RESUMO
Despite much progress in improving graft outcome during cardiac transplantation, chronic allograft vasculopathy (CAV) remains an impediment to long-term graft survival. MicroRNAs (miRNAs) emerged as regulators of the immune response. Here, we aimed to examine the miRNA network involved in CAV. miRNA profiling of heart samples obtained from a murine model of CAV and from cardiac-transplanted patients with CAV demonstrated that miR-21 was most significantly expressed and was primarily localized to macrophages. Interestingly, macrophage depletion with clodronate did not significantly prolong allograft survival in mice, while conditional deletion of miR-21 in macrophages or the use of a specific miR-21 antagomir resulted in indefinite cardiac allograft survival and abrogated CAV. The immunophenotype, secretome, ability to phagocytose, migration, and antigen presentation of macrophages were unaffected by miR-21 targeting, while macrophage metabolism was reprogrammed, with a shift toward oxidative phosphorylation in naïve macrophages and with an inhibition of glycolysis in pro-inflammatory macrophages. The aforementioned effects resulted in an increase in M2-like macrophages, which could be reverted by the addition of L-arginine. RNA-seq analysis confirmed alterations in arginase-associated pathways associated with miR-21 antagonism. In conclusion, miR-21 is overexpressed in murine and human CAV, and its targeting delays CAV onset by reprogramming macrophages metabolism.
Assuntos
Transplante de Coração , MicroRNAs , Aloenxertos , Animais , Rejeição de Enxerto/genética , Rejeição de Enxerto/prevenção & controle , Transplante de Coração/efeitos adversos , Humanos , Macrófagos , Camundongos , MicroRNAs/genéticaRESUMO
BACKGROUND: IgG4-related disease (IgG4-RD) is a fibroinflammatory condition marked by rapid clinical improvement after selective depletion of B lymphocytes with rituximab. This feature suggests that B cells might participate in fibrogenesis and wound healing. OBJECTIVE: In the present work we aimed to demonstrate that B lymphocytes contribute directly to tissue fibrosis in patients with IgG4-RD. METHODS: Total circulating CD19+ B lymphocytes, naive B cells, memory B cells, or plasmablasts from patients with IgG4-RD were cultivated with human fibroblasts. Profibrotic soluble factors and collagen production in cocultures were assessed by using ELISAs and Luminex assays. RNA sequencing and quantitative RT-PCR were used to assess fibroblast activation in the presence of B cells, as well as induction of profibrotic pathways in B-cell subsets. Relevant profibrotic and inflammatory molecules were confirmed in vitro by using functional experiments and on IgG4-RD tissue sections by using multicolor immunofluorescence studies. RESULTS: B cells from patients with IgG4-RD (1) produced the profibrotic molecule platelet-derived growth factor B and stimulated collagen production by fibroblasts; (2) expressed enzymes implicated in extracellular matrix remodeling, such as lysyl oxidase homolog 2; (3) produced the chemotactic factors CCL4, CCL5, and CCL11; and (4) induced production of these same chemokines by activated fibroblasts. Plasmablasts expressed sets of genes implicated in fibroblast activation and proliferation and therefore represent cells with intrinsic profibrotic properties. CONCLUSION: We have demonstrated that B cells contribute directly to tissue fibrosis in patients with IgG4-RD. These unanticipated profibrotic properties of B lymphocytes, particularly plasmablasts, might be relevant for fibrogenesis in patients with other fibroinflammatory disorders and for wound-healing processes in physiologic conditions.
Assuntos
Linfócitos B/patologia , Fibroblastos/patologia , Doença Relacionada a Imunoglobulina G4/patologia , Pâncreas/patologia , Linfócitos B/imunologia , Células Cultivadas , Técnicas de Cocultura , Colágeno/biossíntese , Fibrose/imunologia , Fibrose/patologia , Humanos , Doença Relacionada a Imunoglobulina G4/imunologiaRESUMO
OBJECTIVE: The signals causing the resolution of muscle inflammation are only partially characterized. The long pentraxin PTX3, which modulates leukocyte recruitment and activation, could contribute. METHODS: We analysed the expression of ptx3 after muscle injury and verified whether hematopoietic precursors are a source of the protein. The kinetics of regeneration and leukocytes infiltration, the accumulation of cell remnants and anti-histidyl-t-RNA synthetase autoantibodies were compared in wild-type and ptx3-deficient mice. RESULTS: Ptx3 expression was up-regulated three-five days after injury and restricted to the extracellular matrix. Cellular debris and leukocytes persisted in the muscle of ptx3-deficient mice for a long time after wild-type animals had healed. ptx3-deficient macrophages expressed receptors involved in apoptotic cell clearance and engulfed dead cells in vitro. Accumulation of cell debris in a pro-inflammatory microenvironment was not sufficient to elicit autoantibodies. CONCLUSION: PTX3 generated in response to muscle injury prompts the clearance of debris and the termination of the inflammatory response.
RESUMO
Signals of tissue necrosis, damage-associated molecular patterns (DAMPs), cause inflammation. Leukocytes migrating into injured tissues tonically release DAMPs, including the high mobility group box 1 protein (HMGB1). In the absence of suitable models, the relative role of DAMPs released because of necrosis or leukocyte activation has not, so far, been dissected. We have generated a mouse model lacking Hmgb1 in the hematopoietic system and studied the response to acute sterile injury of the skeletal muscle. Regenerating fibers are significantly less numerous at earlier time points and smaller at the end of the process. Leukocyte Hmgb1 licenses the skeletal muscle to react to hypoxia, to express angiopoietin-2, and to initiate angiogenesis in response to injury. Vascularization of the regenerating tissue is selectively jeopardized in the absence of leukocyte Hmgb1, revealing that it controls the nutrient and oxygen supply to the regenerating tissue. Altogether, our results reveal a novel nonredundant role for leukocyte Hmgb1 in the repair of injured skeletal muscle.
Assuntos
Angiopoietina-2/imunologia , Proteína HMGB1/imunologia , Músculo Esquelético/irrigação sanguínea , Músculo Esquelético/imunologia , Neovascularização Fisiológica/imunologia , Regeneração/imunologia , Angiopoietina-2/genética , Animais , Proteína HMGB1/genética , Leucócitos/imunologia , Camundongos , Camundongos Knockout , Músculo Esquelético/lesões , Neovascularização Fisiológica/genética , Regeneração/genéticaRESUMO
Adult skeletal muscle regeneration results from activation, proliferation, and fusion of muscle stem cells, such as myogenic precursor cells. Macrophages are consistently present in regenerating skeletal muscles and participate into the repair process. The signals involved in the cross-talk between various macrophage populations and myogenic precursor cells have been only partially identified. In this study, we show a key role of inducible NO synthase (iNOS), expressed by classically activated macrophages in the healing of skeletal muscle. We found that, after sterile injury, iNOS expression is required for effective regeneration of the tissue, as myogenic precursor cells in the muscle of injured iNOS(-/-) mice fail to proliferate and differentiate. We also found that iNOS modulates inflammatory cell recruitment: damaged muscles of iNOS(-/-) animals express significantly higher levels of chemokines such as MIP2, MCP1, MIP-1α, and MCP1, and display more infiltrating neutrophils after injury and a persistence of macrophages at later time points. Finally, we found that iNOS expression in the injured muscle is restricted to infiltrating macrophages. To our knowledge, these data thus provide the first evidence that iNOS expression by infiltrating macrophages contributes to muscle regeneration, revealing a novel mechanism of inflammation-dependent muscle healing.
Assuntos
Músculo Esquelético/enzimologia , Músculo Esquelético/lesões , Doenças Musculares/enzimologia , Doenças Musculares/imunologia , Óxido Nítrico Sintase Tipo II/fisiologia , Doença Aguda , Animais , Animais Recém-Nascidos , Movimento Celular/genética , Movimento Celular/imunologia , Modelos Animais de Doenças , Inflamação/enzimologia , Inflamação/imunologia , Inflamação/patologia , Macrófagos/enzimologia , Macrófagos/imunologia , Macrófagos/patologia , Camundongos , Camundongos da Linhagem 129 , Camundongos Endogâmicos C57BL , Camundongos Knockout , Músculo Esquelético/imunologia , Doenças Musculares/patologia , Óxido Nítrico Sintase Tipo I/deficiência , Óxido Nítrico Sintase Tipo I/fisiologia , Óxido Nítrico Sintase Tipo II/deficiência , Óxido Nítrico Sintase Tipo II/genética , Regeneração/genética , Regeneração/imunologia , Cicatrização/genética , Cicatrização/imunologiaRESUMO
Many functional foods and dietary supplements have been reported to be beneficial for the management of dyslipidaemia, one of the major risk factors for CVD. Soluble fibres and legume proteins are known to be a safe and practical approach for cholesterol reduction. The present study aimed at investigating the hypocholesterolaemic effect of the combinations of these bioactive vegetable ingredients and their possible effects on the expression of genes regulating cholesterol homeostasis. A total of six groups of twelve rats each were fed, for 28 d, Nath's hypercholesterolaemic diets, differing in protein and fibre sources, being, respectively, casein and cellulose (control), pea proteins and cellulose (pea), casein and oat fibres (oat), casein and apple pectin (pectin), pea proteins and oat fibres (pea+oat) and pea proteins and apple pectin (pea+pectin). Administration of each vegetable-containing diet was associated with lower total cholesterol concentrations compared with the control. The combinations (pea+oat and pea+pectin) were more efficacious than fibres alone in modulating cholesterolaemia ( - 53 and - 54%, respectively, at 28 d; P< 0·005). In rats fed the diets containing oat fibres or apple pectin, alone or in combination with pea proteins, a lower hepatic cholesterol content (P< 0·005) and higher hepatic mRNA concentrations of CYP7A1 and NTCP were found when compared with the control rats (P< 0·05). In summary, the dietary combinations of pea proteins and oat fibres or apple pectin are extremely effective in lowering plasma cholesterol concentrations in rats and affect cellular cholesterol homeostasis by up-regulating genes involved in hepatic cholesterol turnover.
Assuntos
Colesterol/metabolismo , Fibras na Dieta/metabolismo , Dislipidemias/dietoterapia , Dislipidemias/metabolismo , Pisum sativum/química , Proteínas de Plantas/metabolismo , Animais , Avena/química , Ácidos e Sais Biliares/metabolismo , Doenças Cardiovasculares/prevenção & controle , Caseínas/uso terapêutico , Celulose/uso terapêutico , Homeostase , Fígado/metabolismo , Masculino , Malus/química , Pectinas/uso terapêutico , Ratos , Ratos Sprague-Dawley , Fatores de Risco , Fatores de TempoRESUMO
Th1 cytokines promote monocyte differentiation into proatherogenic M1 macrophages, while Th2 cytokines lead to an "alternative" anti-inflammatory M2 macrophage phenotype. Here we show that in human atherosclerotic lesions, the expression of M2 markers and PPARgamma, a nuclear receptor controlling macrophage inflammation, correlate positively. Moreover, PPARgamma activation primes primary human monocytes into M2 differentiation, resulting in a more pronounced anti-inflammatory activity in M1 macrophages. However, PPARgamma activation does not influence M2 marker expression in resting or M1 macrophages, nor does PPARgamma agonist treatment influence the expression of M2 markers in atherosclerotic lesions, indicating that only native monocytes can be primed by PPARgamma activation to an enhanced M2 phenotype. Furthermore, PPARgamma activation significantly increases expression of the M2 marker MR in circulating peripheral blood mononuclear cells. These data demonstrate that PPARgamma activation skews human monocytes toward an anti-inflammatory M2 phenotype.
Assuntos
Inflamação/metabolismo , Inflamação/patologia , Macrófagos/citologia , Macrófagos/metabolismo , Monócitos/citologia , PPAR gama/metabolismo , Benzofenonas/farmacologia , Biomarcadores , Células Sanguíneas/efeitos dos fármacos , Doenças das Artérias Carótidas/patologia , Diferenciação Celular/efeitos dos fármacos , Células Cultivadas , Células Espumosas/efeitos dos fármacos , Células Espumosas/patologia , Humanos , Macrófagos/efeitos dos fármacos , Monócitos/efeitos dos fármacos , Monócitos/metabolismo , PPAR gama/agonistas , Comunicação Parácrina/efeitos dos fármacos , Fenótipo , Células-Tronco/efeitos dos fármacos , Tirosina/análogos & derivados , Tirosina/farmacologiaRESUMO
Adipose tissue macrophages (ATM) are the major source of visfatin, a visceral fat adipokine upregulated during obesity. Also known to play a role in B cell differentiation (pre-B cell colony-enhancing factor (PBEF)) and NAD biosynthesis (nicotinamide phosphoribosyl transferase (NAMPT)), visfatin has been suggested to play a role in inflammation. Liver X Receptor (LXR) and Peroxisome Proliferator-Activated Receptor (PPAR)γ are nuclear receptors expressed in macrophages controlling the inflammatory response. Recently, we reported visfatin as a PPARγ target gene in human macrophages. In this study, we examined whether LXR regulates macrophage visfatin expression. Synthetic LXR ligands decreased visfatin gene expression in a LXR-dependent manner in human and murine macrophages. The decrease of visfatin mRNA was paralleled by a decrease of protein secretion. Consequently, a modest and transient decrease of NAD(+) concentration was observed. Interestingly, LXR activation decreased the PPARγ-induced visfatin gene and protein secretion in human macrophages. Our results identify visfatin as a gene oppositely regulated by the LXR and PPARγ pathways in human macrophages.
Assuntos
Citocinas/antagonistas & inibidores , Citocinas/genética , Regulação Enzimológica da Expressão Gênica , Macrófagos/enzimologia , Nicotinamida Fosforribosiltransferase/antagonistas & inibidores , Nicotinamida Fosforribosiltransferase/genética , Receptores Nucleares Órfãos/metabolismo , Células Cultivadas , Citocinas/metabolismo , Citocinas/farmacologia , Regulação para Baixo , Expressão Gênica , Humanos , Receptores X do Fígado , NAD/metabolismo , Nicotinamida Fosforribosiltransferase/metabolismo , Receptores Nucleares Órfãos/agonistas , Receptores Nucleares Órfãos/genética , PPAR gama/metabolismoRESUMO
Pancreatic ductal adenocarcinoma (PDAC) is characterized by a prominent stromal reaction that has been variably implicated in both tumor growth and tumor suppression. B-lymphocytes have been recently implicated in PDAC progression but their contribution to the characteristic stromal desmoplasia has never been assessed before. In the present work, we aimed to verify whether B-lymphocytes contribute to stromal cell activation in PDAC. CD19+ B-lymphocytes purified from peripheral blood of patients with PDAC were cultivated in the presence of human pancreatic fibroblasts and cancer-associated fibroblasts. Released pro-fibrotic soluble factors and collagen production were assessed by ELISA and Luminex assays. Quantitative RT-PCR was used to assess fibroblast activation in the presence of B cells. The expression of selected pro-fibrotic and inflammatory molecules was confirmed on PDAC tissue sections by multi-color immunofluorescence studies. We herein demonstrate that B-cells from PDAC patients (i) produce the pro-fibrotic molecule PDGF-B and stimulate collagen production by fibroblasts; (ii) express enzymes implicated in extracellular matrix remodeling including LOXL2; and (iii) produce the chemotactic factors CCL-4, CCL-5, and CCL-11. In addition we demonstrate that circulating plasmablasts are expanded in the peripheral blood of patients with PDAC, stimulate collagen production by fibroblasts, and infiltrate pancreatic lesions. Our results indicate that PDAC is characterized by perturbations of the B-cell compartment with expansion of B-lymphocyte subsets that directly contribute to the stromal reaction observed at disease site. These findings provide an additional rationale for modulating B-cell activity in patients with pancreatic cancer.
Assuntos
Carcinoma Ductal Pancreático , Neoplasias Pancreáticas , Linfócitos B , Humanos , Pâncreas , Células EstromaisRESUMO
Macrophages play a central role in host defense against pathogen microbes by recognizing bacterial components, resulting in the activation of an arsenal of anti-microbial effectors. Toll-like receptor (TLR)-4 mediates the recognition of lipopolysaccharide, a pathogen-associated molecular pattern from gram-negative bacteria. Activation of the TLR-4 signaling pathway by lipopolysaccharide increases antibacterial effects by inducing secretion of cytokines that activate an immune inflammatory response and by generating bactericidal reactive oxygen species via the NADPH oxidase system. Liver X Receptors (LXRs) are nuclear receptors controlling cholesterol homeostasis and inflammation in macrophages. In addition, LXRs are critical for macrophage survival and play a role in the innate immune response in the mouse. In this study, we investigated whether LXR activation also regulates host defense mechanisms in human macrophages. In primary human macrophages, oxidized LDL and synthetic LXR ligands increased TLR-4 gene expression. Transient transfection assays, gel shift and chromatin immunoprecipitation analysis indicated that LXRs induce human TLR-4 promoter activity by binding to a DR4-type LXR response element. LXR induction of TLR-4 mRNA was followed by an induction of TLR-4 protein expression. Moreover, although short-term pretreatment with LXR agonists significantly reduced the inflammatory response induced by lipopolysaccharide, pretreatment of macrophages for 48 hours with LXR agonists resulted in an enhanced lipopolysaccharide response. Finally, LXR activation increased reactive oxygen species generation by enhancing the expression of NADPH oxidase subunits. These data provide evidence for an immunomodulatory function of LXRs in human macrophages via mechanisms distinct from those previously identified in mouse macrophages.
Assuntos
Proteínas de Ligação a DNA/metabolismo , Lipopolissacarídeos/farmacologia , Macrófagos/metabolismo , Receptores Citoplasmáticos e Nucleares/metabolismo , Animais , Células Cultivadas , Proteínas de Ligação a DNA/agonistas , Proteínas de Ligação a DNA/genética , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/imunologia , Humanos , Mediadores da Inflamação/agonistas , Mediadores da Inflamação/metabolismo , Mediadores da Inflamação/fisiologia , Lipopolissacarídeos/antagonistas & inibidores , Receptores X do Fígado , Macrófagos/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Receptores Nucleares Órfãos , Receptores Citoplasmáticos e Nucleares/agonistas , Receptores Citoplasmáticos e Nucleares/genética , Receptor 4 Toll-Like/biossíntese , Receptor 4 Toll-Like/genéticaRESUMO
Peroxisome proliferator-activated receptors (PPARs) and (liver X receptors) LXRs are ligand-activated transcription factors that control lipid and glucose metabolism, as well as the inflammatory response. Because the macrophage plays an important role in host defense and immunoinflammatory pathologies, particular attention has been paid to the role of PPARs and LXRs in the control of macrophage gene expression and function. Research over the last few years has revealed important roles for PPAR-alpha, PPAR-gamma, and LXRs in macrophage inflammation and cholesterol homeostasis with consequences for atherosclerosis development. In this review we will discuss the role of these transcription factors in the control of macrophage activities, with particular attention to species-differences in macrophage function control by PPARs and LXR between rodents and humans.
Assuntos
Proteínas de Ligação a DNA/fisiologia , Macrófagos/fisiologia , PPAR alfa/fisiologia , PPAR gama/fisiologia , Receptores Citoplasmáticos e Nucleares/fisiologia , Animais , Hemostasia/fisiologia , Humanos , Receptores X do Fígado , Camundongos , Modelos Animais , Receptores Nucleares Órfãos , Especificidade da EspécieRESUMO
OBJECTIVE: Liver X receptors (LXRs) are oxysterol-activated nuclear receptors regulating reverse cholesterol transport, in part by modulating cholesterol efflux from macrophages to apoAI and HDL via the ABCA1 and ABCG1/ABCG4 pathways. Moreover, LXR activation increases intracellular cholesterol trafficking via the induction of NPC1 and NPC2 expression. However, implication of LXRs in the selective uptake of cholesteryl esters from lipoproteins in human macrophages has never been reported. METHODS AND RESULTS: Our results show that (1) selective CE uptake from HDL(3) is highly efficient in human monocyte-derived macrophages; (2) surprisingly, HDL(3)-CE uptake is strongly increased by LXR activation despite antiatherogenic effects of LXRs; (3) HDL(3)-CE uptake increase is not linked to SR-BI expression modulation but it is dependent of proteoglycan interactions; (4) HDL(3)-CE uptake increase is associated with increased expression and secretion of apoE and LPL, two proteins interacting with proteoglycans; (5) HDL(3)-CE uptake increase depends on the integrity of raft domains and is associated with an increased caveolin-1 expression. CONCLUSIONS: Our study identifies a new role for LXRs in the control of cholesterol homeostasis in human macrophages. LXR activation results in enhanced dynamic intracellular cholesterol fluxes through an increased CE uptake from HDL and leads to an increased cholesterol availability to efflux to apoAI and HDL.
Assuntos
Ésteres do Colesterol/metabolismo , Proteínas de Ligação a DNA/metabolismo , Lipoproteínas HDL/metabolismo , Macrófagos/metabolismo , Receptores Citoplasmáticos e Nucleares/metabolismo , Apolipoproteína A-I/metabolismo , Aterosclerose/etiologia , Aterosclerose/metabolismo , Transporte Biológico Ativo , Caveolina 1/genética , Caveolina 1/metabolismo , Células Cultivadas , HDL-Colesterol/metabolismo , Proteínas de Ligação a DNA/agonistas , Humanos , Hidrocarbonetos Fluorados/farmacologia , Receptores X do Fígado , Macrófagos/citologia , Macrófagos/efeitos dos fármacos , Microdomínios da Membrana/metabolismo , Receptores Nucleares Órfãos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptores Citoplasmáticos e Nucleares/agonistas , Receptores Depuradores Classe B/genética , Receptores Depuradores Classe B/metabolismo , Sulfonamidas/farmacologiaRESUMO
OBJECTIVE: Macrophages play a central role in the immune response against infectious organisms. Once activated, macrophages secrete proinflammatory cytokines and chemokines. Interleukin (IL)-8 and related CXC chemokines play a role in the recruitment and activation of phagocytes acting through CXCR1 and CXCR2 receptors. The nuclear receptor peroxisome proliferator-activated receptor (PPAR) gamma exerts antiinflammatory properties in macrophages, by inhibiting cytokine and CC chemokine production. In this study, we investigated whether PPAR-gamma also plays a role in the regulation of the CXC chemokine pathway. METHODS AND RESULTS: Synthetic PPAR-gamma ligands increase CXCR2 but not CXCR1 gene expression in a PPAR-gamma-dependent manner in primary human macrophages in vitro and in atherosclerotic plaques in vivo. The increase of CXCR2 mRNA was paralleled by an increase in membrane protein expression. EMSA, ChIP, and transient transfection assays indicate that PPAR-gamma activates the CXCR2 promoter by binding to a PPAR response element (PPRE). Finally, human macrophages acquire responsiveness to the CXCR2 ligands (IL-8 and Grobeta), as measured by superoxide anion production, after induction of CXCR2 expression by PPAR-gamma ligands. CONCLUSIONS: Our results provide a novel mechanism via which PPAR-gamma can enhance the immune response in human macrophages.
Assuntos
Macrófagos/metabolismo , PPAR gama/metabolismo , Receptores de Interleucina-8B/metabolismo , Animais , Aterosclerose/metabolismo , Aterosclerose/patologia , Células COS , Quimiocina CXCL2/farmacologia , Chlorocebus aethiops , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Interleucina-8/farmacologia , Macrófagos/efeitos dos fármacos , Macrófagos/patologia , RNA Mensageiro/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Receptores de Interleucina-8B/genética , Transdução de Sinais , Superóxidos/metabolismoRESUMO
A role of the nuclear receptor Rev-erbalpha in the regulation of transcription pathways involving other nuclear receptors is emerging. Indeed, Rev-erbalpha is a negative regulator of transcription by binding to overlapping response elements shared with various nuclear receptors, including the peroxisome proliferator-activated receptors and the retinoid-related orphan receptor alpha (RORalpha). Here, we show that Rev-erbalpha is expressed in primary human macrophages and that its expression is induced by synthetic ligands for the liver X receptors (LXRs), which control cholesterol homeostasis, inflammation, and the immune response in macrophages. LXRalpha binds to a specific response element in the human Rev-erbalpha promoter, thus inducing Rev-erbalpha transcriptional expression. Interestingly, Rev-erbalpha does not influence basal or LXR-regulated cholesterol homeostasis. However, Rev-erbalpha overexpression represses the induction of toll-like receptor (TLR)-4 by LXR agonists, whereas Rev-erbalpha silencing by short interfering RNA results in enhanced TLR-4 expression upon LXR activation. Electrophoretic mobility shift, chromatin immunoprecipitation, and transient transfection experiments demonstrate that Rev-erbalpha represses human TLR-4 promoter activity by binding as a monomer to a RevRE site overlapping with the LXR response element site in the TLR-4 promoter. These data identify Rev-erbalpha as a new LXR target gene, inhibiting LXR-induction of TLR-4 in a negative transcriptional feedback loop, but not cholesterol homeostasis gene expression.
Assuntos
Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Retroalimentação Fisiológica , Macrófagos/metabolismo , Receptores Citoplasmáticos e Nucleares/genética , Receptores Citoplasmáticos e Nucleares/metabolismo , Sequência de Bases , Células Cultivadas , Colesterol/metabolismo , Dimerização , Retroalimentação Fisiológica/efeitos dos fármacos , Humanos , Lipopolissacarídeos/farmacologia , Receptores X do Fígado , Ativação de Macrófagos/efeitos dos fármacos , Macrófagos/efeitos dos fármacos , Modelos Genéticos , Dados de Sequência Molecular , Membro 1 do Grupo D da Subfamília 1 de Receptores Nucleares , Receptores Nucleares Órfãos , Ligação Proteica/efeitos dos fármacos , Proteínas Repressoras/metabolismo , Elementos de Resposta/genética , Receptor X Retinoide alfa/metabolismo , Receptor 4 Toll-Like/genética , Ativação Transcricional/efeitos dos fármacos , Fator de Necrose Tumoral alfa/metabolismoRESUMO
The biological activities of a protein isolate from lupin (Lupinus albus) were studied in a rabbit model of atherosclerosis. Focal plaque development was induced at both common carotid arteries by perivascular injury. After surgery, animals were fed three different diets for 90 d, all with 1% cholesterol, 15% SFA and 20% protein; the protein source was casein (CAS), lupin proteins (LUP) or 50% CAS+50% LUP (CAS+LUP). Lower cholesterolaemia was detected in the LUP v. the CAS group at 60 and 90 d of treatment (-40·3 and -33·5%, respectively; P<0·05). Cryosection analyses of the carotids indicated a significant reduction in focal lesion progression in the LUP v. the CAS group (-37·4%; P<0·05). In summary, in a rabbit model of atherosclerosis, a protein isolate from L. albus reduced cholesterolaemia and exerted a remarkable protective activity against atherosclerosis progression.
Assuntos
Aterosclerose/tratamento farmacológico , Hiperlipidemias/tratamento farmacológico , Lupinus/química , Proteínas de Plantas/uso terapêutico , Animais , Modelos Animais de Doenças , CoelhosRESUMO
OBJECTIVE: Toll-like receptor 7 (TLR-7), TLR-8, and interferon (IFN)-induced genes are expressed in patients with idiopathic inflammatory myositis. This study was undertaken to investigate whether their activation influences the natural history of the disease. METHODS: Experimental autoimmune myositis was induced in mice by injection of the amino-terminal portion of the murine histidyl-transfer RNA synthetase (HisRS). Disease was compared in the presence or the absence of the TLR-7/8 agonist R-848 in wild-type mice and in mice that fail to express the IFNα/ß receptor (IFNα/ßR-null mice). RESULTS: Experimental autoimmune myositis induced by a single intramuscular immunization with HisRS spontaneously abated after 7-8 weeks. In contrast, levels of anti-HisRS autoantibodies, endomysial/perimysial leukocyte infiltration, and myofiber regeneration persisted at the end of the follow-up period (22 weeks after immunization) in mice immunized with HisRS in the presence of R-848. Myofiber major histocompatibility complex (MHC) class I molecules were detectable only in mice immunized with both HisRS and R-848. MHC up-regulation occurred early and in muscles that were not directly injected with HisRS. Muscle MHC expression paralleled with leukocyte infiltration. MHC class I molecules were selectively up-regulated in myotubes challenged with R-848 in vitro. Type I IFN was necessary for the prolonged autoantibody response and for the spreading of the autoimmune response, as demonstrated using IFNα/ßR-null mice. Muscle infiltration was maintained in the injected muscle up to the end of the follow-up period. CONCLUSION: TLR-7/8 activation is necessary to induce and maintain a systemic autoimmune response targeting the skeletal muscle. This experimental autoimmune myositis model reproduces many characteristics of human idiopathic inflammatory myopathies and may represent a tool for preclinical studies.
Assuntos
Imidazóis/metabolismo , Miosite/imunologia , Doença Autoimune do Sistema Nervoso Experimental/imunologia , Receptor 7 Toll-Like/agonistas , Receptor 8 Toll-Like/agonistas , Animais , Autoanticorpos/sangue , Autoanticorpos/imunologia , Modelos Animais de Doenças , Progressão da Doença , Histidina-tRNA Ligase , Complexo Principal de Histocompatibilidade/imunologia , Camundongos , Músculo Esquelético/imunologia , Miosite/sangue , Miosite/induzido quimicamente , Doença Autoimune do Sistema Nervoso Experimental/sangue , Doença Autoimune do Sistema Nervoso Experimental/induzido quimicamenteRESUMO
Muscle injury induces a classical inflammatory response in which cells of the innate immune system rapidly invade the tissue. Macrophages are prominently involved in this response and required for proper healing, as they are known to be important for clearing cellular debris and supporting satellite cell differentiation. Here, we sought to assess the role of the adaptive immune system in muscle regeneration after acute damage. We show that T lymphocytes are transiently recruited into the muscle after damage and appear to exert a pro-myogenic effect on muscle repair. We observed a decrease in the cross-sectional area of regenerating myofibers after injury in Rag2-/- γ-chain-/- mice, as compared to WT controls, suggesting that T cell recruitment promotes muscle regeneration. Skeletal muscle infiltrating T lymphocytes were enriched in CD4+CD25+FOXP3+ cells. Direct exposure of muscle satellite cells to in vitro induced Treg cells effectively enhanced their expansion, and concurrently inhibited their myogenic differentiation. In vivo, the recruitment of Tregs to acutely injured muscle was limited to the time period of satellite expansion, with possibly important implications for situations in which inflammatory conditions persist, such as muscular dystrophies and inflammatory myopathies. We conclude that the adaptive immune system, in particular T regulatory cells, is critically involved in effective skeletal muscle regeneration. Thus, in addition to their well-established role as regulators of the immune/inflammatory response, T regulatory cells also regulate the activity of skeletal muscle precursor cells, and are instrumental for the proper regeneration of this tissue.
Assuntos
Linhagem da Célula , Fatores de Transcrição Forkhead/metabolismo , Músculo Esquelético/lesões , Músculo Esquelético/patologia , Regeneração , Células Satélites de Músculo Esquelético/patologia , Linfócitos T/imunologia , Animais , Antígenos CD/metabolismo , Diferenciação Celular , Proliferação de Células , Camundongos Endogâmicos C57BL , Linfócitos T Reguladores/imunologiaRESUMO
Macrophages are one of the first barriers of host defence against pathogens. Beyond their role in innate immunity, macrophages play increasingly defined roles in orchestrating the healing of various injured tissues. Perturbations of macrophage function and/or activation may result in impaired regeneration and fibrosis deposition as described in several chronic pathological diseases. Heterogeneity and plasticity have been demonstrated to be hallmarks of macrophages. In response to environmental cues they display a proinflammatory (M1) or an alternative anti-inflammatory (M2) phenotype. A lot of evidence demonstrated that after acute injury M1 macrophages infiltrate early to promote the clearance of necrotic debris, whereas M2 macrophages appear later to sustain tissue healing. Whether the sequential presence of two different macrophage populations results from a dynamic shift in macrophage polarization or from the recruitment of new circulating monocytes is a subject of ongoing debate. In this paper, we discuss the current available information about the role that different phenotypes of macrophages plays after injury and during the remodelling phase in different tissue types, with particular attention to the skeletal muscle.
Assuntos
Ativação de Macrófagos/imunologia , Macrófagos/imunologia , Músculo Esquelético/imunologia , Músculo Esquelético/lesões , Miosite/imunologia , Regeneração/imunologia , Cicatrização/imunologia , Adaptação Fisiológica/imunologia , Animais , Humanos , Modelos Imunológicos , Músculo Esquelético/patologiaRESUMO
In the absence of a clear indication from previous studies, a rat study was designed to evaluate a possible hypolipidaemic effect of Lupinus angustifolius (blue lupin) proteins. Rats were fed for 28days Nath's hypercholesterolaemic diets containing 20% casein or blue lupin proteins. After 14 and 28days of dietary treatment, blue-lupin-fed rats had markedly lower plasma total cholesterol levels than rats fed casein (-53.0% and -55.3%, respectively, p<0.0005). No significant differences were instead observed for triglyceride and HDL-cholesterol levels between the two groups. Lupin-protein-fed rats displayed higher hepatic mRNA levels of SREBP-2, a major transcriptional regulator of intracellular cholesterol levels, and CYP7A1, the rate-limiting enzyme in bile acid biosynthesis (p<0.05). In conclusion, the present study demonstrates a marked cholesterol-lowering activity of proteins from L. angustifolius in rats. Moreover, blue lupin proteins appear to affect cellular lipid homeostasis by up-regulating SREBP-2 and CYP7A1 genes.
RESUMO
Obesity is a low-grade chronic inflammatory disease associated with an increased number of macrophages (adipose tissue macrophages) in adipose tissue. Within the adipose tissue, adipose tissue macrophages are the major source of visfatin/pre-B-cell colony-enhancing factor/nicotinamide phosphoribosyl transferase. The nuclear receptor peroxisome proliferator-activated receptor gamma (PPARgamma) exerts anti-inflammatory effects in macrophages by inhibiting cytokine production and enhancing alternative differentiation. In this study, we investigated whether PPARgamma modulates visfatin expression in murine (bone marrow-derived macrophage) and human (primary human resting macrophage, classical macrophage, alternative macrophage or adipose tissue macrophage) macrophage models and pre-adipocyte-derived adipocytes. We show that synthetic PPARgamma ligands increase visfatin gene expression in a PPARgamma-dependent manner in primary human resting macrophages and in adipose tissue macrophages, but not in adipocytes. The threefold increase of visfatin mRNA was paralleled by an increase of protein expression (30%) and secretion (30%). Electrophoretic mobility shift assay experiments and transient transfection assays indicated that PPARgamma induces visfatin promoter activity in human macrophages by binding to a DR1-PPARgamma response element. Finally, we show that PPARgamma ligands increase NAD(+) production in primary human macrophages and that this regulation is dampened in the presence of visfatin small interfering RNA or by the visfatin-specific inhibitor FK866. Taken together, our results suggest that PPARgamma regulates the expression of visfatin in macrophages, leading to increased levels of NAD(+).