant tolerance is needed. We have shown that cotransplantation with myeloid-derived suppressor cells (MDSC) effectively protect islet allografts from rejection without requirement of immunosuppression. This study was to investigate the underlying mechanisms. METHODS: MDSC were generated by addition of hepatic stellate cells from various stain mice into dendritic cell (DC) culture. The quality of MDSC was monitored by phenotype and function analyses. MDSC mixed with islet allografts were transplanted into diabetic recipients. T-cell response was analyzed after transplantation by using flow and histochemical analyses, and was compared with islet alone and islet/DC transplant groups. B7-H1 knockout mice were used to determine the role of B7-H1 on MDSC in regulation of T-cell response. RESULTS: Cotransplantation with MDSC (not DC) effectively protected islet allografts without requirement of immunosuppression. This is associated with attenuation of CD8 T cells in the grafts and marked expansion of regulatory T (Treg) cells, which contributed to MDSC-induced T-cell hyporesponsiveness. Antigen-specific Treg cells were prone to accumulate in lymphoid organs close to the grafts. Both in vitro and in vivo data demonstrated that B7-H1 was absolutely required for MDSC to exert immune regulatory activity and induction of Treg cells. CONCLUSION: The described approach holds great clinical application potential and may overcome the limitation of requiring chronic administration of immunosuppression in cell transplants. Understanding the underlying mechanisms will facilitate the development of this novel therapeutic strategy.