RESUMO
Antibody-secreting plasma cells (PCs) arise rapidly during adaptive immunity to control infections. The early PCs are retained within the reactive lymphoid organ where their localization and homeostasis rely on extrinsic factors, presumably produced by local niche cells. While myeloid cells have been proposed to form those niches, the contribution by colocalizing stromal cells has remained unclear. Here, we characterized a subset of fibroblastic reticular cells (FRCs) that forms a dense meshwork throughout medullary cords of lymph nodes (LNs) where PCs reside. This medullary FRC type is shown to be anatomically, phenotypically, and functionally distinct from T zone FRCs, both in mice and humans. By using static and dynamic imaging approaches, we provide evidence that medullary FRCs are the main cell type in contact with PCs guiding them in their migration. Medullary FRCs also represent a major local source of the PC survival factors IL-6, BAFF, and CXCL12, besides also producing APRIL. In vitro, medullary FRCs alone or in combination with macrophages promote PC survival while other LN cell types do not have this property. Thus, we propose that this FRC subset, together with medullary macrophages, forms PC survival niches within the LN medulla, and thereby helps in promoting the rapid development of humoral immunity, which is critical in limiting early pathogen spread.
Assuntos
Formação de Anticorpos , Homeostase/imunologia , Linfonodos/imunologia , Plasmócitos/imunologia , Animais , Fator Ativador de Células B/imunologia , Quimiocina CXCL12/imunologia , Interleucina-6/imunologia , Linfonodos/citologia , Masculino , Camundongos , Plasmócitos/citologia , Células Estromais/citologia , Células Estromais/imunologiaRESUMO
INTRODUCTION: Previous studies have shown increased expression of stromal markers in synovial tissue (ST) of patients with established rheumatoid arthritis (RA). Here, ST expression of stromal markers in early arthritis in relationship to diagnosis and prognostic outcome was studied. METHODS: ST from 56 patients included in two different early arthritis cohorts and 7 non-inflammatory controls was analysed using immunofluorescence to detect stromal markers CD55, CD248, fibroblast activation protein (FAP) and podoplanin. Diagnostic classification (gout, psoriatic arthritis, unclassified arthritis (UA), parvovirus associated arthritis, reactive arthritis and RA), disease outcome (resolving vs persistent) and clinical variables were determined at baseline and after follow-up, and related to the expression of stromal markers. RESULTS: We observed expression of all stromal markers in ST of early arthritis patients, independent of diagnosis or prognostic outcome. Synovial expression of FAP was significantly higher in patients developing early RA compared to other diagnostic groups and non-inflammatory controls. In RA FAP protein was expressed in both lining and sublining layers. Podoplanin expression was higher in all early inflammatory arthritis patients than controls, but did not differentiate diagnostic outcomes. Stromal marker expression was not associated with prognostic outcomes of disease persistence or resolution. There was no association with clinical or sonographic variables. CONCLUSIONS: Stromal cell markers CD55, CD248, FAP and podoplanin are expressed in ST in the earliest stage of arthritis. Baseline expression of FAP is higher in early synovitis patients who fulfil classification criteria for RA over time. These results suggest that significant fibroblast activation occurs in RA in the early window of disease.
Assuntos
Artrite/metabolismo , Células Estromais/metabolismo , Membrana Sinovial/metabolismo , Adulto , Idoso , Antígenos CD/metabolismo , Antígenos de Neoplasias/metabolismo , Artrite/diagnóstico , Biomarcadores/metabolismo , Antígenos CD55/metabolismo , Progressão da Doença , Endopeptidases , Feminino , Fibroblastos/metabolismo , Gelatinases/metabolismo , Humanos , Masculino , Glicoproteínas de Membrana/metabolismo , Proteínas de Membrana/metabolismo , Pessoa de Meia-Idade , Prognóstico , Serina Endopeptidases/metabolismo , Sinovite/diagnóstico , Sinovite/metabolismoRESUMO
BACKGROUND: We investigated two distinct synovial fibroblast populations that were located preferentially in the lining or sub-lining layers and defined by their expression of either podoplanin (PDPN) or CD248, and explored their ability to undergo self-assembly and transmigration in vivo. METHODS: Synovial fibroblasts (SF) were cultured in vitro and phenotypic changes following stimulation with interleukin (IL)-1ß, tumor necrosis factor (TNF)-α, and transforming growth factor (TGF)-ß1 were examined. To examine the phenotype of SF in vivo, a severe combined immunodeficiency (SCID) human-mouse model of cartilage destruction was utilised. RESULTS: SF in the lining layer in rheumatoid arthritis (RA) expressed high levels of PDPN compared to the normal synovium, whereas CD248 expression was restricted to sub-lining layer cells. TNF-α or IL1 stimulation in vitro resulted in an increased expression of PDPN. In contrast, stimulation with TGF-ß1 induced CD248 expression. In the SCID human-mouse model, rheumatoid SF recapitulated the expression of PDPN and CD248. Fibroblasts adjacent to cartilage expressed PDPN, and attached to, invaded, and degraded cartilage. PDPN+ CD248- SF preceded the appearance of PDPN- CD248+ cells in contralateral implants. CONCLUSIONS: We have identified two distinct SF populations identified by expression of either PDPN or CD248 which are located within different anatomical compartments of the inflamed synovial membrane. These markers discriminate between SF subsets with distinct biological properties. As PDPN-expressing cells are associated with early fibroblast migration and cartilage erosion in vivo, we propose that PDPN-expressing cells may be an attractive therapeutic target in RA.
Assuntos
Artrite Reumatoide/patologia , Fibroblastos/citologia , Membrana Sinovial/citologia , Idoso , Animais , Antígenos CD/metabolismo , Antígenos de Neoplasias/metabolismo , Cartilagem Articular/metabolismo , Diferenciação Celular/fisiologia , Citocinas/metabolismo , Feminino , Citometria de Fluxo , Xenoenxertos , Humanos , Masculino , Glicoproteínas de Membrana/metabolismo , Camundongos , Camundongos SCID , Microscopia de Fluorescência , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase , Migração Transendotelial e Transepitelial/fisiologiaRESUMO
INTRODUCTION: Monocytic cells play a central role in the aetiology of rheumatoid arthritis, and manipulation of the activation of these cells is an approach currently under investigation to discover new therapies for this and associated diseases. CD148 is a transmembrane tyrosine phosphatase that is highly expressed in monocytes and macrophages and, since this family of molecules plays an important role in the regulation of cell activity, CD148 is a potential target for the manipulation of macrophage activation. For any molecule to be considered a therapeutic target, it is important for it to be increased in activity or expression during disease. METHODS: We have investigated the expression of CD148 in two murine models of arthritis and in joints from rheumatoid arthritis (RA) patients using real-time PCR, immunohistochemistry, and studied the effects of proinflammatory stimuli on CD148 activity using biochemical assays. RESULTS: We report that CD148 mRNA is upregulated in diseased joints of mice with collagen-induced arthritis. Furthermore, we report that in mice CD148 protein is highly expressed in infiltrating monocytes of diseased joints, with a small fraction of T cells also expressing CD148. In human arthritic joints both T cells and monocytes expressed high levels of CD148, however, we show differential expression of CD148 in T cells and monocytes from normal human peripheral blood compared to peripheral blood from RA and both normal and RA synovial fluid. Finally, we show that synovial fluid from rheumatoid arthritis patients suppresses CD148 phosphatase activity. CONCLUSIONS: CD148 is upregulated in macrophages and T cells in human RA samples, and its activity is enhanced by treatment with tumour necrosis factor alpha (TNFα), and reduced by synovial fluid or oxidising conditions. A greater understanding of the role of CD148 in chronic inflammation may lead to alternative therapeutic approaches to these diseases.
Assuntos
Artrite Experimental/genética , Artrite Reumatoide/genética , Perfilação da Expressão Gênica , Leucócitos Mononucleares/metabolismo , Animais , Artrite Experimental/metabolismo , Artrite Experimental/patologia , Artrite Reumatoide/metabolismo , Artrite Reumatoide/patologia , Humanos , Peróxido de Hidrogênio/farmacologia , Imuno-Histoquímica , Articulações/metabolismo , Articulações/patologia , Leucócitos Mononucleares/patologia , Macrófagos/metabolismo , Macrófagos/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos DBA , Microscopia Confocal , Monócitos/metabolismo , Monócitos/patologia , Oxidantes/farmacologia , Proteínas Tirosina Fosfatases Classe 3 Semelhantes a Receptores/genética , Proteínas Tirosina Fosfatases Classe 3 Semelhantes a Receptores/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Líquido Sinovial/metabolismo , Linfócitos T/efeitos dos fármacos , Linfócitos T/metabolismo , Linfócitos T/patologia , Fator de Necrose Tumoral alfa/farmacologia , Regulação para CimaRESUMO
Aseptic lymphocyte-dominated vasculitis-associated lesion (ALVAL) has been used to describe the histological lesion associated with metal-on-metal (M-M) bearings. We tested the hypothesis that the lymphoid aggregates, associated with ALVAL lesions resemble tertiary lymphoid organs (TLOs). Histopathological changes were examined in the periprosthetic tissue of 62 M-M hip replacements requiring revision surgery, with particular emphasis on the characteristics and pattern of the lymphocytic infiltrate. Immunofluorescence and immunohistochemistry were used to study the classical features of TLOs in cases where large organized lymphoid follicles were present. Synchrotron X-ray fluorescence (XRF) measurements were undertaken to detect localisation of implant derived ions/particles within the samples. Based on type of lymphocytic infiltrates, three different categories were recognised; diffuse aggregates (51%), T cell aggregates (20%), and organised lymphoid aggregates (29%). Further investigation of tissues with organised lymphoid aggregates showed that these tissues recapitulate many of the features of TLOs with T cells and B cells organised into discrete areas, the presence of follicular dendritic cells, acquisition of high endothelial venule like phenotype by blood vessels, expression of lymphoid chemokines and the presence of plasma cells. Co-localisation of implant-derived metals with lymphoid aggregates was observed. These findings suggest that in addition to the well described general foreign body reaction mediated by macrophages and a T cell mediated type IV hypersensitivity response, an under-recognized immunological reaction to metal wear debris involving B cells and the formation of tertiary lymphoid organs occurs in a distinct subset of patients with M-M implants.
Assuntos
Articulação do Quadril/patologia , Prótese de Quadril , Linfonodos/patologia , Próteses Articulares Metal-Metal , Fator Ativador de Células B/metabolismo , Linfócitos B/metabolismo , Linfócitos B/patologia , Proliferação de Células , Sobrevivência Celular , Quimiocina CCL21/metabolismo , Quimiocina CXCL13/metabolismo , Células Endoteliais/patologia , Articulação do Quadril/cirurgia , Humanos , Íons , Linfócitos T/metabolismo , Linfócitos T/patologia , Membro 13 da Superfamília de Ligantes de Fatores de Necrose Tumoral/metabolismo , Vênulas/patologiaRESUMO
CD248 (endosialin) is a transmembrane glycoprotein that is dynamically expressed on pericytes and fibroblasts during tissue development, tumour neovascularization and inflammation. Its role in tissue remodelling is associated with increased stromal cell proliferation and migration. We show that CD248 is also uniquely expressed by human, but not mouse (C57BL/6), CD8(+) naive T cells. CD248 is found only on CD8(+) CCR7(+) CD11a(low) naive T cells and on CD8 single-positive T cells in the thymus. Transfection of the CD248 negative T-cell line MOLT-4 with CD248 cDNA surprisingly reduced cell proliferation. Knock-down of CD248 on naive CD8 T cells increased cell proliferation. These data demonstrate opposing functions for CD248 on haematopoietic (CD8(+)) versus stromal cells and suggests that CD248 helps to maintain naive CD8(+) human T cells in a quiescent state.
Assuntos
Antígenos CD/metabolismo , Antígenos de Neoplasias/metabolismo , Linfócitos T CD8-Positivos/imunologia , Células Estromais/imunologia , Animais , Western Blotting , Proliferação de Células , Citometria de Fluxo , Humanos , CamundongosRESUMO
Ectopic or tertiary lymphoid tissues (TLTs) are often induced at sites of chronic inflammation. They typically contain various hematopoietic cell types, high endothelial venules, and follicular dendritic cells; and are organized in lymph node-like structures. Although fibroblastic stromal cells may play a role in TLT induction and persistence, they have remained poorly defined. Herein, we report that TLTs arising during inflammation in mice and humans in a variety of tissues (eg, pancreas, kidney, liver, and salivary gland) contain stromal cell networks consisting of podoplanin(+) T-zone fibroblastic reticular cells (TRCs), distinct from follicular dendritic cells. Similar to lymph nodes, TRCs were present throughout T-cell-rich areas and had dendritic cells associated with them. They expressed lymphotoxin (LT) ß receptor (LTßR), produced CCL21, and formed a functional conduit system. In rat insulin promoter-CXCL13-transgenic pancreas, the maintenance of TRC networks and conduits was partially dependent on LTßR and on lymphoid tissue inducer cells expressing LTßR ligands. In conclusion, TRCs and conduits are hallmarks of secondary lymphoid organs and of well-developed TLTs, in both mice and humans, and are likely to act as important scaffold and organizer cells of the T-cell-rich zone.
Assuntos
Tecido Linfoide/metabolismo , Animais , Quimiocina CXCL13/genética , Células Dendríticas/metabolismo , Fibroblastos/metabolismo , Humanos , Inflamação , Insulina/genética , Ligantes , Linfonodos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Pancrelipase/metabolismo , Regiões Promotoras Genéticas , Ratos , Células Estromais/citologiaRESUMO
CD248 is a cell surface receptor that specifically identifies fibroblasts and pericytes during development and in association with cancer and inflammation. However, its function is poorly defined and its role in lymphoid organs not studied. Here, we used (4-hydroxy-3-nitrophenyl)acetyl chicken gamma-globulin immunisation and mice lacking CD248 to study whether CD248 modulates popliteal LN (pLN) expansion and subsequent immune responses. We have found that CD248 is required for complete pLN expansion but not for co-ordination of B and T cell compartmentalisation or antibody production following (4-hydroxy-3-nitrophenyl)acetyl chicken gamma-globulin immunisation. In vitro, we show that CD248 expression in human MG63 stromal cells and mouse embryonic fibroblasts leads to a pro-proliferative and pro-migratory phenotype. This correlates with a proliferating CD248(+) population observed in vivo during pLN expansion. Taken together, these data highlight a role for CD248 in secondary lymphoid organ remodelling during adaptive immune responses.
Assuntos
Antígenos CD/metabolismo , Antígenos de Neoplasias/metabolismo , Fibroblastos/metabolismo , Linfonodos/imunologia , Proteínas de Neoplasias/metabolismo , Pericitos/metabolismo , Células Estromais/metabolismo , Animais , Formação de Anticorpos/genética , Antígenos CD/genética , Antígenos CD/imunologia , Antígenos de Neoplasias/imunologia , Linfócitos B/imunologia , Biomarcadores/metabolismo , Comunicação Celular/genética , Linhagem Celular , Movimento Celular/genética , Proliferação de Células , Galinhas , Fibroblastos/imunologia , Fibroblastos/patologia , Haptenos/imunologia , Humanos , Imunização , Linfonodos/crescimento & desenvolvimento , Linfonodos/patologia , Camundongos , Camundongos Knockout , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/imunologia , Nitrofenóis/imunologia , Pericitos/imunologia , Pericitos/patologia , Fenilacetatos/imunologia , Células Estromais/imunologia , Células Estromais/patologia , Linfócitos T/imunologia , gama-Globulinas/imunologiaRESUMO
Hematopoiesis is a carefully controlled process that is regulated by complex networks of transcription factors that are, in part, controlled by signals resulting from ligand binding to cell-surface receptors. To further understand hematopoiesis, we have compared gene expression profiles of human erythroblasts, megakaryocytes, B cells, cytotoxic and helper T cells, natural killer cells, granulocytes, and monocytes using whole genome microarrays. A bioinformatics analysis of these data was performed focusing on transcription factors, immunoglobulin superfamily members, and lineage-specific transcripts. We observed that the numbers of lineage-specific genes varies by 2 orders of magnitude, ranging from 5 for cytotoxic T cells to 878 for granulocytes. In addition, we have identified novel coexpression patterns for key transcription factors involved in hematopoiesis (eg, GATA3-GFI1 and GATA2-KLF1). This study represents the most comprehensive analysis of gene expression in hematopoietic cells to date and has identified genes that play key roles in lineage commitment and cell function. The data, which are freely accessible, will be invaluable for future studies on hematopoiesis and the role of specific genes and will also aid the understanding of the recent genome-wide association studies.
Assuntos
Células da Medula Óssea/fisiologia , Diferenciação Celular/genética , Expressão Gênica , Atlas como Assunto , Linhagem da Célula , Células Cultivadas , Citometria de Fluxo , Perfilação da Expressão Gênica , Hematopoese , Humanos , Análise de Sequência com Séries de Oligonucleotídeos , Fatores de Transcrição/metabolismoRESUMO
INTRODUCTION: A surprising feature of the inflammatory infiltrate in rheumatoid arthritis is the accumulation of neutrophils within synovial fluid and at the pannus cartilage boundary. Recent findings suggest that a distinct subset of IL-17-secreting T-helper cells (TH17 cells) plays a key role in connecting the adaptive and innate arms of the immune response and in regulating neutrophil homeostasis. We therefore tested the hypothesis that synovial fibroblasts bridge the biological responses that connect TH17 cells to neutrophils by producing neutrophil survival factors following their activation with IL-17. METHODS: IL-17-expressing cells in the rheumatoid synovium, and IL-17-expressing cells in the peripheral blood, and synovial fluid were examined by confocal microscopy and flow cytometry, respectively. Peripheral blood neutrophils were cocultured either with rheumatoid arthritis synovial fibroblasts (RASF) or with conditioned medium from RASF that had been pre-exposed to recombinant human IL-17, TNFalpha or a combination of the two cytokines. Neutrophils were harvested and stained with the vital mitochondrial dye 3,3'-dihexyloxacarbocyanine iodide before being enumerated by flow cytometry. RESULTS: TH17-expressing CD4+ cells were found to accumulate within rheumatoid synovial tissue and in rheumatoid arthritis synovial fluid. RASF treated with IL-17 and TNFalpha (RASFIL-17/TNF) effectively doubled the functional lifespan of neutrophils in coculture. This was entirely due to soluble factors secreted from the fibroblasts. Specific depletion of granulocyte-macrophage colony-stimulating factor from RASFIL-17/TNF-conditioned medium demonstrated that this cytokine accounted for approximately one-half of the neutrophil survival activity. Inhibition of phosphatidylinositol-3-kinase and NF-kappaB pathways showed a requirement for both signalling pathways in RASFIL-17/TNF-mediated neutrophil rescue. CONCLUSION: The increased number of neutrophils with an extended lifespan found in the rheumatoid synovial microenvironment is partly accounted for by IL-17 and TNFalpha activation of synovial fibroblasts. TH17-expressing T cells within the rheumatoid synovium are likely to contribute significantly to this effect.
Assuntos
Artrite Reumatoide/imunologia , Fibroblastos/metabolismo , Fator Estimulador de Colônias de Granulócitos e Macrófagos/metabolismo , Interleucina-17/metabolismo , Neutrófilos/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Adulto , Idoso , Artrite Reumatoide/metabolismo , Linfócitos T CD4-Positivos/metabolismo , Sobrevivência Celular/fisiologia , Feminino , Citometria de Fluxo , Humanos , Masculino , Microscopia Confocal , Pessoa de Meia-Idade , Membrana Sinovial/citologia , Membrana Sinovial/imunologia , Membrana Sinovial/metabolismo , Subpopulações de Linfócitos T/metabolismoRESUMO
Stromal cells such as fibroblasts play an important role in defining tissue-specific responses during the resolution of inflammation. We hypothesized that this involves tissue-specific regulation of glucocorticoids, mediated via differential regulation of the enzyme 11beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD1). Expression, activity and function of 11beta-HSD1 was assessed in matched fibroblasts derived from various tissues (synovium, bone marrow and skin) obtained from patients with rheumatoid arthritis or osteoarthritis. 11beta-HSD1 was expressed in fibroblasts from all tissues but mRNA levels and enzyme activity were higher in synovial fibroblasts (2-fold and 13-fold higher mRNA levels in dermal and synovial fibroblasts, respectively, relative to bone marrow). Expression and activity of the enzyme increased in all fibroblasts following treatment with tumour necrosis factor-alpha or IL-1beta (bone marrow: 8-fold and 37-fold, respectively, compared to vehicle; dermal fibroblasts: 4-fold and 14-fold; synovial fibroblasts: 7-fold and 31-fold; all P < 0.01 compared with vehicle). Treatment with IL-4 or interferon-gamma was without effect, and there was no difference in 11beta-HSD1 expression between fibroblasts (from any site) obtained from patients with rheumatoid arthritis or osteoarthritis. In the presence of 100 nmol/l cortisone, IL-6 production--a characteristic feature of synovial derived fibroblasts--was significantly reduced in synovial but not dermal or bone marrow fibroblasts. This was prevented by co-treatment with an 11beta-HSD inhibitor, emphasizing the potential for autocrine activation of glucocorticoids in synovial fibroblasts. These data indicate that differences in fibroblast-derived glucocorticoid production (via the enzyme 11beta-HSD1) between cells from distinct anatomical locations may play a key role in the predeliction of certain tissues to develop persistent inflammation.
Assuntos
11-beta-Hidroxiesteroide Desidrogenase Tipo 1/metabolismo , Artrite Reumatoide/enzimologia , Fibroblastos/enzimologia , Osteoartrite/enzimologia , 11-beta-Hidroxiesteroide Desidrogenase Tipo 1/genética , Artrite Reumatoide/metabolismo , Medula Óssea/enzimologia , Medula Óssea/metabolismo , Medula Óssea/patologia , Células Cultivadas , Cortisona/farmacologia , Glucocorticoides/biossíntese , Humanos , Interleucina-1beta/farmacologia , Interleucina-6/antagonistas & inibidores , Interleucina-6/biossíntese , Osteoartrite/metabolismo , RNA Mensageiro/metabolismo , Pele/enzimologia , Pele/metabolismo , Pele/patologia , Membrana Sinovial/enzimologia , Membrana Sinovial/metabolismo , Membrana Sinovial/patologia , Fator de Necrose Tumoral alfa/farmacologiaRESUMO
In hepatitis C virus (HCV) infection the immune response is ineffective, leading to chronic hepatitis and liver damage. Primed CD8 T cells are critical for antiviral immunity and subsets of circulating CD8 T cells have been defined in blood but these do not necessarily reflect the clonality or differentiation of cells within tissue. Current models divide primed CD8 T cells into effector and memory cells, further subdivided into central memory (CCR7+, L-selectin+), recirculating through lymphoid tissues and effector memory (CCR7-, L-selectin-) mediating immune response in peripheral organs. We characterized CD8 T cells derived from organ donors and patients with end-stage HCV infection to show that: 1) all liver-infiltrating CD8 T cells express high levels of CD11a, indicating the effective absence of naive CD8 T cells in the liver. 2) The liver contains distinct subsets of primed CD8+ T cells including a population of CCR7+ L-selectin- cells, which does not reflect current paradigms. The expression of CCR7 by these cells may be induced by the hepatic microenvironment to facilitate recirculation. 3) The CCR7 ligands CCL19 and CCL21 are present on lymphatic, vascular, and sinusoidal endothelium in normal liver and in patients with HCV infection. We suggest that the recirculation of CCR7+/L-selectin- intrahepatic CD8 T cells to regional lymphoid tissue will be facilitated by CCL19 and CCL21 on hepatic sinusoids and lymphatics. This centripetal pathway of migration would allow restimulation in lymph nodes, thereby promoting immune surveillance in normal liver and renewal of effector responses in chronic viral infection.
Assuntos
Linfócitos T CD8-Positivos/imunologia , Movimento Celular/imunologia , Hepatite C Crônica/imunologia , Hepatócitos/imunologia , Memória Imunológica/imunologia , Imunofenotipagem , Receptores de Retorno de Linfócitos/biossíntese , Subpopulações de Linfócitos T/imunologia , Antígeno CD11a/biossíntese , Antígeno CD11a/sangue , Linfócitos T CD8-Positivos/metabolismo , Linfócitos T CD8-Positivos/patologia , Quimiocina CCL19 , Quimiocinas CC/sangue , Quimiocinas CC/metabolismo , Criança , Hepacivirus/imunologia , Hepatite C Crônica/metabolismo , Hepatite C Crônica/patologia , Hepatócitos/citologia , Hepatócitos/patologia , Humanos , Antígenos Comuns de Leucócito/biossíntese , Antígenos Comuns de Leucócito/sangue , Ligantes , Cirrose Hepática/imunologia , Cirrose Hepática/metabolismo , Antígeno-1 Associado à Função Linfocitária/biossíntese , Antígeno-1 Associado à Função Linfocitária/sangue , Receptores CCR7 , Receptores CXCR4/biossíntese , Receptores CXCR4/sangue , Receptores CXCR4/metabolismo , Receptores de Quimiocinas/sangue , Receptores de Quimiocinas/metabolismo , Receptores de Retorno de Linfócitos/sangue , Subpopulações de Linfócitos T/citologia , Subpopulações de Linfócitos T/metabolismo , Subpopulações de Linfócitos T/patologia , Membro 7 da Superfamília de Receptores de Fatores de Necrose Tumoral/biossíntese , Membro 7 da Superfamília de Receptores de Fatores de Necrose Tumoral/sangueRESUMO
ART4 (CD297) is a member of the family of toxin-related ADP-ribosyltransferases (ARTs) and is the carrier of the Dombrock blood group alloantigens (Do). Two mouse monoclonal antibodies (MIMA-52 and MIMA-53), and two rat monoclonal antibodies (N0NI-B4 and NONI-B63) were obtained following immunization of mice with human Do/ART4-transfected cells and of rats with human Do/ART4 cDNA, respectively. All four mAbs recognize Do/ART4-transfected Jurkat cells but not untransfected cells by FACS analysis. Staining of Do/ART4-transfected cells by these mAbs was reduced following treatment of cells with PI-PLC, confirming that Do/ART4 is anchored in the cell membrane by linkage to glycosylphosphatidylinositol as predicted from its amino acid sequence. The four mAbs did not react with Gy(a-) (Dombrock null) erythrocytes but agglutinated other red blood cells. By flow cytometric analysis, all mAbs reacted prominently with erythrocytes, and weakly with peripheral blood monocytes and splenic macrophages, but not with B-lymphocytes or T-lymphocytes. The mAbs reacted weakly also with human umbilical vein endothelial cells and the basophilic leukemia KU-812. Immunohistology revealed staining of epithelia and endothelia on sections of tonsils. In FACS analyses NONI-B4 competed with MIMA-52 for binding to Do/ART4-transfected cells and erythrocytes, whereas NONI-B63 competed with MIMA-53. Neither of the mAbs reacted with mouse ART4-transfected cells, but NONI-B63 and MIMA-53 did react with a mouse/human ART4 chimera, indicating that the epitope recognized by these mAbs lies in the C-terminal half of the protein.
Assuntos
ADP Ribose Transferases/imunologia , Proteínas de Ciclo Celular/imunologia , Proteínas de Membrana/imunologia , Proteínas de Neoplasias/imunologia , ADP Ribose Transferases/genética , ADP Ribose Transferases/metabolismo , Sequência de Aminoácidos , Animais , Anticorpos Monoclonais/biossíntese , Anticorpos Monoclonais/imunologia , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Reações Cruzadas , Endotélio Vascular/imunologia , Eritrócitos/imunologia , Proteínas Ligadas por GPI , Humanos , Macrófagos/imunologia , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Dados de Sequência Molecular , Monócitos/imunologia , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Tonsila Palatina/metabolismo , Ratos , Alinhamento de Sequência , Baço/imunologia , Veias Umbilicais/imunologiaRESUMO
The paradigm for tissue specific homing of leukocytes is the "area code" hypothesis, which predicts that a specific combination of adhesive interactions and chemokine signals from the endothelium directs leukocyte migration into specific tissue sites. This area code hypothesis has been supported by studies from previous HLDA workshops where endothelial specific cell antigens have been studied. Similarly, a clear haematopoietic "stem cell code" comprising the chemokine SDF-1 (CXCL12) and the adhesion receptor VCAM-1 (CD106) has been shown to contribute to the stem cell niche within bone marrow [K. Tokoyoda, T. Egawa, T. Sugiyama, B.I. Chai, T. Nagasawa, Cellular niches controlling B lymphocyte behaviour within bone marrow during development, Immunity 20 (2004) 707-718]. HLDA 7 included a section devoted to stem cell antigens, which began to define additional antigens important in these processes. During the course of HLDA 8 we have extended these observations to determine whether a more global stromal address code defined by fibroblasts, exists in variety of different tissues [G. Parsonage, A.D. Filer, O. Haworth, G.B. Nash, G.E. Rainger, M. Salmon, C.D. Buckley, A stromal area postcode defined by fibroblasts, Trends Immunol. 26 (2005) 150-156]. The stromal cell section in HLDA 8 was designed to complement the malignant cell, endothelial cell, and stem cell/progenitor cell sections. Seven new CD numbers were assigned to antibodies included in this section at the HLDA 8 Workshop meeting held during December 2004.
Assuntos
Células Estromais/imunologia , Anticorpos Monoclonais/imunologia , Especificidade de Anticorpos , Antígenos CD/análise , Células Endoteliais/imunologia , Citometria de Fluxo , Humanos , Imuno-Histoquímica , Neoplasias/imunologia , Células-Tronco/imunologiaRESUMO
Here we report that B cell receptor (BCR) engagement rapidly improves the capacity of CD20 to be accessed by cognate antibody at model Burkitt's lymphoma cell surfaces. None of eight other surface molecules demonstrated such BCR-dependent enhancement of ligand binding while the quantity of accessible CD20 remained unchanged on either CD19 or CD40 engagement. Neither the actin-depolymerizing agent cytochalasin D nor inhibitors targeting signalling pathways associated with the BCR attenuated the CD20 increase that could be uncoupled from BCR endocytosis. Instead, a role for lipid rafts was indicated both from the inhibitory actions of cholesterol-sequestering methyl-beta-cyclodextrin and direct analysis of CD20 redistribution using sucrose density gradients and confocal microscopy. Whether such observations could find application in CD20-directed therapies where success can be compromised by otherwise low-level expression of target antigen is discussed.