RESUMO
Alloreactive memory T cells have been implicated as central drivers of transplant rejection. Perplexingly, innate cytokines, such as IL-6, IL-1ß, and IL-12, are also associated with rejection of organ transplants. However, the pathways of innate immune activation in allogeneic transplantation are unclear. While the role of microbial and cell death products has been previously described, we identified alloreactive memory CD4 T cells as the primary triggers of innate inflammation. Memory CD4 T cells engaged MHC II-mismatched dendritic cells (DCs), leading to the production of innate inflammatory cytokines. This innate inflammation was independent of several pattern recognition receptors and was primarily driven by TNF superfamily ligands expressed by alloreactive memory CD4 T cells. Blocking of CD40L and TNFα resulted in dampened inflammation, and mice genetically deficient in these molecules exhibited prolonged survival of cardiac allografts. Furthermore, myeloid cell and CD8 T cell infiltration into cardiac transplants was compromised in both CD40L- and TNFα-deficient recipients. Strikingly, we found that priming of naive alloreactive CD8 T cells was dependent on licensing of DCs by memory CD4 T cells. This study unravels the key mechanisms by which alloreactive memory CD4 T cells contribute to destructive pathology and transplant rejection.
Assuntos
Linfócitos T CD4-Positivos , Linfócitos T CD8-Positivos , Células Dendríticas , Rejeição de Enxerto , Transplante de Coração , Imunidade Inata , Inflamação , Animais , Rejeição de Enxerto/imunologia , Camundongos , Células Dendríticas/imunologia , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD4-Positivos/imunologia , Inflamação/imunologia , Imunidade Inata/imunologia , Camundongos Endogâmicos C57BL , Ligante de CD40/imunologia , Ligante de CD40/metabolismo , Células T de Memória/imunologia , Camundongos Knockout , Fator de Necrose Tumoral alfa/metabolismo , Fator de Necrose Tumoral alfa/imunologia , Citocinas/metabolismo , Citocinas/imunologiaRESUMO
Antigen-activated lymphocytes undergo extraordinarily rapid cell division in the course of immune responses. We hypothesized that this unique aspect of lymphocyte biology leads to unusual genomic stress in recently antigen-activated lymphocytes and that targeted manipulation of DNA damage-response (DDR) signaling pathways would allow for selective therapeutic targeting of pathological T cells in disease contexts. Consistent with these hypotheses, we found that activated mouse and human T cells display a pronounced DDR in vitro and in vivo. Upon screening a variety of small-molecule compounds, we found that potentiation of p53 (via inhibition of MDM2) or impairment of cell cycle checkpoints (via inhibition of CHK1/2 or WEE1) led to the selective elimination of activated, pathological T cells in vivo. The combination of these strategies [which we termed "p53 potentiation with checkpoint abrogation" (PPCA)] displayed therapeutic benefits in preclinical disease models of hemophagocytic lymphohistiocytosis and multiple sclerosis, which are driven by foreign antigens or self-antigens, respectively. PPCA therapy targeted pathological T cells but did not compromise naive, regulatory, or quiescent memory T-cell pools, and had a modest nonimmune toxicity profile. Thus, PPCA is a therapeutic modality for selective, antigen-specific immune modulation with significant translational potential for diverse immune-mediated diseases.
Assuntos
Dano ao DNA/imunologia , Doenças do Sistema Imunitário/terapia , Animais , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Pontos de Checagem do Ciclo Celular/imunologia , Modelos Animais de Doenças , Avaliação Pré-Clínica de Medicamentos , Encefalomielite Autoimune Experimental/imunologia , Encefalomielite Autoimune Experimental/terapia , Etoposídeo/administração & dosagem , Humanos , Doenças do Sistema Imunitário/imunologia , Ativação Linfocitária , Linfo-Histiocitose Hemofagocítica/imunologia , Linfo-Histiocitose Hemofagocítica/terapia , Camundongos , Camundongos da Linhagem 129 , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Esclerose Múltipla/imunologia , Esclerose Múltipla/terapia , Proteínas Proto-Oncogênicas c-mdm2/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-mdm2/imunologia , Transdução de Sinais/imunologia , Linfócitos T/efeitos dos fármacos , Linfócitos T/imunologia , Linfócitos T/patologia , Proteína Supressora de Tumor p53/imunologiaRESUMO
The current clinical approach for treating autoimmune diseases is to broadly blunt immune responses as a means of preventing autoimmune pathology. Among the major side effects of this strategy are depressed beneficial immunity and increased rates of infections and tumors. Using the experimental autoimmune encephalomyelitis model for human multiple sclerosis, we report a novel alternative approach for purging autoreactive T cells that spares beneficial immunity. The moderate and temporally limited use of etoposide, a topoisomerase inhibitor, to eliminate encephalitogenic T cells significantly reduces the onset and severity of experimental autoimmune encephalomyelitis, dampens cytokine production and overall pathology, while dramatically limiting the off-target effects on naive and memory adaptive immunity. Etoposide-treated mice show no or significantly ameliorated pathology with reduced antigenic spread, yet have normal T cell and T-dependent B cell responses to de novo antigenic challenges as well as unimpaired memory T cell responses to viral rechallenge. Thus, etoposide therapy can selectively ablate effector T cells and limit pathology in an animal model of autoimmunity while sparing protective immune responses. This strategy could lead to novel approaches for the treatment of autoimmune diseases with both enhanced efficacy and decreased treatment-associated morbidities.
Assuntos
Encefalomielite Autoimune Experimental/imunologia , Depleção Linfocítica , Linfócitos T/imunologia , Animais , Apoptose/efeitos dos fármacos , Apoptose/imunologia , Linfócitos B/imunologia , Linfócitos T CD4-Positivos/efeitos dos fármacos , Linfócitos T CD4-Positivos/imunologia , Progressão da Doença , Encefalomielite Autoimune Experimental/tratamento farmacológico , Encefalomielite Autoimune Experimental/patologia , Etoposídeo/administração & dosagem , Etoposídeo/farmacologia , Feminino , Memória Imunológica , Ativação Linfocitária/efeitos dos fármacos , Vírus da Coriomeningite Linfocítica/imunologia , Camundongos , Esclerose Múltipla/tratamento farmacológico , Esclerose Múltipla/imunologia , Esclerose Múltipla/patologia , Recidiva , Linfócitos T/efeitos dos fármacosRESUMO
Preserving endogenous insulin production is clinically advantageous and remains a vital unmet challenge in the treatment and reversal of type 1 diabetes. Although broad immunosuppression has had limited success in prolonging the so-called remission period, it comes at the cost of compromising beneficial immunity. Here, we used a novel strategy to specifically deplete the activated diabetogenic T cells that drive pathogenesis while preserving not only endogenous insulin production but also protective immunity. Effector T (Teff) cells, such as diabetogenic T cells, are naturally poised on the edge of apoptosis because of activation-induced DNA damage that stresses the p53 regulation of the cell cycle. We have found that using small molecular inhibitors that further potentiate p53 while inhibiting the G2/M cell cycle checkpoint control drives apoptosis of activated T cells in vivo. When delivered at the onset of disease, these inhibitors significantly reduce diabetogenic Teff cells, prolong remission, preserve functional islets, and protect islet allografts while leaving naive, memory, and regulatory T-cell populations functionally untouched. Thus, the targeted manipulation of p53 and cell cycle checkpoints represents a new therapeutic modality for the preservation of islet ß-cells in new-onset type 1 diabetes or after islet transplant.
Assuntos
Diabetes Mellitus Tipo 1/tratamento farmacológico , Inibidores Enzimáticos/uso terapêutico , Pâncreas/imunologia , Linfócitos T/imunologia , Animais , Diabetes Mellitus Tipo 1/imunologia , Inibidores Enzimáticos/farmacologia , Imidazóis/farmacologia , Imidazóis/uso terapêutico , Camundongos , Camundongos Endogâmicos NOD , Pâncreas/efeitos dos fármacos , Piperazinas/farmacologia , Piperazinas/uso terapêutico , Pirazóis/farmacologia , Pirazóis/uso terapêutico , Pirimidinas/farmacologia , Pirimidinas/uso terapêutico , Pirimidinonas , Linfócitos T/efeitos dos fármacos , Tiofenos/farmacologia , Tiofenos/uso terapêutico , Ureia/análogos & derivados , Ureia/farmacologia , Ureia/uso terapêuticoRESUMO
Glucagon-like peptide-1 (GLP-1) is an intestinal hormone that is secreted during meal absorption and is essential for normal glucose homeostasis. However, the relatively low plasma levels and rapid metabolism of GLP-1 raise questions as to whether direct endocrine action on target organs, such as islet cells, account for all of its effects on glucose tolerance. Recently, an alternative neural pathway initiated by sensors in the hepatic portal region has been proposed to mediate GLP-1 activity. We hypothesized that visceral afferent neurons in the portal bed express the GLP-1 receptor (GLP-1r) and regulate glucose tolerance. Consistent with this hypothesis, GLP-1r mRNA was present in the nodose ganglia, and nerve terminals innervating the portal vein contained the GLP-1r. Rats given an intraportal infusion of the GLP-1r antagonist, [des-His(1),Glu(9)] exendin-4, in a low dose, had glucose intolerance, with a 53% higher glucose excursion compared with a vehicle-infused control group. Infusion of [des-His(1),Glu(9)] exendin-4 at an identical rate into the jugular vein had no effect on glucose tolerance, demonstrating that this dose of GLP-1r antagonist did not affect blood glucose due to spillover into the systemic circulation. These studies demonstrate that GLP-1r are present on nerve terminals in the hepatic portal bed and that GLP-1 antagonism localized to this region impairs glucose tolerance. These data are consistent with an important component of neural mediation of GLP-1 action.
Assuntos
Peptídeo 1 Semelhante ao Glucagon/fisiologia , Glucose/fisiologia , Terminações Nervosas/metabolismo , Veia Porta/inervação , Receptores de Glucagon/fisiologia , Animais , Glicemia/efeitos dos fármacos , Glicemia/metabolismo , Linhagem Celular , Relação Dose-Resposta a Droga , Exenatida , Receptor do Peptídeo Semelhante ao Glucagon 1 , Intolerância à Glucose/induzido quimicamente , Teste de Tolerância a Glucose , Infusões Intravenosas , Fígado/irrigação sanguínea , Masculino , Gânglio Nodoso/metabolismo , Peptídeos/administração & dosagem , Peptídeos/farmacologia , Veia Porta/efeitos dos fármacos , Veia Porta/metabolismo , Ratos , Ratos Sprague-Dawley , Receptores de Glucagon/antagonistas & inibidores , Receptores de Glucagon/genética , Receptores de Glucagon/metabolismo , Peçonhas/administração & dosagem , Peçonhas/farmacologiaRESUMO
A frequently debated question for studies involving the measurement of stress hormones in rodents is the optimal method for collecting blood with minimal stress to the animal. Some investigators prefer the implantation of indwelling catheters to allow for frequent sampling. Others argue that the implantation of a catheter creates a chronic stress to the animal that confounds stress hormone measures and therefore rely on tail vein sampling. Moreover, some investigators measure hormones in trunk blood samples obtained after anesthesia, a practice that may itself raise hormone levels. To address these controversies, we 1) compared plasma ACTH and corticosterone (Cort) concentrations in pre- and poststress rat blood samples obtained via previously implanted vena cava catheters, tail vein nicks, or clipping the tip off the tail and 2) compared plasma ACTH and Cort in rat blood samples obtained by decapitation with and without anesthesia. Rats sampled via indwelling catheters displayed lower prestress ACTH levels than those sampled by tail vein nick if the time to acquire samples was not limited; however, elevated basal ACTH was not observed in samples obtained by tail clip or tail nick when the samples were obtained within 3 min. Baseline Cort levels were similar in all groups. After restraint stress, the profile of the plasma ACTH and Cort responses was not affected by sampling method. Decapitation with prior administration of CO2 or pentobarbital sodium increased plasma ACTH levels approximately 13- and 2-fold, respectively, when compared with decapitation without anesthesia. These data indicate that tail vein nicking, tail clipping, or indwelling venous catheters can be used for obtaining plasma for ACTH and Cort during acute stress studies without confounding the measurements. However, the elevation in basal ACTH seen in the tail vein nick group at baseline suggests that sampling needs to be completed rapidly (<3 min) to avoid the initiation of the pituitary stress response. Death by CO2 and pentobarbital sodium injection before trunk blood collection cause significant stress to animals, as reflected in the elevated plasma ACTH levels. These results support the use of either chronic vascular cannulas or sampling from a tail vein. However, collection of blood under pentobarbital sodium or CO2 anesthesia is likely to confound the results of stress studies when ACTH is an important endpoint.