RESUMEN
OBJECTIVE: To define the relationship of synovial B cells to clinical phenotypes at different stages of disease evolution and drug exposure in rheumatoid arthritis (RA). METHODS: Synovial biopsy specimens and demographic and clinical data were collected from 2 RA cohorts (n = 329), one of patients with untreated early RA (n = 165) and one of patients with established RA with an inadequate response to tumor necrosis factor inhibitors (TNFi-IR; n = 164). Synovial tissue was subjected to hematoxylin and eosin and immunohistochemical staining and semiquantitative assessment for the degree of synovitis (on a scale of 0-9) and of CD20+ B cell infiltrate (on a scale of 0-4). B cell scores were validated by digital image analysis and B cell lineage-specific transcript analysis (RNA-Seq) in the early RA (n = 91) and TNFi-IR (n = 127) cohorts. Semiquantitative CD20 scores were used to classify patients as B cell rich (≥2) or B cell poor (<2). RESULTS: Semiquantitative B cell scores correlated with digital image analysis quantitative measurements and B cell lineage-specific transcripts. B cell-rich synovitis was present in 35% of patients in the early RA cohort and 47.7% of patients in the TNFi-IR cohort (P = 0.025). B cell-rich patients showed higher levels of disease activity and seropositivity for rheumatoid factor and anti-citrullinated protein antibody in early RA but not in established RA, while significantly higher histologic synovitis scores in B cell-rich patients were demonstrated in both cohorts. CONCLUSION: We describe a robust semiquantitative histologic B cell score that closely replicates the quantification of B cells by digital or molecular analyses. Our findings indicate an ongoing B cell-rich synovitis, which does not seem to be captured by standard clinimetric assessment, in a larger proportion of patients with established RA than early RA.
Asunto(s)
Artritis Reumatoide/complicaciones , Artritis Reumatoide/tratamiento farmacológico , Linfocitos B , Sinovitis/complicaciones , Sinovitis/genética , Adulto , Anciano , Estudios de Cohortes , Progresión de la Enfermedad , Femenino , Humanos , Masculino , Persona de Mediana Edad , Fenotipo , Sinovitis/inmunologíaRESUMEN
Epistatic interactions between the non-autoimmune strains 129 and C57BL/6 (B6), used for generating gene-targeted animals, can induce a lupus-like disease. Genome-wide scan analyses of testcross progeny between these two strains have identified several lupus susceptibility loci, with the strongest linkage to the production of autoantibodies (auto-Abs) displayed by an interval on chromosome 1 of 129 origin (Sle16). However, the contribution of B6 loci to the lupus phenotype remained unknown. We used a congenic approach to deduce the contribution to the autoimmune traits of the B6 genomic interval on chromosome 3 (Sle18), previously shown to be linked to antinuclear Ab production. This interval, when transferred on a 129 background (a strain termed 129.B6-Sle18), promoted auto-Ab production targeting a broad spectrum of autoantigens, expansion of activated CD4(+)T and B cells and mild glomerulonephritis. Surprisingly, these immunological and serological defects were accompanied by a significant increase in the percentage of regulatory T cells (Tregs; CD4(+) Foxp3(+)). However, these cells, that expressed lower levels of Foxp3, had no impaired regulatory function when tested in vitro. These findings illustrate further the efficacy of congenic dissection for functional characterisation of individual lupus susceptibility loci and highlight the contribution of loci derived from non-autoimmune strains to the disease pathogenesis.
Asunto(s)
Autoanticuerpos/biosíntesis , Cromosomas de los Mamíferos , Predisposición Genética a la Enfermedad , Lupus Eritematoso Sistémico/genética , Lupus Eritematoso Sistémico/inmunología , Animales , Autoanticuerpos/genética , Autoanticuerpos/inmunología , Epistasis Genética , Lupus Eritematoso Sistémico/patología , Ratones , Ratones Congénicos , Ratones Endogámicos C57BLRESUMEN
BACKGROUND: It has been suggested that defective handling of apoptotic cells by macrophages plays a key role in the development of systemic lupus erythematosus (SLE). The relative contribution of intrinsic defects and serum factors remains controversial. OBJECTIVE: To compare monocytes from SLE patients, patients with rheumatoid arthritis, and healthy controls for their ability to differentiate in vitro into macrophages and to bind/engulf apoptotic cells. METHODS: Peripheral blood derived monocytes from healthy donors or from patients with SLE or rheumatoid arthritis were allowed to differentiate into macrophages. The in vitro uptake of apoptotic cells by macrophages was evaluated by a flow cytometry assay that allowed discrimination between binding and internalisation. RESULTS: Monocytes from SLE and rheumatoid patients showed a striking defect in adherence to plastic compared with healthy donors. Absence or heat inactivation of serum resulted in a reduction in the binding and engulfment of apoptotic cells by macrophages. Macrophages from rheumatoid and SLE patients had similar percentages of apoptotic cells bound to their surface compared with normal controls. However, macrophages from SLE patients showed a significant defect in the internalisation of apoptotic cells compared with those from healthy controls, even in the presence of normal human serum. CONCLUSIONS: Monocytes from patients with SLE and rheumatoid arthritis have a similar defect in their capacity to adhere to plastic. However, only macrophages from SLE patients showed an impaired ability to engulf apoptotic cells, which indicates that an intrinsic cellular defect may be responsible for this phenomenon.
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Artritis Reumatoide/patología , Lupus Eritematoso Sistémico/patología , Macrófagos/fisiología , Adolescente , Adulto , Anciano , Análisis de Varianza , Apoptosis , Estudios de Casos y Controles , Adhesión Celular , Separación Celular , Células Cultivadas , Femenino , Citometría de Flujo , Humanos , Células Jurkat/patología , Células Jurkat/efectos de la radiación , Masculino , Persona de Mediana Edad , Fagocitosis , Rayos Ultravioleta/efectos adversosRESUMEN
The New Zealand Black (NZB) mouse strain is a model of autoimmune haemolytic anaemia (AHA) and systemic lupus erythematosus (SLE), characterized by the production of anti-red blood cell (RBC) antibodies and anti-nuclear antibodies (ANA), respectively. A linkage analysis was carried out in an (NZB x BALB/c) F(2) cross in order to identify loci involved in the production of both anti-RBC IgM and IgG antibodies. These regions of linkage were compared with linkage data to ANA from the same cohort and other linkage analyses involving New Zealand mice. Four previously described NZB loci linked to anti-RBC antibodies were confirmed, and eight novel loci linked to this trait were also mapped: five of which were of NZB origin, and three derived from the non-autoimmune BALB/c background. A comparison between loci linked with anti-RBC antibodies and ANA demonstrated many that co-localize, suggesting the presence of genes that result in the general breaking of tolerance to self-antigen. Furthermore, the observation that some loci were associated only with the anti-RBC response suggests an antigen specific mechanism in addition to a general breaking of tolerance. A locus linked with anti-RBC antibodies and ANA on distal chromosome 7 in this cohort is orthologous to one on the q arm of human chromosome 11, a region linked to AHA and ANA in human SLE.
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Anemia Hemolítica Autoinmune/inmunología , Formación de Anticuerpos/inmunología , Eritrocitos/inmunología , Lupus Eritematoso Sistémico/inmunología , Anemia Hemolítica Autoinmune/genética , Anemia Hemolítica Autoinmune/patología , Animales , Anticuerpos Antinucleares/genética , Anticuerpos Antinucleares/inmunología , Formación de Anticuerpos/genética , Cromosomas de los Mamíferos/genética , Cromosomas de los Mamíferos/inmunología , Ligamiento Genético/genética , Ligamiento Genético/inmunología , Inmunoglobulina G/sangre , Inmunoglobulina M/sangre , Hígado/inmunología , Hígado/patología , Lupus Eritematoso Sistémico/genética , Lupus Eritematoso Sistémico/patología , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos NZBRESUMEN
An increased number of apoptotic bodies have been detected in glomeruli of non-nephritic kidneys of C1q-deficient mice. In these mice an in vivo impaired uptake of apoptotic cells by peritoneal macrophages was also demonstrated. Here we investigated whether C1q plays a role in the in vitro clearance of apoptotic cells by glomerular mesangial cells. Phagocytosis was assessed using a novel flow cytometric assay that was validated by immunofluorescence studies. The uptake of apoptotic cells by mesangial cells, measured as percentage of mesangial cells ingesting apoptotic cells, was approximately 25%, 10% and 10% for a T cell lymphoma line (RMA), thymocytes and neutrophils, respectively. The uptake reached a plateau phase after 3 h, was specific for apoptotic cells and was mediated by serum but not by complement components C1q or C3. The phagocytosis of apoptotic cells was significantly inhibited by Arg-Gly-Asp-Ser (RGDS), a peptide capable of blocking the interaction of thrombospondin with CD36 or the vitronectin receptor. Pretreatment of the mesangial cells with dexamethasone (200 nm) but not with LPS increased the uptake markedly. These findings indicate that murine mesangial cells are capable of taking up syngeneic apoptotic cells, although much less efficiently than professional phagocytic cells. They also show that serum proteins other than complement components mediate the removal of apoptotic cells by murine mesangial cells in vitro.
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Proteínas Sanguíneas/metabolismo , Complemento C1q , Mesangio Glomerular/citología , Fagocitosis , Animales , Apoptosis , Complemento C3 , Dexametasona/farmacología , Citometría de Flujo , Mesangio Glomerular/efectos de los fármacos , Glucocorticoides/farmacología , Leucocitos , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Neutrófilos , Oligopéptidos/farmacología , Estimulación Química , Células Tumorales CultivadasRESUMEN
Cryoglobulin activity associated with murine immunoglobulin G3 (IgG3) has been shown to play a significant role in the development of murine lupuslike glomerulonephritis. A fraction, but not all, IgG3 monoclonal antibodies are capable of inducing a severe acute lupuslike glomerulonephritis as a result of direct localization of IgG3 cryoglobulins, suggesting the importance of qualitative features of cryoglobulins in their nephritogenic activities. Here a remarkable difference is shown in the renal pathogenicity of 2 murine IgG3 monoclonal cryoglobulins, identical in the amino acid sequences of their heavy and light chains but different in galactosylation patterns of oligosaccharide side chains because of their synthesis in different myeloma cells. The antibody lacking the capacity to induce severe glomerulonephritis displayed an increased proportion of galactosylated heavy chains. Changes in conformation, as revealed by gel filtration analysis, reduced cryoglobulin activity, and accelerated clearance could account for the lack of the renal pathogenicity of the more galactosylated variant. This observation provides a direct demonstration for the role of IgG galactosylation in the pathogenic potential of cryoglobulins. (Blood. 2001;97:3537-3543)
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Anticuerpos Monoclonales/química , Crioglobulinas/química , Galactosa/metabolismo , Glomerulonefritis/inmunología , Inmunoglobulina G/química , Animales , Anticuerpos Monoclonales/metabolismo , Línea Celular , Crioglobulinas/metabolismo , ADN Complementario/química , Hibridomas/inmunología , Inmunoglobulina G/genética , Inmunoglobulina G/metabolismo , Cadenas Pesadas de Inmunoglobulina/química , Cadenas Pesadas de Inmunoglobulina/genética , Cadenas Pesadas de Inmunoglobulina/metabolismo , Glomérulos Renales/inmunología , Glomérulos Renales/patología , Tasa de Depuración Metabólica , Ratones , Ratones Endogámicos BALB C , Ácido N-Acetilneuramínico/análisis , Oligosacáridos/química , Conformación Proteica , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Análisis de Secuencia de ADN , Relación Estructura-Actividad , TransfecciónAsunto(s)
Anemia Hemolítica Autoinmune/inmunología , Autoanticuerpos/inmunología , Enfermedades Autoinmunes/inmunología , Isotipos de Inmunoglobulinas/inmunología , Animales , Anticuerpos Monoclonales/inmunología , Afinidad de Anticuerpos , Proteínas del Sistema Complemento/inmunología , Modelos Animales de Enfermedad , Eritrocitos/inmunología , Humanos , Cambio de Clase de Inmunoglobulina , Fragmentos Fab de Inmunoglobulinas/inmunología , Fragmentos Fc de Inmunoglobulinas/inmunología , Ratones , Ratones Endogámicos , Receptores de IgG/inmunologíaRESUMEN
TWEAK is a newly identified member of the Tumor Necrosis Factor (TNF) family of proteins which are involved in many immunoinflammatory mechanisms. The putative role of TWEAK in inflammation was analyzed in mice treated with lipopolysaccharide (LPS), a strong inducer of the immuno-inflammatory responses. TWEAK mRNA rapidly disappeared in all the tissues tested. Analysis of LPS-treated thioglycolate-elicited peritoneal macrophages revealed that the rapid loss of TWEAK mRNA was due to its active destabilization. In chronic pathologies like autoimmune hemolytic anemia in the NZB mouse strain or systemic lupus erythematosus (SLE) in the BXSB mouse strain, TWEAK mRNA was shown to be reduced concomitantly to the development of chronic autoimmune diseases. These results demonstrated that TWEAK mRNA, contrary to TNF mRNA, is stable, ubiquitously distributed in tissues, and is down-regulated after LPS treatment or in chronic inflammation, suggesting that TWEAK could be an important factor, along with TNF, in acute and chronic inflammations.
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Proteínas Portadoras/genética , Regulación hacia Abajo , Inflamación/genética , ARN Mensajero/genética , Enfermedad Aguda , Animales , Proteínas Reguladoras de la Apoptosis , Enfermedades Autoinmunes/genética , Enfermedad Crónica , Citocina TWEAK , Lipopolisacáridos/administración & dosificación , Macrófagos Peritoneales/metabolismo , Ratones , Factores de Necrosis TumoralRESUMEN
The BXSB murine strain spontaneously develops an autoimmune syndrome with features of systemic lupus erythematosus (SLE) that affects males much earlier than females, due to the presence of an as yet unidentified mutant gene located on its Y chromosome, designated Yaa (Y-linked autoimmune acceleration). The Yaa gene by itself is unable to induce significant autoimmune responses in mice without an apparent SLE background, while it can induce and accelerate the development of an SLE in combination with autosomal susceptibility alleles present in lupus-prone mice. Although the genes encoded within or closely linked to the MHC locus play an important role in the development or protection of SLE, the MHC effect can be completely masked by the presence of the Yaa gene in mice highly predisposed to SLE. The role of the Yaa gene for the acceleration of SLE is apparently two-fold; it enhances overall autoimmune responses against autoantigens to which mice respond relatively weakly, and promotes Th 1 responses against autoantigens to which mice respond relatively well, leading to the production of more pathogenic autoantibodies, i.e., FcgammaR-fixing IgG2a and cryoglobulin IgG3 autoantibodies. Yaa+ - Yaa- double bone marrow chimera experiments revealed that the Yaa defect is expressed in B cells, but not in T cells, and that T cells from non-autoimmune mice are capable of providing help for autoimmune responses by collaborating Yaa+ B cells. We speculate that the Yaa defect may decrease the threshold for antigen receptor-dependent stimulation, leading to the triggering and excessive stimulation of autoreactive T and B cells.
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Modelos Animales de Enfermedad , Lupus Eritematoso Sistémico/genética , Animales , Síndrome Antifosfolípido/etiología , Autoanticuerpos/biosíntesis , Autoinmunidad , Predisposición Genética a la Enfermedad , Antígenos H-2/genética , Humanos , Ratones , Cromosoma YRESUMEN
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 acquired immunodeficiency syndrome (MAIDS) is characterized by lymphoproliferation, polyclonal B cell activation resulting in the production of autoantibodies, and a progressive immunodeficiency. These are all hallmarks of some autoimmune diseases. Yaa is a Y-chromosome-linked gene that accelerates autoimmune diseases in some autoimmune-prone strains of mice. To further elucidate a possible relationship with autoimmunity, the effect of the Yaa gene on MAIDS was investigated. Analysis of phenotypic and functional disease parameters revealed that Yaa does not accelerate MAIDS disease. This is probably due to the generalized activation of most or all lymphoid cells in MAIDS, which cannot be enhanced by the Yaa gene. This result is in accordance with the selective enhancing effect of the Yaa gene on the immune response against self and foreign antigens in a specific genetic background. It suggests that the autoimmune response associated with MAIDS is a secondary phenomenon. Interestingly, even in wild-type C57BL/6 mice, autoantibody production may contribute overproportionally to the hypergammaglobulinemia associated with MAIDS.
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Autoinmunidad/genética , Síndrome de Inmunodeficiencia Adquirida del Murino/inmunología , Mutación , Cromosoma Y/inmunología , Animales , Anticuerpos Antinucleares/inmunología , Linfocitos B/inmunología , Línea Celular , Células Cultivadas , ADN de Cadena Simple/inmunología , Ratones , Ratones Endogámicos C57BL , Ratones Mutantes , Síndrome de Inmunodeficiencia Adquirida del Murino/genética , Síndrome de Inmunodeficiencia Adquirida del Murino/fisiopatología , Bazo/citología , Antígenos Thy-1/inmunologíaRESUMEN
To assess the potency of low-affinity anti-red blood cell (RBC) autoantibodies in the induction of anemia, we generated an immunoglobulin (Ig)G2a class-switch variant of a 4C8 IgM anti-mouse RBC autoantibody, and compared its pathogenic potential with that of its IgM isotype and a high-affinity 34-3C IgG2a autoantibody. The RBC-binding activity of the 4C8 IgG2a variant was barely detectable, at least 1,000 times lower than that of its IgM isotype, having a high-binding avidity, and that of the 34-3C IgG2a monoclonal antibody (mAb). This low-affinity feature of the 4C8 mAb was consistent with the lack of detection of opsonized RBCs in the circulating blood from the 4C8 IgG2a-injected mice. However, the 4C8 IgG2a variant was highly pathogenic, as potent as its IgM isotype and the 34-3C IgG2a mAb, due to its capacity to interact with Fc receptors involved in erythrophagocytosis. In addition, our results indicated that the pentameric form of the low-affinity IgM isotype, by promoting the binding and agglutination of RBCs, is critical for its pathogenic activity. Demonstration of the remarkably high pathogenic potency of low-affinity autoantibodies, if combined with appropriate heavy chain effector functions, highlights the critical role of the Ig heavy chain constant regions, but the relatively minor role of autoantigen-binding affinities, in autoimmune hemolytic anemia.