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Tumor evasion has recently been associated with a novel member of the B7 family, HERV-H LTR-associating 2 (HHLA2), which is mostly overexpressed in PDL-1neg tumors. HHLA2 can either induce a costimulation signal when bound to CD28H or inhibit it by binding to KIR3DL3 on T- and NK cells. Given the broad distribution of CD28H expression on NK cells and its role, we compared two monoclonal antibodies targeting this novel NK-cell engager in this study. We show that targeting CD28H at a specific epitope not only strongly activates Ca2+ flux but also results in NK-cell activation. CD28H-activated NK cells further display increased cytotoxic activity against hematopoietic cell lines and bypass HHLA2 and HLA-E inhibitory signals. Additionally, scRNA-seq analysis of clear cell renal cancer cells revealed that HHLA2+ clear cell renal cancer cell tumors were infiltrated with CD28H+ NK cells, which could be targeted by finely chosen anti-CD28H Abs.
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Presensitized patients awaiting a kidney transplant have a lower graft survival and a longer waiting time because of the limited number of potential donors and the higher risk of antibody-mediated rejection (AMR), particularly in the early posttransplant period, because of preformed donor-specific antibodies binding major histocompatibility complex (MHC) molecules expressed by the graft endothelium followed by the activation of the complement. Advances in kidney preservation techniques allow the development of ex vivo treatment of transplants. We hypothesized that masking MHC ex vivo before transplantation could help to prevent early AMR in presensitized recipients. We evaluated a strategy of MHC I masking by an antibody during ex vivo organ perfusion in a porcine model of kidney transplantation in alloimmunized recipients. Methods: Through the in vitro calcein-release assay and flow cytometry, we evaluated the protective effect of a monoclonal anti-swine leukocyte antigen class I antibody (clone JM1E3) against alloreactive IgG complement-dependent cytotoxicity toward donor endothelial cells. Kidneys perfused ex vivo with JM1E3 during hypothermic machine perfusion were transplanted to alloimmunized recipients. Results: In vitro incubation of endothelial cells with JM1E3 decreased alloreactive IgG cytotoxicity (mean complement-dependent cytotoxicity index [% of control condition] with 1 µg/mL 74.13% ± 35.26 [calcein assay] and 66.88% ± 33.46 [cytometry]), with high interindividual variability. After transplantation, acute AMR occurred in all recipients on day 1, with signs of complement activation (C5b-9 staining) as soon as 1 h after transplantation, despite effective JM1E3 binding on graft endothelium. Conclusions: Despite a partial protective effect of swine leukocyte antigen I masking with JM1E3 in vitro, ex vivo perfusion of the kidney with JM1E3 before transplantation was not sufficient alone at preventing or delaying AMR in highly sensitized recipients.
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The critical role of neutrophils in pathological inflammation, notably in various autoimmune disorders, is currently the focus of renewed interest. Here, we demonstrate for the first time that activation of neutrophils with various inflammatory stimuli induces the release of extracellular vesicles (EVs) that are internalized by endothelial cells (ECs), thus leading to the transfer of miR-223, miR-142-3p and miR-451 and subsequent endothelial damage. Indeed, while miR-223 has little effect on EC responses, we show that the induced expression of miR-142-3p and miR-451 in ECs results in profound cell damage, especially in inflammatory conditions, characterized by a dramatic increase in cell apoptosis, impaired angiogenic repair responses, and the induction of IL-6, IL-8, CXCL10 and CXCL11 expression. We show that the strong deleterious effect of miR-142-3p may be due in part to its ability to block the activation of ERK1/2 and eNOS-mediated signals in ECs. miR-142-3p also inhibits the expression of RAC1, ROCK2 and CLIC4, three genes that are critical for EC migration and angiogenic responses. Importantly, miR-223, miR-142-3p and miR-451 are markedly increased in kidney biopsies from patients with active ANCA-associated vasculitis, a severe autoimmune disease that is prototypical of a neutrophil-induced microvascular damage. Taken together, our results suggest that miR-142-3p and miR-451 released in EVs by activated neutrophils can target EC to trigger an inflammatory cascade and induce direct vascular damage, and that therapeutic strategies based on the inhibition of these miRNAs in ECs will have implications for neutrophil-mediated inflammatory diseases.
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Vesículas Extracelulares , MicroARNs , Canales de Cloruro/metabolismo , Células Endoteliales/metabolismo , Vesículas Extracelulares/metabolismo , Humanos , Inflamación/metabolismo , MicroARNs/genética , Neutrófilos/metabolismoRESUMEN
T cell exclusion causes resistance to cancer immunotherapies via immune checkpoint blockade (ICB). Myeloid cells contribute to resistance by expressing signal regulatory protein-α (SIRPα), an inhibitory membrane receptor that interacts with ubiquitous receptor CD47 to control macrophage phagocytosis in the tumor microenvironment. Although CD47/SIRPα-targeting drugs have been assessed in preclinical models, the therapeutic benefit of selectively blocking SIRPα, and not SIRPγ/CD47, in humans remains unknown. We report a potent synergy between selective SIRPα blockade and ICB in increasing memory T cell responses and reverting exclusion in syngeneic and orthotopic tumor models. Selective SIRPα blockade stimulated tumor nest T cell recruitment by restoring murine and human macrophage chemokine secretion and increased anti-tumor T cell responses by promoting tumor-antigen crosspresentation by dendritic cells. However, nonselective SIRPα/SIRPγ blockade targeting CD47 impaired human T cell activation, proliferation, and endothelial transmigration. Selective SIRPα inhibition opens an attractive avenue to overcoming ICB resistance in patients with elevated myeloid cell infiltration in solid tumors.
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Memoria Inmunológica , Inmunoterapia , Neoplasias Mamarias Experimentales/terapia , Proteínas de Neoplasias/inmunología , Receptores Inmunológicos/inmunología , Linfocitos T/inmunología , Animales , Femenino , Neoplasias Mamarias Experimentales/inmunología , Neoplasias Mamarias Experimentales/patología , Ratones , Ratones Endogámicos BALB C , Proteínas de Neoplasias/genética , Receptores Inmunológicos/genética , Linfocitos T/patologíaRESUMEN
BACKGROUND: Identifying biomarkers to predict kidney transplant failure and to define new therapeutic targets requires more comprehensive understanding of the immune response to chronic allogeneic stimulation. METHODS: We investigated the frequency and function of CD8+ T cell subsets-including effector memory (EM) and terminally differentiated EM (TEMRA) CD8+ T cells-in blood samples from 284 kidney transplant recipients recruited 1 year post-transplant and followed for a median of 8.3 years. We also analyzed CD8+ T cell reactivity to donor-specific PBMCs in 24 patients who had received living-donor kidney transplants. RESULTS: Increased frequency of circulating TEMRA CD8+ T cells at 1 year post-transplant associated with increased risk of graft failure during follow-up. This association remained after adjustment for a previously reported composite of eight clinical variables, the Kidney Transplant Failure Score. In contrast, increased frequency of EM CD8+ T cells associated with reduced risk of graft failure. A distinct TEMRA CD8+ T cell subpopulation was identified that was characterized by expression of FcγRIIIA (CD16) and by high levels of proinflammatory cytokine secretion and cytotoxic activity. Although donor-specific stimulation induced a similar rapid, early response in EM and TEMRA CD8+ T cells, CD16 engagement resulted in selective activation of TEMRA CD8+ T cells, which mediated antibody-dependent cytotoxicity. CONCLUSIONS: At 1 year post-transplant, the composition of memory CD8+ T cell subsets in blood improved prediction of 8-year kidney transplant failure compared with a clinical-variables score alone. A subpopulation of TEMRA CD8+ T cells displays a novel dual mechanism of activation mediated by engagement of the T-cell receptor or of CD16. These findings suggest that TEMRA CD8+ T cells play a pivotal role in humoral and cellular rejection and reveal the potential value of memory CD8+ T cell monitoring for predicting risk of kidney transplant failure.
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Linfocitos T CD8-positivos , Rechazo de Injerto/etiología , Supervivencia de Injerto , Fallo Renal Crónico/sangre , Fallo Renal Crónico/cirugía , Trasplante de Riñón , Adulto , Anciano , Biomarcadores/sangre , Estudios de Cohortes , Femenino , Rechazo de Injerto/sangre , Rechazo de Injerto/diagnóstico , Humanos , Memoria Inmunológica , Masculino , Persona de Mediana Edad , Valor Predictivo de las Pruebas , Curva ROC , Medición de Riesgo , Resultado del TratamientoRESUMEN
Myeloid-derived suppressor cells (MDSC) are a heterogeneous population of immature hematopoietic precursors known to suppress immune responses. Interaction of SIRP alpha (SIRPα), expressed by myeloid cells, with the ubiquitous receptor CD47 is an important immune checkpoint of the innate response regulating macrophages and dendritic cells functions. We previously described that MDSC expressing SIRPα accumulated after transplantation and maintained kidney allograft tolerance. However, the role of the SIRPα/CD47 axis on MDSC function remained unknown. Here, we found that blocking SIRPα or CD47 with monoclonal antibodies (mAbs) induced differentiation of MDSC into myeloid cells overexpressing MHC class II, CD86 costimulatory molecule and increased secretion of macrophage-recruiting chemokines (eg, MCP-1). Using a model of long-term kidney allograft tolerance sustained by MDSC, we observed that administration of blocking anti-SIRPα or CD47 mAbs induced graft dysfunction and rejection. Loss of tolerance came along with significant decrease of MDSC and increase in MCP-1 concentration in the periphery. Graft histological and transcriptomic analyses revealed an inflammatory (M1) macrophagic signature at rejection associated with overexpression of MCP-1 mRNA and protein in the graft. These findings indicate that the SIRPα-CD47 axis regulates the immature phenotype and chemokine secretion of MDSC and contributes to the induction and the active maintenance of peripheral acquired immune tolerance.
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Antígeno CD47/metabolismo , Rechazo de Injerto/inmunología , Trasplante de Riñón/efectos adversos , Células Mieloides/inmunología , Células Supresoras de Origen Mieloide/inmunología , Receptores Inmunológicos/metabolismo , Tolerancia al Trasplante/inmunología , Animales , Anticuerpos Monoclonales/administración & dosificación , Antígeno CD47/antagonistas & inhibidores , Antígeno CD47/inmunología , Quimiocinas , Rechazo de Injerto/patología , Supervivencia de Injerto/inmunología , Células Mieloides/citología , Ratas , Receptores Inmunológicos/antagonistas & inhibidores , Receptores Inmunológicos/inmunologíaRESUMEN
It remains unknown what causes inflammatory bowel disease (IBD), including signaling networks perpetuating chronic gastrointestinal inflammation in Crohn's disease (CD) and ulcerative colitis (UC), in humans. According to an analysis of up to 500 patients with IBD and 100 controls, we report that key transcripts of the IL-7 receptor (IL-7R) pathway are accumulated in inflamed colon tissues of severe CD and UC patients not responding to either immunosuppressive/corticosteroid, anti-TNF, or anti-α4ß7 therapies. High expression of both IL7R and IL-7R signaling signature in the colon before treatment is strongly associated with nonresponsiveness to anti-TNF therapy. While in mice IL-7 is known to play a role in systemic inflammation, we found that in humans IL-7 also controlled α4ß7 integrin expression and imprinted gut-homing specificity on T cells. IL-7R blockade reduced human T cell homing to the gut and colonic inflammation in vivo in humanized mouse models, and altered effector T cells in colon explants from UC patients grown ex vivo. Our findings show that failure of current treatments for CD and UC is strongly associated with an overexpressed IL-7R signaling pathway and point to IL-7R as a relevant therapeutic target and potential biomarker to fill an unmet need in clinical IBD detection and treatment.
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Colitis Ulcerosa/metabolismo , Colon/metabolismo , Enfermedad de Crohn/metabolismo , Receptores de Interleucina-7/metabolismo , Linfocitos T/citología , Factor de Necrosis Tumoral alfa/antagonistas & inhibidores , Adolescente , Adulto , Anciano , Animales , Colon/patología , Citocinas/metabolismo , Endoscopía , Femenino , Perfilación de la Expresión Génica , Regulación de la Expresión Génica , Enfermedad Injerto contra Huésped/metabolismo , Humanos , Inflamación , Integrinas/metabolismo , Mucosa Intestinal/metabolismo , Leucocitos Mononucleares/citología , Masculino , Ratones , Ratones Endogámicos NOD , Ratones SCID , Persona de Mediana Edad , Transducción de Señal , Adulto JovenRESUMEN
Targeting the expansion of pathogenic memory immune cells is a promising therapeutic strategy to prevent chronic autoimmune attacks. Here we investigate the therapeutic efficacy and mechanism of new anti-human IL-7Rα monoclonal antibodies (mAb) in non-human primates and show that, depending on the target epitope, a single injection of antagonistic anti-IL-7Rα mAbs induces a long-term control of skin inflammation despite repeated antigen challenges in presensitized monkeys. No modification in T cell numbers, phenotype, function or metabolism is observed in the peripheral blood or in response to polyclonal stimulation ex vivo. However, long-term in vivo hyporesponsiveness is associated with a significant decrease in the frequency of antigen-specific T cells producing IFN-γ upon antigen restimulation ex vivo. These findings indicate that chronic antigen-specific memory T cell responses can be controlled by anti-IL-7Rα mAbs, promoting and maintaining remission in T-cell mediated chronic inflammatory diseases.
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Anticuerpos Monoclonales/uso terapéutico , Memoria Inmunológica/efectos de los fármacos , Inflamación/tratamiento farmacológico , Receptores de Interleucina-7/antagonistas & inhibidores , Linfocitos T/inmunología , Animales , Enfermedad Crónica , Supresión Clonal/inmunología , Modelos Animales de Enfermedad , Humanos , Memoria Inmunológica/inmunología , Inflamación/inmunología , Interferón gamma/inmunología , Papio , Receptores de Interleucina-7/agonistas , Receptores de Interleucina-7/inmunología , Transducción de Señal/efectos de los fármacos , Piel/inmunología , Piel/patologíaRESUMEN
Rat and human CD4+ and CD8+ Tregs expressing low levels of CD45RC have strong immunoregulatory properties. We describe here that human CD45 isoforms are nonredundant and identify distinct subsets of cells. We show that CD45RC is not expressed by CD4+ and CD8+ Foxp3+ Tregs, while CD45RA/RB/RO are. Transient administration of a monoclonal antibody (mAb) targeting CD45RC in a rat cardiac allotransplantation model induced transplant tolerance associated with inhibition of allogeneic humoral responses but maintained primary and memory responses against cognate antigens. Anti-CD45RC mAb induced rapid death of CD45RChigh T cells through intrinsic cell signaling but preserved and potentiated CD4+ and CD8+ CD45RClow/- Tregs, which are able to adoptively transfer donor-specific tolerance to grafted recipients. Anti-CD45RC treatment results in distinct transcriptional signature of CD4+ and CD8+ CD45RClow/- Tregs. Finally, we demonstrate that anti-human CD45RC treatment inhibited graft-versus-host disease (GVHD) in immune-humanized NSG mice. Thus, short-term anti-CD45RC is a potent therapeutic candidate to induce transplantation tolerance in human.
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Anticuerpos Monoclonales/administración & dosificación , Enfermedad Injerto contra Huésped/tratamiento farmacológico , Antígenos Comunes de Leucocito/metabolismo , Linfocitos T Reguladores/inmunología , Animales , Anticuerpos Monoclonales/inmunología , Anticuerpos Monoclonales/farmacología , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD8-positivos/inmunología , Células Cultivadas , Modelos Animales de Enfermedad , Factores de Transcripción Forkhead/metabolismo , Enfermedad Injerto contra Huésped/inmunología , Trasplante de Corazón , Humanos , Inmunidad Humoral/efectos de los fármacos , Ratones , Ratas , Tolerancia al TrasplanteRESUMEN
Polyclonal xenogenic IgGs, although having been used in the prevention and cure of severe infectious diseases, are highly immunogenic, which may restrict their usage in new applications such as Ebola hemorrhagic fever. IgG glycans display powerful xenogeneic antigens in humans, for example α1-3 Galactose and the glycolyl form of neuraminic acid Neu5Gc, and IgGs deprived of these key sugar epitopes may represent an advantage for passive immunotherapy. In this paper, we explored whether low immunogenicity IgGs had a protective effect on a guinea pig model of Ebola virus (EBOV) infection. For this purpose, a double knock-out pig lacking α1-3 Galactose and Neu5Gc was immunized against virus-like particles displaying surface EBOV glycoprotein GP. Following purification from serum, hyper-immune polyclonal IgGs were obtained, exhibiting an anti-EBOV GP titer of 1:100,000 and a virus neutralizing titer of 1:100. Guinea pigs were injected intramuscularly with purified IgGs on day 0 and day 3 post-EBOV infection. Compared to control animals treated with IgGs from non-immunized double KO pigs, the anti-EBOV IgGs-treated animals exhibited a significantly prolonged survival and a decreased virus load in blood on day 3. The data obtained indicated that IgGs lacking α1-3 Galactose and Neu5Gc, two highly immunogenic epitopes in humans, have a protective effect upon EBOV infection.
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Anticuerpos Antivirales/sangre , Vacunas contra el Virus del Ébola/uso terapéutico , Galactosa/deficiencia , Fiebre Hemorrágica Ebola/prevención & control , Inmunoglobulina G/inmunología , Ácidos Neuramínicos/metabolismo , Proteínas del Envoltorio Viral/inmunología , Animales , Anticuerpos Antiidiotipos/inmunología , Vacunas contra el Virus del Ébola/inmunología , Ebolavirus/inmunología , Cobayas , Fiebre Hemorrágica Ebola/sangre , Fiebre Hemorrágica Ebola/inmunología , Masculino , Porcinos , Vacunación , Carga ViralRESUMEN
In transplantation tolerance, numerous regulatory populations have the capacity to inhibit allograft rejection; however, their compensatory capacities have never been clearly evidenced. We have previously demonstrated that the tolerogenic effect mediated by CD8(+)CD45RC(low) regulatory T cells (Tregs) in a model of organ transplantation with CD40Ig could be abrogated by permanent depletion of CD8(+) cells that resulted in allograft rejection in half of the recipients. This result demonstrated that CD8(+) Tregs were essential, but also that half of the recipients still survived indefinitely. We also demonstrated that no other regulatory populations, besides CD8(+) Tregs, could induce and maintain allograft tolerance in CD40Ig-treated tolerant animals. In the current study, we analyzed the mechanisms that arose following CD8(+) Treg depletion and allowed establishment of networks of new regulatory cells to maintain allograft survival. We identified regulatory B cells (Bregs) and regulatory myeloid cells (RegMCs) as being responsible of the maintenance of the long-term allograft survival. We demonstrated that both regulatory cell subsets efficiently inhibited antidonor immune responses in adoptively transferred recipients. Although Bregs were induced, they were not essential for the maintenance of the graft as demonstrated in IgM-deficient recipients. In addition, we showed that RegMCs were the most suppressive and acted alone, whereas Bregs activity was associated with increased suppressive activity of other subsets in adoptively transferred recipients. Altogether, to our knowledge, we demonstrated in this study for the first time the emergence of both Bregs and RegMCs following Tregs depletion and highlighted the importance of regulatory cell networks and their synergistic potential in transplantation.
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Linfocitos B Reguladores/inmunología , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD8-positivos/inmunología , Rechazo de Injerto/inmunología , Trasplante de Corazón , Células Mieloides/inmunología , Traslado Adoptivo , Animales , Linfocitos B Reguladores/trasplante , Comunicación Celular , Células Cultivadas , Inmunomodulación , Células Mieloides/trasplante , Ratas , Ratas Endogámicas Lew , Tolerancia al TrasplanteRESUMEN
T cell depletion is commonly used in organ transplantation for immunosuppression; however, a restoration of T cell homeostasis following depletion leads to increased memory T cells, which may promote transplant rejection. The cytokine IL-7 is important for controlling lymphopoiesis under both normal and lymphopenic conditions. Here, we investigated whether blocking IL-7 signaling with a mAb that targets IL-7 receptor α (IL-7Rα) alone or following T cell depletion confers an advantage for allograft survival in murine transplant models. We found that IL-7R blockade alone induced indefinite pancreatic islet allograft survival if anti-IL-7R treatment was started 3 weeks before graft. IL-7R blockade following anti-CD4- and anti-CD8-mediated T cell depletion markedly prolonged skin allograft survival. Furthermore, IL-7 inhibition in combination with T cell depletion synergized with either CTLA-4Ig administration or suboptimal doses of tacrolimus to induce long-term skin graft acceptance in this stringent transplant model. Together, these therapies inhibited T cell reconstitution, decreased memory T cell numbers, increased the relative frequency of Tregs, and abrogated both cellular and humoral alloimmune responses. Our data suggest that IL-7R blockade following T cell depletion has potential as a robust, immunosuppressive therapy in transplantation.
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Supervivencia de Injerto/inmunología , Terapia de Inmunosupresión/métodos , Depleción Linfocítica , Receptores de Interleucina-7/antagonistas & inhibidores , Linfocitos T/inmunología , Abatacept , Aloinjertos , Animales , Anticuerpos Monoclonales/administración & dosificación , Inmunidad Celular , Inmunidad Humoral , Inmunoconjugados/administración & dosificación , Inmunosupresores/administración & dosificación , Trasplante de Islotes Pancreáticos/inmunología , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Mutantes , Ratones Desnudos , Trasplante de Piel , Tacrolimus/administración & dosificación , Inmunología del Trasplante , Tolerancia al Trasplante/inmunologíaRESUMEN
Antagonist antibodies targeting CD28 have been proposed as an alternative to the use of CD80/86 antagonists to modulate T cell responses in autoimmunity and transplantation. Advantages would be the blockade of CD28-mediated co-stimulatory signals without impeding the co-inhibitory signals dependent on CD80 interactions with CTLA-4 and PD-L1 that are important for the control of immune responses and for the function of regulatory T cells. Anti-CD28 antibodies are candidate antagonists only if they prevent access to the CD80/86 ligands without simultaneously stimulating CD28 itself, a process that is believed to depend on receptor multimerization. In this study, we evaluated the impact of different formats of a potentially antagonist anti-human CD28 antibody on T cell activation. In particular, we examined the role of valency and of the presence of an Fc domain, two components that might affect receptor multimerization either directly or in the presence of accessory cells expressing Fc receptors. Among monovalent (Fab', scFv), divalent (Fab'2), monovalent-Fc (Fv-Fc) and divalent-Fc (IgG) formats, only the monovalent formats showed consistent absence of induced CD28 multimerization and absence of associated activation of phosphoinositol-3-kinase, and clear antagonist properties in T cell stimulation assays. In contrast, divalent antibodies showed agonist properties that resulted in cell proliferation and cytokine release in an Fc-independent manner. Conjugation of monovalent antibodies with polyethylene glycol, α-1-antitrypsin or an Fc domain significantly extended their in vivo half-life without modifying their antagonist properties. In conclusion, these data indicate that monovalency is mandatory for maintaining the antagonistic activity of anti-CD28 monoclonal antibodies.
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Anticuerpos Monoclonales/inmunología , Antígenos CD28/antagonistas & inhibidores , Anticuerpos de Dominio Único/inmunología , Linfocitos T/inmunología , Animales , Anticuerpos Monoclonales/farmacocinética , Anticuerpos Monoclonales/farmacología , Antígenos CD28/inmunología , Femenino , Semivida , Humanos , Inmunoconjugados/inmunología , Fragmentos Fab de Inmunoglobulinas/química , Fragmentos Fab de Inmunoglobulinas/inmunología , Células Jurkat , Activación de Linfocitos , Prueba de Cultivo Mixto de Linfocitos , Ratones , Polietilenglicoles/química , Anticuerpos de Dominio Único/químicaRESUMEN
Myeloid-derived suppressor cells (MDSC) are a heterogeneous population of immature cells that are believed to inhibit immune responses in the contexts of cancer and organ transplantation, in association with regulatory T cells (Treg). However, the way in which MDSC cooperate with Treg remains elusive. In this study, we used DNA microarrays to analyze gene expression in blood-derived MDSC from rat recipients of kidney allografts. We found CCL5 (Rantes), a chemotactic C-C motif 5 chemokine, to be strongly downregulated after treatment with a tolerizing regimen. The amount of CCL5 protein was also lower in the plasma of tolerant recipients, whereas intragraft CCL5 was unchanged. Because CCL5 is chemotactic for Treg, we hypothesized that a gradient of CCL5 between the graft and peripheral blood might contribute to the intragraft localization of Treg in tolerant animals. To test this hypothesis, we treated tolerant rat recipients of kidney allografts with recombinant rat CCL5 to restore normal plasma concentrations. This led to a strong reduction in intragraft Treg monitored by immunohistofluorescence and by quantitative real-time PCR measurement of Foxp3 mRNA. Ultimately, this treatment led to an increase in serum creatinine concentrations and to kidney graft rejection after about a month. The kidney function of syngeneic grafts was not affected by a similar administration of CCL5. These data highlight the contribution of MDSC to the establishment of a graft-to-periphery CCL5 gradient in tolerant kidney allograft recipients, which controls recruitment of Treg to the graft where they likely contribute to maintaining tolerance.
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Quimiocina CCL5/inmunología , Trasplante de Riñón , Riñón/inmunología , Células Mieloides/inmunología , Linfocitos T Reguladores/inmunología , Tolerancia al Trasplante , Trasplantes , Animales , Línea Celular , Quimiocina CCL5/metabolismo , Factores de Transcripción Forkhead/biosíntesis , Factores de Transcripción Forkhead/inmunología , Perfilación de la Expresión Génica , Regulación de la Expresión Génica/inmunología , Masculino , Ratones , Células Mieloides/metabolismo , Análisis de Secuencia por Matrices de Oligonucleótidos , ARN Mensajero/biosíntesis , ARN Mensajero/inmunología , Ratas , Ratas Endogámicas Lew , Linfocitos T Reguladores/metabolismo , Trasplante HomólogoRESUMEN
Transplantation is the treatment of choice for patients with end-stage organ failure. Its success is limited by side effects of immunosuppressive drugs, such as inhibitors of the calcineurin pathway that prevent rejection by reducing synthesis of interleukin-2 by T cells. Moreover, none of the existing drugs efficiently prevent the eventual rejection of the organ. Blocking the CD28-mediated T cell costimulation pathway is a nontoxic alternative immunosuppression strategy that is now achieved by blockade of CD80/86, the receptor for CD28 on antigen-presenting cells. However, interaction of CD80/86 with cytotoxic T lymphocyte-associated antigen 4 (CTLA-4) is required for immune regulation. Therefore, CD28 blockade, instead of CD80/86 blockade, might preserve regulatory signals mediated by CTLA-4 and preserve immune regulation. By using monovalent antibodies, we identified true CD28 antagonists that induced CTLA-4-dependent decreased T cell function compatible with regulatory T (Treg) cell suppression. In transplantation experiments in primates, blocking CD28 augmented intragraft and peripheral blood Treg cells, induced molecular signatures of immune regulation, and prevented graft rejection and vasculopathy in synergy with calcineurin inhibition. These findings suggest that targeting costimulation blockade at CD28 preserves CTLA-4-dependent immune regulation and promotes allograft survival.
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Antígenos CD/metabolismo , Antígenos CD28/metabolismo , Trasplante de Órganos/métodos , Animales , Antígeno B7-1/biosíntesis , Antígeno B7-2/biosíntesis , Antígeno CTLA-4 , Calcineurina/metabolismo , Supervivencia de Injerto , Humanos , Sistema Inmunológico , Linfocitos/citología , Macaca fascicularis , Papio , Linfocitos T Reguladores/citologíaRESUMEN
The immune tolerance to rat kidney allografts induced by a perioperative treatment with anti-CD28 Abs is associated with a severe unresponsiveness of peripheral blood cells to donor Ags. In this model, we identified an accumulation in the blood of CD3(-)class II(-)CD11b(+)CD80/86(+) plastic-adherent cells that additionally expressed CD172a as well as other myeloid markers. These cells were able to inhibit proliferation, but not activation, of effector T cells and to induce apoptosis in a contact-dependent manner. Their suppressive action was found to be under the control of inducible NO synthase, an enzyme also up-regulated in tolerated allografts. Based on these features, these cells can be defined as myeloid-derived suppressor cells (MDSC). Interestingly, CD4(+)CD25(high)FoxP3(+) regulatory T cells were insensitive in vitro to MDSC-mediated suppression. Although the adoptive transfer of MDSC failed to induce kidney allograft tolerance in recently transplanted recipients, the maintenance of tolerance after administration of anti-CD28 Abs was found to be dependent on the action of inducible NO synthase. These results suggest that increased numbers of MDSC can inhibit alloreactive T cell proliferation in vivo and that these cells may participate in the NO-dependent maintenance phase of tolerance.
Asunto(s)
Diferenciación Celular/inmunología , Movimiento Celular/inmunología , Tolerancia Inmunológica , Trasplante de Riñón/inmunología , Células Mieloides/citología , Células Mieloides/inmunología , Animales , Antígeno B7-1/biosíntesis , Antígeno B7-2/biosíntesis , Antígeno CD11b/biosíntesis , Adhesión Celular , Células Cultivadas , Pruebas Inmunológicas de Citotoxicidad , Inmunofenotipificación , Masculino , Modelos Inmunológicos , Células Mieloides/metabolismo , Óxido Nítrico Sintasa de Tipo II/biosíntesis , Óxido Nítrico Sintasa de Tipo II/fisiología , Ratas , Ratas Endogámicas Lew , Bazo/citología , Bazo/inmunología , Bazo/patología , Linfocitos T Reguladores/citología , Linfocitos T Reguladores/inmunologíaRESUMEN
BACKGROUND: In heart allograft in the rat, a sustained costimulation blockade with CTLA4Ig prevents alloreactive T-cell activation and promotes a long-term graft survival through the action of tolerogeneic dendritic cells. It is unclear whether similar mechanisms might occur after xenotransplantation. To test that hypothesis, we have analyzed the action of CTLA4Ig in a model of CD4(+)T cell-mediated xenograft rejection. METHODS: Hamster hearts were transplanted into LEW.1A rats receiving an accommodation-inducing treatment consisting of a short course administration of LF15-0195 and a daily administration of cyclosporine A (CSA). To achieve long-term delivery of CTLA4Ig, an intravenous administration of an adenovirus vector coding for mouse CTLA4Ig (Ad-CTLA4Ig) was added to the accommodation induction protocol. On day 40 post-transplantation, rejection was induced by CSA withdrawal. In other xenograft recipients, CD28/B7 costimulation was inhibited at that time only by injections of CTLA4Ig or anti-CD28 antibodies. Graft survival, immunohistology, as well as development of antibodies and regulatory cells were examined. RESULTS: Xenografts survived 6 days after CSA withdrawal in controls and were rejected, as previously described, through the action of CD4(+) xenoreactive T cells. Interfering with CD28/B7 costimulation inhibited this xenoreactive T cell response and delayed rejection to day 10. In recipients that had received Ad-CTLA4Ig, survival was prolonged to day 19 and this was accompanied by the appearance of regulatory cells exhibiting non-donor-specific suppressive activity dependent on IL-2, NO, and IDO. These regulatory cells were different from those previously identified after Ad-CTLA4Ig administration in heart allograft in the rat. In these recipients, rejection occurred as a consequence of an evoked anti-donor IgM response and complement activation and not of a cellular rejection as complement inhibition with cobra venom factor further prolonged xenograft survival. CONCLUSION: CD28/B7 blockade delays CD4(+) T cell-mediated rejection after CSA withdrawal in accommodated recipients of hamster heart xenografts. In addition, a sustained expression of CTLA4Ig has the potential of inducing cellular regulatory mechanisms. However, such treatment does not prevent the development of xenoreactive IgM antibodies that participate in vascular rejection processes in a complement-dependent manner.
Asunto(s)
Enfermedades Cardiovasculares/inmunología , Cricetinae , Rechazo de Injerto/inmunología , Trasplante de Corazón/inmunología , Ratas , Linfocitos T Reguladores/inmunología , Trasplante Heterólogo/inmunología , Abatacept , Adenoviridae/genética , Animales , Antígeno B7-1/inmunología , Antígenos CD28/inmunología , Enfermedades Cardiovasculares/metabolismo , Enfermedades Cardiovasculares/patología , Enfermedades Cardiovasculares/cirugía , Células Cultivadas , Ciclosporina/farmacología , Rechazo de Injerto/prevención & control , Supervivencia de Injerto/inmunología , Trasplante de Corazón/efectos adversos , Inmunoconjugados/genética , Inmunoconjugados/inmunología , Inmunoconjugados/metabolismo , Masculino , Bazo/citología , Linfocitos T Reguladores/metabolismoRESUMEN
Recent studies suggest that particular dendritic cells (DC) subpopulations may be tolerogenic. To test the capacity of different DC subpopulations to modulate allograft rejection, we generated two distinct populations of rat bone marrow-derived DCs (BMDC) with low doses of GM-CSF and IL-4. The non-adherent population (nBMDC), which are the 'classical' DCs was able to stimulate naive allogeneic T cells and could be induced to completely mature using various stimuli. In contrast, the adherent population (aBMDC), which displayed an immature phenotype, was unable to stimulate T cells and was more resistant to maturation. We found that syngeneic aBMDCs, injected one day before transplantation, induced significant prolongation of heart allograft survival and decreased anti-donor humoral and cellular responses. Similarly, syngeneic aBMDCs inhibited T-cell responses to KLH in the spleen but not in lymph node in a KLH immunization model without graft. This effect was not antigen specific and could be reversed using an inhibitor of inducible nitric oxide synthase. This compartmentalized inhibition could be in part explained by the fact that the majority of syngeneic adherent cells administered intravenously were found in the spleen with some of them reaching the T-cell areas. These data suggest that syngeneic aBMDCs can modulate immune responses.