RESUMO
The CD4-binding site (CD4bs) is a conserved epitope on HIV-1 envelope (Env) that can be targeted by protective broadly neutralizing antibodies (bnAbs). HIV-1 vaccines have not elicited CD4bs bnAbs for many reasons, including the occlusion of CD4bs by glycans, expansion of appropriate naive B cells with immunogens, and selection of functional antibody mutations. Here, we demonstrate that immunization of macaques with a CD4bs-targeting immunogen elicits neutralizing bnAb precursors with structural and genetic features of CD4-mimicking bnAbs. Structures of the CD4bs nAb bound to HIV-1 Env demonstrated binding angles and heavy-chain interactions characteristic of all known human CD4-mimicking bnAbs. Macaque nAb were derived from variable and joining gene segments orthologous to the genes of human VH1-46-class bnAb. This vaccine study initiated in primates the B cells from which CD4bs bnAbs can derive, accomplishing the key first step in the development of an effective HIV-1 vaccine.
Assuntos
Vacinas contra a AIDS , HIV-1 , Animais , Humanos , Anticorpos Amplamente Neutralizantes , Antígenos CD4 , Moléculas de Adesão Celular , HIV-1/fisiologia , Macaca , Vacinas contra a AIDS/imunologiaRESUMO
SARS-CoV-2-neutralizing antibodies (NAbs) protect against COVID-19. A concern regarding SARS-CoV-2 antibodies is whether they mediate disease enhancement. Here, we isolated NAbs against the receptor-binding domain (RBD) or the N-terminal domain (NTD) of SARS-CoV-2 spike from individuals with acute or convalescent SARS-CoV-2 or a history of SARS-CoV infection. Cryo-electron microscopy of RBD and NTD antibodies demonstrated function-specific modes of binding. Select RBD NAbs also demonstrated Fc receptor-γ (FcγR)-mediated enhancement of virus infection in vitro, while five non-neutralizing NTD antibodies mediated FcγR-independent in vitro infection enhancement. However, both types of infection-enhancing antibodies protected from SARS-CoV-2 replication in monkeys and mice. Three of 46 monkeys infused with enhancing antibodies had higher lung inflammation scores compared to controls. One monkey had alveolar edema and elevated bronchoalveolar lavage inflammatory cytokines. Thus, while in vitro antibody-enhanced infection does not necessarily herald enhanced infection in vivo, increased lung inflammation can rarely occur in SARS-CoV-2 antibody-infused macaques.
Assuntos
Anticorpos Neutralizantes/imunologia , SARS-CoV-2/fisiologia , Glicoproteína da Espícula de Coronavírus/imunologia , Animais , Anticorpos Antivirais/imunologia , Líquido da Lavagem Broncoalveolar/química , COVID-19/patologia , COVID-19/virologia , Citocinas/metabolismo , Feminino , Haplorrinos , Humanos , Pulmão/patologia , Pulmão/virologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Domínios Proteicos , RNA Guia de Cinetoplastídeos/metabolismo , Receptores de IgG/metabolismo , SARS-CoV-2/isolamento & purificação , Glicoproteína da Espícula de Coronavírus/química , Carga Viral , Replicação ViralRESUMO
MRL/lpr mice typically succumb to immune complex-mediated nephritis within the first year of life. However, MRL/lpr mice that only secrete IgM Abs because of activation-induced deaminase deficiency (AID-/-MRL/lpr mice) experienced a dramatic increase in survival. Further crossing of these mice to those incapable of making secretory IgM (µS mice) generated mice lacking any secreted Abs but with normal B cell receptors. Both strains revealed no kidney pathology, yet Ab-deficient mice still experienced high mortality. In this article, we report Ab-deficient MRL/lpr mice progressed to high-grade T cell lymphoma that can be reversed with injection of autoreactive IgM Abs or following adoptive transfer of IgM-secreting MRL/lpr B cells. Anti-nuclear Abs, particularly anti-dsDNA IgM Abs, exhibited tumor-killing activities against a murine T cell lymphoma cell line. Passive transfers of autoreactive IgM Abs into p53-deficient mice increased survival by delaying onset of T cell lymphoma. The lymphoma originated from a double-negative aberrant T cell population seen in MRL/lpr mice and most closely resembled human anaplastic large cell lymphoma. Combined, these results strongly implicate autoreactive IgM Abs in protection against T cell lymphoma.
Assuntos
Transferência Adotiva/métodos , Anticorpos Antinucleares/administração & dosagem , Citidina Desaminase/deficiência , Imunoglobulina M/administração & dosagem , Imunoglobulina M/deficiência , Linfoma Anaplásico de Células Grandes/imunologia , Linfoma Anaplásico de Células Grandes/terapia , Animais , Autoimunidade/genética , Linfócitos B/imunologia , Citidina Desaminase/genética , Modelos Animais de Doenças , Imunoglobulina M/genética , Linfoma Anaplásico de Células Grandes/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos MRL lpr , Camundongos Knockout , Linfócitos T/imunologia , Resultado do Tratamento , Proteína Supressora de Tumor p53/deficiência , Proteína Supressora de Tumor p53/genéticaRESUMO
Apoptotic debris, autoantibody, and IgG-immune complexes (ICs) have long been implicated in the inflammation associated with systemic lupus erythematosus (SLE); however, it remains unclear whether they initiate immune-mediated events that promote disease. In this study, we show that PBMCs from SLE patients experiencing active disease, and hematopoietic cells from lupus-prone MRL/lpr and NZM2410 mice accumulate markedly elevated levels of surface-bound nuclear self-antigens. On dendritic cells (DCs) and macrophages (MFs), the self-antigens are part of IgG-ICs that promote FcγRI-mediated signal transduction. Accumulation of IgG-ICs is evident on ex vivo myeloid cells from MRL/lpr mice by 10 wk of age and steadily increases prior to lupus nephritis. IgG and FcγRI play a critical role in disease pathology. Passive transfer of pathogenic IgG into IgG-deficient MRL/lpr mice promotes the accumulation of IgG-ICs prior to significant B cell expansion, BAFF secretion, and lupus nephritis. In contrast, diminishing the burden IgG-ICs in MRL/lpr mice through deficiency in FcγRI markedly improves these lupus pathologies. Taken together, our findings reveal a previously unappreciated role for the cell surface accumulation of IgG-ICs in human and murine lupus.
Assuntos
Apoptose , Células Sanguíneas/imunologia , Células Dendríticas/imunologia , Lúpus Eritematoso Sistêmico/imunologia , Macrófagos/imunologia , Adulto , Animais , Autoantígenos/imunologia , Autoantígenos/metabolismo , Fator Ativador de Células B/metabolismo , Feminino , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos MRL lpr , Camundongos Knockout , Pessoa de Meia-Idade , Receptores de IgG/genética , Adulto JovemRESUMO
To test the hypothesis that DNA polymerase ζ participates in Ig hypermutation, we generated two mouse models of Pol ζ function: a B cell-specific conditional knockout and a knock-in strain with a Pol ζ mutagenesis-enhancing mutation. Pol ζ-deficient B cells had a reduction in mutation frequency at Ig loci in the spleen and in Peyer's patches, whereas knock-in mice with a mutagenic Pol ζ displayed a marked increase in mutation frequency in Peyer's patches, revealing a pattern that was similar to mutations in yeast strains with a homologous mutation in the gene encoding the catalytic subunit of Pol ζ. Combined, these data are best explained by a direct role for DNA polymerase ζ in Ig hypermutation.
Assuntos
Hipermutação Somática de Imunoglobulina/genética , Hipermutação Somática de Imunoglobulina/imunologia , Animais , Linfócitos B/enzimologia , Linfócitos B/imunologia , Linfócitos B/patologia , DNA Polimerase Dirigida por DNA/deficiência , DNA Polimerase Dirigida por DNA/genética , DNA Polimerase Dirigida por DNA/fisiologia , Ativação Enzimática/genética , Ativação Enzimática/imunologia , Técnicas de Introdução de Genes , Rearranjo Gênico de Cadeia Pesada de Linfócito B , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Modelos AnimaisRESUMO
A major goal of HIV-1 vaccine development is the induction of broadly neutralizing antibodies (bnAbs). Although success has been achieved in initiating bnAb B cell lineages, design of boosting immunogens that select for bnAb B cell receptors with improbable mutations required for bnAb affinity maturation remains difficult. Here, we demonstrate a process for designing boosting immunogens for a V3-glycan bnAb B cell lineage. The immunogens induced affinity-matured antibodies by selecting for functional improbable mutations in bnAb precursor knockin mice. Moreover, we show similar success in prime and boosting with nucleoside-modified mRNA-encoded HIV-1 envelope trimer immunogens, with improved selection by mRNA immunogens of improbable mutations required for bnAb binding to key envelope glycans. These results demonstrate the ability of both protein and mRNA prime-boost immunogens for selection of rare B cell lineage intermediates with neutralizing breadth after bnAb precursor expansion, a key proof of concept and milestone toward development of an HIV-1 vaccine.
Assuntos
Vacinas contra a AIDS , Anticorpos Neutralizantes , Linfócitos B , Anticorpos Anti-HIV , HIV-1 , Vacinas contra a AIDS/imunologia , Vacinas contra a AIDS/genética , Animais , Anticorpos Anti-HIV/imunologia , HIV-1/imunologia , HIV-1/genética , Camundongos , Anticorpos Neutralizantes/imunologia , Linfócitos B/imunologia , Humanos , Infecções por HIV/imunologia , Infecções por HIV/prevenção & controle , Anticorpos Amplamente Neutralizantes/imunologia , Mutação , Desenvolvimento de Vacinas , Imunização Secundária , Produtos do Gene env do Vírus da Imunodeficiência Humana/imunologia , Produtos do Gene env do Vírus da Imunodeficiência Humana/genéticaRESUMO
We recently identified a novel germinal center GTPase, SLIP-GC, that localizes to replication factories in B cells and that, when reduced, induces DNA breaks in lymphoma B cell lines in an activation-induced deaminase (AID)-dependent manner. Herein, we generated mice deficient in SLIP-GC and examined the impact of SLIP-GC deficiency in immunoglobulin hypermutation and class switch recombination, both AID-dependent mechanisms. SLIP-GC-deficient mice experienced a substantial increase in mutations at G:C base pairs at the region downstream of JH4 in the immunoglobulin heavy chain locus. This change was reflected in the overall mutation frequency, and it was associated with an increase in transitions from G:C base pairs, a hallmark of AID-mediated deamination during replication. In addition, G:C transitions at non-immunoglobulin loci also increased in these mice. Given the intracellular localization of SLIP-GC to sites of replicating DNA, these results suggest that SLIP-GC protects replicating DNA from AID-mediated deamination of cytosines in both strands.
Assuntos
Citidina Desaminase/metabolismo , GTP Fosfo-Hidrolases/biossíntese , GTP Fosfo-Hidrolases/genética , Imunoglobulinas/genética , Hipermutação Somática de Imunoglobulina/genética , Animais , Linfócitos B/citologia , Linfócitos B/metabolismo , Citidina Desaminase/genética , Citosina/química , Análise Mutacional de DNA , Replicação do DNA , Genótipo , Centro Germinativo/metabolismo , Switching de Imunoglobulina , Ativação Linfocitária , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Modelos Genéticos , Nódulos Linfáticos Agregados/metabolismoRESUMO
Targeting germline (gl-) precursors of broadly neutralizing antibodies (bNAbs) is acknowledged as an important strategy for HIV-1 vaccines. The VRC01-class of bNAbs is attractive because of its distinct genetic signature. However, VRC01-class bNAbs often require extensive somatic hypermutation, including rare insertions and deletions. We describe a BG505 SOSIP trimer, termed GT1.2, to optimize binding to gl-CH31, the unmutated common precursor of the CH30-34 bNAb lineage that acquired a large CDRH1 insertion. The GT1.2 trimer activates gl-CH31 naive B cells in knock-in mice, and B cell responses could be matured by selected boosting immunogens to generate cross-reactive Ab responses. Next-generation B cell sequencing reveals selection for VRC01-class mutations, including insertions in CDRH1 and FWR3 at positions identical to VRC01-class bNAbs, as well as CDRL1 deletions and/or glycine substitutions to accommodate the N276 glycan. These results provide proof of concept for vaccine-induced affinity maturation of B cell lineages that require rare insertions and deletions.
Assuntos
Soropositividade para HIV , HIV-1 , Camundongos , Animais , Anticorpos Amplamente Neutralizantes , Anticorpos Neutralizantes , HIV-1/genética , Anticorpos Anti-HIV , VacinaçãoRESUMO
OBJECTIVE: We previously generated MRL/lpr mice deficient in activation-induced deaminase (AID) that lack isotype switching and immunoglobulin hypermutation. These mice have high levels of unmutated (germline) autoreactive IgM, yet they experienced an increase in survival and an improvement in lupus nephritis that exceeded that of MRL/lpr mice lacking IgG. The purpose of the present study was to test the hypothesis that high levels of germline autoreactive IgM in these mice confer protection against lupus nephritis. METHODS: Autoreactive IgM antibodies of various specificities, including antibodies against double-stranded DNA (dsDNA), from AID-deficient MRL/lpr mice were given to asymptomatic MRL/lpr mice, and the levels of cytokines, proteinuria, immune complex deposition in the kidneys, and glomerulonephritis were examined. Novel AID-deficient MRL/lpr mice that lack any antibodies were generated for comparison to AID-deficient MRL/lpr mice that secrete only IgM. RESULTS: Treatment with IgM anti-dsDNA resulted in a dramatic improvement in lupus nephritis. Other autoreactive IgM antibodies, such as antiphospholipid and anti-Sm, did not alter the pathologic changes. Secretion of proinflammatory cytokines by macrophages and the levels of inflammatory cells and apoptotic debris in the kidneys were lower in mice receiving IgM anti-dsDNA. Protective IgM derived from AID-deficient MRL/lpr mice displayed a distinct B cell repertoire, with a bias toward members of the V(H) 7183 family. CONCLUSION: IgM anti-dsDNA protected MRL/lpr mice from lupus nephritis, likely by stopping the inflammatory cascade leading to kidney damage. A distinct repertoire of V(H) usage in IgM anti-dsDNA hybridomas from AID-deficient mice suggests that there is enrichment of a dedicated B cell population that secretes unmutated protective IgM in these mice.
Assuntos
Anticorpos/metabolismo , Citidina Desaminase/deficiência , DNA/imunologia , Imunoglobulina M/metabolismo , Lúpus Eritematoso Sistêmico/complicações , Lúpus Eritematoso Sistêmico/imunologia , Nefrite Lúpica/prevenção & controle , Animais , Apoptose/efeitos dos fármacos , Apoptose/fisiologia , Linfócitos B/imunologia , Linfócitos B/metabolismo , Citidina Desaminase/genética , Citidina Desaminase/metabolismo , Modelos Animais de Doenças , Hibridomas/imunologia , Hibridomas/metabolismo , Imunoglobulina M/farmacologia , Rim/imunologia , Rim/patologia , Lúpus Eritematoso Sistêmico/metabolismo , Nefrite Lúpica/metabolismo , Nefrite Lúpica/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos MRL lpr , Camundongos Knockout , Proteinúria/metabolismoRESUMO
SARS-CoV-2 neutralizing antibodies (NAbs) protect against COVID-19. A concern regarding SARS-CoV-2 antibodies is whether they mediate disease enhancement. Here, we isolated NAbs against the receptor-binding domain (RBD) and the N-terminal domain (NTD) of SARS-CoV-2 spike from individuals with acute or convalescent SARS-CoV-2 or a history of SARS-CoV-1 infection. Cryo-electron microscopy of RBD and NTD antibodies demonstrated function-specific modes of binding. Select RBD NAbs also demonstrated Fc receptor-γ (FcγR)-mediated enhancement of virus infection in vitro , while five non-neutralizing NTD antibodies mediated FcγR-independent in vitro infection enhancement. However, both types of infection-enhancing antibodies protected from SARS-CoV-2 replication in monkeys and mice. Nonetheless, three of 31 monkeys infused with enhancing antibodies had higher lung inflammation scores compared to controls. One monkey had alveolar edema and elevated bronchoalveolar lavage inflammatory cytokines. Thus, while in vitro antibody-enhanced infection does not necessarily herald enhanced infection in vivo , increased lung inflammation can occur in SARS-CoV-2 antibody-infused macaques.
RESUMO
We previously reported that activation-induced deaminase (AID) heterozygous MRL/lpr mice have substantially lower levels of serum anti-dsDNA autoantibodies than AID wild-type littermates. Given the known functions of AID, here we examined whether this decrease in pathogenic autoantibodies in the heterozygotes was the result of a defect in class switch recombination, somatic hypermutation, or both. We report significant impairment of switch recombination to most isotypes except immunoglobulin G3 (IgG3) in vitro. However, serum levels of IgG were similar to AID wild-type levels even in very young mice. Mutation accumulation in the B cells from Peyer's patches also revealed reduced somatic hypermutation in the heterozygotes. Unlike the switch defect, the hypermutation defect probably resulted in an in vivo effect because the serum IgG antibodies from the heterozygotes were of strikingly lower affinity to dsDNA than serum IgG antibodies from wild-type littermates. This suggests that the somatic hypermutation defect resulted in impaired affinity maturation of autoantibodies in these mice and explains the low levels of specific anti-dsDNA antibodies in the heterozygotes. This correlated with a delay in the development of kidney damage. These results imply that AID levels impact the class switch recombination and somatic hypermutation mechanisms and directly implicate affinity maturation of autoantibodies in autoimmunity.
Assuntos
Autoanticorpos/biossíntese , Citidina Desaminase/genética , Nefrite Lúpica/imunologia , Animais , Anticorpos Antinucleares/imunologia , Afinidade de Anticorpos/genética , Afinidade de Anticorpos/imunologia , Linfócitos B/imunologia , Células Cultivadas , DNA/imunologia , Heterozigoto , Imunoglobulina G/sangue , Imunoglobulina G/imunologia , Nefrite Lúpica/genética , Nefrite Lúpica/patologia , Ativação Linfocitária/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos MRL lpr , Mutação , Nódulos Linfáticos Agregados/imunologia , RNA Mensageiro/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Hipermutação Somática de Imunoglobulina/imunologiaRESUMO
The basis for mutations at A:T base pairs in immunoglobulin hypermutation and defining how AID interacts with the DNA of the immunoglobulin locus are major aspects of the immunoglobulin mutator mechanism where questions remain unanswered. Here, we examined the pattern of mutations generated in mice deficient in various DNA repair proteins implicated in A:T mutation and found a previously unappreciated bias at G:C base pairs in spectra from mice simultaneously deficient in DNA mismatch repair and uracil DNA glycosylase. This suggests a strand-biased DNA transaction for AID delivery which is then masked by the mechanism that introduces A:T mutations. Additionally, we asked if any of the known components of the A:T mutation machinery underscore the basis for the paucity of A:T mutations in the Burkitt lymphoma cell lines, Ramos and BL2. Ramos and BL2 cells were proficient in MSH2/MSH6-mediated mismatch repair, and express high levels of wild-type, full-length DNA polymerase eta. In addition, Ramos cells have high levels of uracil DNA glycosylase protein and are proficient in base excision repair. These results suggest that Burkitt lymphoma cell lines may be deficient in an unidentified factor that recruits the machinery necessary for A:T mutation or that AID-mediated cytosine deamination in these cells may be processed by conventional base excision repair truncating somatic hypermutation at the G:C phase. Either scenario suggests that cytosine deamination by AID is not enough to trigger A:T mutation, and that additional unidentified factors are required for full spectrum hypermutation in vivo.
Assuntos
Linfoma de Burkitt/genética , Reparo de Erro de Pareamento de DNA , Enzimas Reparadoras do DNA/genética , Nucleotídeos/genética , Hipermutação Somática de Imunoglobulina/genética , Trifosfato de Adenosina/genética , Animais , Linhagem Celular Tumoral , Citidina Desaminase/metabolismo , Citidina Trifosfato/genética , Guanosina Trifosfato/genética , Humanos , Camundongos , Mutação , Nucleotídeos de Timina/genéticaRESUMO
The events required for the induction of broad neutralizing antibodies (bnAbs) following HIV-1 envelope (Env) vaccination are unknown, and their induction in animal models as proof of concept would be critical. Here, we describe the induction of plasma antibodies capable of neutralizing heterologous primary (tier 2) HIV-1 strains in one macaque and two rabbits. Env immunogens were designed to induce CD4 binding site (CD4bs) bnAbs, but surprisingly, the macaque developed V1V2-glycan bnAbs. Env immunization of CD4bs bnAb heavy chain rearrangement (VHDJH) knockin mice similarly induced V1V2-glycan neutralizing antibodies (nAbs), wherein the human CD4bs VH chains were replaced with mouse rearrangements bearing diversity region (D)-D fusions, creating antibodies with long, tyrosine-rich HCDR3s. Our results show that Env vaccination can elicit broad neutralization of tier 2 HIV-1, demonstrate that V1V2-glycan bnAbs are more readily induced than CD4bs bnAbs, and define VH replacement and diversity region fusion as potential mechanisms for generating V1V2-glycan bnAb site antibodies.
Assuntos
Vacinas contra a AIDS/imunologia , Anticorpos Neutralizantes/imunologia , Anticorpos Anti-HIV/imunologia , HIV-1/imunologia , Sequência de Aminoácidos , Animais , Modelos Animais de Doenças , Epitopos/química , Epitopos/imunologia , Imunização , Macaca mulatta , Camundongos , Polissacarídeos/imunologia , Multimerização Proteica , Coelhos , Produtos do Gene env do Vírus da Imunodeficiência Humana/imunologiaRESUMO
A strategy for HIV-1 vaccine development is to define envelope (Env) evolution of broadly neutralizing antibodies (bnAbs) in infection and to recreate those events by vaccination. Here, we report host tolerance mechanisms that limit the development of CD4-binding site (CD4bs), HCDR3-binder bnAbs via sequential HIV-1 Env vaccination. Vaccine-induced macaque CD4bs antibodies neutralize 7% of HIV-1 strains, recognize open Env trimers, and accumulate relatively modest somatic mutations. In naive CD4bs, unmutated common ancestor knock-in mice Env+B cell clones develop anergy and partial deletion at the transitional to mature B cell stage, but become Env- upon receptor editing. In comparison with repetitive Env immunizations, sequential Env administration rescue anergic Env+ (non-edited) precursor B cells. Thus, stepwise immunization initiates CD4bs-bnAb responses, but immune tolerance mechanisms restrict their development, suggesting that sequential immunogen-based vaccine regimens will likely need to incorporate strategies to expand bnAb precursor pools.
Assuntos
Anticorpos Neutralizantes/biossíntese , Linfócitos B/imunologia , Anticorpos Anti-HIV/biossíntese , HIV-1/imunologia , Produtos do Gene env do Vírus da Imunodeficiência Humana/imunologia , Vacinas contra a AIDS/imunologia , Animais , Anticorpos Neutralizantes/química , Anticorpos Neutralizantes/genética , Linfócitos B/citologia , Sítios de Ligação de Anticorpos , Antígenos CD4/metabolismo , Linhagem da Célula/imunologia , Anergia Clonal , Feminino , Técnicas de Introdução de Genes , Anticorpos Anti-HIV/química , Anticorpos Anti-HIV/genética , Humanos , Tolerância Imunológica , Imunização/métodos , Macaca mulatta , Masculino , Camundongos , Camundongos Transgênicos , Modelos MolecularesAssuntos
Vacinas contra a AIDS/genética , Afinidade de Anticorpos/genética , Linfócitos B/imunologia , Anticorpos Anti-HIV/imunologia , HIV-1/imunologia , Seleção Genética , Produtos do Gene env do Vírus da Imunodeficiência Humana/genética , Vacinas contra a AIDS/imunologia , Animais , Epitopos/genética , Técnicas de Introdução de Genes , Humanos , Macaca , Camundongos , Camundongos Mutantes , Mutação , Produtos do Gene env do Vírus da Imunodeficiência Humana/imunologiaRESUMO
B cells contribute to autoimmunity both as secretors of pathogenic antibodies and through the activation of autoreactive T cells. B cells and antibodies acquire higher affinity to self-antigen through a process known as immunoglobulin hypermutation or SHM. The contribution of SHM to pathogenic antibody development in lupus has been established in various autoimmune mouse models and by examining antibodies from patients. However, its role in the antibody-independent contribution of B cells to autoimmunity has not been examined. Herein, we generate lupus-prone MRL/lpr mice with a limited IgM-only B cell repertoire, no secreted antibodies and no SHM. This enabled us to isolate the role of somatic hypermutation in B cell-mediated autoimmunity. We found that SHM-deficiency correlated with a reduction in autoreactive B cells, a decrease in T cell activation and a decrease in kidney lymphocytic infiltration. These data establish AID as an important contributor to the antibody-independent role of B cells in autoimmunity.
Assuntos
Autoimunidade/imunologia , Linfócitos B/imunologia , Citidina Desaminase/genética , Citidina Desaminase/metabolismo , Hipermutação Somática de Imunoglobulina/genética , Animais , Autoanticorpos/imunologia , Autoantígenos/imunologia , Autoimunidade/genética , Citidina Desaminase/imunologia , Imunoglobulina M/metabolismo , Ativação Linfocitária , Camundongos , Camundongos Endogâmicos MRL lprRESUMO
We generated MRL/lpr mice deficient in activation-induced deaminase (AID). Because AID is required for Ig hypermutation and class switch recombination, these mice lack hypermutated IgG Abs. Unlike their AID wild-type littermates, AID-deficient MRL/lpr mice not only lacked autoreactive IgG Abs but also experienced a dramatic increase in the levels of autoreactive IgM. This phenotype in AID-deficient mice translated into a significant reduction in glomerulonephritis, minimal mononuclear cell infiltration in the kidney, and a dramatic increase in survival to levels comparable to those previously reported for MRL/lpr mice completely lacking B cells and well below those of mice lacking secreted Abs. Therefore, this study wherein littermates with either high levels of autoreactive IgM or autoreactive IgG were directly examined proves that autoreactive IgM Abs alone are not sufficient to promote kidney disease in MRL/lpr mice. In addition, the substantial decrease in mortality combined with a dramatic increase in autoreactive IgM Abs in AID-deficient MRL/lpr mice suggest that autoreactive IgM Abs might not only fail to promote nephritis but may also provide a protective role in MRL/lpr mice. This novel mouse model containing high levels of autoreactive, unmutated IgM Abs will help delineate the contribution of autoreactive IgM to autoimmunity.
Assuntos
Citidina Desaminase/deficiência , Citidina Desaminase/genética , Nefrite Lúpica/enzimologia , Nefrite Lúpica/prevenção & controle , Animais , Autoanticorpos/biossíntese , Autoanticorpos/metabolismo , Subpopulações de Linfócitos B/enzimologia , Subpopulações de Linfócitos B/imunologia , Subpopulações de Linfócitos B/patologia , Movimento Celular/genética , Movimento Celular/imunologia , Citidina Desaminase/biossíntese , Imunoglobulina G/biossíntese , Imunoglobulina G/metabolismo , Imunoglobulina M/biossíntese , Imunoglobulina M/metabolismo , Rim/enzimologia , Rim/patologia , Leucócitos Mononucleares/enzimologia , Leucócitos Mononucleares/patologia , Nefrite Lúpica/imunologia , Nefrite Lúpica/mortalidade , Contagem de Linfócitos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos MRL lpr , Camundongos Knockout , Especificidade de Órgãos/genética , Especificidade de Órgãos/imunologia , Especificidade da Espécie , Análise de Sobrevida , Subpopulações de Linfócitos T/enzimologia , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/patologiaRESUMO
Recent studies have described the development of distinct functional subsets of macrophages in association with cancer, autoimmune disease, and chronic infections. Based on the ability of Th1 vs Th2 cytokines to promote opposing activities in macrophages, it has been proposed that macrophages develop into either type 1 inflammatory or type 2 anti-inflammatory subsets. As an alternative to the concept of subset development, we propose that macrophages, in response to changes in their tissue environment, can reversibly and progressively change the pattern of functions that they express. As demonstrated herein, macrophages can reversibly shift their functional phenotype through a multitude of patterns in response to changes in cytokine environment. Macrophages display distinct functional patterns after treatment with IFN-gamma, IL-12, IL-4, or IL-10 and additional functional patterns are displayed depending on whether the cytokine is present alone or with other cytokines and whether the cytokines are added before or concomitantly with the activating stimulus (LPS). Sequential treatment of macrophages with multiple cytokines results in a progression through multiple functional phenotypes. This ability to adapt to changing cytokine environments has significant in vivo relevance, as evidenced by the demonstration that macrophage functional phenotypes established in vivo in aged or tumor-bearing mice can be altered by changing their microenvironment. A concept of functional adaptivity is proposed that has important implications for therapeutic targeting of macrophages in chronic diseases that result in the dominance of particular functional phenotypes of macrophages that play a significant role in disease pathology.