RESUMEN
The costimulatory receptor 4-1BB is expressed on activated immune cells, including activated T cells. Antibodies targeting 4-1BB enhance the proliferation and survival of antigen-stimulated T cells in vitro and promote CD8 T cell-dependent anti-tumor immunity in pre-clinical cancer models. We found that T regulatory (Treg) cells infiltrating human or murine tumors expressed high amounts of 4-1BB. Intra-tumoral Treg cells were preferentially depleted by anti-4-1BB mAbs in vivo. Anti-4-1BB mAbs also promoted effector T cell agonism to promote tumor rejection. These distinct mechanisms were competitive and dependent on antibody isotype and FcγR availability. Administration of anti-4-1BB IgG2a, which preferentially depletes Treg cells, followed by either agonistic anti-4-1BB IgG1 or anti-PD-1 mAb augmented anti-tumor responses in multiple solid tumor models. An antibody engineered to optimize both FcγR-dependent Treg cell depleting capacity and FcγR-independent agonism delivered enhanced anti-tumor therapy. These insights into the effector mechanisms of anti-4-1BB mAbs lay the groundwork for translation into the clinic.
Asunto(s)
Anticuerpos Monoclonales/farmacología , Linfocitos T CD8-positivos/inmunología , Linfocitos T CD8-positivos/metabolismo , Inmunomodulación/efectos de los fármacos , Neoplasias/inmunología , Neoplasias/metabolismo , Linfocitos T Reguladores/inmunología , Linfocitos T Reguladores/metabolismo , Miembro 9 de la Superfamilia de Receptores de Factores de Necrosis Tumoral/antagonistas & inhibidores , Animales , Expresión Génica , Humanos , Inmunoglobulina G/farmacología , Linfocitos Infiltrantes de Tumor/inmunología , Linfocitos Infiltrantes de Tumor/metabolismo , Ratones , Ratones Noqueados , Neoplasias/genética , Miembro 9 de la Superfamilia de Receptores de Factores de Necrosis Tumoral/genética , Miembro 9 de la Superfamilia de Receptores de Factores de Necrosis Tumoral/metabolismoRESUMEN
Infection with viruses, such as the lactate dehydrogenase-elevating virus (LDV), is known to trigger the onset of autoimmune anemia through the enhancement of the phagocytosis of autoantibody-opsonized erythrocytes by activated macrophages. Type I interferon receptor-deficient mice show enhanced anemia, which suggests a protective effect of these cytokines, partly through the control of type II interferon production. The development of anemia requires the expression of Fcγ receptors (FcγR) I, III, and IV. Whereas LDV infection decreases FcγR III expression, it enhances FcγR I and IV expression in wild-type animals. The LDV-associated increase in the expression of FcγR I and IV is largely reduced in type I interferon receptor-deficient mice, through both type II interferon-dependent and -independent mechanisms. Thus, the regulation of the expression of FcγR I and IV, but not III, by interferons may partly explain the exacerbating effect of LDV infection on anemia that results from the enhanced phagocytosis of IgG autoantibody-opsonized erythrocytes.
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Anemia Hemolítica Autoinmune/inmunología , Infecciones por Arterivirus/inmunología , Interferones/metabolismo , Virus Elevador de Lactato Deshidrogenasa/inmunología , Receptores de IgG/metabolismo , Anemia Hemolítica Autoinmune/virología , Animales , Infecciones por Arterivirus/virología , Interacciones Huésped-Patógeno , Ratones Endogámicos C57BL , Ratones Noqueados , FagocitosisRESUMEN
OBJECTIVE: Macrophage interleukin (IL)-10 signalling plays a critical role in the maintenance of a regulatory phenotype that prevents the development of IBD. We have previously found that anti-tumour necrosis factor (TNF) monoclonal antibodies act through Fcγ-receptor (FcγR) signalling to promote repolarisation of proinflammatory intestinal macrophages to a CD206+ regulatory phenotype. The role of IL-10 in anti-TNF-induced macrophage repolarisation has not been examined. DESIGN: We used human peripheral blood monocytes and mouse bone marrow-derived macrophages to study IL-10 production and CD206+ regulatory macrophage differentiation. To determine whether the efficacy of anti-TNF was dependent on IL-10 signalling in vivo and in which cell type, we used the CD4+CD45Rbhigh T-cell transfer model in combination with several genetic mouse models. RESULTS: Anti-TNF therapy increased macrophage IL-10 production in an FcγR-dependent manner, which caused differentiation of macrophages to a more regulatory CD206+ phenotype in vitro. Pharmacological blockade of IL-10 signalling prevented the induction of these CD206+ regulatory macrophages and diminished the therapeutic efficacy of anti-TNF therapy in the CD4+CD45Rbhigh T-cell transfer model of IBD. Using cell type-specific IL-10 receptor mutant mice, we found that IL-10 signalling in macrophages but not T cells was critical for the induction of CD206+ regulatory macrophages and therapeutic response to anti-TNF. CONCLUSION: The therapeutic efficacy of anti-TNF in resolving intestinal inflammation is critically dependent on IL-10 signalling in macrophages.
Asunto(s)
Enfermedades Inflamatorias del Intestino/tratamiento farmacológico , Interleucina-10/metabolismo , Macrófagos/metabolismo , Transducción de Señal , Factor de Necrosis Tumoral alfa/antagonistas & inhibidores , Adulto , Animales , Anticuerpos Monoclonales , Enfermedad de Crohn/tratamiento farmacológico , Enfermedad de Crohn/metabolismo , Modelos Animales de Enfermedad , Femenino , Humanos , Enfermedades Inflamatorias del Intestino/metabolismo , Mucosa Intestinal/metabolismo , Macrófagos/efectos de los fármacos , Masculino , Ratones , Ratones Noqueados , Persona de Mediana Edad , Transducción de Señal/efectos de los fármacos , Adulto JovenRESUMEN
Infection of C57BL/6 mice with lymphocytic choriomeningitis virus (LCMV) strain Armstrong (Arm) induces an acute infection with rapid virus clearance by CD8+ T cells independently of CD4+ T cell help. Residual viral antigen may, however, persist for a prolonged time. Here, we demonstrate that mice that had been transiently depleted of CD4+ T cells during acute LCMV Arm infection generated high levels of virus-specific IgG antibodies (Ab) after viral clearance. Robust induction of LCMV-specific IgG after transient CD4+ T cell depletion was dependent on Fcγ receptors but not on the complement receptors CD21/CD35. In contrast to the potent production of LCMV-specific IgG, the generation of LCMV-specific isotype-switched memory B cells after transient CD4+ T cell depletion was considerably reduced. Moreover, mice depleted of CD4+ T cells during acute infection were strongly impaired in generating a secondary LCMV-specific B cell response upon LCMV rechallenge. In conclusion, our data indicate that LCMV antigen depots after viral clearance were capable of inducing high levels of virus-specific IgG. They failed, however, to induce robust virus-specific B cell memory revealing a previously unappreciated dichotomy of specific Ab production and memory cell formation after priming with residual antigen.
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Anticuerpos Antivirales/inmunología , Antígenos Virales/inmunología , Linfocitos B/inmunología , Linfocitos B/metabolismo , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD4-Positivos/metabolismo , Coriomeningitis Linfocítica/inmunología , Virus de la Coriomeningitis Linfocítica/inmunología , Animales , Biomarcadores , Inmunoglobulina G/inmunología , Inmunoglobulina G/metabolismo , Memoria Inmunológica , Inmunofenotipificación , Depleción Linfocítica , Coriomeningitis Linfocítica/virología , Ratones , Ratones Noqueados , Células Plasmáticas/inmunología , Células Plasmáticas/metabolismo , Linfocitos T Citotóxicos/inmunologíaRESUMEN
AbstractMonocytes are evolutionally conserved innate immune cells that play essential roles for the protection of the host against pathogens and also produce several inflammatory cytokines. Thus, the aberrant functioning of monocytes may affect not only host defense but also the development of inflammatory diseases. Monocytes are a heterogeneous population with phenotypical and functional differences. Most recent studies have shown that monocytes are divided into three subsets, namely classical, intermediate and non-classical subsets, both in humans and mice. Accumulating evidence showed that monocyte activation is associated with the disease progression in autoimmune diseases, such as systemic lupus erythematosus (SLE) and rheumatoid arthritis (RA). However, it remains to be determined how monocytes contribute to the disease process and which subset is involved. In this review, we discuss the pathogenic role of monocyte subsets in SLE and RA on the basis of current studies by ourselves and others to shed light on the suitability of monocyte-targeted therapies in these diseases.
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Artritis Reumatoide/inmunología , Lupus Eritematoso Sistémico/inmunología , Monocitos/inmunología , Animales , HumanosRESUMEN
C57BL/6 (B6).FcγRIIb-/- Yaa mice spontaneously develop lethal lupus nephritis. To define the cell type-specific role of FcγRIIb in Yaa-associated lupus, we established B cell- (CD19Cre Yaa), myeloid cell- (C/EBPαCre Yaa), and dendritic cell- (DC) (CD11cCre Yaa) specific FcγRIIb-deficient B6.Yaa mouse strains. CD19Cre Yaa mice developed milder lupus than B6.FcγRIIb-/- Yaa mice, indicating that FcγRIIb deficiency on B cells is not sufficient for the development of severe disease. Surprisingly, C/EBPαCre Yaa mice also showed autoantibody production and mild lupus similar to that in CD19Cre Yaa mice, whereas CD11cCre Yaa mice stayed disease free. These observations indicate that FcγRIIb deficiency in B cells and myeloid cells, but not DCs, contributes to the severe disease in B6.FcγRIIb-/- Yaa mice. Flow cytometric analysis showed that the frequency of peripheral Gr-1- but not Gr-1+ monocyte was increased in B6.FcγRIIb-/- Yaa and C/EBPαCre Yaa but not CD19Cre Yaa mice, suggesting a link between FcγRIIb deficiency on myeloid cells and the high frequency of Gr-1- monocytes. RNA sequencing revealed that compared with Gr-1+ monocytes, Gr-1- monocytes expressed higher levels of the B cell-stimulating cytokines BSF-3, IL-10, and IL-1ß, the DC markers CD11c, CD83, and Adamdec1, and the antiapoptotic factors Bcl2 and Bcl6. In conclusion, in Yaa-associated lupus nephritis, FcγRIIb on B cells and myeloid cells modulates B cell activation via different but synergistic pathways. Gr-1- monocytes are the most likely candidate myeloid cells involved.
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Linfocitos B/fisiología , Células Dendríticas/fisiología , Nefritis Lúpica/inmunología , Células Mieloides/fisiología , Receptores de IgG/metabolismo , Animales , Antígenos CD19/genética , Antígenos CD19/metabolismo , Autoanticuerpos/metabolismo , Células Cultivadas , Susceptibilidad a Enfermedades , Humanos , Activación de Linfocitos , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Receptores de IgG/genéticaRESUMEN
Therapy with tumor-specific Abs is common in the clinic but has limited success against solid malignancies. We aimed at improving the efficacy of this therapy by combining a tumor-specific Ab with immune-activating compounds. In this study, we demonstrate in the aggressive B16F10 mouse melanoma model that concomitant application of the anti-TRP1 Ab (clone TA99) with TLR3-7/8 or -9 ligands, and IL-2 strongly enhanced tumor control in a therapeutic setting. Depletion of NK cells, macrophages, or CD8+ T cells all mitigated the therapeutic response, showing a coordinated immune rejection by innate and adaptive immune cells. FcγRs were essential for the therapeutic effect, with a dominant role for FcγRI and a minor role for FcγRIII and FcγRIV. FcγR expression on NK cells and granulocytes was dispensable, indicating that other tumoricidal functions of NK cells were involved and implicating that FcγRI, -III, and -IV exerted their activity on macrophages. Indeed, F4/80+Ly-6C+ inflammatory macrophages in the tumor microenvironment displayed high levels of these receptors. Whereas administration of the anti-TRP1 Ab alone reduced the frequency of these macrophages, the combination with a TLR agonist retained these cells in the tumor microenvironment. Thus, the addition of innate stimulatory compounds, such as TLR ligands, to tumor-specific Ab therapy could greatly enhance its efficacy in solid cancers via optimal exploitation of FcγRs.
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Anticuerpos/uso terapéutico , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Inmunoterapia/métodos , Interleucina-2/uso terapéutico , Células Asesinas Naturales/inmunología , Macrófagos/inmunología , Melanoma/terapia , Receptores de IgG/metabolismo , Animales , Antígenos de Neoplasias/inmunología , Modelos Animales de Enfermedad , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Inmunización , Masculino , Melanoma/inmunología , Melanoma Experimental , Glicoproteínas de Membrana/inmunología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Oxidorreductasas/inmunología , Receptores de IgG/genética , Receptores Toll-Like/agonistasRESUMEN
By their interaction with IgG immune complexes, FcγR and complement link innate and adaptive immunity, showing functional redundancy. In complement-deficient mice, IgG downstream effector functions are often impaired, as well as adaptive immunity. Based on a variety of model systems using FcγR-knockout mice, it has been concluded that FcγRs are also key regulators of innate and adaptive immunity; however, several of the model systems underpinning these conclusions suffer from flawed experimental design. To address this issue, we generated a novel mouse model deficient for all FcγRs (FcγRI/II/III/IV-/- mice). These mice displayed normal development and lymphoid and myeloid ontogeny. Although IgG effector pathways were impaired, adaptive immune responses to a variety of challenges, including bacterial infection and IgG immune complexes, were not. Like FcγRIIb-deficient mice, FcγRI/II/III/IV-/- mice developed higher Ab titers but no autoantibodies. These observations indicate a redundant role for activating FcγRs in the modulation of the adaptive immune response in vivo. We conclude that FcγRs are downstream IgG effector molecules with a restricted role in the ontogeny and maintenance of the immune system, as well as the regulation of adaptive immunity.
RESUMEN
Immunomodulatory antibodies blocking interactions of coinhibitory receptors to their ligands such as CTLA-4, PD1 and PD-L1 on immune cells have shown impressive therapeutic efficacy in clinical studies. The therapeutic effect of these antibodies is mainly mediated by reactivating antitumor T cell immune responses. Detailed analysis of anti-CTLA4 antibody therapy revealed that an optimal therapeutic efficacy also requires binding to Fc receptors for IgG, FcγR, mediating depletion of intratumoral regulatory T cells. Here, we investigated the role of Fc binding in anti-PD-L1 antibody therapy in the MC38 C57BL/6 and CT26 BALB/c colon adenocarcinoma tumor models. In the MC38 tumor model, all IgG subclasses anti-PD-L1 showed similar therapeutic efficacy when compared to each other in either wild-type mice or in mice deficient for all FcγR. In contrast, in the CT26 tumor model, anti-PD-L1 mIgG2a, the IgG subclass with the highest affinity for activating FcγR, showed stronger therapeutic efficacy than other IgG subclasses. This was associated with a reduction of a myeloid cell subset with high expression of PD-L1 in the tumor microenvironment. This subclass preference for mIgG2a was lost in C57BL/6 × BALB/c F1 mice, indicating that the genetic background of the host may determine the additional clinical benefit of the high affinity antibody subclasses. Based on these data, we conclude that FcγR are not crucial for anti-PD-L1 antibody therapy but might play a role in some tumor models.
Asunto(s)
Adenocarcinoma , Antineoplásicos Inmunológicos/farmacología , Antígeno B7-H1/antagonistas & inhibidores , Neoplasias del Colon , Receptores de IgG , Animales , Anticuerpos Monoclonales , Modelos Animales de Enfermedad , Inmunoterapia/métodos , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BLRESUMEN
Dendritic cells (DCs) are specialized in Ag engulfment via a wide variety of uptake receptors on their cell surface. In the present study we investigated Ag uptake and presentation of in vivo-formed Ag-Ab complexes by i.v. injecting mice with Ag-specific Abs followed by the cognate Ag. We show by this natural Ab-mediated Ag targeting system that uptake by splenic APC subsets is severely hampered in mice lacking complement factor C1q (C1qa-/-). Moreover, no detectable Ag cross-presentation by CD8α+ DCs from C1qa-/- mice was found. On the contrary, Ag uptake was not hampered by APCs in FcγRI/II/III/IV-deficient (FcγR quadruple-/-) mice, and the cross-presentation ability of CD8α+ DCs was not affected. In conclusion, we show that C1q rather than FcγRs controls the Ab-mediated Ag uptake and its presentation by spleen APC subsets to T cells.
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Presentación de Antígeno , Complejo Antígeno-Anticuerpo/inmunología , Complemento C1q/inmunología , Células Dendríticas/inmunología , Inmunidad Adaptativa , Animales , Antígenos CD8/inmunología , Complemento C1q/deficiencia , Complemento C1q/genética , Reactividad Cruzada , Ratones , Ratones Endogámicos C57BL , Receptores de IgG/inmunologíaRESUMEN
BACKGROUND & AIMS: Although tumor necrosis factor (TNF) antagonists reduce many clinical features of inflammatory bowel disease, complete mucosal healing occurs in fewer than 50% of patients. The Fc-region of monoclonal antibodies against TNF has immunosuppressive properties via effects on macrophage polarization. We examined the interaction between the anti-TNF Fc-region and Fcγ receptors (FcγR), and whether the absence of the Fc core fucose (which increases binding to FcγRIIIa) increases the efficacy of anti-TNF in mice with colitis. METHODS: We generated Rag1-/- mice that lack all activating FcγRs (FcγRI, FcγRIII, and FcγRIV; called FcγR-/-Rag1-/- mice). We produced hypo-fucosylated antibodies against mouse and human TNF (adalimumab). Colitis was induced in mice by transfer of CD4+CD45RBhi to FcγR-/-Rag1-/- or Rag1-/- littermates; mice were given different antibodies against TNF or isotype (control) antibodies and disease activity index scores were determined. Colon tissues were collected and analyzed by histology. Human peripheral blood mononuclear cells (PBMCs) were isolated from blood of healthy donors. T-cell proliferation and proportions of CD206+ (immune regulatory) macrophages were measured in mixed lymphocyte reactions. Human PBMCs were genotyped for FCGR3A158 (the FcγRIIIa-158F allotype displays a lower Fc binding affinity) using the TaqMan single nucleotide polymorphism genotype assay. RESULTS: Rag1-/- mice with colitis given anti-TNF had near complete mucosal healing and Rag1-/- mice given an isotype control antibody developed severe colitis. In contrast, FcγR-/-Rag1-/- mice were refractory to the effects of anti-TNF: their histological colitis scores were as severe as those from FcγR-/-Rag1-/- mice given a control antibody. Colons from Rag1-/- mice that received anti-TNF had an increased number of CD206+ macrophages compared with Rag1-/- mice given control antibody; in FcγR-/-Rag1-/- mice given anti-TNF these numbers were as low as FcγR-/-Rag1-/- given the control antibody. In human PBMCs, anti-TNF increased the number of CD206+ macrophages: this required expression of FcγRIIIa; numbers of these cells were reduced in PBMCs with the low-affinity FcγRIIIa-158F genotype. A hypo-fucosylated form of adalimumab bound human FcγRIIIa with a higher affinity than control adalimumab. When hypo-fucosylated adalimumab was added to PBMCs, a larger number of CD206+ macrophages formed and T-cell proliferation was reduced, compared with addition of a control adalimumab. Hypo-fucosylated adalimumab increased the number of CD206+ macrophages in PMBCs that expressed the low-affinity FcγRIIIa. In mice with colitis, hypo-fucosylated anti-TNF significantly increased the number of CD206+ macrophages in the colon compared with control anti-TNF and was more effective in reducing colitis severity as measured by histology. CONCLUSIONS: In a study of the in vitro and in vivo mechanisms of anti-TNF, we found FcγR engagement by anti-TNF to be required for reduction of colitis in mice and development of CD206+ macrophages. A hypo-fucosylated form of anti-TNF binds FcγRIIIa with higher affinity and induces development of CD206+ macrophages in human PBMCs, especially PBMCs that express low-affinity FcγRIIIa. Hypo-fucosylated anti-TNF might be more effective in patients with inflammatory bowel disease.
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Adalimumab/farmacología , Anticuerpos Monoclonales/farmacología , Colitis/tratamiento farmacológico , Colon/efectos de los fármacos , Inmunosupresores/farmacología , Mucosa Intestinal/efectos de los fármacos , Factor de Necrosis Tumoral alfa/antagonistas & inhibidores , Traslado Adoptivo , Animales , Proliferación Celular/efectos de los fármacos , Células Cultivadas , Colitis/genética , Colitis/inmunología , Colitis/metabolismo , Colon/inmunología , Colon/metabolismo , Colon/patología , Modelos Animales de Enfermedad , Predisposición Genética a la Enfermedad , Proteínas de Homeodominio/genética , Proteínas de Homeodominio/metabolismo , Mucosa Intestinal/inmunología , Mucosa Intestinal/metabolismo , Mucosa Intestinal/patología , Lectinas Tipo C/inmunología , Lectinas Tipo C/metabolismo , Activación de Linfocitos/efectos de los fármacos , Macrófagos/efectos de los fármacos , Macrófagos/inmunología , Macrófagos/metabolismo , Receptor de Manosa , Lectinas de Unión a Manosa/inmunología , Lectinas de Unión a Manosa/metabolismo , Ratones Endogámicos C57BL , Ratones Noqueados , Fenotipo , Receptores de Superficie Celular/inmunología , Receptores de Superficie Celular/metabolismo , Receptores de IgG/deficiencia , Receptores de IgG/genética , Linfocitos T/efectos de los fármacos , Linfocitos T/inmunología , Linfocitos T/metabolismo , Linfocitos T/trasplante , Factores de Tiempo , Factor de Necrosis Tumoral alfa/inmunología , Cicatrización de Heridas/efectos de los fármacosRESUMEN
Helminth parasites can cause considerable damage when migrating through host tissues, thus making rapid tissue repair imperative to prevent bleeding and bacterial dissemination particularly during enteric infection. However, how protective type 2 responses targeted against these tissue-disruptive multicellular parasites might contribute to homeostatic wound healing in the intestine has remained unclear. Here, we observed that mice lacking antibodies (Aid-/-) or activating Fc receptors (Fcrg-/-) displayed impaired intestinal repair following infection with the murine helminth Heligmosomoides polygyrus bakeri (Hpb), whilst transfer of immune serum could partially restore chemokine production and rescue wound healing in Aid-/- mice. Impaired healing was associated with a reduced expression of CXCR2 ligands (CXCL2/3) by macrophages (MΦ) and myofibroblasts (MF) within intestinal lesions. Whilst antibodies and helminths together triggered CXCL2 production by MΦ in vitro via surface FcR engagement, chemokine secretion by intestinal MF was elicited by helminths directly via Fcrg-chain/dectin2 signaling. Blockade of CXCR2 during Hpb challenge infection reproduced the delayed wound repair observed in helminth infected Aid-/- and Fcrg-/- mice. Finally, conditioned media from human MΦ stimulated with infective larvae of the helminth Ascaris suum together with immune serum, promoted CXCR2-dependent scratch wound closure by human MF in vitro. Collectively our findings suggest that helminths and antibodies instruct a chemokine driven MΦ-MF crosstalk to promote intestinal repair, a capacity that may be harnessed in clinical settings of impaired wound healing.
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Anticuerpos Antihelmínticos/inmunología , Intestinos/inmunología , Macrófagos/inmunología , Miofibroblastos/inmunología , Nematospiroides dubius/inmunología , Receptores de Interleucina-8B/inmunología , Infecciones por Strongylida/inmunología , Animales , Anticuerpos Antihelmínticos/genética , Humanos , Intestinos/parasitología , Intestinos/patología , Macrófagos/patología , Ratones , Ratones Noqueados , Miofibroblastos/patología , Receptores de Interleucina-8B/genética , Infecciones por Strongylida/genética , Infecciones por Strongylida/patologíaRESUMEN
Infections with intestinal helminths severely impact on human and veterinary health, particularly through the damage that these large parasites inflict when migrating through host tissues. Host immunity often targets the motility of tissue-migrating helminth larvae, which ideally should be mimicked by anti-helminth vaccines. However, the mechanisms of larval trapping are still poorly defined. We have recently reported an important role for Abs in the rapid trapping of tissue-migrating larvae of the murine parasite Heligmosomoides polygyrus bakeri. Trapping was mediated by macrophages (MΦ) and involved complement, activating FcRs, and Arginase-1 (Arg1) activity. However, the receptors and Ab isotypes responsible for MΦ adherence and Arg1 induction remained unclear. Using an in vitro coculture assay of H. polygyrus bakeri larvae and bone marrow-derived MΦ, we now identify CD11b as the major complement receptor mediating MΦ adherence to the larval surface. However, larval immobilization was largely independent of CD11b and instead required the activating IgG receptor FcγRI (CD64) both in vitro and during challenge H. polygyrus bakeri infection in vivo. FcγRI signaling also contributed to the upregulation of MΦ Arg1 expression in vitro and in vivo. Finally, IgG2a/c was the major IgG subtype from early immune serum bound by FcγRI on the MΦ surface, and purified IgG2c could trigger larval immobilization and Arg1 expression in MΦ in vitro. Our findings reveal a novel role for IgG2a/c-FcγRI-driven MΦ activation in the efficient trapping of tissue-migrating helminth larvae and thus provide important mechanistic insights vital for anti-helminth vaccine development.
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Anticuerpos Antihelmínticos/inmunología , Antígeno CD11b/metabolismo , Helmintiasis Animal/inmunología , Helmintiasis Animal/metabolismo , Helmintos/inmunología , Receptores de IgG/metabolismo , Animales , Arginasa/genética , Arginasa/metabolismo , Proteínas del Sistema Complemento/inmunología , Proteínas del Sistema Complemento/metabolismo , Expresión Génica , Helmintiasis Animal/genética , Sueros Inmunes/inmunología , Inmunoglobulina G/inmunología , Inmunoglobulina G/metabolismo , Interleucina-33 , Interleucinas/metabolismo , Larva , Activación de Macrófagos/inmunología , Macrófagos/inmunología , Ratones Noqueados , Modelos Biológicos , Unión Proteica , Receptores de Interleucina-4/genética , Receptores de Interleucina-4/metabolismo , Transducción de SeñalRESUMEN
IgM anti-mouse platelet autoantibodies cause thrombocytopenia by mediating uptake of opsonized thrombocytes, whereas IgM anti-erythrocyte autoantibodies induce anemia through a phagocytosis-independent cell destruction. In this article, we show that infection with lactate dehydrogenase-elevating virus, a benign mouse arterivirus, exacerbates the pathogenicity of IgM anti-platelet, but not anti-erythrocyte autoantibodies. To define the role of Fcα/µ receptor (Fcα/µR) in IgM-mediated thrombocytopenia and anemia, we generated mice deficient for this receptor. These animals were resistant to IgM autoantibody-mediated thrombocytopenia, but not anemia. However, the lactate dehydrogenase-elevating virus-induced exacerbation of thrombocytopenia was not associated with enhanced Fcα/µR expression on macrophages. These results indicate that Fcα/µR is required for the pathogenicity of IgM anti-platelet autoantibodies but is not sufficient to explain the full extent of the disease in virally infected animals.
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Autoanticuerpos/inmunología , Plaquetas/inmunología , Eritrocitos/inmunología , Inmunoglobulina M/inmunología , Receptores Fc/fisiología , Animales , Infecciones por Arterivirus/inmunología , Virus Elevador de Lactato Deshidrogenasa , Ratones , Ratones Endogámicos C57BL , Trombocitopenia/etiologíaRESUMEN
The uptake of Ag-Ab immune complexes (IC) after the ligation of activating FcγR on dendritic cells (DC) leads to 100 times more efficient Ag presentation than the uptake of free Ags. FcγRs were reported to facilitate IC uptake and simultaneously induce cellular activation that drives DC maturation and mediates efficient T cell activation. Activating FcγRs elicit intracellular signaling via the ITAM domain of the associated FcRγ-chain. Studies with FcRγ-chain knockout (FcRγ(-/-)) mice reported FcRγ-chain ITAM signaling to be responsible for enhancing both IC uptake and DC maturation. However, FcRγ-chain is also required for surface expression of activating FcγRs, hampering the dissection of ITAM-dependent and independent FcγR functions in FcRγ(-/-) DCs. In this work, we studied the role of FcRγ-chain ITAM signaling using DCs from NOTAM mice that express normal surface levels of activating FcγR, but lack functional ITAM signaling. IC uptake by bone marrow-derived NOTAM DCs was reduced compared with wild-type DCs, but was not completely absent as in FcRγ(-/-) DCs. In NOTAM DCs, despite the uptake of ICs, both MHC class I and MHC class II Ag presentation was completely abrogated similar to FcRγ(-/-) DCs. Secretion of cytokines, upregulation of costimulatory molecules, and Ag degradation were abrogated in NOTAM DCs in response to FcγR ligation. Cross-presentation using splenic NOTAM DCs and prolonged incubation with OVA-IC was also abrogated. Interestingly, in this setup, proliferation of CD4(+) OT-II cells was induced by NOTAM DCs. We conclude that FcRγ-chain ITAM signaling facilitates IC uptake and is essentially required for cross-presentation, but not for MHC class II Ag presentation.
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Linfocitos T CD4-Positivos/inmunología , Células Dendríticas/inmunología , Receptores de IgG/metabolismo , Animales , Complejo Antígeno-Anticuerpo/inmunología , Antígenos CD/metabolismo , Diferenciación Celular/genética , Células Cultivadas , Reactividad Cruzada/genética , Endocitosis/genética , Antígenos de Histocompatibilidad Clase I/metabolismo , Antígenos de Histocompatibilidad Clase II/metabolismo , Activación de Linfocitos/genética , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Ratones Transgénicos , Estructura Terciaria de Proteína/genética , Receptores de IgG/genética , Transducción de SeñalRESUMEN
Synthetic long peptides (SLP) are a promising vaccine modality to induce therapeutic T cell responses in patients with chronic infections and tumors. We studied different vaccine formulations in mice using SLP derived from carcinoembryonic Ag. We discovered that one of the SLP contains a linear Ab epitope in combination with a CD4 epitope. Repeated vaccination with this carcinoembryonic Ag SLP in mice shows improved T cell responses and simultaneously induced high titers of peptide-specific Abs. These Abs resulted in unexpected anaphylaxis after a third or subsequent vaccinations with the SLP when formulated in saline. Administration of low SLP doses in the slow-release vehicle IFA prevented the anaphylaxis after repeated vaccination. This study underscores both the immunogenicity of SLP vaccination, for inducing T cell as well as B cell responses, and the necessity of safe administration routes.
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Anafilaxia/prevención & control , Antígeno Carcinoembrionario/farmacología , Epítopos de Linfocito B/farmacología , Inmunoglobulina G/inmunología , Péptidos/farmacología , Vacunas/farmacología , Anafilaxia/inmunología , Animales , Antígeno Carcinoembrionario/inmunología , Preparaciones de Acción Retardada/farmacología , Epítopos de Linfocito B/inmunología , Femenino , Ratones , Ratones Noqueados , Péptidos/inmunología , Vacunación/métodosRESUMEN
Extensive analysis of a variety of arthritis models in germline KO mice has revealed that all four receptors for the Fc part of IgG (FcγR) play a role in the disease process. However, their precise cell type-specific contribution is still unclear. In this study, we analyzed the specific role of the inhibiting FcγRIIb on B lymphocytes (using CD19Cre mice) and in the myeloid cell compartment (using C/EBPαCre mice) in the development of arthritis induced by immunization with either bovine or chicken collagen type II. Despite their comparable anti-mouse collagen autoantibody titers, full FcγRIIb knockout (KO), but not B cell-specific FcγRIIb KO, mice showed a significantly increased incidence and severity of disease compared with wild-type control mice when immunized with bovine collagen. When immunized with chicken collagen, disease incidence was significantly increased in pan-myeloid and full FcγRIIb KO mice, but not in B cell-specific KO mice, whereas disease severity was only significantly increased in full FcγRIIb KO mice compared with incidence and severity in wild-type control mice. We conclude that, although anti-mouse collagen autoantibodies are a prerequisite for the development of collagen-induced arthritis, their presence is insufficient for disease development. FcγRIIb on myeloid effector cells, as a modulator of the threshold for downstream Ab effector pathways, plays a dominant role in the susceptibility to collagen-induced arthritis, whereas FcγRIIb on B cells, as a regulator of Ab production, has a minor effect on disease susceptibility.
Asunto(s)
Artritis Experimental/inmunología , Autoanticuerpos/inmunología , Linfocitos B/inmunología , Células Mieloides/inmunología , Receptores de IgG/inmunología , Animales , Artritis Experimental/genética , Artritis Experimental/patología , Autoanticuerpos/genética , Linfocitos B/patología , Bovinos , Pollos , Colágeno Tipo II/inmunología , Ratones , Ratones Noqueados , Células Mieloides/patología , Especificidad de Órganos/genética , Especificidad de Órganos/inmunología , Receptores de IgG/genéticaRESUMEN
Immunomodulatory mAbs, led by the anti-CTLA4 mAb ipilimumab, are an exciting new class of drugs capable of promoting anticancer immunity and providing durable control of some tumors. Close analysis of a number of agents has revealed a critical yet variable role for Fcγ receptors in their efficacy. In this article, we reveal that agonistic anti-CD40 mAbs have an absolute requirement for cross-linking by inhibitory FcγRIIB when used systemically to treat established BCL1 syngeneic lymphoma, and therapy is lost when using a mouse IgG2a mAb not cross-linked by FcγRIIB. Furthermore, in FcγRIIB-deficient mice the lymphoma itself can provide FcγRIIB to cross-link anti-CD40 on neighboring cells, and only when this is blocked does therapy fail. The dependence on FcγRIIB for immunostimulatory activity was not absolute, however, because when anti-CD40 mAbs were administered systemically with the TLR3 agonist polyinosinic:polycytidylic acid or were given subcutaneously, activatory FcγR could also provide cross-linking. Using this mechanistic insight, we designed multimeric forms of anti-CD40 mAb with intrinsic FcγR-independent activity that were highly effective in the treatment of lymphoma-bearing mice. In conclusion, FcγR-independent anti-CD40 activation is a viable strategy in vivo. These findings have important translational implications, as humans, unlike mice, do not have IgG that binds strongly to FcγRIIB; therefore FcγR-independent derivatives represent an attractive therapeutic option.
Asunto(s)
Anticuerpos Monoclonales/uso terapéutico , Antígenos CD40/inmunología , Linfoma/terapia , Multimerización de Proteína/inmunología , Receptores de IgG/inmunología , Animales , Anticuerpos Monoclonales/inmunología , Linfocitos B/inmunología , Linfocitos T CD8-positivos/inmunología , Proliferación Celular , Células Cultivadas , Inmunoglobulina G/inmunología , Inmunoglobulina G/uso terapéutico , Inmunoterapia , Linfoma/inmunología , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Noqueados , Poli I-C/uso terapéutico , Receptores de IgG/genética , Resonancia por Plasmón de Superficie , Receptor Toll-Like 3/agonistasRESUMEN
MOTIVATION: Advances in sequencing technologies and computational algorithms have enabled the study of genomic variants to dissect their functional consequence. Despite this unprecedented progress, current tools fail to reliably detect and characterize more complex allelic variants, such as short tandem repeats (STRs). We developed TSSV as an efficient and sensitive tool to specifically profile all allelic variants present in targeted loci. Based on its design, requiring only two short flanking sequences, TSSV can work without the use of a complete reference sequence to reliably profile highly polymorphic, repetitive or uncharacterized regions. RESULTS: We show that TSSV can accurately determine allelic STR structures in mixtures with 10% representation of minor alleles or complex mixtures in which a single STR allele is shared. Furthermore, we show the universal utility of TSSV in two other independent studies: characterizing de novo mutations introduced by transcription activator-like effector nucleases (TALENs) and profiling the noise and systematic errors in an IonTorrent sequencing experiment. TSSV complements the existing tools by aiding the study of highly polymorphic and complex regions and provides a high-resolution map that can be used in a wide range of applications, from personal genomics to forensic analysis and clinical diagnostics. AVAILABILITY AND IMPLEMENTATION: We have implemented TSSV as a Python package that can be installed through the command-line using pip install TSSV command. Its source code and documentation are available at https://pypi.python.org/pypi/tssv and http://www.lgtc.nl/tssv.
Asunto(s)
Alelos , Genómica/métodos , Repeticiones de Microsatélite , Programas Informáticos , Algoritmos , Desoxirribonucleasas/metabolismo , Distrofina/genética , Femenino , Genoma Humano , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Masculino , Mutación , Análisis de Secuencia de ADNRESUMEN
Approximately one-third of the world's population suffers from chronic helminth infections with no effective vaccines currently available. Antibodies and alternatively activated macrophages (AAM) form crucial components of protective immunity against challenge infections with intestinal helminths. However, the mechanisms by which antibodies target these large multi-cellular parasites remain obscure. Alternative activation of macrophages during helminth infection has been linked to signaling through the IL-4 receptor alpha chain (IL-4Rα), but the potential effects of antibodies on macrophage differentiation have not been explored. We demonstrate that helminth-specific antibodies induce the rapid trapping of tissue migrating helminth larvae and prevent tissue necrosis following challenge infection with the natural murine parasite Heligmosomoides polygyrus bakeri (Hp). Mice lacking antibodies (JH (-/-)) or activating Fc receptors (FcRγ(-/-)) harbored highly motile larvae, developed extensive tissue damage and accumulated less Arginase-1 expressing macrophages around the larvae. Moreover, Hp-specific antibodies induced FcRγ- and complement-dependent adherence of macrophages to larvae in vitro, resulting in complete larval immobilization. Antibodies together with helminth larvae reprogrammed macrophages to express wound-healing associated genes, including Arginase-1, and the Arginase-1 product L-ornithine directly impaired larval motility. Antibody-induced expression of Arginase-1 in vitro and in vivo occurred independently of IL-4Rα signaling. In summary, we present a novel IL-4Rα-independent mechanism of alternative macrophage activation that is antibody-dependent and which both mediates anti-helminth immunity and prevents tissue disruption caused by migrating larvae.