RESUMO
OBJECTIVE: Inflammatory macrophages promote the development of atherosclerosis. We have identified the adaptor protein Dab2 (disabled homolog 2) as a regulator of phenotypic polarization in macrophages. The absence of Dab2 in myeloid cells promotes an inflammatory phenotype, but the impact of myeloid Dab2 deficiency on atherosclerosis has not been shown. APPROACH AND RESULTS: To determine the role of myeloid Dab2 in atherosclerosis, Ldlr-/- mice were reconstituted with either Dab2-positive or Dab2-deficient bone marrow and fed a western diet. Consistent with our previous finding that Dab2 inhibits NFκB (nuclear factor κ-light-chain-enhancer of activated B cells) signaling in macrophages, Ldlr-/- mice reconstituted with Dab2-deficient bone marrow had increased systemic inflammation as evidenced by increased serum IL-6 (interleukin-6) levels and increased inflammatory cytokine expression levels in liver. Serum lipid levels were significantly lower in Ldlr-/- mice reconstituted with Dab2-deficient bone marrow, and further examination of livers from these mice revealed drastically increased inflammatory tissue damage and massive infiltration of immune cells. Surprisingly, the atherosclerotic lesion burden in Ldlr-/- mice reconstituted with Dab2-deficient bone marrow was decreased compared with Ldlr-/- mice reconstituted with wild-type bone marrow. Further analysis of aortic root sections revealed increased macrophage content and evidence of increased apoptosis in lesions from Ldlr-/- mice reconstituted with Dab2-deficient bone marrow but no difference in collagen or α-smooth muscle actin content. CONCLUSIONS: Dab2 deficiency in myeloid cells promotes inflammation in livers and atherosclerotic plaques in a mouse model of atherosclerosis. Nevertheless, decreased serum lipids as a result of massive inflammatory liver damage may preclude an appreciable increase in atherosclerotic lesion burden in mice reconstituted with Dab2-deficient bone marrow.
Assuntos
Proteínas Adaptadoras de Transporte Vesicular/deficiência , Aorta/metabolismo , Doenças da Aorta/metabolismo , Aterosclerose/metabolismo , Hepatite/metabolismo , Fígado/metabolismo , Macrófagos/metabolismo , Placa Aterosclerótica , Receptores de LDL/deficiência , Proteínas Adaptadoras de Transdução de Sinal , Proteínas Adaptadoras de Transporte Vesicular/genética , Animais , Aorta/patologia , Doenças da Aorta/genética , Doenças da Aorta/patologia , Proteínas Reguladoras de Apoptose , Aterosclerose/genética , Aterosclerose/patologia , Caspases/metabolismo , Dieta Hiperlipídica , Modelos Animais de Doenças , Progressão da Doença , Feminino , Hepatite/genética , Hepatite/patologia , Humanos , Interleucina-6/sangue , Células Jurkat , Lipídeos/sangue , Fígado/patologia , Macrófagos/patologia , Camundongos Endogâmicos C57BL , NF-kappa B/metabolismo , Fenótipo , Receptores de LDL/genética , Transdução de Sinais , Triglicerídeos/metabolismoRESUMO
Toll-like receptors (TLRs) sense pathogen-associated molecular patterns to activate the production of inflammatory mediators. TLR4 recognizes lipopolysaccharide (LPS) and drives the secretion of inflammatory cytokines, often contributing to sepsis. We report that transient receptor potential melastatin-like 7 (TRPM7), a non-selective but Ca2+-conducting ion channel, mediates the cytosolic Ca2+ elevations essential for LPS-induced macrophage activation. LPS triggered TRPM7-dependent Ca2+ elevations essential for TLR4 endocytosis and the subsequent activation of the transcription factor IRF3. In a parallel pathway, the Ca2+ signaling initiated by TRPM7 was also essential for the nuclear translocation of NFκB. Consequently, TRPM7-deficient macrophages exhibited major deficits in the LPS-induced transcriptional programs in that they failed to produce IL-1ß and other key pro-inflammatory cytokines. In accord with these defects, mice with myeloid-specific deletion of Trpm7 are protected from LPS-induced peritonitis. Our study highlights the importance of Ca2+ signaling in macrophage activation and identifies the ion channel TRPM7 as a central component of TLR4 signaling.
Assuntos
Cálcio/metabolismo , Ativação de Macrófagos/efeitos dos fármacos , Canais de Cátion TRPM/metabolismo , Receptor 4 Toll-Like/metabolismo , Animais , Técnicas de Cultura de Células , Endocitose/efeitos dos fármacos , Feminino , Citometria de Fluxo , Imunofluorescência , Regulação da Expressão Gênica , Técnicas de Genotipagem , Immunoblotting , Fator Regulador 3 de Interferon/metabolismo , Lipopolissacarídeos/farmacologia , Macrófagos/metabolismo , Masculino , Camundongos , NF-kappa B/metabolismo , Técnicas de Patch-Clamp , Reação em Cadeia da Polimerase em Tempo Real , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia , Canais de Cátion TRPM/genéticaRESUMO
Acute and chronic tissue injury results in the generation of a myriad of environmental cues that macrophages respond to by changing their phenotype and function. This phenotypic regulation is critical for controlling tissue inflammation and resolution. Here, we have identified the adaptor protein disabled homolog 2 (DAB2) as a regulator of phenotypic switching in macrophages. Dab2 expression was upregulated in M2 macrophages and suppressed in M1 macrophages isolated from both mice and humans, and genetic deletion of Dab2 predisposed macrophages to adopt a proinflammatory M1 phenotype. In mice with myeloid cell-specific deletion of Dab2 (Dab2fl/fl Lysm-Cre), treatment with sublethal doses of LPS resulted in increased proinflammatory gene expression and macrophage activation. Moreover, chronic high-fat feeding exacerbated adipose tissue inflammation, M1 polarization of adipose tissue macrophages, and the development of insulin resistance in DAB2-deficient animals compared with controls. Mutational analyses revealed that DAB2 interacts with TNF receptor-associated factor 6 (TRAF6) and attenuates IκB kinase ß-dependent (IKKß-dependent) phosphorylation of Ser536 in the transactivation domain of NF-κB p65. Together, these findings reveal that DAB2 is critical for controlling inflammatory signaling during phenotypic polarization of macrophages and suggest that manipulation of DAB2 expression and function may hold therapeutic potential for the treatment of acute and chronic inflammatory disorders.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/biossíntese , Proteínas Adaptadoras de Transporte Vesicular/biossíntese , Tecido Adiposo/metabolismo , Regulação da Expressão Gênica , Macrófagos/metabolismo , Paniculite/metabolismo , Proteínas Supressoras de Tumor/biossíntese , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transporte Vesicular/genética , Tecido Adiposo/patologia , Animais , Proteínas Reguladoras de Apoptose , Linhagem Celular , Células HEK293 , Humanos , Quinase I-kappa B/genética , Quinase I-kappa B/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Macrófagos/patologia , Camundongos , Camundongos Knockout , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Paniculite/genética , Paniculite/patologia , Fator de Transcrição RelA/genética , Fator de Transcrição RelA/metabolismo , Proteínas Supressoras de Tumor/genéticaRESUMO
Extracellular ATP is an important signaling molecule throughout the inflammatory cascade, serving as a danger signal that causes activation of the inflammasome, enhancement of immune cell infiltration, and fine-tuning of several signaling cascades including those important for the resolution of inflammation. Recent studies demonstrated that ATP can be released from cells in a controlled manner through pannexin (Panx) channels. Panx1-mediated ATP release is involved in inflammasome activation and neutrophil/macrophage chemotaxis, activation of T cells, and a role for Panx1 in inducing and propagating inflammation has been demonstrated in various organs, including lung and the central and peripheral nervous system. The recognition and clearance of dying cells and debris from focal points of inflammation is critical in the resolution of inflammation, and Panx1-mediated ATP release from dying cells has been shown to recruit phagocytes. Moreover, extracellular ATP can be broken down by ectonucleotidases into ADP, AMP, and adenosine, which is critical in the resolution of inflammation. Together, Panx1, ATP, purinergic receptors, and ectonucleotidases contribute to important feedback loops during the inflammatory response, and thus represent promising candidates for new therapies.