RESUMO
Understanding the factors that impede immune responses to persistent viruses is essential in designing therapies for HIV infection. Mice infected with LCMV clone-13 have persistent high-level viremia and a dysfunctional immune response. Interleukin-7, a cytokine that is critical for immune development and homeostasis, was used here to promote immunity toward clone-13, enabling elucidation of the inhibitory pathways underlying impaired antiviral immune response. Mechanistically, IL-7 downregulated a critical repressor of cytokine signaling, Socs3, resulting in amplified cytokine production, increased T cell effector function and numbers, and viral clearance. IL-7 enhanced thymic output to expand the naive T cell pool, including T cells that were not LCMV specific. Additionally, IL-7 promoted production of cytoprotective IL-22 that abrogated liver pathology. The IL-7-mediated effects were dependent on endogenous IL-6. These attributes of IL-7 have profound implications for its use as a therapeutic in the treatment of chronic viral diseases.
Assuntos
Interleucina-7/uso terapêutico , Coriomeningite Linfocítica/imunologia , Vírus da Coriomeningite Linfocítica/fisiologia , Animais , Antígenos de Diferenciação/metabolismo , Regulação para Baixo , Fatores de Transcrição Forkhead/metabolismo , Humanos , Interleucina-6/imunologia , Interleucina-7/imunologia , Camundongos , Receptor de Morte Celular Programada 1 , Proteínas Recombinantes/metabolismo , Proteína 3 Supressora da Sinalização de Citocinas , Proteínas Supressoras da Sinalização de Citocina/metabolismo , Linfócitos T/imunologiaRESUMO
BACKGROUND: A major challenge to adeno-associated virus (AAV)-mediated gene therapy is the presence of anti-AAV capsid neutralizing antibodies (NAbs), which can block viral vector transduction even at very low titers. In the present study, we examined the ability of a combination immunosuppression (IS) treatment with bortezomib and a mouse-specific CD20 monoclonal antibody to suppress anti-AAV NAbs and enable readministration of AAV vectors of the same capsid in mice. METHODS: An AAV8 vector (AAV8-CB-hGAA) that ubiquitously expresses human α-glucosidase was used for initial gene therapy and a second AAV8 vector (AAV8-LSP-hSEAP) that contains a liver-specific promoter to express human secreted embryonic alkaline phosphatase (hSEAP) was used for AAV readministration. Plasma samples were used for determination of anti-AAV8 NAb titers. Cells isolated from whole blood, spleen, and bone marrow were analyzed for B-cell depletion by flow cytometry. The efficiency of AAV readministration was determined by the secretion of hSEAP in blood. RESULTS: In näive mice, an 8-week IS treatment along with AAV8-CB-hGAA injection effectively depleted CD19+ B220+ B cells from blood, spleen, and bone marrow and prevented the formation of anti-AAV8 NAbs. Following administration of AAV8-LSP-hSEAP, increasing levels of hSEAP were detected in blood for up to 6 weeks, indicating successful AAV readministration. In mice pre-immunized with AAV8-CB-hGAA, comparison of IS treatment for 8, 12, 16, and 20 weeks revealed that the 16-week IS treatment demonstrated the highest plasma hSEAP level following AAV8-LSP-hSEAP readministration. CONCLUSIONS: Our data suggest that this combination treatment is an effective IS approach that will allow retreatment of patients with AAV-mediated gene therapy. A combination IS treatment with bortezomib and a mouse-specific CD20 monoclonal antibody effectively suppressed anti-AAV NAbs in naïve mice and in mice with pre-existing antibodies, allowing successful readministration of the same AAV capsid vector.
Assuntos
Anticorpos Neutralizantes , Doença de Depósito de Glicogênio Tipo II , Humanos , Camundongos , Animais , Bortezomib/farmacologia , Bortezomib/uso terapêutico , Capsídeo , Anticorpos Antivirais , Vetores Genéticos/genética , Retratamento , Anticorpos Monoclonais/farmacologia , Anticorpos Monoclonais/uso terapêutico , Dependovirus/genéticaRESUMO
Non-Hodgkin lymphoma (NHL) is the most commonly diagnosed hematologic cancer of adults in the United States, with the vast majority of NHLs deriving from malignant B lymphocytes that express cell surface CD20. CD20 immunotherapy (rituximab) is widely used to treat NHL, even though the initial effectiveness of rituximab varies widely among patients and typically wanes over time. The mechanisms through which lymphomas initially resist or gain resistance to immunotherapy are not well established. To address this, a preclinical mouse model system was developed to comprehensively identify lymphoma transcriptomic changes that confer resistance to CD20 immunotherapy. The generation of spontaneous primary and familial lymphomas revealed that sensitivity to CD20 immunotherapy was not regulated by differences in CD20 expression, prior exposure to CD20 immunotherapy, or serial in vivo passage. An unbiased forward exome screen of these primary lymphomas was used to validate the utility of this expansive lymphoma cohort, which revealed that increased lymphoma galectin-1 (Gal-1) expression strongly correlated with resistance to immunotherapy. Genetically induced lymphoma Gal-1 expression ablated antibody-dependent lymphoma phagocytosis in vitro and lymphoma sensitivity to CD20 immunotherapy in vivo. Human NHLs also express elevated Gal-1 compared with nonmalignant lymphocytes, demonstrating the ability of this preclinical model system to identify molecular targets that could be relevant to human therapy. This study therefore established a powerful preclinical model system that permits the comprehensive identification of the dynamic lymphoma molecular network that drives resistance to immunotherapy.
Assuntos
Antígenos CD20/genética , Resistencia a Medicamentos Antineoplásicos , Galectina 1/fisiologia , Imunoterapia/métodos , Linfoma de Células B/imunologia , Animais , Antineoplásicos/uso terapêutico , Modelos Animais de Doenças , Proteínas de Fluorescência Verde/metabolismo , Hemizigoto , Humanos , Macrófagos/citologia , Camundongos , Camundongos Endogâmicos C57BL , Microscopia de Fluorescência , Fagocitose , Rituximab/uso terapêuticoRESUMO
B cells regulate immune responses by producing antigen-specific antibodies. However, specific B-cell subsets can also negatively regulate T-cell immune responses, and have been termed regulatory B cells. Human and mouse regulatory B cells (B10 cells) with the ability to express the inhibitory cytokine interleukin-10 (IL-10) have been identified. Although rare, B10 cells are potent negative regulators of antigen-specific inflammation and T-cell-dependent autoimmune diseases in mice. How B10-cell IL-10 production and regulation of antigen-specific immune responses are controlled in vivo without inducing systemic immunosuppression is unknown. Using a mouse model for multiple sclerosis, here we show that B10-cell maturation into functional IL-10-secreting effector cells that inhibit in vivo autoimmune disease requires IL-21 and CD40-dependent cognate interactions with T cells. Moreover, the ex vivo provision of CD40 and IL-21 receptor signals can drive B10-cell development and expansion by four-million-fold, and generate B10 effector cells producing IL-10 that markedly inhibit disease symptoms when transferred into mice with established autoimmune disease. The ex vivo expansion and reinfusion of autologous B10 cells may provide a novel and effective in vivo treatment for severe autoimmune diseases that are resistant to current therapies.
Assuntos
Autoimunidade/imunologia , Linfócitos B Reguladores/imunologia , Interleucinas/imunologia , Linfócitos T/imunologia , Animais , Antígenos CD19/genética , Antígenos CD19/metabolismo , Linfócitos B Reguladores/citologia , Linfócitos B Reguladores/metabolismo , Antígenos CD40/imunologia , Antígenos CD40/metabolismo , Antígenos CD5/metabolismo , Divisão Celular , Modelos Animais de Doenças , Encefalomielite Autoimune Experimental/imunologia , Encefalomielite Autoimune Experimental/patologia , Feminino , Antígenos de Histocompatibilidade Classe II/imunologia , Humanos , Interleucina-10/biossíntese , Interleucina-10/imunologia , Interleucina-10/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Esclerose Múltipla/imunologia , Esclerose Múltipla/patologia , Receptores de Interleucina-21/imunologia , Receptores de Interleucina-21/metabolismoRESUMO
L-selectin is a cell adhesion molecule that tethers free-flowing leukocytes from the blood to luminal vessel walls, facilitating the initial stages of their emigration from the circulation toward an extravascular inflammatory insult. Following shear-resistant adhesion to the vessel wall, L-selectin has frequently been reported to be rapidly cleaved from the plasma membrane (known as ectodomain shedding), with little knowledge of the timing or functional consequence of this event. Using advanced imaging techniques, we observe L-selectin shedding occurring exclusively as primary human monocytes actively engage in transendothelial migration (TEM). Moreover, the shedding was localized to transmigrating pseudopods within the subendothelial space. By capturing monocytes in midtransmigration, we could monitor the subcellular distribution of L-selectin and better understand how ectodomain shedding might contribute to TEM. Mechanistically, L-selectin loses association with calmodulin (CaM; a negative regulator of shedding) specifically within transmigrating pseudopods. In contrast, L-selectin/CaM interaction remained intact in nontransmigrated regions of monocytes. We show phosphorylation of L-selectin at Ser 364 is critical for CaM dissociation, which is also restricted to the transmigrating pseudopod. Pharmacological or genetic inhibition of L-selectin shedding significantly increased pseudopodial extensions in transmigrating monocytes, which potentiated invasive behavior during TEM and prevented the establishment of front/back polarity for directional migration persistence once TEM was complete. We conclude that L-selectin shedding directly regulates polarity in transmigrated monocytes, which affirms an active role for this molecule in driving later stages of the multistep adhesion cascade.
Assuntos
Polaridade Celular , Selectina L/metabolismo , Monócitos/citologia , Sequência de Aminoácidos , Adesão Celular , Movimento Celular , Citoplasma/metabolismo , Transferência Ressonante de Energia de Fluorescência , Proteínas de Fluorescência Verde/metabolismo , Células Endoteliais da Veia Umbilical Humana , Humanos , Inflamação , Leucócitos/metabolismo , Microscopia Eletrônica de Transmissão , Microscopia de Vídeo , Dados de Sequência Molecular , Monócitos/metabolismo , Fosforilação , Serina/químicaRESUMO
While B cells are traditionally regarded as promoters of the immune response via antibody secretion and pro-inflammatory cytokine production, recent studies have also confirmed an important role for B-cell-mediated negative regulation of immunity. Tremendous advances in the characterization of the mechanisms by which regulatory B cells function has led to the identification of a novel subset of regulatory B cells known as B10 cells, which regulate immune responses through the production of the anti-inflammatory cytokine interleukin-10 (IL-10). B10 cells are best defined by their functional ability to produce IL-10, as they are not confined to any particular phenotypic subset. B10 cells function in an antigen-specific manner that requires cognate interactions with T cells in vivo to regulate immune responses and have been demonstrated to be potent regulators of allergic and autoimmune disease, cancer, infection, and transplant rejection. Importantly, the recent discovery of human B10 cells has accelerated this field to the forefront of clinical research where the possibility of harnessing the regulatory potential of B10 cells for treatment of aberrant immune responses and diseases may become feasible.
Assuntos
Subpopulações de Linfócitos B/imunologia , Subpopulações de Linfócitos B/metabolismo , Interleucina-10/biossíntese , Animais , Doenças Autoimunes/imunologia , Doenças Autoimunes/metabolismo , Autoimunidade/imunologia , Linfócitos B Reguladores/imunologia , Linfócitos B Reguladores/metabolismo , Humanos , Imunidade/fisiologia , Imunomodulação , Camundongos , FenótipoRESUMO
B cells mediate multiple functions that influence immune and inflammatory responses. In this study, T cell-mediated inflammation was exaggerated in CD19-deficient (Cd19(-/-)) mice and wild-type mice depleted of CD20(+) B cells, whereas inflammation was substantially reduced in mice with hyperactive B cells as a result of CD19 overexpression (hCD19Tg). These inflammatory responses were negatively regulated by a unique CD1d(hi)CD5(+) B cell subset that was absent in Cd19(-/-) mice, represented only 1%-2% of spleen B220(+) cells in wild-type mice, but was expanded to approximately 10% of spleen B220(+) cells in hCD19Tg mice. Adoptive transfer of these CD1d(hi)CD5(+) B cells normalized inflammation in wild-type mice depleted of CD20(+) B cells and in Cd19(-/-) mice. Remarkably, IL-10 production was restricted to this CD1d(hi)CD5(+) B cell subset, with IL-10 production diminished in Cd19(-/-) mice, yet increased in hCD19Tg mice. Thereby, CD1d(hi)CD5(+) B cells represent a unique subset of potent regulatory B cells.
Assuntos
Subpopulações de Linfócitos B/imunologia , Citocinas/metabolismo , Inflamação/imunologia , Interleucina-10/metabolismo , Linfócitos T/imunologia , Animais , Antígenos CD1/análise , Antígenos CD1/imunologia , Antígenos CD1d , Subpopulações de Linfócitos B/metabolismo , Antígenos CD5/análise , Antígenos CD5/imunologia , Citocinas/imunologia , Imunofenotipagem , Inflamação/metabolismo , Interleucina-10/imunologia , Depleção Linfocítica , Camundongos , Camundongos Mutantes , Camundongos Transgênicos , Cavidade Peritoneal/citologia , Fenótipo , Baço/citologia , Baço/imunologia , Linfócitos T/metabolismoRESUMO
B cells are commonly thought to enhance inflammatory immune responses. However, specific regulatory B cell subsets recently were identified that downregulate adaptive and innate immunity, inflammation, and autoimmunity through diverse molecular mechanisms. In both mice and humans, a rare, but specific, subset of regulatory B cells is functionally characterized by its capacity to produce IL-10, a potent inhibitory cytokine. For clarity, this regulatory B cell subset has been labeled as B10 cells, because their ability to downregulate immune responses and inflammatory disease is fully attributable to IL-10, and their absence or loss exacerbates disease symptoms in mouse models. This review preferentially focuses on what is known about mouse B10 cell development, phenotype, and effector function, as well as on mechanistic studies that demonstrated their functional importance during inflammation, autoimmune disease, and immune responses.
Assuntos
Linfócitos B Reguladores/imunologia , Interleucina-10/imunologia , Animais , HumanosRESUMO
We have previously shown that polyclonal natural IgM protects mice from renal ischemia/reperfusion injury (IRI) by inhibiting the reperfusion inflammatory response. We hypothesized that a potential mechanism involved IgM modulation of dendritic cells (DC), as we observed high IgM binding to splenic DC. To test this hypothesis, we pretreated bone marrow-derived DC (BMDC) with polyclonal murine or human IgM prior to LPS activation and demonstrated that 0.5 × 10(6) IgM/LPS-pretreated BMDC, when injected into wild-type C57BL/6 mice 24 h before renal ischemia, protect mice from developing renal IRI. We show that this switching of LPS-activated BMDC to a regulatory phenotype requires modulation of BMDC function that is mediated by IgM binding to nonapoptotic BMDC receptors. Regulatory BMDC require IL-10 and programmed death 1 as well as downregulation of CD40 and p65 NF-κB phosphorylation to protect in renal IRI. Blocking the programmed death ligand 1 binding site just before i.v. injection of IgM/LPS-pretreated BMDC or using IL-10 knockout BMDC fails to induce protection. Similarly, IgM/LPS-pretreated BMDC are rendered nonprotective by increasing CD40 expression and phosphorylation of p65 NF-κB. How IgM/LPS regulatory BMDC suppress in vivo ischemia-induced innate inflammation remains to be determined. However, we show that suppression is dependent on other in vivo regulatory mechanisms in the host, that is, CD25(+) T cells, B cells, IL-10, and circulating IgM. There was no increase in Foxp3(+) regulatory T cells in the spleen either before or after renal IRI. Collectively, these findings show that natural IgM anti-leukocyte Abs can switch BMDC to a regulatory phenotype despite the presence of LPS that ordinarily induces BMDC maturation.
Assuntos
Células da Medula Óssea/imunologia , Células Dendríticas/imunologia , Imunoglobulina M/imunologia , Inflamação/imunologia , Rim/irrigação sanguínea , Traumatismo por Reperfusão/prevenção & controle , Animais , Linfócitos B/imunologia , Antígenos CD40/metabolismo , Células Cultivadas , Interleucina-10/imunologia , Lipopolissacarídeos/imunologia , Ativação Linfocitária/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fosforilação , Receptor de Morte Celular Programada 1/imunologia , Traumatismo por Reperfusão/imunologia , Transdução de Sinais/imunologia , Linfócitos T/imunologia , Fator de Transcrição RelA/metabolismoRESUMO
OBJECTIVE: To determine the serum levels of nucleosome in patients with systemic sclerosis (SSc) and relate the results to the clinical features of SSc. METHODS: Serum nucleosome levels in 91 patients with SSc were examined by ELISA. The expression of Toll-like receptor (TLR) 9 in T and B cells was quantified by flow cytometric intracellular protein analysis. The effects of nucleosomes on lymphocytes were also analysed. Moreover, we assessed the effects of nucleosomes on fibrosis, using wild type and CD19-deficient bleomycin-treated mice, an experimental model for human SSc. RESULTS: Serum nucleosome levels were elevated in SSc compared with healthy controls and correlated positively with the extent of skin and pulmonary fibrosis and immunological abnormalities. The retrospective longitudinal analysis showed the serum nucleosome levels to be attenuated during the follow-up period. TLR9, which can be stimulated by nucleosome expression was upregulated in the affected T and B cells of patients with SSc. Moreover, nucleosome stimulation strongly increased interleukin (IL)-4 and IL-17 expression of T cells, B-cell IgG production and proliferation of lymphocytes in SSc compared with those in healthy controls. In bleomycin-induced SSc model mice, serum nucleosome levels were elevated compared with control mice. Furthermore, nucleosomes increased IgG production and proliferation of mouse B cells. Although TLR9 expression was similar between wild type and CD19-deficient splenic B cells, CD19 deficiency reduced these nucleosome effects. CONCLUSION: These results suggest that nucleosomes and its signalling in B and T cells contribute to disease development in SSc via TLR9.
Assuntos
Linfócitos B/metabolismo , Nucleossomos/metabolismo , Escleroderma Sistêmico/sangue , Linfócitos T/metabolismo , Animais , Linfócitos B/imunologia , Bleomicina , Estudos de Casos e Controles , Modelos Animais de Doenças , Feminino , Humanos , Imunoglobulina G/metabolismo , Interleucina-17/metabolismo , Interleucina-4/metabolismo , Estudos Longitudinais , Masculino , Camundongos , Estudos Retrospectivos , Escleroderma Sistêmico/induzido quimicamente , Escleroderma Sistêmico/imunologia , Transdução de Sinais , Linfócitos T/imunologia , Receptor Toll-Like 9/metabolismoRESUMO
Desmoglein 3 is a transmembrane component of desmosome complexes that mediate epidermal cell-to-cell adhesion and tissue integrity. Antibody blockade of desmoglein 3 function in pemphigus vulgaris patients leads to skin blistering (acantholysis) and oral mucosa lesions. Desmoglein 3 deficiency in mice leads to a phenotype characterized by cyclic alopecia in addition to the dramatic skin and mucocutaneous acantholysis observed in pemphigus patients. In this study, mice that developed an overt squeaky (sqk) phenotype were identified with obstructed airways, cyclic hair loss, and severe immunodeficiency subsequent to the development of oral lesions and malnutrition. Single-nucleotide polymorphism-based quantitative trait loci mapping revealed a genetic deletion that resulted in expression of a hypomorphic desmoglein 3 protein with a truncation of an extracellular cadherin domain. Because hypomorphic expression of a truncated desmoglein 3 protein led to a spectrum of severe pathology not observed in mice deficient in desmoglein 3, similar human genetic alterations may also disrupt desmosome function and induce a disease course distinct from pathogenesis of pemphigus vulgaris.
Assuntos
Alopecia/genética , Desmogleína 3/genética , Síndromes de Imunodeficiência/genética , Desnutrição/genética , Alopecia/imunologia , Alopecia/metabolismo , Animais , Desmogleína 3/metabolismo , Síndromes de Imunodeficiência/imunologia , Síndromes de Imunodeficiência/metabolismo , Queratinócitos/imunologia , Queratinócitos/metabolismo , Queratinócitos/patologia , Masculino , Desnutrição/imunologia , Desnutrição/metabolismo , Camundongos , Fenótipo , Polimorfismo de Nucleotídeo Único , Locos de Características Quantitativas , Deleção de Sequência , Pele/imunologia , Pele/metabolismo , Pele/patologiaRESUMO
B cells are known to instigate and promulgate immune responses by producing antibodies and presenting antigens to T cells. However, a rare but potent B-cell subset in both humans and mice is capable of inhibiting immune responses through the production of the anti-inflammatory cytokine IL-10. Regulatory B cells do not express any unique combination of surface markers but instead represent a small population of B cells that have acquired the unique ability to produce IL-10. This numerically rare B-cell subset is therefore functionally referred to as 'B10 cells' to reflect both their molecular program and the fact that their anti-inflammatory effects in models of autoimmunity, infection and cancer are solely attributable to IL-10 production. As with most B cells, B10 cell development and function appear to be predominantly, if not exclusively, driven by antigen-receptor signals. Once generated, B10 cells respond to both innate and adaptive immune signals, with a requirement for antigen-specific local interactions with T cells to induce IL-10 production and to provide optimal immune suppression in mouse models of autoimmune disease. B10 cells therefore provide an antigen-specific mechanism for delivering IL-10 locally to sites of immune activation and inflammation. The ability of B10 cells to regulate innate and adaptive immune responses makes them an ideal therapeutic target for the treatment of many immune-related disorders.
Assuntos
Apresentação de Antígeno , Linfócitos B Reguladores/imunologia , Dermatite de Contato/imunologia , Imunidade Humoral , Interleucina-10/imunologia , Animais , Linfócitos B Reguladores/patologia , Comunicação Celular , Dermatite de Contato/genética , Dermatite de Contato/patologia , Modelos Animais de Doenças , Regulação da Expressão Gênica/imunologia , Humanos , Imunidade Inata , Interleucina-10/genética , Camundongos , Receptores de Antígenos de Linfócitos B/genética , Receptores de Antígenos de Linfócitos B/imunologia , Transdução de Sinais , Baço/imunologia , Baço/patologia , Linfócitos T/imunologia , Linfócitos T/patologiaRESUMO
OBJECTIVE: Abdominal aortic aneurysm (AAA) is widespread among elderly people and results in progressive expansion and rupture of the aorta with high mortality. Macrophages, which are the main population observed within the site of aneurysm, are thought to derive from circulating monocytes although no direct evidence has been provided to date. In this study, we were particularly interested in understanding the trafficking behavior of monocyte subsets in AAA and their role in disease pathogenesis. APPROACH AND RESULTS: Using bone marrow transplantation in Apoe(-/-) mice, we showed that circulating monocytes give rise to abdominal aortic macrophages in hypercholesterolemic mice submitted to angiotensin II (AngII). Detailed monitoring of monocyte compartmentalization revealed that lymphocyte antigen 6C(high) and lymphocyte antigen 6C(low) monocytes transiently increase in blood early after AngII infusion and differentially infiltrate the abdominal aorta. The splenic reservoir accounted for the mobilization of the 2 monocyte subsets after 3 days of AngII infusion. Spleen removal or lymphocyte deficiency in Apoe(-/-) Rag2(-/-) mice similarly impaired early monocyte increase in blood in response to AngII and protected against AAA development, independently of blood pressure. Reconstitution of Apoe(-/-) Rag2(-/-) mice with total splenocytes but not with B-cell-depleted splenocytes restored monocyte mobilization in response to AngII and enhanced susceptibility to AAA. CONCLUSIONS: Taken together, the data show that lymphocyte antigen 6C(high) and lymphocyte antigen 6C(low) monocytes are mobilized from the spleen in response to AngII. Intriguingly, the process is dependent on the presence of B cells and significantly contributes to the development of AAA and the occurrence of aortic rupture.
Assuntos
Angiotensina II , Aneurisma da Aorta Abdominal/induzido quimicamente , Apolipoproteínas E/deficiência , Movimento Celular , Hipercolesterolemia/complicações , Monócitos/metabolismo , Baço/metabolismo , Animais , Antígenos Ly/metabolismo , Aneurisma da Aorta Abdominal/genética , Aneurisma da Aorta Abdominal/imunologia , Aneurisma da Aorta Abdominal/metabolismo , Aneurisma da Aorta Abdominal/patologia , Apolipoproteínas E/genética , Linfócitos B/imunologia , Linfócitos B/metabolismo , Transplante de Medula Óssea , Células Cultivadas , Proteínas de Ligação a DNA/deficiência , Proteínas de Ligação a DNA/genética , Modelos Animais de Doenças , Hipercolesterolemia/genética , Hipercolesterolemia/metabolismo , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Monócitos/imunologia , Baço/imunologia , Esplenectomia , Fatores de TempoRESUMO
Selectins are a family of adhesion receptors designed for efficient leukocyte tethering to the endothelium under shear. As a key property to resist premature bond disruption, selectin adhesiveness is enhanced by tensile forces that promote the conversion of a bent into an extended conformation of the N-terminal lectin and epidermal growth factor-like domains. Conformation-specific Abs have been invaluable in deciphering the activation mechanism of integrins, but similar reagents are not available for selectins. In this study, we show that the anti-human L-selectin mAbs DREG-55 and LAM1-5 but not DREG-56, DREG-200, or LAM1-1 heterotropically modulate adhesion presumably by stabilizing the extended receptor conformation. Force-free affinity assays, flow chamber, and microkinetic studies reveal a ligand-specific modulation of L-selectin affinity by DREG-55 mAb, resulting in a dramatic decrease of rolling velocity under flow. Furthermore, secondary tethering of polymorphonuclear cells was blocked by DREG-200 but significantly boosted by DREG-55 mAb. The results emphasize the need for a new classification for selectin Abs and introduce the new concept of heterotropic modulation of receptor function.
Assuntos
Anticorpos Monoclonais/imunologia , Migração e Rolagem de Leucócitos/imunologia , Neutrófilos/imunologia , Selectinas/metabolismo , Sequência de Aminoácidos , Anticorpos/imunologia , Adesão Celular/imunologia , Linhagem Celular , Humanos , Células Jurkat , Selectinas/imunologia , Alinhamento de SequênciaRESUMO
B cells provide humoral protection against pathogens and promote cellular immunity through diverse nonclassical effector functions. To assess B cell function in promoting T cell homeostasis, mature B cells were either acutely or chronically depleted in mice using CD20 mAb. Acute B cell depletion in either 2- or 4-mo-old mice significantly reduced spleen and lymph node CD4(+) and CD8(+) T cell numbers, including naive, activated, and Foxp3(+)CD25(+)CD4(+) regulatory T cell subsets. The numbers of IFN-γ- and TNF-α-producing T cells were also significantly reduced. Chronic B cell depletion for 6 mo in aged naive mice resulted in a 40-70% reduction in activated CD4(+) and CD8(+) T cell numbers and 20-50% reductions in IFN-γ-producing T cells. Therefore, B cells were necessary for maintaining naive CD4(+) and CD8(+) T cell homeostasis for subsequent optimal T cell expansion in young and old mice. To determine the significance of this finding, a week of B cell depletion in 4-mo-old mice was followed by acute viral infection with lymphocytic choriomeningitis virus Armstrong. Despite their expansion, activated and cytokine-producing CD4(+) and CD8(+) T cell numbers were still significantly reduced 1 wk later. Moreover, viral peptide-specific CD4(+) and CD8(+) T cell numbers and effector cell development were significantly reduced in mice lacking B cells, whereas lymphocytic choriomeningitis virus titers were dramatically increased. Thus, T cell function is maintained in B cell-depleted mice, but B cells are required for optimal CD4(+) and CD8(+) T cell homeostasis, activation, and effector development in vivo, particularly during responses to acute viral infection.
Assuntos
Linfócitos B/imunologia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Homeostase/imunologia , Coriomeningite Linfocítica/imunologia , Vírus da Coriomeningite Linfocítica/imunologia , Doença Aguda , Animais , Linfócitos B/metabolismo , Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD8-Positivos/metabolismo , Proliferação de Células , Citometria de Fluxo , Fatores de Transcrição Forkhead/imunologia , Fatores de Transcrição Forkhead/metabolismo , Interações Hospedeiro-Patógeno/imunologia , Interferon gama/imunologia , Interferon gama/metabolismo , Subunidade alfa de Receptor de Interleucina-2/imunologia , Subunidade alfa de Receptor de Interleucina-2/metabolismo , Linfonodos/imunologia , Linfonodos/metabolismo , Contagem de Linfócitos , Depleção Linfocítica , Coriomeningite Linfocítica/virologia , Vírus da Coriomeningite Linfocítica/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Baço/imunologia , Baço/metabolismo , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/metabolismo , Fator de Necrose Tumoral alfa/imunologia , Fator de Necrose Tumoral alfa/metabolismoRESUMO
Chronic graft-versus-host disease (cGVHD) is an increasingly frequent cause of morbidity and mortality of allogeneic hematopoietic stem-cell transplantation. Sclerodermatous cGVHD (Scl-cGVHD) is characterized by fibrosis and autoimmune features resembling those of systemic sclerosis (SSc). Transplantation of B10.D2 bone marrow and splenocytes into irradiated BALB/c mice is an established model of human Scl-cGVHD. To examine the role of B cells in Scl-cGVHD, CD19-deficient (CD19(-/-)) mice were used as donors or recipients. CD19(-/-) donors induced more severe Scl-cGVHD than wild-type donors, but use of CD19(-/-) recipients resulted in no significant differences compared with wild-type recipients. Moreover, CD19 deficiency on donor B cells resulted in the expansion of splenic interleukin (IL) -6-producing monocytes/macrophages, cytotoxic CD8(+) T cells, and Th1 cells during the early stage of disease and increased the infiltration of T cells, TGF-ß-producing monocytes/macrophages, and Th2 cells into the skin in the later stage of Scl-cGVHD. IL-10-producing regulatory B cells (B10 cells) were not reconstituted by CD19(-/-) donor cells, and early adoptive transfer of B10 cells attenuated the augmented manifestations of CD19(-/-) donor-induced Scl-cGVHD. Therefore, donor-derived B10 cells have a suppressive role in Scl-cGVHD development, warranting future investigation of regulatory B-cell-based therapy for treatment of Scl-cGVHD and SSc.
Assuntos
Linfócitos B Reguladores/imunologia , Transplante de Medula Óssea/imunologia , Doença Enxerto-Hospedeiro/imunologia , Escleroderma Sistêmico/imunologia , Pele/imunologia , Transferência Adotiva , Animais , Antígenos CD19/genética , Antígenos CD19/imunologia , Antígenos CD1d/análise , Antígenos CD1d/imunologia , Transplante de Medula Óssea/efeitos adversos , Antígeno CD11b/análise , Antígeno CD11b/imunologia , Antígenos CD5/análise , Antígenos CD5/imunologia , Doença Crônica , Feminino , Fibrose/imunologia , Fibrose/patologia , Deleção de Genes , Doença Enxerto-Hospedeiro/patologia , Humanos , Interleucina-6/imunologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Monócitos/imunologia , Escleroderma Sistêmico/patologia , Pele/patologia , Linfócitos T/imunologiaRESUMO
The immune system has evolved into two main arms: the primitive innate arm that is the first line of defense but relatively short-lived and broad acting; and the advanced adaptive arm that generates immunological memory, allowing rapid, specific recall responses. T cell-independent type-2 (TI-2) antigens (Ags) invoke innate immune responses. However, due to its 'at the ready' nature, how the innate arm of the immune system maintains tolerance to potentially abundant host TI-2 Ags remains elusive. Therefore, it is important to define the mechanisms that establish innate immune tolerance. This review highlights recent insights into B cell tolerance to theoretical self TI-2 Ags, and examines how the B cell-restricted sialic acid binding Ig-like lectins (Siglecs), CD22 and Siglec-G, might contribute to this process.
Assuntos
Linfócitos B/imunologia , Tolerância Imunológica , Lectinas/imunologia , Receptores de Antígenos de Linfócitos B/imunologia , Lectina 2 Semelhante a Ig de Ligação ao Ácido Siálico/imunologia , Animais , Doenças Autoimunes/imunologia , HumanosRESUMO
The spleen regulatory B cell subset with the functional capacity to express IL-10 (B10 cells) modulates both immune responses and autoimmune disease severity. However, the peritoneal cavity also contains relatively high frequencies of functionally defined IL-10-competent B10 cells. In this study, peritoneal cavity B10 cells shared similar cell surface phenotypes with their spleen counterparts. However, peritoneal cavity B10 cells were 10-fold more frequent among B cells than occurred within the spleen, intestinal tract, or mesenteric lymph nodes and were present at higher proportions among the phenotypically defined peritoneal B1a > B1b > B2 cell subpopulations. The development or localization of B10 cells within the peritoneal cavity was not dependent on the presence of commensal microbiota, T cells, IL-10 or B10 cell IL-10 production, or differences between their fetal liver or adult bone marrow progenitor cell origins. The BCR repertoire of peritoneal cavity B10 cells was diverse, as occurs in the spleen, and predominantly included germline-encoded VH and VL regions commonly found in either the conventional or B1 B cell compartments. Thereby, the capacity to produce IL-10 appears to be an intrinsic functional property acquired by clonally diverse B cells. Importantly, IL-10 production by peritoneal cavity B cells significantly reduced disease severity in spontaneous and induced models of colitis by regulating neutrophil infiltration, colitogenic CD4(+) T cell activation, and proinflammatory cytokine production during colitis onset. Thus, the numerically small B10 cell subset within the peritoneal cavity has regulatory function and is important for maintaining homeostasis within gastrointestinal tissues and the immune system.
Assuntos
Linfócitos B Reguladores/imunologia , Linfócitos T CD4-Positivos/imunologia , Colite/imunologia , Cavidade Peritoneal/citologia , Transferência Adotiva , Animais , Modelos Animais de Doenças , Ensaio de Imunoadsorção Enzimática , Imunofluorescência , Interferon gama/imunologia , Interleucina-10/biossíntese , Interleucina-10/imunologia , Subpopulações de Linfócitos/imunologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos KnockoutRESUMO
Pathogens use numerous methods to subvert host immune responses, including the modulation of host IL-10 production by diverse cell types. However, the B cell sources of IL-10 and their overall influence on innate and cellular immune responses have not been well characterized during infections. Using Listeria as a model pathogen, infection drove the acute expansion of a small subset of regulatory B cells (B10 cells) that potently suppress inflammation and autoimmunity through the production of IL-10. Unexpectedly, spleen bacteria loads were 92-97% lower in B10 cell-deficient CD19(-/-) mice, in mice depleted of mature B cells, and in mice treated with CD22 mAb to preferentially deplete B10 cells before infection. By contrast, the adoptive transfer of wild-type B10 cells reduced bacterial clearance by 38-fold in CD19(-/-) mice through IL-10-dependent pathways. B10 cell depletion using CD22 mAb significantly enhanced macrophage phagocytosis of Listeria and their production of IFN-γ, TNF-α, and NO ex vivo. Accelerated bacteria clearance following B10 cell depletion significantly reduced Ag-specific CD4(+) T cell proliferation and cytokine production, but did not alter CD8(+) T cell responses. B10 cell regulatory function during innate immune responses was nonetheless dependent on cognate interactions with CD4(+) T cells because B10 cells deficient in IL-10, MHC-II, or IL-21R expression did not influence Listeria clearance. Thus, Listeria manipulates immune responses through a strategy of immune evasion that involves the preferential expansion of endogenous B10 cells that regulate the magnitude and duration of both innate and cellular immune responses.