RESUMO
Regulatory B (Breg) cells have been shown to play a critical role in immune homeostasis and in autoimmunity models. We have recently demonstrated that combined anti-T cell immunoglobulin domain and mucin domain-1 and anti-CD45RB antibody treatment results in tolerance to full MHC-mismatched islet allografts in mice by generating Breg cells that are necessary for tolerance. Breg cells are antigen-specific and are capable of transferring tolerance to untreated, transplanted animals. Here, we demonstrate that adoptively transferred Breg cells require the presence of regulatory T (Treg) cells to establish tolerance, and that adoptive transfer of Breg cells increases the number of Treg cells. Interaction with Breg cells in vivo induces significantly more Foxp3 expression in CD4(+) CD25(-) T cells than with naive B cells. We also show that Breg cells express the TGF-ß associated latency-associated peptide and that Breg-cell mediated graft prolongation post-adoptive transfer is abrogated by neutralization of TGF-ß activity. Breg cells, like Treg cells, demonstrate preferential expression of both C-C chemokine receptor 6 and CXCR3. Collectively, these findings suggest that in this model of antibody-induced transplantation tolerance, Breg cells promote graft survival by promoting Treg-cell development, possibly via TGF-ß production.
Assuntos
Subpopulações de Linfócitos B/imunologia , Sobrevivência de Enxerto/imunologia , Transplante das Ilhotas Pancreáticas/imunologia , Linfócitos T Reguladores/imunologia , Fator de Crescimento Transformador beta/imunologia , Tolerância ao Transplante , Transferência Adotiva , Aloenxertos , Animais , Subpopulações de Linfócitos B/patologia , Fatores de Transcrição Forkhead/genética , Fatores de Transcrição Forkhead/imunologia , Regulação da Expressão Gênica/genética , Regulação da Expressão Gênica/imunologia , Sobrevivência de Enxerto/genética , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Receptores CCR6/genética , Receptores CCR6/imunologia , Receptores CXCR3/genética , Receptores CXCR3/imunologia , Linfócitos T Reguladores/patologia , Fator de Crescimento Transformador beta/genéticaRESUMO
The mechanism by which regulatory T (Treg) cells suppress the immune response is not well defined. A recent study has shown that ß-catenin prolongs Treg cell survival. Because ß-catenin is regulated by glycogen synthase kinase 3ß (GSK-3ß)-directed phosphorylation, we focused on GSK-3ß and the role it plays in Treg cell function. Inhibition of GSK-3ß led to increased suppression activity by Treg cells. Inhibitor-treated Treg cells exhibited prolonged FoxP3 expression and increased levels of ß-catenin and of the antiapoptotic protein Bcl-xL. Systemic administration of GSK-3ß inhibitor resulted in prolonged islet survival in an allotransplant mouse model. Our data suggest that GSK-3ß could be a useful target in developing strategies designed to increase the stability and function of Treg cells for inducing allotransplant tolerance or treating autoimmune conditions.
Assuntos
Inibidores Enzimáticos/farmacologia , Quinase 3 da Glicogênio Sintase/antagonistas & inibidores , Tolerância Imunológica/efeitos dos fármacos , Linfócitos T Reguladores/enzimologia , Animais , Doenças Autoimunes/tratamento farmacológico , Doenças Autoimunes/enzimologia , Doenças Autoimunes/imunologia , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/imunologia , Fatores de Transcrição Forkhead/imunologia , Fatores de Transcrição Forkhead/metabolismo , Quinase 3 da Glicogênio Sintase/imunologia , Quinase 3 da Glicogênio Sintase/metabolismo , Glicogênio Sintase Quinase 3 beta , Tolerância Imunológica/imunologia , Ilhotas Pancreáticas/imunologia , Ilhotas Pancreáticas/metabolismo , Transplante das Ilhotas Pancreáticas/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C3H , Fosforilação/efeitos dos fármacos , Fosforilação/imunologia , Linfócitos T Reguladores/imunologia , Transplante Homólogo , Proteína bcl-X/imunologia , Proteína bcl-X/metabolismo , beta Catenina/imunologia , beta Catenina/metabolismoRESUMO
Involvement of Treg in transplant tolerance has been demonstrated in multiple models. During the active process of graft rejection, these regulatory cells are themselves regulated and inactivated, a process termed counter-regulation. We hypothesize that ligation of the costimulatory molecule glucocorticoid-induced TNF receptor-related protein (GITR) on Treg inhibits their ability to promote graft survival, and by blocking GITR ligation graft survival can be prolonged. To this aim, we have designed a soluble GITR fusion protein (GITR-Fc), which binds GITR ligand and inhibits activation of GITR. Here, we show that GITR-Fc prolonged mouse skin graft survival, and this prolongation is dependent on Treg. In a full MHC-mismatched skin graft setting, GITR-Fc significantly improved graft survival when used in combination with MR1, anti-CD40L, while GITR-Fc alone did not demonstrate graft prolongation. These results demonstrate that disruption of binding of GITR with GITR ligand may be an important strategy in prolonging allograft survival.
Assuntos
Sobrevivência de Enxerto/efeitos dos fármacos , Imunossupressores/uso terapêutico , Receptores de Fator de Crescimento Neural/imunologia , Receptores do Fator de Necrose Tumoral/imunologia , Transplante de Pele/imunologia , Linfócitos T Reguladores/imunologia , Transplante Homólogo/imunologia , Inibidores do Fator de Necrose Tumoral , Transferência Adotiva , Animais , Anticorpos Monoclonais/administração & dosagem , Anticorpos Monoclonais/farmacologia , Anticorpos Monoclonais/uso terapêutico , Células Apresentadoras de Antígenos/efeitos dos fármacos , Células Apresentadoras de Antígenos/imunologia , Ligação Competitiva , Ligante de CD40 , Proteína Relacionada a TNFR Induzida por Glucocorticoide , Antígenos de Histocompatibilidade Classe I , Humanos , Tolerância Imunológica/efeitos dos fármacos , Imunossupressores/administração & dosagem , Imunossupressores/farmacologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Antígenos de Histocompatibilidade Menor , Receptores de Fator de Crescimento Neural/genética , Receptores do Fator de Necrose Tumoral/genética , Proteínas Recombinantes de Fusão/administração & dosagem , Proteínas Recombinantes de Fusão/farmacologia , Proteínas Recombinantes de Fusão/uso terapêutico , Fatores de Necrose Tumoral/imunologiaRESUMO
BACKGROUND: Allogeneic tolerance can be reliably obtained with monoclonal antibody therapy targeting CD45RB. Although regulatory T cells play an important role in the mechanism, we have recently demonstrated the active participation of host B lymphocytes. After anti-CD45RB therapy, B lymphocytes demonstrate phenotypic alterations that include up-regulation of CD54 (intercellular adhesion molecule [ICAM]-1). We have investigated the hypothesis that alteration in ICAM-1 expression is required for tolerance induction. MATERIALS AND METHODS: Recipients of heterotopic allogeneic cardiac grafts (C3H donors into B6 recipients) were treated with anti-CD45RB, anti-ICAM, anti-lymphocyte function-associated antigen-1 (LFA), or the combination of these agents. These data were extended by performing allogeneic cardiac transplants into ICAM or LFA recipients treated with a 5-day course of anti-CD45RB. Finally, B-cell-deficient animals were reconstituted with ICAM splenocytes to create a recipient with a selective deficiency of ICAM-1 restricted to the B-cell compartment. RESULTS: Anti-CD45RB alone or the combination of anti-LFA/anti-ICAM reliably induced transplantation tolerance. However, the triple combination was routinely unsuccessful and induced long-term graft survival in no recipients. ICAM-deficient or LFA-deficient recipients were also resistant to tolerance induced by anti-CD45RB. Finally, transfer of control splenocytes to B-cell-deficient recipients permitted anti-CD45RB-induced tolerance, whereas transfer of ICAM cells was unable to support tolerance induction. CONCLUSIONS: Expression of ICAM-1 by B lymphocytes and interaction with LFA-1 form a central aspect of transplantation tolerance induced by anti-CD45RB therapy. These data further elucidate the cellular mechanisms used by B lymphocytes in the induction of transplantation tolerance.
Assuntos
Linfócitos B/imunologia , Transplante de Coração/imunologia , Molécula 1 de Adesão Intercelular/fisiologia , Antígenos Comuns de Leucócito/imunologia , Antígeno-1 Associado à Função Linfocitária/fisiologia , Linfócitos T/imunologia , Tolerância ao Transplante/imunologia , Animais , Anticorpos/imunologia , Citometria de Fluxo , Molécula 1 de Adesão Intercelular/genética , Antígenos Comuns de Leucócito/genética , Antígeno-1 Associado à Função Linfocitária/genética , Camundongos , Camundongos Endogâmicos C3H , Camundongos Endogâmicos C57BL , Camundongos Knockout , Baço/imunologia , Transplante Homólogo/imunologiaRESUMO
INTRODUCTION: The relative contributions of CD4+ and CD8+ T cells to transplant rejection remain unknown. The authors integrated a previous model of CD4-mediated graft rejection with a complementary model of CD8-mediated rejection to directly compare the function of graft-reactive CD4+ and CD8+ lymphocytes in vivo in a model where rejection requires transgenic T cells. These studies allow direct comparison of CD4 and CD8 T cell responses to the same antigen without the confounding effects of T cell depletion or homeostatic proliferation. MATERIALS AND METHODS: Clone 4 and TS1 mice possess MHC class I- and II-restricted CD8+ and CD4+ T cells, respectively, which express transgenic T cell receptors that recognize the influenza hemagglutinin antigen (HA). We compared the in vivo response of CFSE-labeled, HA-specific transgenic CD8+ and CD4+ T cells after adoptive transfer into syngeneic BALB/c mice grafted with HA-expressing skin. RESULTS: As in the authors' CD4+ model, HA104 skin was consistently rejected by both Clone 4 mice (n=9, MST: 14.2) and by 5 x 10(5) Clone 4 lymphocytes transferred to naive BALB/c hosts that do not otherwise reject HA+ grafts. Rejection correlated with extensive proliferation of either graft-reactive T cell subset in the draining lymph nodes, and antigen-specific CD4+ and CD8+ cells acquired effector function and proliferated with similar kinetics. CONCLUSIONS: These data extend the authors' unique transgenic transplantation model to the investigation of CD8 T cell function. The initial results confirm fundamental functional similarity between the CD4 and CD8 T cell subsets and provide insight into the considerable redundancy underlying T cell mechanisms mediating allograft rejection.
Assuntos
Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Rejeição de Enxerto/imunologia , Transplante de Pele/imunologia , Animais , Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD8-Positivos/metabolismo , Sobrevivência de Enxerto/imunologia , Hemaglutininas/imunologia , Hemaglutininas/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Transgênicos , Receptores de Antígenos de Linfócitos T/imunologia , Receptores de Antígenos de Linfócitos T/metabolismo , Transplante Homólogo/imunologiaRESUMO
We previously demonstrated that T-regs inhibit proliferation of graft-reactive T cells in the draining lymph node (DLN), suggesting that this site may be important for regulation. TCR transgenic mice (TS1) specific for viral hemagglutinin (HA) provided antigen-specific T cells for adoptive transfer into syngeneic Balb/c hosts bearing HA+ skin grafts. T-regs were obtained from (TS1xHA28)F1 mice known to have an expanded population of HA-specific T-regs. To determine whether the lymph node is an independent site of suppression, we developed a model in which donor antigen that migrates from the allograft to the DLN drives T-cell activation after graft removal. T-regs that did not encounter the allograft itself remained able to inhibit graft antigen-specific T-cell proliferation in the DLN. Alloantigen-induced regulation can occur in the absence of the graft. This finding identifies the DLN as a potentially critical site of regulation in the early posttransplant period.
Assuntos
Linfonodos/imunologia , Linfócitos T Reguladores/imunologia , Transplante Homólogo/imunologia , Animais , Terapia de Imunossupressão , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Transgênicos , Receptores de Interleucina-2/sangue , Transplante de Pele/imunologia , Tolerância ao Transplante/imunologiaRESUMO
Selection of an optimal donor pancreas is the first key task for successful islet isolation. We conducted a retrospective multicenter study in 11 centers in North America to develop an islet donor scoring system using donor variables. The data set consisting of 1,056 deceased donors was used for development of a scoring system to predict islet isolation success (defined as postpurification islet yield >400,000 islet equivalents). With the aid of univariate logistic regression analyses, we developed the North American Islet Donor Score (NAIDS) ranging from 0 to 100 points. The c index in the development cohort was 0.73 (95% confidence interval 0.70-0.76). The success rate increased proportionally as the NAIDS increased, from 6.8% success in the NAIDS < 50 points to 53.7% success in the NAIDS ≥ 80 points. We further validated the NAIDS using a separate set of data consisting of 179 islet isolations. A comparable outcome of the NAIDS was observed in the validation cohort. The NAIDS may be a useful tool for donor pancreas selection in clinical practice. Apart from its utility in clinical decision making, the NAIDS may also be used in a research setting as a standardized measurement of pancreas quality.
Assuntos
Transplante das Ilhotas Pancreáticas/métodos , Adolescente , Adulto , Idoso , Criança , Pré-Escolar , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Pâncreas/cirurgia , Estudos Retrospectivos , Doadores de Tecidos , Adulto JovemRESUMO
Type 2 diabetic subjects manifest both disordered insulin action and abnormalities in their pancreatic islet cells. Whether the latter represents a primary defect or is a consequence of the former is unknown. To examine the beta-cell mass and function of islets from type 2 diabetic patients directly, we isolated islets from pancreata of type 2 diabetic cadaveric donors (n = 14) and compared them with islets from normal donors (n = 14) matched for age, BMI, and cold ischemia time. The total recovered islet mass from type 2 diabetic pancreata was significantly less than that from nondiabetic control subjects (256,260 islet equivalents [2,588 IEq/g pancreas] versus 597,569 islet equivalents [6,037 IEq/g pancreas]). Type 2 diabetic islets were also noted to be smaller on average, and histologically, islets from diabetic patients contained a higher proportion of glucagon-producing alpha-cells. In vitro study of islet function from diabetic patients revealed an abnormal glucose-stimulated insulin release response in perifusion assays. In addition, in comparison with normal islets, an equivalent number of type 2 diabetic islets failed to reverse hyperglycemia when transplanted to immunodeficient diabetic mice. These results provide direct evidence for abnormalities in the islets of type 2 diabetic patients that may contribute to the pathogenesis of the disease.
Assuntos
Diabetes Mellitus Tipo 2/patologia , Ilhotas Pancreáticas/patologia , Idade de Início , Índice de Massa Corporal , Diabetes Mellitus Tipo 2/fisiopatologia , Humanos , Ilhotas Pancreáticas/citologia , Ilhotas Pancreáticas/fisiologia , Transplante das Ilhotas Pancreáticas/fisiologia , Pessoa de Meia-Idade , Tamanho do Órgão , Seleção de Pacientes , Valores de Referência , Estudos RetrospectivosRESUMO
Recent improvements in isolated islet transplantation indicate that this therapy may ultimately prove applicable to patients with type I diabetes. An obstacle preventing widespread application of islet transplantation is an insufficient supply of cadaveric pancreata. Non-heart-beating donors (NHBDs) are generally not deemed suitable for whole-organ pancreas donation and could provide a significant source of pancreata for islet transplantation. Isolated pancreatic islets prepared from 10 NHBDs were compared with those procured from 10 brain-dead donors (BDDs). The success of the isolation for the two groups was analyzed for preparation purity, quality, and recovered islet mass. The function of NHBD and BDD islets was evaluated using in vitro and in vivo assays. On the basis of the results of this analysis, an NHBD isolated islet allograft was performed in a type I diabetic. The recovery of islets from NHBDs was comparable to that of control BDDs. In vitro assessment of NHBD islet function revealed function-equivalent BDD islets, and NHBD islets transplanted to non-obese diabetic-severe combined immunodeficient (NOD-SCID) mice efficiently reversed diabetes. Transplantation of 446,320 islet equivalents (IEq) (8,500 IEq/kg of recipient body weight) from a single NHBD successfully reversed the diabetes of a type I diabetic recipient. Normally functioning pancreatic islets can be isolated successfully from NHBDs. A single donor transplant from an NHBD resulted in a state of stable insulin independence in a type I diabetic recipient. These results indicate that NHBDs may provide an as yet untapped source of pancreatic tissue for preparation of isolated islets for clinical transplantation.
Assuntos
Transplante das Ilhotas Pancreáticas/métodos , Doadores de Tecidos , Adulto , Animais , Morte Encefálica , Cadáver , Diabetes Mellitus Tipo 1/fisiopatologia , Diabetes Mellitus Tipo 1/terapia , Humanos , Ilhotas Pancreáticas/fisiologia , Camundongos , Camundongos SCID , Pessoa de Meia-IdadeRESUMO
BACKGROUND: Examination of the in vivo activation and function of CD4+ T cells in response to allografts may advance our understanding of the rejection process. We analyzed the capacity of transgenic class II-restricted CD4 T cells to reject skin, cardiac, and islet transplants. METHODS: TS1 mice possess a high frequency of CD4+ T cells specific for the immunodominant epitope of the viral hemagglutinin (HA) protein. We analyzed the kinetics of rejection of skin, heart, and islet grafts by naïve and sensitized TS1 mice and by adoptively transferred TS1 lymphocytes. RESULTS: Rejection of heart transplants was more rapid than skin grafts (mean survival time, 12.9 vs. 26.6 days), and islet grafts survived indefinitely in TS1 mice. These findings may be partly attributable to the supranormal frequency of HA-reactive cells in TS1 mice. In support of this, we found that adoptive transfer of 5 x 10(5) TS1 lymphocytes to Balb/c hosts effected consistent rejection of HA-bearing skin transplants, whereas a significantly greater number (3 x 10(6)) was required for heart transplant rejection. The in vivo proliferative response of HA-specific T cells to heart and skin was found to be robust and predominantly localized to the draining lymph nodes. CONCLUSION: We developed a model of allograft rejection in which the responding T cells and relevant graft antigen are specifically defined. Adoptive transfer of carboxy-fluorescein succinimidyl ester-labeled transgenic T cells allowed us to visualize a robust proliferative response in vivo to heart and skin allografts, which in both cases was localized to regional lymph nodes.
Assuntos
Rejeição de Enxerto/etiologia , Transplante de Coração , Antígenos de Histocompatibilidade Classe II/metabolismo , Transplante das Ilhotas Pancreáticas , Receptores de Antígenos de Linfócitos T/metabolismo , Transplante de Pele , Transferência Adotiva , Animais , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Divisão Celular , Suscetibilidade a Doenças , Hemaglutininas Virais/metabolismo , Memória Imunológica , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Transgênicos , Linfócitos T/patologia , Linfócitos T/transplante , Transplante HomólogoRESUMO
BACKGROUND: Isolated islet transplantation with infusions from two to three donor pancreata and Edmonton immunosuppression consistently achieves insulin independence in patients with type 1 diabetes. The success of this protocol has been attributed to a novel combination of immunosuppressive agents and avoidance of steroids; however, the outcome of islet transplantation may differ in kidney transplant recipients who are already immunosuppressed. METHODS: We compared the metabolic outcomes and graft survival of islet transplantation in our program where nine patients underwent islet transplantation alone treated with Edmonton immunosuppression and eight patients received islet after kidney (IAK) transplants under standard kidney transplant immunosuppression often including steroids. RESULTS: Transplants in the IAK and islet transplantation alone setting demonstrated similar islet potency (islet equivalents/unit insulin reduction) and recipients from both groups routinely gained insulin independence, functional islet mass, and duration of graft survival, however, seemed superior in the IAK group. CONCLUSIONS: These results suggest that better islet graft function and survival may be attained using non-Edmonton rather than Edmonton immunosuppression and can include maintenance steroid therapy.
Assuntos
Transplante das Ilhotas Pancreáticas/métodos , Transplante das Ilhotas Pancreáticas/fisiologia , Transplante de Rim/métodos , Adulto , Glicemia/metabolismo , Peptídeo C/sangue , Diabetes Mellitus Tipo 1/tratamento farmacológico , Diabetes Mellitus Tipo 1/metabolismo , Diabetes Mellitus Tipo 1/cirurgia , Feminino , Hemoglobinas Glicadas/metabolismo , Sobrevivência de Enxerto/fisiologia , Humanos , Terapia de Imunossupressão/métodos , Insulina/administração & dosagem , Masculino , Pessoa de Meia-Idade , Fatores de TempoRESUMO
BACKGROUND: Many models of transplant tolerance have been found to depend on the induction of regulatory T cells (Tregs). Innate immune signals are known to suppress Tregs thereby augmenting immunity by abrogating Treg function. Such signals may also provide a barrier to transplantation tolerance mediated by Tregs. A number of cell surface molecules expressed by Tregs have been found to inhibit Treg activity, the best characterized of which is the glucocorticoid-induced tumor necrosis factor receptor-related (GITR) protein. METHODS: By using an adoptive transfer model of allograft rejection, we can study the effects of inflammation and antigen-specific Tregs on graft survival. Inflammation resulting from the transplant procedure counter-regulates the suppressor activity of Tregs. To assess whether Treg activity could be enhanced by blocking GITR signaling, we compared the capacity of Tregs to prolong the survival of grafts in the presence or absence of activation-inducible TNF receptor (AITRL)-Fc, a novel construct that binds GITR. RESULTS: We report that interruption of GITR-GITR ligand (GITRL) binding by AITRL-Fc resulted in long-term Treg-dependent acceptance of skin grafts in the setting of innate immune signals that otherwise interfere with Treg activity. CONCLUSIONS: Inflammation and other innate immune signals may activate antigen presenting cells to upregulate GITRL. GITR-GITRL interaction is one pathway by which antigen presenting cells may enhance the adaptive response to foreign antigen by counter-regulating Tregs and by costimulating effector T cells. By blocking this interaction with AITRL-Fc, one can sustain the benefit conferred by graft-protective Tregs.
Assuntos
Sobrevivência de Enxerto/imunologia , Receptores de Fator de Crescimento Neural/antagonistas & inibidores , Receptores do Fator de Necrose Tumoral/antagonistas & inibidores , Transplante de Pele/imunologia , Linfócitos T Reguladores/imunologia , Transplante Homólogo/imunologia , Animais , Linfócitos T CD4-Positivos/imunologia , Cruzamentos Genéticos , Epitopos/imunologia , Citometria de Fluxo , Proteína Relacionada a TNFR Induzida por Glucocorticoide , Humanos , Terapia de Imunossupressão , Inflamação/imunologia , Linfócitos/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Transgênicos , Receptores de Fator de Crescimento Neural/fisiologia , Receptores do Fator de Necrose Tumoral/fisiologia , Transdução de Sinais/imunologia , Transdução de Sinais/fisiologia , Transplante de Pele/mortalidade , Taxa de Sobrevida , Fatores de Necrose Tumoral/imunologiaRESUMO
Selective interference with the CD45RB isoform by mAb (anti-CD45RB) reliably induces donor-specific tolerance. Although previous studies suggest participation of regulatory T cells, a mechanistic understanding of anti-CD45RB-induced tolerance is lacking. We report herein the unexpected finding that tolerance induced by this agent is not established in B cell-deficient mice but can be recovered by preemptive B lymphocyte transfer to B cell-deficient hosts. Using B cells from genetically modified donors to reconstitute B cell-deficient recipients, we evaluate the role of B lymphocyte-expressed CD45RB, T cell costimulatory molecules, and the production of Abs in this novel tolerance mechanism. Our data document an Ab-induced tolerance regimen that is uniquely B lymphocyte-dependent and suggest mechanistic contributions to tolerance development from the B cell compartment through interactions with T cells.
Assuntos
Anticorpos Monoclonais/administração & dosagem , Subpopulações de Linfócitos B/imunologia , Transplante de Coração/imunologia , Antígenos Comuns de Leucócito/imunologia , Tolerância ao Transplante/imunologia , Animais , Anticorpos Monoclonais/fisiologia , Subpopulações de Linfócitos B/citologia , Subpopulações de Linfócitos B/transplante , Comunicação Celular/genética , Comunicação Celular/imunologia , Proliferação de Células , Imunofenotipagem , Antígenos Comuns de Leucócito/biossíntese , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C3H , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/metabolismo , Tolerância ao Transplante/genéticaRESUMO
Targeting of the CD45RB isoform by mAb (anti-CD45RB) effectively induces donor-specific tolerance to allografts. The immunological mechanisms underlying the tolerant state remain unclear although some studies have suggested the involvement of regulatory T cells (T-regs). Although their generative pathway remains undefined, tolerance promoting T-regs induced by systemic anti-CD45RB treatment have been assumed to originate in the peripheral immune system. We demonstrate herein that separable effects on the peripheral and central immune compartments mediate graft survival induced by anti-CD45RB administration. In the absence of the thymus, anti-CD45RB therapy is not tolerogenic though it retains peripheral immunosuppressive activity. The thymus is required for anti-CD45RB to produce indefinite graft survival and donor-specific tolerance, and this effect is accomplished through thymic production of donor-specific T-regs. These data reveal for the first time an Ab-based tolerance regimen that relies on the central tolerance pathway.
Assuntos
Anticorpos Monoclonais/administração & dosagem , Rejeição de Enxerto/imunologia , Linfócitos T Reguladores/imunologia , Timo/citologia , Timo/imunologia , Tolerância ao Transplante/imunologia , Transferência Adotiva , Animais , Diferenciação Celular/imunologia , Células Cultivadas , Rejeição de Enxerto/prevenção & controle , Transplante de Coração/imunologia , Injeções Intraperitoneais , Antígenos Comuns de Leucócito/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C3H , Camundongos Endogâmicos C57BL , Camundongos SCID , Camundongos TransgênicosRESUMO
Transplantation of allogeneic islets can correct the metabolic abnormalities of Type I diabetes. Limited availability of donor pancreas tissues restricts the application of this therapeutic modality to a subset of eligible recipients. In an attempt to expand the utility of available donor human pancreas tissue, we developed a method to stimulate the proliferation of insulin-secreting beta-cells within human islets. A lentiviral vector was used to introduce into human islets chimeric signaling receptors that are activated to stimulate cell proliferation through interactions with a small-molecule drug called a chemical inducer of dimerization (CID). In vitro exposure of vector-transduced human islets to the CID expanded the number of cells and increased regulated insulin secretion. Transplantation of the regenerated islets into diabetic immunodeficient mice, followed by in vivo administration of the CID, corrected hyperglycemia. This strategy has the potential to reduce the quantity of human islets required for treatment of patients with Type I diabetes.
Assuntos
Ilhotas Pancreáticas/efeitos dos fármacos , Ilhotas Pancreáticas/patologia , Regeneração/efeitos dos fármacos , Animais , Glicemia/metabolismo , Proliferação de Células/efeitos dos fármacos , Diabetes Mellitus/sangue , Diabetes Mellitus/patologia , Diabetes Mellitus/terapia , Dimerização , Humanos , Ilhotas Pancreáticas/metabolismo , Transplante das Ilhotas Pancreáticas , Camundongos , Técnicas de Cultura de Órgãos , Fatores de Tempo , Transplante HeterólogoRESUMO
While great advances have been made in the success of islet transplantation to cure autoimmune diabetes, this protocol remains limited by our inability to induce donor-specific tolerance within the recipient. The profound resistance of the NOD mouse to tolerance-inducing regimens that are routinely successful in other strains further defines the imposing barriers that must be surmounted. Herein, we have assessed the utility of anti-CD45RB therapy to induce tolerance to allografts in C57BL/6 and NOD-strain mice. We find that, as with other therapies, NOD mice are also resistant to this manipulation, despite robust tolerance induction in the comparison strain. Analysis of cell surface markers revealed a number of changes within the B lymphocyte compartment following contact with antibody and alloantigen in the B6 strain. The absence of reciprocal changes within the NOD lymphocyte compartment suggests that B cells might contribute to the mechanism of action of this therapy and to the resistance to immunological tolerance noted in the NOD strain.
Assuntos
Sobrevivência de Enxerto/imunologia , Tolerância Imunológica , Transplante das Ilhotas Pancreáticas/imunologia , Antígenos Comuns de Leucócito/imunologia , Animais , Anticorpos/uso terapêutico , Diabetes Mellitus Tipo 1/cirurgia , Imunossupressores/uso terapêutico , Transplante das Ilhotas Pancreáticas/patologia , Camundongos , Camundongos Endogâmicos C3H , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos NOD , Modelos Animais , Proteína Tirosina Fosfatase não Receptora Tipo 1 , Especificidade da Espécie , Fatores de Tempo , Transplante Homólogo/imunologiaRESUMO
Regulatory T cells preserve tolerance to peripheral self-Ags and may control the response to allogeneic tissues to promote transplantation tolerance. Although prior studies have demonstrated prolonged allograft survival in the presence of regulatory T cells (T-reg), data documenting the capacity of these cells to promote tolerance in immunocompetent transplant models are lacking, and the mechanism of suppression in vivo remains unclear. We used a TCR transgenic model of allograft rejection to characterize the in vivo activity of CD4(+)CD25(+) T-reg. We demonstrate that graft Ag-specific T-reg effectively intercede in the rejection response of naive T cells to established skin allografts. Furthermore, CFSE labeling demonstrates impaired proliferation of naive graft Ag-specific T cells in the draining lymph node in the presence of T-reg. These results confirm the efficacy of T-reg in promoting graft survival and suggest that their suppressive action is accomplished in part through inhibition of proliferation.
Assuntos
Antígenos CD4/análise , Sobrevivência de Enxerto , Ativação Linfocitária , Receptores de Interleucina-2/análise , Subpopulações de Linfócitos T/imunologia , Animais , Antígenos CD , Antígenos de Diferenciação/fisiologia , Antígeno CTLA-4 , Rejeição de Enxerto/prevenção & controle , Interleucina-10/fisiologia , Camundongos , Camundongos Endogâmicos BALB C , Transplante de Pele , Fator de Crescimento Transformador beta/fisiologia , Transplante HomólogoRESUMO
Transplantation tolerance is induced reliably in experimental animals following intrathymic inoculation with the relevant donor strain Ags; however, the immunological mechanisms responsible for the induction and maintenance of the tolerant state remain unknown. We investigated these mechanisms using TCR transgenic mice (TS1) that carry T cells specific for an immunodominant, MHC class II-restricted peptide (S1) of the influenza PR8 hemagglutinin (HA) molecule. We demonstrated that TS1 mice reject skin grafts that have transgene-encoded HA molecules (HA104) as their sole antigenic disparity and that intrathymic but not i.v. inoculation of TS1 mice with S1 peptide induces tolerance to HA-expressing skin grafts. Intrathymic peptide inoculation was associated with a dose-dependent reduction in T cells bearing high levels of TCR specific for HA. However, this reduction was both incomplete and transient, with a full recovery of S1-specific thymocytes by 4 wk. Peptide inoculation into the thymus also resulted in the generation of immunoregulatory T cells (CD4+CD25+) that migrated to the peripheral lymphoid organs. Adoptive transfer experiments using FACS sorted CD4+CD25- and CD4+CD25+ T cells from tolerant mice revealed that the former but not the latter maintain the capacity to induce rejection of HA bearing skin allografts in syngeneic hosts. Our results suggest that both clonal frequency reduction in the thymus and immunoregulatory T cells exported from the thymus are critical to transplantation tolerance induced by intrathymic Ag inoculation.