RESUMO
Elucidation of the immune requirements for control or elimination of retroviral infection remains an important aim. We studied the induction of adaptive immunity to neonatal infection with a murine retrovirus, under conditions leading to immunological tolerance. We found that the absence of either maternal or offspring adaptive immunity permitted efficient vertical transmission of the retrovirus. Maternal immunodeficiency allowed the retrovirus to induce central Th cell tolerance in the infected offspring. In turn, this compromised the offspring's ability to mount a protective Th cell-dependent B cell response. However, in contrast to T cells, offspring B cells were not centrally tolerized and retained their ability to respond to the infection when provided with T cell help. Thus, escape of retrovirus-specific B cells from deletional tolerance offers the opportunity to induce protective retroviral immunity by restoration of retrovirus-specific T cell help, suggesting similar T cell immunotherapies for persistent viral infections.
Assuntos
Transferência Adotiva , Linfócitos B/imunologia , Transmissão Vertical de Doenças Infecciosas/prevenção & controle , Vírus da Leucemia Murina/imunologia , Leucemia Experimental/prevenção & controle , Infecções por Retroviridae/prevenção & controle , Linfócitos T/imunologia , Infecções Tumorais por Vírus/prevenção & controle , Animais , Animais Recém-Nascidos , Linfócitos B/transplante , Linfócitos B/virologia , Células Cultivadas , Tolerância Central , Feminino , Leucemia Experimental/imunologia , Masculino , Exposição Materna/efeitos adversos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Gravidez , Infecções por Retroviridae/imunologia , Infecções por Retroviridae/transmissão , Linfócitos T/transplante , Linfócitos T/virologia , Infecções Tumorais por Vírus/imunologia , Infecções Tumorais por Vírus/transmissãoRESUMO
Monocytes are heterogeneous effector cells involved in the maintenance and restoration of tissue integrity. However, their response to hyperlipidemia remains poorly understood. Here, we report that in the presence of elevated levels of triglyceride-rich lipoproteins, induced by administration of poloxamer 407, the blood numbers of non-classical Ly6C/Gr1(low) monocytes drop, while the number of bone marrow progenitors remains similar. We observed an increased crawling and retention of the Gr1(low) monocytes at the endothelial interface and a marked accumulation of CD68(+) macrophages in several organs. Hypertriglyceridemia was accompanied by an increased expression of tissue, and plasma CCL4 and blood Gr1(low) monocyte depletion involved a pertussis-toxin-sensitive receptor axis. Collectively, these findings demonstrate that a triglyceride-rich environment can alter blood monocyte distribution, promoting the extravasation of Gr1(low) cells. The behavior of these cells in response to dyslipidemia highlights the significant impact that high levels of triglyceride-rich lipoproteins may have on innate immune cells.
Assuntos
Antígenos Ly/metabolismo , Lipoproteínas/metabolismo , Monócitos/metabolismo , Triglicerídeos/metabolismo , Animais , Camundongos , Camundongos Endogâmicos C57BLRESUMO
A role for complement, particularly the classical pathway, in the regulation of immune responses is well documented. Deficiencies in C1q or C4 predispose to autoimmunity, while deficiency in C3 affects the suppression of contact sensitization and generation of oral tolerance. Complement components including C3 have been shown to be required for both B-cell and T-cell priming. The mechanisms whereby complement can mediate these diverse regulatory effects are poorly understood. Our previous work, using the mouse minor histocompatibility (HY) model of skin graft rejection, showed that both C1q and C3 were required for the induction of tolerance following intranasal peptide administration. By comparing tolerance induction in wild-type C57BL/6 and C1q-, C3-, C4- and C5-deficient C57BL/6 female mice, we show here that the classical pathway components including C3 are required for tolerance induction, whereas C5 plays no role. C3-deficient mice failed to generate a functional regulatory T (Treg) -dendritic cell (DC) tolerogenic loop required for tolerance induction. This was related to the inability of C3-deficient DC to up-regulate the arginine-consuming enzyme, inducible nitric oxide synthase (Nos-2), in the presence of antigen-specific Treg cells and peptide, leading to reduced Treg cell generation. Our findings demonstrate that the classical pathway and C3 play a critical role in the peptide-mediated induction of tolerance to HY by modulating DC function.
Assuntos
Proteínas do Sistema Complemento/genética , Células Dendríticas/imunologia , Antígeno H-Y/imunologia , Tolerância Imunológica/efeitos dos fármacos , Peptídeos/farmacologia , Linfócitos T Reguladores/imunologia , Administração Intranasal , Animais , Proteínas do Sistema Complemento/imunologia , Células Dendríticas/citologia , Feminino , Antígeno H-Y/genética , Tolerância Imunológica/genética , Camundongos , Camundongos Knockout , Óxido Nítrico Sintase Tipo II/genética , Óxido Nítrico Sintase Tipo II/imunologia , Peptídeos/imunologia , Linfócitos T Reguladores/citologiaRESUMO
Apoptotic cells are a source of autoantigens and impairment of their removal contributes to the development of autoimmunity in C1q deficiency. However, the lack of complement component 3 (C3), the predominant complement opsonin, does not predispose to autoimmunity, suggesting a modifying role of C3 in disease pathogenesis. To explore this hypothesis, here we investigated the role of C3 in the T-cell response to apoptotic cell-associated antigens. By comparing the phagosome maturation and the subsequent MHC class II presentation of a peptide derived from the internalized cargo between C3-deficient or C3-sufficient dendritic cells, we found that C3 deficiency accelerated the fusion of the apoptotic cargo with lysosomes. As a result, C3 deficiency led to impaired antigen-specific T-cell proliferation in vitro and in vivo. Notably, preopsonization of the apoptotic cells with C3 activation fragments rectified the trafficking and T-cell stimulation defects. These data indicate that activated C3 may act as a "chaperone" in the intracellular processing of an apoptotic cargo and, thus, may modulate the T-cell response to self-antigens displayed on dying cells.
Assuntos
Apoptose , Autoantígenos/imunologia , Complemento C3/metabolismo , Endocitose , Proteínas Opsonizantes/metabolismo , Linfócitos T/imunologia , Animais , Camundongos , Camundongos Knockout , Receptor 4 Toll-Like/genética , Receptor 4 Toll-Like/imunologiaRESUMO
Endogenous retroviruses are implicated in murine lupus nephritis. They provide a source of nephritogenic retroviral gp70-anti-gp70 immune complexes through the production of serum gp70 protein and anti-gp70 autoantibodies as a result of the activation of TLR7. The Sgp (serum gp70 production) loci identified in lupus-prone mice play distinct roles for the expression of different classes of endogenous retroviruses, as Sgp3 regulates the transcription of xenotropic, polytropic and modified polytropic (mPT) viruses, and Sgp4 the transcription of only xenotropic viruses. In the present study, we extended these analyses to a third locus, Sgp5, using BALB/c mice congenic for the NZW-derived Sgp5 allele and also explored the possible interaction of Sgp3 and Sgp4 loci to promote the expression of endogenous retroviruses and serum gp70. The analysis of Sgp5 BALB/c congenic mice demonstrated that the Sgp5 locus enhanced the expression of xenotropic and mPT viruses, thereby upregulating the production of serum gp70. These data indicate a distinct action of the Sgp5 locus on the expression of endogenous retroviruses, as compared with two other Sgp loci. Moreover, comparative analysis of C57BL/6 double congenic mice for Sgp3 and Sgp4 loci with single congenic mice revealed that Sgp3 and Sgp4 acted synergistically to elevate the transcription of the potentially replication-competent Xmv18 provirus and the production of serum gp70. This indicates that the combined effect of three different Sgp loci markedly enhance the expression of endogenous retroviruses and their gene product, serum gp70, thereby contributing to the formation of nephritogenic gp70-anti-gp70 immune complexes in murine lupus.
Assuntos
Retrovirus Endógenos/genética , Glicoproteínas/genética , Nefrite Lúpica/genética , Nefrite Lúpica/virologia , Chaperonas Moleculares/genética , Animais , Complexo Antígeno-Anticorpo/metabolismo , Retrovirus Endógenos/imunologia , Glicoproteínas/imunologia , Nefrite Lúpica/imunologia , Masculino , Glicoproteínas de Membrana/metabolismo , Camundongos , Camundongos Congênicos , Camundongos Endogâmicos BALB C , Chaperonas Moleculares/imunologia , Provírus/genética , Provírus/imunologia , RNA/genética , RNA Viral/genética , Receptor 7 Toll-Like/metabolismo , Regulação para Cima , Proteínas do Envelope Viral/genética , Proteínas do Envelope Viral/imunologiaRESUMO
The envelope glycoprotein, gp70, of endogenous retroviruses represents one of the major nephritogenic autoantigens implicated in murine systemic lupus erythematosus. Among different endogenous retroviruses (ecotropic, xenotropic and polytropic), lupus-prone mice express remarkably high levels of modified polytropic (mPT) retroviruses, which are controlled by the Sgp3 (serum gp70 production) locus. To define the contribution of the Sgp3 locus derived from lupus-prone mice to the expression of the specific mPT proviruses, the genetic origin of different mPT viruses expressed in livers and thymi of wild-type and Sgp3 congenic C57BL/6 mice was determined through clonal analysis of their transcripts. Among 13 mPT proviruses present in the C57BL/6 genome, only 3 proviruses (Mpmv6, Mpmv10 and Mpmv13) were selectively but differentially expressed in livers and thymi. This was likely a result of co-regulated expression with host genes because of their integration in the same transcriptional direction. In contrast, Sgp3 induced the steady-state expression of an additional select group of mPT proviruses and, after stimulation of TLR7, the highly upregulated expression of a potentially replication-competent mPT virus Mpmv4. These results indicated that the expression of distinct subpopulations of mPT retroviruses was regulated by Sgp3- and TLR7-dependent mechanisms. The induction of potentially replication-competent mPT viruses and the upregulation of one such virus after stimulation with TLR7 in Sgp3 congenic mice further highlight the implication of Sgp3 in autoimmune responses against nephritogenic serum gp70 through the activation of TLR7.
Assuntos
Retrovirus Endógenos/efeitos dos fármacos , Retrovirus Endógenos/genética , Glicoproteínas/metabolismo , Lúpus Eritematoso Sistêmico/genética , Chaperonas Moleculares/metabolismo , Receptor 7 Toll-Like/agonistas , Adenina/análogos & derivados , Adenina/farmacologia , Animais , Replicação do DNA , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Genoma , Glicoproteínas/genética , Fígado/metabolismo , Lúpus Eritematoso Sistêmico/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos NZB , Chaperonas Moleculares/genética , Provírus/genética , Timo/metabolismo , Transcrição Gênica/efeitos dos fármacosRESUMO
The envelope glycoprotein gp70 of endogenous retroviruses implicated in murine lupus nephritis is secreted by hepatocytes and its expression is controlled by Sgp3 (serum gp70 production 3) and Sgp4 loci derived from lupus-prone mice. Among three different endogenous retroviruses (ecotropic, xenotropic and polytropic), xenotropic viruses are considered to be the major source of serum gp70. Although the abundance of xenotropic viral gp70 RNA in livers was up-regulated by the presence of these two Sgp loci, it has not yet been clear whether Sgp3 and Sgp4 regulate the expression of a fraction or multiple xenotropic viruses present in mouse genome. To address this question, we determined the genetic origin of xenotropic viral sequences expressed in wild-type and two different Sgp congenic C57BL/6 mice. Among 14 xenotropic proviruses present in the C57BL/6 genome, only two proviruses (Xmv10 and Xmv14) were actively transcribed in wild-type C57BL/6 mice. In contrast, Sgp3 enhanced the transcription of Xmv10 and induced the transcription of three additional xenotropic viruses (Xmv15, Xmv17 and Xmv18), while Sgp4 induced the expression of a different xenotropic virus (Xmv13). Notably, stimulation of TLR7 in Sgp3 congenic C57BL/6 mice led to a highly enhanced expression of potentially replication-competent Xmv18. These results indicated that Sgp3 and Sgp4 independently regulated the transcription of distinct and restricted sets of xenotropic viruses in trans, thereby promoting the production of nephritogenic gp70 autoantigens. Furthermore, the induced expression of potentially replication-competent xenotropic viruses by Sgp3 may contribute to the development of autoimmune responses against gp70 through the activation of TLR7.
Assuntos
Glicoproteínas/metabolismo , Nefrite Lúpica/genética , Chaperonas Moleculares/metabolismo , Infecções por Retroviridae/genética , Proteínas do Envelope Viral/metabolismo , Vírus Relacionado ao Vírus Xenotrópico da Leucemia Murina/fisiologia , Animais , Autoanticorpos/sangue , Regulação Viral da Expressão Gênica/imunologia , Produtos do Gene env/sangue , Produtos do Gene env/genética , Produtos do Gene env/metabolismo , Glicoproteínas/genética , Nefrite Lúpica/etiologia , Nefrite Lúpica/imunologia , Nefrite Lúpica/virologia , Masculino , Camundongos , Camundongos Congênicos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos NZB , Chaperonas Moleculares/genética , Chaperonas Moleculares/imunologia , Infecções por Retroviridae/complicações , Infecções por Retroviridae/imunologia , Infecções por Retroviridae/virologia , Ativação Transcricional/imunologia , Proteínas do Envelope Viral/genética , Proteínas do Envelope Viral/imunologia , Vírus Relacionado ao Vírus Xenotrópico da Leucemia Murina/patogenicidadeRESUMO
Circulating monocytes are divided into two major, phenotypically and functionally distinct subsets: Gr-1(hi) "inflammatory" and Gr-1(lo) "resting" monocytes. One of the unique cellular abnormalities in lupus-prone mice is monocytosis, which is characterized by a selective expansion of Gr-1(lo) monocytes and dependent on the expression of stimulatory IgG Fc receptors (FcγR). We speculated that IgG immune complexes containing nuclear antigens could stimulate Gr-1(hi) monocytes through interaction with FcγRs and then TLR7 and TLR9, thereby promoting the maturation towards Gr-1(lo) monocytes. In the present study, we assessed this hypothesis by analyzing effects of TLR9 or TLR7 agonist on monocytes in vivo. The analysis of various surface markers differentially expressed on both subsets of monocytes in combination with selective depletion of either subset revealed that within 48 h after injection of the TLR9 agonist CpG, approximately one third of Gr-1(hi) monocytes became phenotypically identical to Gr-1(lo) monocytes. In addition, we observed approximately two-fold increases in the total monocyte population 8-24 h after injection of CpG. Moreover, the activation of TLR9 resulted in an increased expression of stimulatory FcγRIV relative to inhibitory FcγRIIB on monocytes, thereby enhancing their responsiveness to IgG immune complexes. Essentially identical results were obtained after stimulation of TLR7 with a synthetic agonist (1V136). Our results indicate that the activation of TLR7 and TLR9 not only induced the maturation of a fraction of Gr-1(hi) monocytes towards Gr-1(lo) monocytes but also promoted the overall generation of monocytes, thereby supporting the critical role of TLR7 and TLR9 for the development of monocytosis in lupus-prone mice.
Assuntos
Inflamação/patologia , Lúpus Eritematoso Sistêmico/patologia , Monócitos/imunologia , Oligodesoxirribonucleotídeos/administração & dosagem , Receptores de IgG/imunologia , Receptor 7 Toll-Like/imunologia , Receptor Toll-Like 9/imunologia , Animais , Contagem de Células Sanguíneas , Diferenciação Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Modelos Animais de Doenças , Feminino , Citometria de Fluxo , Deleção de Genes , Inflamação/imunologia , Inflamação/metabolismo , Procedimentos de Redução de Leucócitos , Lúpus Eritematoso Sistêmico/imunologia , Lúpus Eritematoso Sistêmico/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Monócitos/citologia , Monócitos/efeitos dos fármacos , Monócitos/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Receptores de IgG/deficiência , Receptores de IgG/genética , Receptor 7 Toll-Like/agonistas , Receptor 7 Toll-Like/deficiência , Receptor 7 Toll-Like/genética , Receptor Toll-Like 9/agonistas , Receptor Toll-Like 9/deficiência , Receptor Toll-Like 9/genéticaRESUMO
The endogenous retroviral envelope glycoprotein, gp70, implicated in murine lupus nephritis is secreted by hepatocytes, and its expression is largely regulated by the Sgp3 (serum gp70 production 3) locus derived from lupus-prone mice. Because of the localization of the macroH2A1 gene encoding macroH2A histone variants within the Sgp3 interval and of an up-regulated transcription of endogenous retroviral sequences in macroH2A1-deficient C57BL/6 (B6) mice, we investigated whether macroH2A1 is a candidate gene for Sgp3. macroH2A1-deficient B6 mice carrying the 129-derived Sgp3 locus, which was co-transferred with the 129 macroH2A1 mutant gene, displayed increased levels of serum gp70 and hepatic retroviral gp70 RNAs comparable to those of B6.NZB-Sgp3 congenic mice bearing the Sgp3 locus of lupus-prone NZB mice. In contrast, the abundance of retroviral gp70 RNAs in macroH2A1-deficient 129 mice was not elevated at all as compared with wild-type 129 mice. Furthermore, Sgp3 subcongenic B6 mice devoid of the NZB-derived macroH2A1 gene displayed an Sgp3 phenotype identical to that of B6.NZB-Sgp3 congenic mice carrying the NZB-derived macroH2A1 gene, thus excluding macroH2A1 as a candidate Sgp3 gene. Collectively, our data indicate that enhanced transcription of endogenous retroviral sequences observed in macroH2A1-deficient B6 mice is not a result of the macroH2A1 mutation, but due to the presence of the 129-derived Sgp3 locus. In contrast, the effect of a macroH2A1 knockout mutation on the expression of several non-retroviral cellular genes was very similar on the B6 and 129 backgrounds, indicating that these effects were due to the macroH2A1 knockout.
Assuntos
Retrovirus Endógenos/fisiologia , Hepatócitos/metabolismo , Lúpus Eritematoso Sistêmico/genética , Nefrite Lúpica/genética , Proteínas do Envelope Viral/metabolismo , Animais , Modelos Animais de Doenças , Regulação da Expressão Gênica , Glicoproteínas/genética , Hepatócitos/patologia , Histonas/genética , Humanos , Lúpus Eritematoso Sistêmico/imunologia , Nefrite Lúpica/imunologia , Camundongos , Camundongos da Linhagem 129 , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos NZB , Chaperonas Moleculares/genética , Proteínas do Envelope Viral/genéticaRESUMO
The endogenous retroviral envelope glycoprotein, gp70, implicated in murine systemic lupus erythematosus (SLE), has been considered to be a product of xenotropic, polytropic (PT) and modified PT (mPT) endogenous retroviruses. It is secreted by hepatocytes like an acute phase protein, but its response is under a genetic control. Given critical roles of TLR7 and TLR9 in the pathogenesis of SLE, we assessed their contribution to the acute phase expression of serum gp70, and defined a pivotal role of the Sgp3 (serum gp70 production 3) and Sgp4 loci in this response. Our results demonstrated that serum levels of gp70 were up-regulated in lupus-prone NZB mice injected with TLR7 or TLR9 agonist at levels comparable to those induced by injection of IL-1, IL-6 or TNF. In addition, studies of C57BL/6 Sgp3 and/or Sgp4 congenic mice defined the major roles of these two loci in up-regulated production of serum gp70 during acute phase responses. Finally, the analysis of Sgp3 congenic mice strongly suggests the presence of at least two distinct genetic factors in the Sgp3 interval, one of which controlled the basal-level expression of xenotropic, PT and mPT gp70 and the other which controlled the up-regulated production of xenotropic and mPT gp70 during acute phase responses. Our results uncovered an additional pathogenic role of TLR7 and TLR9 in murine lupus nephritis by promoting the expression of nephritogenic gp70 autoantigen. Furthermore, they revealed the involvement of multiple regulatory genes for the expression of gp70 autoantigen under steady-state and inflammatory conditions in lupus-prone mice.
Assuntos
Proteínas de Fase Aguda/biossíntese , Reação de Fase Aguda/genética , Autoantígenos/biossíntese , Retrovirus Endógenos/genética , Glicoproteínas/biossíntese , Glicoproteínas/metabolismo , Lúpus Eritematoso Sistêmico/genética , Chaperonas Moleculares/metabolismo , Proteínas Virais/biossíntese , Proteínas de Fase Aguda/genética , Proteínas de Fase Aguda/imunologia , Reação de Fase Aguda/sangue , Reação de Fase Aguda/complicações , Reação de Fase Aguda/imunologia , Reação de Fase Aguda/virologia , Animais , Autoantígenos/sangue , Autoantígenos/genética , Autoantígenos/imunologia , Citocinas/administração & dosagem , Modelos Animais de Doenças , Feminino , Regulação Viral da Expressão Gênica , Glicoproteínas/sangue , Glicoproteínas/genética , Glicoproteínas/imunologia , Humanos , Lipopolissacarídeos/administração & dosagem , Lúpus Eritematoso Sistêmico/sangue , Lúpus Eritematoso Sistêmico/complicações , Lúpus Eritematoso Sistêmico/imunologia , Lúpus Eritematoso Sistêmico/virologia , Nefrite Lúpica/etiologia , Glicoproteínas de Membrana/imunologia , Glicoproteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos NZB , Camundongos Transgênicos , Chaperonas Moleculares/genética , Oligodesoxirribonucleotídeos/administração & dosagem , Receptor 7 Toll-Like/imunologia , Receptor 7 Toll-Like/metabolismo , Receptor Toll-Like 9/imunologia , Receptor Toll-Like 9/metabolismo , Proteínas Virais/sangue , Proteínas Virais/genética , Proteínas Virais/imunologiaRESUMO
Systemic lupus erythematosus (SLE) is an autoimmune disorder characterized by B cell hyperactivity leading to the production of various autoantibodies and subsequent development of glomerulonephritis, i.e. lupus nephritis. Among the principal targets of the autoantibodies produced in murine SLE are nucleic acid-protein complexes and the envelope glycoprotein gp70 of endogenous retroviruses. Recent studies have revealed that the innate receptor TLR7 plays a pivotal role in the development of a wide variety of autoimmune responses against DNA- and RNA-containing nuclear antigens, while TLR9 rather plays a protective role. In addition, the regulation of autoimmune responses against endogenous retroviral gp70 by TLR7 suggests the implication of endogenous retroviruses in this autoimmune response. Moreover, the demonstration that TLR7 is involved in the acute phase expression of serum gp70 uncovers an additional pathogenic role of TLR7 in murine lupus nephritis by promoting the expression of nephritogenic gp70 autoantigen. Clearly, the eventual identification of endogenous retroviruses implicated in murine SLE and of mouse genes regulating their production could provide a clue for the potential role of endogenous retroviruses in human SLE.
Assuntos
Retrovirus Endógenos/fisiologia , Lúpus Eritematoso Sistêmico/virologia , Animais , Retrovirus Endógenos/imunologia , Humanos , Lúpus Eritematoso Sistêmico/fisiopatologia , Camundongos , Receptor 7 Toll-Like/metabolismo , Proteínas do Envelope Viral/metabolismoRESUMO
EndoS from Streptococcus pyogenes is an immunomodulating enzyme that specifically hydrolyzes glycans from human immunoglobulin G and thereby affects antibody effector functions. Autoimmune hemolytic anemia is caused by antibody-mediated red blood cell (RBC) destruction and often resists treatment with corticosteroids that also cause frequent adverse effects. We show here that anti-RhD (anti-D) and rabbit anti-human-RBC antibodies (anti-RBC) mediated destruction of RBC, ie, phagocytosis, complement activation, and hemolysis in vitro and in vivo was inhibited by EndoS. Phagocytosis by monocytes in vitro was inhibited by pretreatment of anti-D with EndoS before sensitization of RBCs and abrogated by direct addition of EndoS to blood containing sensitized RBCs. The toxic effects of monocytes stimulated with anti-D-sensitized RBCs, as measured by interleukin-8 secretion and oxygen metabolite production, was restrained by EndoS. Agglutination of RBCs and complement-mediated hemolysis in vitro in whole human blood caused by rabbit anti-RBCs was inhibited by EndoS. Development of anemia in mice caused by a murine anti-RBC immunoglobulin G2a monoclonal autoantibody and complement activation and erythrophagocytosis by Kupffer cells in the liver were reduced by EndoS. Our data indicate that EndoS is a potential therapeutic agent that might be evaluated as an alternative to current treatment regimens against antibody-mediated destruction of RBCs.
Assuntos
Anemia Hemolítica Autoimune , Proteínas de Bactérias/farmacologia , Glicosídeo Hidrolases/farmacologia , Hemólise/efeitos dos fármacos , Hemólise/imunologia , Imunoglobulina G/metabolismo , Anemia Hemolítica Autoimune/tratamento farmacológico , Anemia Hemolítica Autoimune/imunologia , Anemia Hemolítica Autoimune/metabolismo , Animais , Proteínas de Bactérias/metabolismo , Complemento C1q/metabolismo , Eritrócitos/efeitos dos fármacos , Eritrócitos/imunologia , Eritrócitos/metabolismo , Glicosídeo Hidrolases/metabolismo , Humanos , Imunoglobulina G/imunologia , Interleucina-8/metabolismo , Isoanticorpos/imunologia , Isoanticorpos/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Monócitos/efeitos dos fármacos , Monócitos/imunologia , Monócitos/metabolismo , Oxigênio/metabolismo , Fagocitose/efeitos dos fármacos , Fagocitose/imunologia , Coelhos , Imunoglobulina rho(D)RESUMO
Growth arrest-specific gene 6 (Gas6) is expressed in antigen-presenting cells and endothelial cells (ECs) but not in T cells. When wild-type (WT) or Gas6(-/-) mice received allogeneic non-T cell-depleted bone marrow cells, hepatic graft-versus-host disease (GVHD) was alleviated in Gas6(-/-) recipients regardless of donor genotype, but not in WT recipients. T-cell infiltration was more prominent and diffuse in WT than in Gas6(-/-) recipients' liver. When mice received 0.5 x 10(6) allogeneic T cells with T cell-depleted allogeneic bone marrow, clinical signs indicated that GVHD was less severe in Gas6(-/-) than in WT recipients, as shown by a significant improvement of the survival and reduced liver GVHD. These data demonstrate that donor cells were not involved in the protection mechanism. In addition, lack of Gas6 in antigen-presenting cells did not affect WT or Gas6(-/-) T-cell proliferation. We therefore assessed the response of WT or Gas6(-/-) ECs to tumor necrosis factor-alpha. Lymphocyte transmigration was less extensive through Gas6(-/-) than WT ECs and was not accompanied by increases in adhesion molecule levels. Thus, the lack of Gas6 in ECs impaired donor T-cell transmigration into the liver, providing a rationale for considering Gas6 pathway as a potential nonimmunosuppressive target to minimize GVHD in patients receiving allogeneic hematopoietic stem cell transplantation.
Assuntos
Doença Enxerto-Hospedeiro/prevenção & controle , Transplante de Células-Tronco Hematopoéticas/efeitos adversos , Peptídeos e Proteínas de Sinalização Intercelular/deficiência , Fígado/imunologia , Animais , Separação Celular , Quimiotaxia de Leucócito/genética , Quimiotaxia de Leucócito/imunologia , Células Endoteliais/metabolismo , Citometria de Fluxo , Doença Enxerto-Hospedeiro/genética , Doença Enxerto-Hospedeiro/imunologia , Imuno-Histoquímica , Peptídeos e Proteínas de Sinalização Intercelular/genética , Fígado/patologia , Ativação Linfocitária/imunologia , Teste de Cultura Mista de Linfócitos , Camundongos , Camundongos Knockout , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Linfócitos T/citologia , Linfócitos T/imunologia , Linfócitos T/metabolismo , Transplante HomólogoRESUMO
Systemic lupus erythematosus (SLE) is an autoimmune disorder characterized by B cell hyperactivity leading to the production of various autoantibodies and subsequent development of glomerulonephritis, i.e. lupus nephritis. Among the principal targets of autoantibodies produced in murine SLE are nucleic acid-protein complexes, such as chromatin and ribonucleoproteins, and the envelope glycoprotein gp70 of endogenous retroviruses. The preferential production of these autoantibodies is apparently promoted by the presence of genetic abnormalities leading to defects in the elimination of apoptotic cells and to an enhanced expression of endogenous retroviruses. Moreover, recent studies revealed that the innate receptors TLR7 and TLR9 are critically involved in the activation of dendritic cells and autoreactive B cells through the recognition of endogenous DNA- or RNA-containing antigens and subsequent development of autoimmune responses against nuclear autoantigens. Furthermore, the regulation of autoimmune responses against endogenous retroviral gp70 by TLR7 suggested the implication of endogenous retroviruses in this autoimmune response. Clearly, further elucidation of the precise molecular role of TLR7 and TLR9 in the development of autoimmune responses will help to develop novel therapeutic strategies and targets for SLE.
Assuntos
Linfócitos B/imunologia , Células Dendríticas/imunologia , Lúpus Eritematoso Sistêmico/imunologia , Glicoproteínas de Membrana/imunologia , Receptor 7 Toll-Like/imunologia , Receptor Toll-Like 9/imunologia , Animais , Autoanticorpos/sangue , Autoantígenos/imunologia , Linfócitos B/metabolismo , Células Dendríticas/metabolismo , Modelos Animais de Doenças , Retrovirus Endógenos/imunologia , Retrovirus Endógenos/metabolismo , Lúpus Eritematoso Sistêmico/metabolismo , Ativação Linfocitária/imunologia , Glicoproteínas de Membrana/metabolismo , Camundongos , Receptor 7 Toll-Like/metabolismo , Receptor Toll-Like 9/metabolismo , Proteínas Virais/imunologia , Proteínas Virais/metabolismoRESUMO
OBJECTIVE: Lupus-prone BXSB mice develop monocytosis characterized by selective accumulation of the Gr-1- monocyte subset. The aim of this study was to explore the possible role of activating IgG Fc receptors (FcgammaR) in the development of monocytosis and to characterize the functional phenotype of the Gr-1- subset that accumulates in lupus-prone mice bearing the NZB-type defective Fcgr2b allele for the inhibitory FcgammaRIIB. METHODS: The development of monocytosis was analyzed in BXSB and anti-IgG2a rheumatoid factor-transgenic C57BL/6 mice deficient in activating FcgammaR. Moreover, we assessed the expression levels of activating FcgammaR and inhibitory FcgammaRIIB on Gr-1+ and Gr-1- monocyte subsets in C57BL/6 mice bearing the C57BL/6-type or the NZB-type Fcgr2b allele. RESULTS: We observed monocytosis with expansion of the Gr-1- subset in anti-IgG2a-transgenic C57BL/6 mice expressing IgG2a, but not in those lacking IgG2a. Moreover, monocytosis barely developed in BXSB and anti-IgG2a-transgenic C57BL/6 mice deficient in activating FcgammaR. The Gr-1- subset that accumulated in lupus-prone mice displayed a unique hyperactive phenotype. It expressed very low levels of inhibitory FcgammaRIIB, due to the presence of the NZB-type Fcgr2b allele, but high levels of activating FcgammaRIV. This was in contrast to high levels of FcgammaRIIB expression and no FcgammaRIV expression on the Gr-1+ subset. CONCLUSION: Our results demonstrated a critical role of activating FcgammaR in the development of monocytosis and in the expansion of a Gr-1-FcgammaRIIB(low)FcgammaRIV+ hyperactive monocyte subset in lupus-prone mice. Our findings further highlight the importance of the NZB-type Fcgr2b susceptibility allele in murine lupus, the presence of which induces increased production of hyperactive monocytes as well as dysregulated activation of autoreactive B cells.
Assuntos
Lúpus Eritematoso Sistêmico/genética , Lúpus Eritematoso Sistêmico/imunologia , Monócitos/imunologia , Receptores de IgG/biossíntese , Receptores de IgG/genética , Animais , Contagem de Células , Proliferação de Células , Modelos Animais de Doenças , Feminino , Citometria de Fluxo , Endogamia , Lúpus Eritematoso Sistêmico/sangue , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Monócitos/patologia , Receptores de Quimiocinas/genética , Receptores de Quimiocinas/metabolismo , Receptores de IgG/deficiência , Receptores de IgG/imunologiaRESUMO
Endogenous retroviruses are implicated in the pathogenesis of systemic lupus erythematosus (SLE). Because four different classes of endogenous retroviruses, i.e., ecotropic, xenotropic, polytropic, or modified polytropic (mPT), are expressed in mice, we investigated the possibility that a particular class of endogenous retroviruses is associated with the development of murine SLE. We observed >15-fold increased expression of mPT env (envelope) RNA in livers of all four lupus-prone mice, as compared with those of nine nonautoimmune strains of mice. This was not the case for the three other classes of retroviruses. Furthermore, we found that in addition to intact mPT transcripts, many strains of mice expressed two defective mPT env transcripts which carry a deletion in the env sequence of the 3' portion of the gp70 surface protein and the 5' portion of the p15E transmembrane protein, respectively. Remarkably, in contrast to nonautoimmune strains of mice, all four lupus-prone mice expressed abundant levels of intact mPT env transcripts, but only low or nondetectable levels of the mutant env transcripts. The Sgp3 (serum gp70 production 3) locus derived from lupus-prone mice was responsible for the selective up-regulation of the intact mPT env RNA. Finally, we observed that single-stranded RNA-specific TLR7 played a critical role in the production of anti-gp70 autoantibodies. These data suggest that lupus-prone mice may possess a unique genetic mechanism responsible for the expression of mPT retroviruses, which could act as a triggering factor through activating TLR7 for the development of autoimmune responses in mice predisposed to SLE.
Assuntos
Retrovirus Endógenos/metabolismo , Produtos do Gene env/metabolismo , Glicoproteínas/genética , Glicoproteínas/metabolismo , Lúpus Eritematoso Sistêmico/metabolismo , Chaperonas Moleculares/genética , Chaperonas Moleculares/metabolismo , RNA Viral/genética , Regulação para Cima , Alelos , Sequência de Aminoácidos , Animais , Sequência de Bases , Retrovirus Endógenos/genética , Produtos do Gene env/genética , Glicoproteínas de Membrana/metabolismo , Camundongos , Dados de Sequência Molecular , Mutação/genética , Alinhamento de Sequência , Receptor 7 Toll-Like/metabolismo , Transcrição Gênica/genéticaRESUMO
Replacement of aspartic acid by alanine at position 265 (D265A) in mouse IgG1 results in a complete loss of interaction between this isotype and low-affinity IgG Fc receptors (Fc gammaRIIB and Fc gammaRIII). However, it has not yet been defined whether the D265A substitution could exhibit similar effects on the interaction with two other Fc gammaR (Fc gammaRI and Fc gammaRIV) and on the activation of complement. To address this question, 34-3C anti-RBC IgG2a and IgG2b switch variants bearing the D265A mutation were generated, and their effector functions and in vivo pathogenicity were compared with those of the respective wild-type Abs. The introduction of the D265A mutation almost completely abolished the binding of 34-3C IgG2a and IgG2b to all four classes of Fc gammaR and the activation of complement. Consequently, these mutants were hardly pathogenic. Although oligosaccharide side chains of these mutants were found to contain higher levels of sialic acids than those of wild-type Abs, the analysis of enzymatically desialylated D265A variants ruled out the possibility that very poor Fc-associated effector functions of the D265A mutants were due to an increased level of the mutant Fc sialylation. Thus, our results demonstrate that aspartic acid at position 265 is a residue critically implicated in triggering the Fc-associated effector functions of IgG, probably by defining a crucial three-dimensional structure of the Fc region.
Assuntos
Ácido Aspártico/fisiologia , Ativação do Complemento/imunologia , Imunoglobulina G/metabolismo , Alanina/genética , Substituição de Aminoácidos/genética , Substituição de Aminoácidos/fisiologia , Anemia Hemolítica Autoimune/genética , Anemia Hemolítica Autoimune/imunologia , Animais , Anticorpos Monoclonais/toxicidade , Ácido Aspártico/genética , Autoanticorpos/toxicidade , Ativação do Complemento/genética , Eritrócitos/imunologia , Imunoglobulina G/química , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , Isoformas de Proteínas/química , Isoformas de Proteínas/deficiência , Isoformas de Proteínas/genética , Isoformas de Proteínas/fisiologia , Estrutura Terciária de Proteína/genética , Estrutura Terciária de Proteína/fisiologia , Receptores Fc/química , Receptores Fc/deficiência , Receptores Fc/genética , Receptores Fc/fisiologia , Ácidos Siálicos/genética , Relação Estrutura-AtividadeRESUMO
Four murine IgG subclasses display markedly different Fc-associated effector functions because of their differential binding to three activating IgG Fc receptors (FcgammaRI, FcgammaRIII, and FcgammaRIV) and C1q. Previous analysis of IgG subclass switch variants of 34-3C anti-RBC monoclonal autoantibodies revealed that the IgG1 subclass, which binds only to FcgammaRIII and fails to activate complement, displayed the poorest pathogenic potential. This could be related to the presence of a three amino acid deletion at positions 233-235 in the CH2 domain uniquely found in this subclass. To address this question, IgG1 insertion and IgG2b deletion mutants at positions 233-235 of 34-3C anti-RBC Abs were generated, and their ability to initiate effector functions and their pathogenicity were compared with those of the respective wild-type Abs. The insertion of amino acid residues at positions 233-235 enabled the IgG1 subclass to bind FcgammaRIV but did not improve the binding to C1q. Accordingly, its pathogenicity was enhanced but still inferior to that of IgG2b. In contrast, the IgG2b deletion mutant lost its ability to bind to FcgammaRIV and activate complement. Consequently, its pathogenicity was markedly diminished to a level comparable to that of IgG1. Our results demonstrated that the initiation of FcgammaR- and complement-mediated effector functions of IgG2b was profoundly affected by the three amino acid deletion at positions 233-235, but that this natural three amino acid deletion could only partially explain the poor binding of IgG1 to FcgammaRIV and C1q. This indicates the lack in the IgG1 subclass of as yet unknown motifs promoting efficient interaction with FcgammaRIV and C1q.
Assuntos
Aminoácidos/química , Aminoácidos/genética , Imunoglobulina G/metabolismo , Cadeias Pesadas de Imunoglobulinas/genética , Receptores de IgG/antagonistas & inibidores , Deleção de Sequência , Aminoácidos/metabolismo , Anemia Hemolítica Autoimune/genética , Anemia Hemolítica Autoimune/imunologia , Animais , Afinidade de Anticorpos/genética , Regulação para Baixo/genética , Regulação para Baixo/imunologia , Imunoglobulina G/genética , Cadeias Pesadas de Imunoglobulinas/biossíntese , Cadeias Pesadas de Imunoglobulinas/metabolismo , Região de Troca de Imunoglobulinas , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos NZB , Camundongos Knockout , Mutagênese Sítio-Dirigida , Estrutura Terciária de Proteína/genética , Receptores de IgG/genética , Receptores de IgG/metabolismoRESUMO
The endogenous retroviral envelope glycoprotein, gp70, implicated in murine lupus nephritis is secreted by hepatocytes as an acute phase protein, and it has been thought to be a product of an endogenous xenotropic virus, NZB-X1. However, since endogenous polytropic (PT) and modified polytropic (mPT) viruses encode gp70s that are closely related to xenotropic gp70, these viruses can be additional sources of serum gp70. To better understand the genetic basis of the expression of serum gp70, we analyzed the abundance of xenotropic, PT, or mPT gp70 RNAs in livers and the genomic composition of corresponding proviruses in various strains of mice, including two different Sgp (serum gp70 production) congenic mice. Our results demonstrated that the expression of different viral gp70 RNAs was remarkably heterogeneous among various mouse strains and that the level of serum gp70 production was regulated by multiple structural and regulatory genes. Additionally, a significant contribution of PT and mPT gp70s to serum gp70 was revealed by the detection of PT and mPT, but not xenotropic transcripts in 129 mice, and by a closer correlation of serum levels of gp70 with the abundance of PT and mPT gp70 RNAs than with that of xenotropic gp70 RNA in Sgp3 congenic mice. Furthermore, the injection of lipopolysaccharides selectively up-regulated the expression of xenotropic and mPT gp70 RNAs, but not PT gp70 RNA. Our data indicate that the genetic origin of serum gp70 is more heterogeneous than previously thought, and that distinct retroviral gp70s are differentially regulated in physiological vs inflammatory conditions.
Assuntos
Retrovirus Endógenos/imunologia , Glicoproteínas/biossíntese , Glicoproteínas/genética , Nefrite Lúpica/genética , Animais , Autoantígenos/biossíntese , Autoantígenos/genética , Autoantígenos/imunologia , Retrovirus Endógenos/genética , Glicoproteínas/sangue , Mediadores da Inflamação/sangue , Mediadores da Inflamação/fisiologia , Nefrite Lúpica/sangue , Nefrite Lúpica/patologia , Camundongos , Camundongos Congênicos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos MRL lpr , Camundongos Endogâmicos NZB , Chaperonas Moleculares/biossíntese , Chaperonas Moleculares/genéticaRESUMO
Murine phagocytes express three different activating IgG FcgammaR: FcgammaRI is specific for IgG2a; FcgammaRIII for IgG1, IgG2a, and IgG2b; and FcgammaRIV for IgG2a and IgG2b. Although the role of FcgammaRIII in IgG1 and IgG2a anti-RBC-induced autoimmune hemolytic anemia (AIHA) is well documented, the contribution of FcgammaRI and FcgammaRIV to the development of IgG2a- and IgG2b-induced anemia has not yet been defined. In the present study, using mice deficient in FcgammaRI, FcgammaRIII, and C3, in combination with an FcgammaRIV-blocking mAb, we assessed the respective roles of these three FcgammaR in the development of mild and severe AIHA induced by two different doses (50 and 200 microg) of the IgG2a and IgG2b subclasses of the 34-3C anti-RBC monoclonal autoantibody. We observed that the development of mild anemia induced by a low dose of 34-3C IgG2a autoantibody was highly dependent on FcgammaRIII, while FcgammaRI and FcgammaRIV additionally contributed to the development of severe anemia induced by a high dose of this subclass. In contrast, the development of both mild and severe anemia induced by 34-3C IgG2b was dependent on FcgammaRIII and FcgammaRIV. Our results indicate differential roles of the three activating FcgammaR in IgG2a- and IgG2b-mediated AIHA.