RESUMEN
We induced the passive reverse Arthus reaction to IgG immune complexes (IC) at different tissue sites in mice lacking C3 treated or not with a C5aR-specific antagonist, or in mice lacking mast cells (Kit(W)/Kit(W-v) mice), and compared the inflammatory responses with those in the corresponding wild-type mice. We confirmed that IC inflammation of skin can be mediated largely by mast cells expressing C5aR and FcgammaRIII. In addition, we provided evidence for C3-independent C5aR triggering, which may explain why the cutaneous Arthus reaction develops normally in C3(-/-) mice. Furthermore, some, but not all, of the acute changes associated with the Arthus response in the lung were significantly more intense in normal mice than in C3(-/-) or Kit(W)/Kit(W-v) mice, indicating for C3- and mast cell-dependent and -independent components. Finally, we demonstrated that C3 contributed to the elicitation of neutrophils to alveoli, which corresponded to an increased synthesis of TNF-alpha, macrophage-inflammatory protein-2, and cytokine-induced neutrophil chemoattractant. While mast cells similarly influenced alveolar polymorphonuclear leukocyte influx, the levels of these cytokines remained largely unaffected in mast cell deficiency. Together, the phenotypes of C3(-/-) mice and Kit(W)/Kit(W-v) mice suggest that complement and mast cells have distinct tissue site-specific requirements acting by apparently distinct mechanisms in the initiation of IC inflammation.
Asunto(s)
Antígenos CD/fisiología , Complemento C3/fisiología , Complemento C5a/fisiología , Enfermedades del Complejo Inmune/inmunología , Inmunoglobulina G/fisiología , Pulmón/inmunología , Mastocitos/inmunología , Receptores de Complemento/fisiología , Piel/inmunología , Animales , Reacción de Arthus/inmunología , Reacción de Arthus/metabolismo , Reacción de Arthus/patología , Movimiento Celular/genética , Movimiento Celular/inmunología , Complemento C3/deficiencia , Complemento C3/genética , Citocinas/biosíntesis , Enfermedades del Complejo Inmune/metabolismo , Enfermedades del Complejo Inmune/patología , Pulmón/metabolismo , Pulmón/patología , Masculino , Mastocitos/patología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Ratones Mutantes , Neutrófilos/inmunología , Neutrófilos/patología , Especificidad de Órganos/genética , Especificidad de Órganos/inmunología , Receptor de Anafilatoxina C5a , Piel/metabolismo , Piel/patologíaRESUMEN
We recently demonstrated a codominant role of C5aR and FcgammaRIII in the initiation of IgG immune complex-mediated inflammation in mice. In this study, we investigated the relative contribution of FcgammaRIII in the generation of several cytokines during experimental hypersensitivity pneumonitis/alveolitis in vivo. Induction of immune complex-alveolitis in C57BL/6 mice resulted in strong accumulation of neutrophils into the lung and enhanced chemotactic activity within bronchoalveolar lavage fluid accompanied by an increased production of the proinflammatory cytokines TNF-alpha and IL-1beta as well as the ELR-CXC chemokines macrophage inflammatory protein-2 (MIP-2) and cytokine-induced neutrophil chemoattractant (KC). FcgammaRIII-deficient C57BL/6 mice (FcgammaRIII(-/-)) showed a marked reduction of the inflammatory response due to decreased production of TNF-alpha, IL-1beta, and MIP-2. Results obtained in C57BL/6 mice either lacking the TNF-alpha class I receptor (TNF-alphaRI(-/-)) or treated with neutralizing anti-TNF-alpha mAb demonstrated an essential contribution of TNF-alpha for mediating IL-1beta release, neutrophil influx, and hemorrhage. Surprisingly, MIP-2 and KC chemokine levels remained largely unaffected in TNF-alphaRI(-/-) mice or after functional inhibition of TNF-alpha. These data suggest that in immune complex alveolitis, the activation of FcgammaRIII may induce divergent downstream effector pathways with TNF-alpha acting independently of CXC chemokines to trigger the inflammatory response in C57BL/6 mice.
Asunto(s)
Alveolitis Alérgica Extrínseca/inmunología , Quimiocinas CXC/biosíntesis , Enfermedades del Complejo Inmune/inmunología , Receptores de IgG/fisiología , Factor de Necrosis Tumoral alfa/biosíntesis , Alveolitis Alérgica Extrínseca/patología , Animales , Complejo Antígeno-Anticuerpo/administración & dosificación , Líquido del Lavado Bronquioalveolar/inmunología , Movimiento Celular/inmunología , Quimiocina CXCL2 , Quimiocinas/biosíntesis , Quimiocinas/fisiología , Quimiocinas CXC/fisiología , Quimiotaxis de Leucocito/inmunología , Citocinas/metabolismo , Enfermedades del Complejo Inmune/patología , Inmunoglobulina G/administración & dosificación , Inyecciones Intravenosas , Interleucina-1/biosíntesis , Interleucina-1/metabolismo , Intubación Intratraqueal , Cinética , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Neutrófilos/patología , Ovalbúmina/administración & dosificación , Ovalbúmina/inmunología , Receptores de IgG/deficiencia , Receptores de IgG/genética , Factor de Necrosis Tumoral alfa/metabolismo , Factor de Necrosis Tumoral alfa/fisiologíaRESUMEN
The pathogenic effects of antiplatelet antibodies were investigated in mice. Monoclonal antibodies (mAbs) of different immunoglobulin G subclass directed against mouse GPIIbIIIa, GPIIIa, GPIbalpha, GPIb-IX, GPV, and CD31 were generated and characterized biochemically. MAbs against GPIb-IX, GPV, CD31, and linear epitopes on GPIIIa had mild and transient effects on platelet counts and induced no spontaneous bleeding. Anti-GPIbalpha mAbs induced profound irreversible thrombocytopenia (< 3% of normal) by Fc-independent mechanisms but only had minor effects on hematocrits. In contrast, injection of intact mAbs, but not F(ab)(2) fragments, against conformational epitopes on GPIIbIIIa, induced irreversible thrombocytopenia, acute systemic reactions, hypothermia, decreased hematocrits, and a paradoxical loss of surface GPIIbIIIa on platelets in vivo, the latter suggesting the formation of platelet-derived microparticles. Blockage of platelet-activating factor receptors inhibited the acute reactions, but not thrombocytopenia, loss of GPIIbIIIa, and decreases in hematocrits. Repeated injections of low doses of anti-GPIIbIIIa antibodies resulted in profound thrombocytopenia and bleeding, whereas no acute systemic reactions were observed. These data strongly suggest that the identity of the target antigen recognized by antiplatelet antibodies determines the mechanisms of platelet destruction and the severity of bleeding in mice, the latter depending on previously unrecognized anti-GPIIbIIIa-specific inflammatory mechanisms.
Asunto(s)
Autoantígenos/inmunología , Plaquetas/inmunología , Púrpura Trombocitopénica Idiopática/inmunología , Animales , Anticuerpos Monoclonales/administración & dosificación , Anticuerpos Monoclonales/farmacología , Plaquetas/química , Epítopos , Hemorragia/inmunología , Fragmentos Fc de Inmunoglobulinas/inmunología , Fragmentos Fc de Inmunoglobulinas/farmacología , Inmunoglobulina G/farmacología , Ratones , Fenotipo , Factor de Activación Plaquetaria/farmacología , Molécula-1 de Adhesión Celular Endotelial de Plaqueta/inmunología , Complejo GPIIb-IIIa de Glicoproteína Plaquetaria/inmunología , Complejo GPIb-IX de Glicoproteína Plaquetaria/inmunología , Organismos Libres de Patógenos EspecíficosRESUMEN
Platelet glycoprotein (GP) VI has been proposed as the major collagen receptor for activation of human platelets. Human GPVI belongs to the immunoglobulin superfamily and is noncovalently associated with the FcRgamma chain that is involved in signaling through the receptor. In mice, similar mechanisms seem to exist as platelets from FcRgamma chain-deficient mice do not aggregate in response to collagen. However, the activating collagen receptor on mouse platelets has not been definitively identified. In the current study we examined the function and in vivo expression of GPVI in control and FcRgamma chain-deficient mice with the first monoclonal antibody against GPVI (JAQ1). On wild type platelets, JAQ1 inhibited platelet aggregation induced by collagen but not PMA or thrombin. Cross-linking of bound JAQ1, on the other hand, induced aggregation of wild type but not FcRgamma chain-deficient platelets. JAQ1 stained platelets and megakaryocytes from wild type but not FcRgamma chain-deficient mice. Furthermore, JAQ1 recognized GPVI (approximately 60 kDa) in immunoprecipitation and Western blot experiments with wild type but not FcRgamma chain-deficient platelets. These results strongly suggest that GPVI is the collagen receptor responsible for platelet activation in mice and demonstrate that the association with the FcRgamma chain is critical for its expression and function.
Asunto(s)
Colágeno/farmacología , Integrinas/metabolismo , Agregación Plaquetaria/fisiología , Glicoproteínas de Membrana Plaquetaria/metabolismo , Receptores de IgG/metabolismo , Animales , Anticuerpos Monoclonales/farmacología , Megacariocitos/metabolismo , Ratones , Ratones Endogámicos , Ratones Mutantes , Glicoproteínas de Membrana Plaquetaria/inmunología , Unión Proteica , Receptores de Colágeno , Receptores de IgG/genéticaRESUMEN
Using three different Fcgamma receptor (FcgammaR)-deficient mouse strains, we examined the induction of autoimmune hemolytic anemia by each of the four immunoglobulin (Ig)G isotype-switch variants of a 4C8 IgM antierythrocyte autoantibody and its relation to the contributions of the two FcgammaR, FcgammaRI, and FcgammaRIII, operative in the phagocytosis of opsonized particles. We found that the four IgG isotypes of this antibody displayed striking differences in pathogenicity, which were related to their respective capacity to interact in vivo with the two phagocytic FcgammaRs, defined as follows: IgG2a > IgG2b > IgG3/IgG1 for FcgammaRI, and IgG2a > IgG1 > IgG2b > IgG3 for FcgammaRIII. Accordingly, the IgG2a autoantibody exhibited the highest pathogenicity, approximately 20-100-fold more potent than its IgG1 and IgG2b variants, respectively, while the IgG3 variant, which displays little interaction with these FcgammaRs, was not pathogenic at all. An unexpected critical role of the low-affinity FcgammaRIII was revealed by the use of two different IgG2a anti-red blood cell autoantibodies, which displayed a striking preferential utilization of FcgammaRIII, compared with the high-affinity FcgammaRI. This demonstration of the respective roles in vivo of four different IgG isotypes, and of two phagocytic FcgammaRs, in autoimmune hemolytic anemia highlights the major importance of the regulation of IgG isotype responses in autoantibody-mediated pathology and humoral immunity.
Asunto(s)
Inmunoglobulina G/genética , Inmunoglobulina G/metabolismo , Receptores de IgG/metabolismo , Anemia Hemolítica Autoinmune/etiología , Anemia Hemolítica Autoinmune/genética , Anemia Hemolítica Autoinmune/inmunología , Animales , Autoanticuerpos/metabolismo , Secuencia de Bases , Cartilla de ADN/genética , Eritrocitos/inmunología , Variación Genética , Isotipos de Inmunoglobulinas/genética , Isotipos de Inmunoglobulinas/metabolismo , Región de Cambio de la Inmunoglobulina/genética , Técnicas In Vitro , Hierro/metabolismo , Hígado/metabolismo , Hígado/patología , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones NoqueadosRESUMEN
Murine low-affinity receptors for IgG, FcgammaRII and FcgammaRIII, differ by their distinct capacities in mediating down-regulation or activation of cellular effector functions, respectively. In this study, antibodies detecting the mouse Ly-17.1 / 2 alloantigen system are demonstrated to be specific for FcgammaRII with no cross-reactivities to other FcgammaR, including FcgammaRIII. Using these FcgammaRII-specific monoclonal antibodies (mAb), the significance of FcgammaRII inhibition of FcgammaRIII was examined in two models of autoantibody [autoimmune hemolytic anemia (AIHA)]- and IgG immune complex-induced (Arthus reaction) inflammation in C57BL / 6 mice in comparison with FcgammaRII(- / -) and FcgammaRIII(- / -) mice. Our results demonstrate that both FcgammaRIII and FcgammaRII contributed to the binding of erythrocytes opsonized with the pathogenic IgG1 autoreactive anti-murine red blood cell antibody 105-2H. However, the functional blocking with anti-FcgammaRII mAb in C57BL / 6 mice and the lack of FcgammaRII expression in FcgammaRII(- / -) mice, which both lowered the threshold level of FcgammaRIII-triggered phagocytosis in vitro, did not results in enhanced disease development of 105-2H mAb-induced AIHA in vivo. This was in sharp contrast to cutaneous Arthus reaction, where FcgammaRIII-mediated activation was inhibited by FcgammaRII. Together these results show that murine AIHA is markedly different from other FcgammaR-dependent inflammatory diseases where FcgammaRIII is normally counterregulated by FcgammaRII.
Asunto(s)
Anemia Hemolítica/inmunología , Reacción de Arthus/inmunología , Receptores de IgG/inmunología , Anemia Hemolítica/genética , Animales , Complejo Antígeno-Anticuerpo/inmunología , Reacción de Arthus/genética , Autoanticuerpos/inmunología , Regulación hacia Abajo , Regulación de la Expresión Génica/inmunología , Inmunoglobulina G/inmunología , Ratones , Ratones Endogámicos C57BL , Receptores de IgG/genéticaRESUMEN
Recent attempts to specify the relative contribution of FcR and complement in various experimental systems of immune complex disease have led to opposing conclusions. As concluded in IgG FcRgamma-/- mice, manifestation of disease is almost exclusively determined by FcgammaR on effector cells, arguing for a minor role of complement. In contrast, data obtained with C5aR-/- mice suggested that, dependent on the tissue site, complement is more important than FcgammaR. In this paper, we demonstrate that, in response to IgG immune complex formation, FcgammaRI/III- and C5aR-mediated pathways are both necessary and only together are they sufficient to trigger the full expression of inflammation in skin and lung. Moreover, both effector systems are not entirely independent, suggesting an interaction between FcgammaR and C5aR. Therefore, FcgammaR-mediated responses can be integrated through C5aR activation, which may explain why these two receptor pathways have previously been considered to dominate each other.
Asunto(s)
Antígenos CD/fisiología , Reacción de Arthus/inmunología , Reacción de Arthus/metabolismo , Complemento C5a/metabolismo , Receptores de Complemento/fisiología , Receptores de IgG/fisiología , Animales , Reacción de Arthus/patología , Inflamación/inmunología , Inflamación/metabolismo , Inflamación/patología , Pulmón/inmunología , Pulmón/metabolismo , Pulmón/patología , Ratones , Ratones Endogámicos C57BL , Ratones Mutantes , Receptor de Anafilatoxina C5a , Receptores de IgG/deficiencia , Receptores de IgG/genética , Piel/inmunología , Piel/metabolismo , Piel/patologíaRESUMEN
The generation of autoantibodies and deposition of immune complexes (ICs) in tissue play a primary role in autoimmune diseases. However, the IC-triggered response consists of complex mechanisms that make it difficult to identify the pathogenesis and develop specific therapy. We clarified here a sequential mechanism for the induction of hypersensitivity angiitis by analyzing the responsible Fc receptor (FcR), effector cells, and mediators in an animal model using FcR-deficient mice. In this model, rheumatoid factor-mediated skin vasculitis was induced in wild-type mice, whereas FcRgamma-deficient mice did not develop the vasculitis. Adoptive transfer of various FcR(+) cells into FcRgamma-deficient mice showed that mast cells but not macrophages derived from wild-type mice triggered skin vasculitis. Mast cells derived from either FcgammaRIII-deficient or tumor necrosis factor (TNF)-deficient mice did not possess the inducibility of skin vasculitis. These results indicate that triggering of vascular inflammation was induced by mast cells through IC binding on FcgammaRIII. TNF produced by such activated mast cells was mainly responsible for the pathogenesis of autoantibody-mediated vasculitis. These findings illustrate the clinical significance of mast cells, Fcgamma receptors, and TNF in IC-induced vasculitis syndrome.
Asunto(s)
Autoanticuerpos/inmunología , Mastocitos/inmunología , Receptores Fc/inmunología , Factor de Necrosis Tumoral alfa/inmunología , Vasculitis/inmunología , Animales , Mastocitos/patología , Ratones , Transducción de Señal/inmunología , Factor de Necrosis Tumoral alfa/biosíntesis , Vasculitis/etiología , Vasculitis/patologíaRESUMEN
NK cells reject non-self hematopoietic bone marrow (BM) grafts via Ly49 receptor-mediated MHC class I-specific recognition and calibration of receptor expression levels. In this paper we investigated how Ly49+ subset frequencies were regulated dependent on MHC class I expression. The development of donor and host Ly49A+ (recognizes H-2Dd and H-2Dk ligands) and Ly49C/I+ (Ly49CBALB/c recognizes H-2Kb, H-2Kd, and H-2Dd, and Ly49CB6 recognizes only H-2Kb) NK cell frequencies were monitored for 120 days in murine-mixed allogeneic BM chimeras. C57BL/6 (H-2b) BM was transplanted into BALB/c (H-2d) mice and vice versa. Peripheral NK cell populations were examined every 5 days. Chimerism was found to be stable with 80-90% donor NK cells. In contrast to syngeneic controls reexpressing pretransplant patterns, donor and host NK cells revealed new and mainly reduced subset frequencies 55 days after allogeneic transplantation. Recipient NK cells acquired these later than donor NK cells. In H-2d --> H-2b chimeras Ly49A+, Ly49C/I+, and Ly49A+/Ly49C/I+ proportions were mainly diminished upon interaction with cognate ligands. Also in H-2b --> H-2d chimeras, Ly49A+ and Ly49A+/Ly49C/I+ subsets were reduced, but there was a transient normalization of Ly49C/I+ proportions in the noncognate host. After 120 days all subsets were reduced. Therefore, down-regulation of developing Ly49A+ and Ly49C/I+ chimeric NK cell frequencies by cognate ligands within 7-8 wk after BM transplantation may be important for successful engraftment.
Asunto(s)
Antígenos Ly , Trasplante de Médula Ósea/inmunología , Antígenos H-2/inmunología , Células Asesinas Naturales/inmunología , Subgrupos Linfocitarios/inmunología , Glicoproteínas de Membrana , Quimera por Trasplante/inmunología , Animales , Tolerancia Inmunológica , Lectinas Tipo C , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Subfamilia A de Receptores Similares a Lectina de Células NK , Receptores Similares a Lectina de Células NK , Trasplante HomólogoRESUMEN
Shock is frequently accompanied by thrombocytopenia. To investigate the pathogenic role of platelets in shock, we examined the in vivo effects of monoclonal antibodies (MoAbs) against mouse platelet membrane proteins. Injection of the platelet-specific MoAb MWReg30 to the fibrinogen receptor (gpIIb/IIIa) rendered mice severely hypothermic within minutes. Isotype-matched control antibodies, even if they also recognized platelet surface antigens, did not induce comparable signs. MWReg30 induced early signs of acute lung injury with increased cellularity in the lung interstitium and rapid engorgement of alveolar septal vessels. Despite this in vivo activity, MWReg30 inhibited rather than stimulated platelet aggregation in vitro. MWReg30-binding to platelets led to phosphorylation of gpIIIa, but did not induce morphological signs of platelet activation. The MWReg30-induced reaction was abolished after treatment with MoAbs 2.4G2 to FcgammaRII/III and was absent in FcgammaRIII-deficient mice, clearly demonstrating the requirement for FcgammaRIII on involved leukocytes. Simultaneous administration of tumor necrosis factor exacerbated, whereas a tolerizing regimen of tumor necrosis factor or bacterial lipopolysaccharide completely prevented the reaction. These data suggest that platelet surface-deposited MWReg30-immune complexes lead to an acute Fc-mediated reaction with pulmonary congestion and life-threatening potential that could serve as an in vivo model of acute lung injury.
Asunto(s)
Anticuerpos Monoclonales/toxicidad , Complejo GPIIb-IIIa de Glicoproteína Plaquetaria/inmunología , Edema Pulmonar/etiología , Choque/etiología , Trombocitopenia/etiología , Animales , Anticuerpos Monoclonales/inmunología , Complejo Antígeno-Anticuerpo/inmunología , Eritema/etiología , Eritema/inmunología , Eritema/fisiopatología , Hipotermia/etiología , Hipotermia/inmunología , Hipotermia/fisiopatología , Lipopolisacáridos/administración & dosificación , Lipopolisacáridos/uso terapéutico , Pulmón/patología , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C3H , Ratones Endogámicos , Ratones Noqueados , Fosforilación/efectos de los fármacos , Agregación Plaquetaria/efectos de los fármacos , Complejo GPIIb-IIIa de Glicoproteína Plaquetaria/fisiología , Procesamiento Proteico-Postraduccional/efectos de los fármacos , Edema Pulmonar/inmunología , Edema Pulmonar/fisiopatología , Receptores de IgG/deficiencia , Receptores de IgG/genética , Receptores de IgG/inmunología , Choque/complicaciones , Choque/fisiopatología , Choque/prevención & control , Organismos Libres de Patógenos Específicos , Trombocitopenia/inmunología , Factor de Necrosis Tumoral alfa/administración & dosificación , Factor de Necrosis Tumoral alfa/uso terapéutico , Factor de Necrosis Tumoral alfa/toxicidadRESUMEN
A C5a-receptor antagonist was selected from human C5a phage display libraries in which the C terminus of des-Arg74-hC5a was mutated. The selected molecule is a competitive C5a receptor antagonist in vitro and in vivo. Signal transduction is interrupted at the level of G-protein activation. In addition, the antagonist does not cause any C5a receptor phosphorylation. Proinflammatory properties such as chemotaxis or lysosomal enzyme release of differentiated U937 cells, as well as C5a-induced changes in intracellular Ca2+ concentration of murine peritoneal macrophages, are inhibited. The in vivo efficacy was evaluated in three different animal models of immune complex diseases in mice, i.e., the reverse passive Arthus reaction in the peritoneum, skin, and lung. The i.v. application of the C5a receptor antagonist abrogated polymorphonuclear neutrophil accumulation in peritoneum and markedly attenuated polymorphonuclear neutrophil migration into the skin and the lung. In a model of intestinal ischemia/reperfusion injury, i.v. administration of the C5a receptor antagonist decreased local and remote tissue injury: bowel wall edema and hemorrhage as well as pulmonary microvascular dysfunction. These data give evidence that C5a is an important mediator triggering the inflammatory sequelae seen in immune complex diseases and ischemia/reperfusion injury. The selected C5a receptor antagonist may prove useful to attenuate the inflammatory response in these disorders.
Asunto(s)
Antígenos CD/química , Bacteriófago M13/inmunología , Complemento C5a/metabolismo , Enfermedades del Complejo Inmune/patología , Biblioteca de Péptidos , Receptores de Complemento/antagonistas & inhibidores , Receptores de Complemento/química , Daño por Reperfusión/patología , Sustitución de Aminoácidos/genética , Animales , Antígenos CD/genética , Reacción de Arthus/inmunología , Reacción de Arthus/patología , Bacteriófago M13/genética , Unión Competitiva/genética , Unión Competitiva/inmunología , Degranulación de la Célula/genética , Degranulación de la Célula/inmunología , Diferenciación Celular/genética , Diferenciación Celular/inmunología , Inhibición de Migración Celular , Movimiento Celular/genética , Movimiento Celular/inmunología , Quimiotaxis de Leucocito/genética , Quimiotaxis de Leucocito/inmunología , Femenino , Humanos , Enfermedades del Complejo Inmune/genética , Enfermedades del Complejo Inmune/inmunología , Pulmón/inmunología , Pulmón/patología , Macrófagos Peritoneales/inmunología , Macrófagos Peritoneales/metabolismo , Ratones , Ratones Endogámicos BALB C , Mutagénesis Sitio-Dirigida , Neutrófilos/inmunología , Peritonitis/genética , Peritonitis/inmunología , Peritonitis/patología , Receptor de Anafilatoxina C5a , Receptores de Complemento/genética , Daño por Reperfusión/genética , Daño por Reperfusión/inmunología , Piel/inmunología , Piel/patología , Células U937RESUMEN
The contributions of Fc receptors (FcRs) for IgG (FcgammaRs) and complement to immune complex (IC)-mediated peritonitis were evaluated in BALB/c-, C57BL/6-, FcRgamma chain-, and FcR type III for IgG (FcgammaRIII)-deficient mice, backcrossed to the C57BL/6 background. In BALB/c mice, but not in C57BL/6 mice, neutrophil migration was markedly attenuated after complement depletion. In mice lacking FcRgamma chain, neutrophil migration was abolished, whereas it was unaffected in FcgammaRIII-deficient mice. Huge amounts of TNF-alpha (TNF) were found in the peritoneal exudate of BALB/c and C57BL/6 mice but were absent in mice lacking FcRgamma chain or FcgammaRIII. Surprisingly, a functional inhibition of TNF in BALB/c and C57BL/6 mice had no effect on neutrophil infiltration. These data provide evidence that in IC peritonitis, the activation of FcR type I for IgG on peritoneal macrophages and the activation of the complement cascade, but not the interaction of ICs with FcgammaRIII and the subsequent release of TNF, initiate the inflammatory response in BALB/c and C57BL/6 mice.
Asunto(s)
Reacción de Arthus/inmunología , Proteínas del Sistema Complemento/inmunología , Macrófagos Peritoneales/inmunología , Peritonitis/inmunología , Receptores de IgG/inmunología , Animales , Reacción de Arthus/genética , Líquido Ascítico/química , Líquido Ascítico/citología , Quimiotaxis de Leucocito , Cruzamientos Genéticos , Citotoxicidad Inmunológica , Antígenos HLA/inmunología , Antígenos de Histocompatibilidad Clase I/inmunología , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Mutantes , Neutrófilos/inmunología , Peritonitis/genética , Especificidad de la Especie , Factor de Necrosis Tumoral alfa/análisis , Antígenos HLA-ERESUMEN
The contribution of either the complement system or the activation of Fc receptors for IgG (FcyRs) to the inflammatory response in immune complex (IC) disease is puzzling. A series of studies has been performed in mice with engineered deficiencies of either FcgammaRs, the complement components C3, C4 or the C5a receptor. In addition, different C5-deficient mice strains have been evaluated. Mice with gene targeted disruption of the gamma-subunit, which mediates surface expression and signal transduction of the high affinity Fc receptor type I for IgG (FcgammaRI), the low affinity receptor Fc receptor type III for IgG (FcgammaRIII) and the high affinity receptor type I for IgE (IgepsilonRI), showed an impaired inflammatory response in the reverse passive Arthus reaction in skin, peritoneum and lung. These data suggest, that the activation of FgammaRs is the initial event triggering the inflammatory cascade in IC disease. On the other hand, C5aR deficient mice are either protected from tissue injury induced by ICs, as in the lung, or the degree of the inflammatory response is markedly attenuated, as in peritoneum and skin. A detailed analysis of data obtained with the different knock-out strains revealed that both the activation of the complement system as well as the activation of different effector cells via FcgammaRs contribute to the inflammatory sequelae leading to tissue destruction in IC disease. The relative contributions of FcgammaRI or FcgammaRIII and the main effector cells through which these receptors mediate their effector functions are tissue dependent. The activation of the C5a receptor pathway appears to be the prominent contribution of the complement system.
Asunto(s)
Reacción de Arthus/etiología , Reacción de Arthus/inmunología , Proteínas del Sistema Complemento/metabolismo , Receptores de IgG/metabolismo , Animales , Antígenos CD/genética , Antígenos CD/metabolismo , Proteínas del Sistema Complemento/deficiencia , Proteínas del Sistema Complemento/genética , Humanos , Enfermedades del Complejo Inmune/etiología , Enfermedades del Complejo Inmune/inmunología , Inflamación/etiología , Inflamación/inmunología , Enfermedades Pulmonares/etiología , Enfermedades Pulmonares/inmunología , Ratones , Ratones Noqueados , Peritonitis/etiología , Peritonitis/inmunología , Receptor de Anafilatoxina C5a , Receptores de Complemento/genética , Receptores de Complemento/metabolismo , Receptores de IgG/deficiencia , Receptores de IgG/genéticaRESUMEN
In autoimmune hemolytic anemia (AIHA), there is accumulating evidence for an involvement of FcgammaR expressed by phagocytic effector cells, but demonstration of a causal relationship between individual FcgammaRs and IgG isotypes for disease development is lacking. Although the relevance of IgG isotypes to human AIHA is limited, we could show a clear IgG isotype dependency in murine AIHA using pathogenic IgG1 (105-2H) and IgG2a (34-3C) autoreactive anti-red blood cell antibodies in mice defective for FcgammaRIII, and comparing the clinical outcome to those in wild-type mice. FcgammaRIII-deficient mice were completely resistent to the pathogenic effects of 105-2H monoclonal antibody, as shown by a lack of IgG1-mediated erythrophagocytosis in vitro and in vivo. In addition, the IgG2a response by 34-3C induced a less severe but persistent AIHA in FcgammaRIII knock-out mice, as documented by a decrease in hematocrit. Blocking studies indicated that the residual anemic phenotype induced by 34-3C in the absence of FcgammaRIII reflects an activation of FcgammaRI that is normally coexpressed with FcgammaRIII on macrophages. Together these results show that the pathogenesis of AIHA through IgG1-dependent erythrophagocytosis is exclusively mediated by FcgammaRIII and further suggest that FcgammaRI, in addition to FcgammaRIII, contributes to this autoimmune disease when other IgG isotypes such as IgG2a are involved.
Asunto(s)
Anemia Hemolítica Autoinmune/inmunología , Eritrocitos/inmunología , Inmunoglobulina G/inmunología , Receptores de IgG/deficiencia , Animales , Autoanticuerpos/inmunología , Citotoxicidad Inmunológica , Humanos , Isotipos de Inmunoglobulinas/inmunología , Ratones , Ratones Noqueados , Fagocitosis , Receptores de IgG/genética , Receptores de IgG/inmunologíaRESUMEN
The deposition of immune complexes, followed by activation of complement and/or Fc receptors and generation of chemoattractants, is the most common feature of human glomerulonephritis. Recently we have shown that primary cultured human glomerular mesangial cells (HMC), which are normally negative for IgG Fc receptors, can be stimulated to express the low-affinity FcgammaRIII-A receptor isoform. In this study we further demonstrate that activation of HMC through IFN-gamma resulted in the functional expression of the high-affinity Fc receptor for IgG (FcgammaRI, CD64). IFN-gamma-dependent induction of classical FcgammaRIa1 mRNA as well as a2, b2 splice variants were evident after 24 h in proliferating HMC and after 48 h in resting HMC. Transcription of FcgammaRI mRNA was also induced by IL-10 in proliferating HMC, whereas other cytokines such as IL-3, transforming growth factor-beta1 and granulocyte-macrophage colony-stimulating factor were not effective. Cell surface expression of FcgammaRI could be detected by flow cytometric analysis after IFN-gamma stimulation and was accompanied by the augmentation of MHC class II and the up-regulation of intercellular adhesion molecule-1 expression. Triggering of HMC by cross-linking FcgammaRI with F(ab')2 fragments of the anti-CD64 monoclonal antibody 22 led to enhanced synthesis of mRNA for the chemokines IL-8 and monocyte chemoattractant protein-1, indicating that the FcgammaRI of HMC is functionally active. These in vitro data suggest that engagement of both FcgammaRI and FcgammaRIII-A on activated HMC through IgG immune complexes may result in an increased chemoattraction of leukocytes into the glomerulus, contributing to the development of glomerulonephritis.
Asunto(s)
Mesangio Glomerular/inmunología , Inmunoglobulina G/inmunología , Interferón gamma/farmacología , Receptores de IgG/agonistas , Receptores de IgG/inmunología , Células Cultivadas , Citocinas/inmunología , Citocinas/farmacología , Humanos , Molécula 1 de Adhesión Intercelular/inmunología , Interferón gamma/inmunología , ARN Mensajero/análisis , ARN Mensajero/biosíntesisRESUMEN
Previously, we have demonstrated that phagocytosis of IgG1-coated particles by macrophages in vitro is impaired by deletion of Fc gamma RIII in mice, suggesting that IgG1 may interact preferentially with Fc gamma RIII. In the present study, the biologic relevance of this observation was addressed by triggering various effector functions of the immune system in Fc gamma RIII(-/-) mice, using panels of mAbs of different IgG subclasses. Both binding and phagocytosis of IgG1-coated sheep or human erythrocytes by Fc gamma RIII(-/-) macrophages in vitro were strongly impaired, indicating that the impaired ingestion of complexed IgG1 by Fc gamma RIII(-/-) macrophages is due to a defect in binding. An in vivo consequence of the defective phagocytosis was observed by resistance of Fc gamma RIII-deficient mice to experimental autoimmune hemolytic anemia, as shown by a lack of IgG1-mediated erythrophagocytosis in vivo by liver macrophages. Furthermore, trapping of soluble IgG1-containing immune complexes by follicular dendritic cells in mesenteric lymph nodes from Fc gamma RIII(-/-) mice was abolished. Whole blood from Fc gamma RIII(-/-) mice was unable to induce lysis of tumor cells in the presence of IgG1 antitumor Abs. Finally, IgG1 mAbs proved unable to mount a passive cutaneous anaphylaxis in Fc gamma RIII(-/-) mice. Together, these results demonstrate that IgG1 complexes, either in particulate or in soluble form, trigger in vitro and in vivo immune effector functions in mice predominantly via Fc gamma RIII.
Asunto(s)
Complejo Antígeno-Anticuerpo/fisiología , Inmunoglobulina G/fisiología , Receptores de IgG/fisiología , Animales , Anticuerpos Monoclonales/farmacología , Citotoxicidad Celular Dependiente de Anticuerpos/inmunología , Complejo Antígeno-Anticuerpo/sangre , Complejo Antígeno-Anticuerpo/metabolismo , Antígenos de Grupos Sanguíneos/inmunología , Neoplasias de la Mama , Células Dendríticas/inmunología , Eritrocitos/inmunología , Eritrocitos/metabolismo , Humanos , Sueros Inmunes/fisiología , Inmunoglobulina G/sangre , Inmunoglobulina G/farmacología , Hígado/inmunología , Macrófagos Peritoneales/inmunología , Macrófagos Peritoneales/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Especificidad de Órganos/inmunología , Anafilaxis Cutánea Pasiva , Fagocitosis/inmunología , Formación de Roseta , Células Tumorales CultivadasRESUMEN
IgG immune complexes are of central importance in the humoral immune system and strongly implicated in the pathogenesis of hematologic and rheumatic autoimmune disorders. Cross-linking of receptors for the Fc domain of IgG antibodies (FcgammaRs) triggers a wide variety of effector functions including phagocytosis, antibody-dependent cellular cytotoxicity, and release of inflammatory mediators, as well as immune complex clearance and regulation of antibody production. In this way, FcgammaR provide an essential feedback between the humoral and cellular immune response. In the past, significant advances have been made in the molecular dissection of FcgammaR function using cellular transfection systems. Current approaches designed to target and change individual FcgammaR genes in mice have given further insight into their specific contributions to systemic processes, also indicating them to be important immunoregulatory receptors involved in various disease states of allergy, autoimmunity, and inflammation. Future work on targeting FcgammaR binding sites in combination with humanized FcgammaR mouse models will lead to novel therapeutic strategies in the treatment of IgG-mediated human disease in which FcgammaR activation plays an integral part.
Asunto(s)
Enfermedades Autoinmunes/inmunología , Hipersensibilidad/inmunología , Receptores de IgG/fisiología , Anemia Hemolítica/inmunología , Animales , Antígenos CD/química , Antígenos CD/genética , Antígenos CD/inmunología , Antígenos CD/fisiología , Glomerulonefritis/inmunología , Humanos , Inmunidad Celular , Inmunoglobulina G/inmunología , Ratones , Ratones Noqueados , Modelos Moleculares , Receptores de IgG/química , Receptores de IgG/genética , Receptores de IgG/inmunología , Vasculitis/inmunologíaRESUMEN
The molecular events governing the differentiation pathway of natural killer (NK) cells are not well understood. The phenotype of mature NK cells is specified by the expression of the low affinity Fc receptor for IgG (human FcgammaRIII, CD16) encoded by the FcgammaRIII-A gene. Here we report that the Pprox promoter (-198/-10) of FcgammaRIII-A stimulated by its own intron enhancer (+10/+712) was only one of the cis-elements that target the expression of a reporter gene in the immature NK cell line, YT. The transcription start sites of the FcgammaRIII-A a2/3 and a5/6 splice alternatives in NK cells were mapped to the medial -1817/-850 FcgammaRIII-A control region. Two promoters, Pmed1 (-942/-850) and Pmed2 (-1376/-1123) resided in this region and controlled for the initiation of these transcript classes encoding the known FcgammaRIII-A receptor protein. Deletion mapping studies demonstrated that the 93 base pairs -942/-850 Pmed1 sequence was sufficient to confer cell type-specific expression in YT cells. The 5' end of Pmed1 (-942 to -921) was required for full promoter function indicating the presence of an important sequence motif recognized by a YT-specific factor. Our data suggest that this motif might be a useful tool for subsequent identification of putative transcription factors uniquely active in YT and NK cells.
Asunto(s)
Células Asesinas Naturales/fisiología , Regiones Promotoras Genéticas , Receptores de IgG/genética , Empalme Alternativo , Secuencia de Bases , Genes , Humanos , Datos de Secuencia Molecular , ARN Mensajero/genética , Eliminación de Secuencia , Transcripción GenéticaRESUMEN
The family of receptors for IgG (Fc gamma R) plays an essential role in antibody-mediated effector functions of the immune system. However, the specific contribution of each of the Fc gamma R classes to in vivo immune reactions is still unclear. Here, we demonstrate that mice deficient for the ligand-binding alpha chain of Fc gamma RIII lack NK cell-mediated antibody-dependent cytotoxicity and phagocytosis of IgG1-coated particles by macrophages. Strikingly, these mice lack IgG-mediated mast cell degranulation, are resistant to IgG-dependent passive cutaneous anaphylaxis, and exhibit an impaired Arthus reaction. These results indicate a prominent role for Fc gamma RIII in inflammatory and anaphylactic responses, making this receptor a potential target in immunotherapy.