RESUMO
Most tissue-resident macrophages (Mφs) are believed to be derived prenatally and are assumed to maintain themselves throughout life by self-proliferation. However, in adult mice we identified a progenitor within bone marrow, early pro-B cell/fraction B, that differentiates into tissue Mφs. These Mφ precursors have non-rearranged B-cell receptor genes and coexpress myeloid (GR1, CD11b, and CD16/32) and lymphoid (B220 and CD19) lineage markers. During steady state, these precursors exit bone marrow, losing Gr1, and enter the systemic circulation, seeding the gastrointestinal system as well as pleural and peritoneal cavities but not the brain. While in these tissues, they acquire a transcriptome identical to embryonically derived tissue-resident Mφs. Similarly, these Mφ precursors also enter sites of inflammation, gaining CD115, F4/80, and CD16/32, and become indistinguishable from blood monocyte-derived Mφs. Thus, we have identified a population of cells within the bone marrow early pro-B cell compartment that possess functional plasticity to differentiate into either tissue-resident or inflammatory Mφs, depending on microenvironmental signals. We propose that these precursors represent an additional source of Mφ populations in adult mice during steady state and inflammation.
Assuntos
Ativação de Macrófagos/fisiologia , Macrófagos/imunologia , Células Precursoras de Linfócitos B/fisiologia , Animais , Linfócitos B/fisiologia , Medula Óssea , Células da Medula Óssea/fisiologia , Homeostase/fisiologia , Inflamação/imunologia , Macrófagos/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Monócitos/imunologia , Células Precursoras de Linfócitos B/imunologia , Células Precursoras de Linfócitos B/metabolismoRESUMO
Acute inflammation is traditionally characterized by polymorphonuclear leukocytes (PMN) influx followed by phagocytosing macrophage (Mφs) that clear injurious stimuli leading to resolution and tissue homeostasis. However, using the peritoneal cavity, we found that although innate immune-mediated responses to low-dose zymosan or bacteria resolve within days, these stimuli, but not hyperinflammatory stimuli, trigger a previously overlooked second wave of leukocyte influx into tissues that persists for weeks. These cells comprise distinct populations of tissue-resident Mφs (resMφs), Ly6c(hi) monocyte-derived Mφs (moMφs), monocyte-derived dendritic cells (moDCs), and myeloid-derived suppressor cells (MDSCs). Postresolution mononuclear phagocytes were observed alongside lymph node expansion and increased numbers of blood and peritoneal memory T and B lymphocytes. The resMφs and moMφs triggered FoxP3 expression within CD4 cells, whereas moDCs drive T-cell proliferation. The resMφs preferentially clear apoptotic PMNs and migrate to lymph nodes to bring about their contraction in an inducible nitric oxide synthase-dependent manner. Finally, moMφs remain in tissues for months postresolution, alongside altered numbers of T cells collectively dictating the magnitude of subsequent acute inflammatory reactions. These data challenge the prevailing idea that resolution leads back to homeostasis and asserts that resolution acts as a bridge between innate and adaptive immunity, as well as tissue reprogramming.
Assuntos
Imunidade Adaptativa/fisiologia , Fagocitose/fisiologia , Imunidade Adaptativa/efeitos dos fármacos , Animais , Linfócitos T CD4-Positivos/citologia , Linfócitos T CD4-Positivos/imunologia , Movimento Celular/efeitos dos fármacos , Movimento Celular/genética , Movimento Celular/imunologia , Proliferação de Células/efeitos dos fármacos , Células Dendríticas/citologia , Células Dendríticas/imunologia , Fatores de Transcrição Forkhead/genética , Fatores de Transcrição Forkhead/imunologia , Inflamação/induzido quimicamente , Inflamação/genética , Inflamação/imunologia , Macrófagos Peritoneais/citologia , Macrófagos Peritoneais/imunologia , Camundongos , Monócitos/citologia , Monócitos/imunologia , Fagocitose/efeitos dos fármacos , Zimosan/toxicidadeRESUMO
BACKGROUND & AIMS: Patients with decompensated cirrhosis are susceptible to infection. Innate immune dysfunction and development of organ failure are considered to underlie this. A rodent model of liver disease sharing these phenotypic features would assist in vivo study of underlying mechanisms and testing of therapeutics. We evaluated three models to identify which demonstrated the greatest clinical and immunological phenotypic similarity to patients with acutely decompensated (AD) cirrhosis. METHODS: We selected Bile Duct Ligation (BDL) rats at 4 weeks, BDL mice at 14 days and Carbon tetrachloride (CCl4 ) mice at 10 weeks (with studies performed 7 days after final CCl4 infection). We examined organ dysfunction, inflammatory response to carrageenan-in-paw, plasma eicosanoid concentrations, macrophage cytokine production and responses to peritoneal infection. RESULTS: Bile duct ligation caused sarcopenia, liver, cardiovascular and renal dysfunction whereas CCl4 mice demonstrated no clinical abnormalities. BDL rodents exhibited depressed response to carrageenan-in-paw unlike CCl4 mice. BDL rats have slightly elevated plasma eicosanoid levels and plasma showed partial PGE2 -mediated immune suppression whereas CCl4 mice did not. Plasma NOx was elevated in patients with acute or chronic liver failure (AoCLF) compared to healthy volunteers and BDL rodents but not CCl4 mice. Elevated nitric oxide (NO) via inducible nitric oxide synthase (iNOS) mediates defective leucocyte trafficking in BDL rodent models. CONCLUSIONS: We conclude that BDL mice and rats are not simply models of cholestatic liver injury but may be used to study mechanisms underlying poor outcome from infection in AD and have identified elevated NO as a potential mediator of depressed leucocyte trafficking.
Assuntos
Colestase/fisiopatologia , Fígado/fisiopatologia , Óxido Nítrico Sintase Tipo II/metabolismo , Óxido Nítrico/sangue , Animais , Ductos Biliares/cirurgia , Tetracloreto de Carbono , Modelos Animais de Doenças , Humanos , Ligadura , Cirrose Hepática/fisiopatologia , Macrófagos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Ratos , Ratos Sprague-DawleyRESUMO
Asthma and allergies are major health concerns in which Ig isotype E plays a pivotal role. Ag-bound IgE drives mast cells and basophils into exocytosis, thereby promoting allergic and potentially anaphylactic reactions. The importance of tightly regulated IgE production is underscored by severe immunological conditions in humans with elevated IgE levels. Cytokines direct IgH class-switching to a particular isotype by initiation of germline transcription (GLT) from isotype-specific intronic (I) promoters. The switch to IgE depends on IL-4, which stimulates GLT of the Iε promoter, but is specifically and strongly impaired in Swap-70(-/-) mice. Although early events in IL-4 signal transduction (i.e., activation of the JAK/STAT6 pathway) do not require SWAP-70, SWAP-70 deficiency results in impaired Iε GLT. The affinity of STAT6 to chromatin is reduced in absence of SWAP-70. Chromatin immunoprecipitation revealed that SWAP-70 binds to Iε and is required for association of STAT6 with Iε. BCL6, known to antagonize STAT6 particularly at Iε, is increased on Iε in absence of SWAP-70. Other promoters bound by BCL6 and STAT6 were found unaffected. We conclude that SWAP-70 controls IgE production through regulation of the antagonistic STAT6 and BCL6 occupancy of Iε. The identification of this mechanism opens new avenues to inhibit allergic reactions triggered by IgE.
Assuntos
Proteínas de Ligação a DNA/metabolismo , Fatores de Troca do Nucleotídeo Guanina/metabolismo , Switching de Imunoglobulina/imunologia , Imunoglobulina E/biossíntese , Proteínas Nucleares/metabolismo , Fator de Transcrição STAT6/metabolismo , Células 3T3 , Animais , Linfócitos B/imunologia , Células Cultivadas , Cromatina/metabolismo , Proteínas de Ligação a DNA/genética , Fatores de Troca do Nucleotídeo Guanina/genética , Hipersensibilidade/imunologia , Switching de Imunoglobulina/genética , Cadeias Pesadas de Imunoglobulinas/imunologia , Cadeias épsilon de Imunoglobulina/metabolismo , Interleucina-4/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Antígenos de Histocompatibilidade Menor , Proteínas Nucleares/genética , Regiões Promotoras Genéticas , Proteínas Proto-Oncogênicas c-bcl-6 , Transdução de Sinais/imunologia , Transcrição GênicaRESUMO
B-cell migration into and within lymphoid tissues is not only central to the humoral immune response but also for the development of malignancies and autoimmunity. We previously demonstrated that SWAP-70, an F-actin-binding, Rho GTPase-interacting protein strongly expressed in activated B cells, is necessary for normal B-cell migration in vivo. SWAP-70 regulates integrin-mediated adhesion and cell attachment. Here we show that upon B-cell activation, SWAP-70 is extensively posttranslationally modified and becomes tyrosine phosphorylated by SYK at position 517. This phosphorylation inhibits binding of SWAP-70 to F-actin. Phospho-site mutants of SWAP-70 disrupt B-cell polarization in a dominant-negative fashion in vitro and impair migration in vivo. After CXCL12 stimulation of B cells SYK becomes activated and SWAP-70 is phosphorylated in a SYK-dependent manner. Use of the highly specific SYK inhibitor BAY61-3606 showed SYK activity is necessary for normal chemotaxis and B-cell polarization in vitro and for entry of B cells into lymph nodes in vivo. These findings demonstrate a novel requirement for SYK in migration and polarization of naive recirculating B cells and show that SWAP-70 is an important target of SYK in this pathway.
Assuntos
Actinas/metabolismo , Linfócitos B/metabolismo , Movimento Celular , Proteínas de Ligação a DNA/fisiologia , Fatores de Troca do Nucleotídeo Guanina/fisiologia , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Proteínas Nucleares/fisiologia , Proteínas Tirosina Quinases/metabolismo , Tirosina/metabolismo , Animais , Linfócitos B/citologia , Western Blotting , Citometria de Fluxo , Imunoprecipitação , Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/isolamento & purificação , Camundongos , Antígenos de Histocompatibilidade Menor , Fosforilação , Proteínas Tirosina Quinases/genética , Proteínas Tirosina Quinases/isolamento & purificação , Quinase SykRESUMO
There is an increasing need to understand the leukocytes and soluble mediators that drive acute inflammation and bring about its resolution in humans. We therefore carried out an extensive characterisation of the cantharidin skin blister model in healthy male volunteers. A novel fluorescence staining protocol was designed and implemented, which facilitated the identification of cell populations by flow cytometry. We observed that at the onset phase, 24 h after blister formation, the predominant cells were CD16hi/CD66b+ PMNs followed by HLA-DR+/CD14+ monocytes/macrophages, CD11c+ and CD141+ dendritic cells as well as Siglec-8+ eosinophils. CD3+ T cells, CD19+ B cells and CD56+ NK cells were also present, but in comparatively fewer numbers. During resolution, 72 h following blister induction, numbers of PMNs declined whilst the numbers of monocyte/macrophages remain unchanged, though they upregulated expression of CD16 and CD163. In contrast, the overall numbers of dendritic cells and Siglec-8+ eosinophils increased. Post hoc analysis of these data revealed that of the inflammatory cytokines measured, TNF-α but not IL-1ß or IL-8 correlated with increased PMN numbers at the onset. Volunteers with the greatest PMN infiltration at onset displayed the fastest clearance rates for these cells at resolution. Collectively, these data provide insight into the cells that occupy acute resolving blister in humans, the soluble mediators that may control their influx as well as the phenotype of mononuclear phagocytes that predominate the resolution phase. Further use of this model will improve our understanding of the evolution and resolution of inflammation in humans, how defects in these over-lapping pathways may contribute to the variability in disease longevity/chronicity, and lends itself to the screen of putative anti-inflammatory or pro-resolution therapies.