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1.
Proc Natl Acad Sci U S A ; 118(38)2021 09 21.
Artigo em Inglês | MEDLINE | ID: mdl-34479991

RESUMO

COVID-19 induces a robust, extended inflammatory "cytokine storm" that contributes to an increased morbidity and mortality, particularly in patients with type 2 diabetes (T2D). Macrophages are a key innate immune cell population responsible for the cytokine storm that has been shown, in T2D, to promote excess inflammation in response to infection. Using peripheral monocytes and sera from human patients with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), and a murine hepatitis coronavirus (MHV-A59) (an established murine model of SARS), we identified that coronavirus induces an increased Mφ-mediated inflammatory response due to a coronavirus-induced decrease in the histone methyltransferase, SETDB2. This decrease in SETDB2 upon coronavirus infection results in a decrease of the repressive trimethylation of histone 3 lysine 9 (H3K9me3) at NFkB binding sites on inflammatory gene promoters, effectively increasing inflammation. Mφs isolated from mice with a myeloid-specific deletion of SETDB2 displayed increased pathologic inflammation following coronavirus infection. Further, IFNß directly regulates SETDB2 in Mφs via JaK1/STAT3 signaling, as blockade of this pathway altered SETDB2 and the inflammatory response to coronavirus infection. Importantly, we also found that loss of SETDB2 mediates an increased inflammatory response in diabetic Mϕs in response to coronavirus infection. Treatment of coronavirus-infected diabetic Mφs with IFNß reversed the inflammatory cytokine production via up-regulation of SETDB2/H3K9me3 on inflammatory gene promoters. Together, these results describe a potential mechanism for the increased Mφ-mediated cytokine storm in patients with T2D in response to COVID-19 and suggest that therapeutic targeting of the IFNß/SETDB2 axis in T2D patients may decrease pathologic inflammation associated with COVID-19.


Assuntos
Coronavirus/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Histona-Lisina N-Metiltransferase/metabolismo , Mediadores da Inflamação/metabolismo , Inflamação/virologia , Macrófagos/metabolismo , Animais , COVID-19/imunologia , Infecções por Coronavirus/genética , Infecções por Coronavirus/imunologia , Síndrome da Liberação de Citocina , Citocinas/metabolismo , Diabetes Mellitus Tipo 2/genética , Feminino , Histona-Lisina N-Metiltransferase/genética , Humanos , Inflamação/metabolismo , Inflamação/fisiopatologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , NF-kappa B/metabolismo , SARS-CoV-2/metabolismo , Transdução de Sinais
2.
J Clin Tuberc Other Mycobact Dis ; 24: 100258, 2021 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-34307905

RESUMO

Objective: There is a clear need for improved biomarkers to diagnose HIV/TB coinfection. Although numerous tests can identify the existence of both of these microbes within the host, a parallel assessment of the host response to HIV/TB coinfection may prove as useful confirmation in cases where microbiological tests are inconclusive. To this end we assessed the levels of Notch ligands found in serum samples of patients with TB, HIV or HIV/TB coinfection. The Notch system is involved in almost every stage of development, including the maturation of the immune response. Upon exposure to a pathogen, the innate immune system will increase expression of Notch ligands Delta-like 1 and Delta-like 4. Previous research has demonstrated that Notch ligand expression is increased on monocytes from patients diagnosed with tuberculosis. We hypothesized that if Notch ligands were present in the peripheral blood of individuals diagnosed with TB, they may serve as a novel marker for infection.Design: Serum samples from patients with HIV, TB or HIV/TB coinfection were compared to serum from uninfected individuals to determine levels of DLL1 and DLL4 in a case controlled study. Methods: DLL1 and DLL4 were measured by ELISA. Linear regression with post tests were used to determine if levels of DLL1 and DLL4 were increased in individuals with HIV/TB coinfection as compared to individuals infected with either HIV or TB or healthy controls. Results: Delta-like 1 and Delta-like 4 were significantly increased in the serum of patients with HIV and HIV/ M. tuberculosis coinfection compared to other groups. Conclusions: Assessment of Notch ligands in peripheral blood may enhance the diagnosis of individuals with active TB that are co-infected with HIV. The study will also need to be validated in in a larger cohort.

3.
J Exp Med ; 218(6)2021 06 07.
Artigo em Inglês | MEDLINE | ID: mdl-33779682

RESUMO

Abdominal aortic aneurysms (AAAs) are a life-threatening disease for which there is a lack of effective therapy preventing aortic rupture. During AAA formation, pathological vascular remodeling is driven by macrophage infiltration, and the mechanisms regulating macrophage-mediated inflammation remain undefined. Recent evidence suggests that an epigenetic enzyme, JMJD3, plays a critical role in establishing macrophage phenotype. Using single-cell RNA sequencing of human AAA tissues, we identified increased JMJD3 in aortic monocyte/macrophages resulting in up-regulation of an inflammatory immune response. Mechanistically, we report that interferon-ß regulates Jmjd3 expression via JAK/STAT and that JMJD3 induces NF-κB-mediated inflammatory gene transcription in infiltrating aortic macrophages. In vivo targeted inhibition of JMJD3 with myeloid-specific genetic depletion (JMJD3f/fLyz2Cre+) or pharmacological inhibition in the elastase or angiotensin II-induced AAA model preserved the repressive H3K27me3 on inflammatory gene promoters and markedly reduced AAA expansion and attenuated macrophage-mediated inflammation. Together, our findings suggest that cell-specific pharmacologic therapy targeting JMJD3 may be an effective intervention for AAA expansion.


Assuntos
Aneurisma da Aorta Abdominal/metabolismo , Histona Desmetilases/metabolismo , Histona Desmetilases com o Domínio Jumonji/metabolismo , Macrófagos/metabolismo , Angiotensina II/farmacologia , Animais , Modelos Animais de Doenças , Inflamação/metabolismo , Mediadores da Inflamação/metabolismo , Macrófagos/efeitos dos fármacos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , NF-kappa B/metabolismo , Regulação para Cima/efeitos dos fármacos , Regulação para Cima/fisiologia
4.
J Biol Chem ; 296: 100235, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33376138

RESUMO

Epigenetic mechanisms that alter heritable gene expression and chromatin structure play an essential role in many biological processes, including liver function. Human MOF (males absent on the first) is a histone acetyltransferase that is globally downregulated in human steatohepatitis. However, the function of MOF in the liver remains unclear. Here, we report that MOF plays an essential role in adult liver. Genetic deletion of Mof by Mx1-Cre in the liver leads to acute liver injury, with increase of lipid deposition and fibrosis akin to human steatohepatitis. Surprisingly, hepatocyte-specific Mof deletion had no overt liver abnormality. Using the in vitro coculturing experiment, we show that Mof deletion-induced liver injury requires coordinated changes and reciprocal signaling between hepatocytes and Kupffer cells, which enables feedforward regulation to augment inflammation and apoptotic responses. At the molecular level, Mof deletion induced characteristic changes in metabolic gene programs, which bore noticeable similarity to the molecular signature of human steatohepatitis. Simultaneous deletion of Mof in both hepatocytes and macrophages results in enhanced expression of inflammatory genes and NO signaling in vitro. These changes, in turn, lead to apoptosis of hepatocytes and lipotoxicity. Our work highlights the importance of histone acetyltransferase MOF in maintaining metabolic liver homeostasis and sheds light on the epigenetic dysregulation in liver pathogenesis.


Assuntos
Histona Acetiltransferases/genética , Inflamação/metabolismo , Hepatopatias/genética , Fígado/lesões , Óxido Nítrico/genética , Apoptose/genética , Cromatina/genética , Epigênese Genética/genética , Fígado Gorduroso/genética , Fígado Gorduroso/metabolismo , Fígado Gorduroso/patologia , Deleção de Genes , Regulação da Expressão Gênica/genética , Hepatócitos/metabolismo , Hepatócitos/patologia , Histona Acetiltransferases/química , Humanos , Inflamação/genética , Inflamação/patologia , Lipídeos/efeitos adversos , Lipídeos/genética , Fígado/metabolismo , Fígado/patologia , Hepatopatias/metabolismo , Hepatopatias/patologia , Macrófagos/metabolismo , Macrófagos/patologia , Óxido Nítrico/metabolismo , Transdução de Sinais/genética
5.
JCI Insight ; 5(17)2020 09 03.
Artigo em Inglês | MEDLINE | ID: mdl-32879137

RESUMO

Macrophages are a primary immune cell involved in inflammation, and their cell plasticity allows for transition from an inflammatory to a reparative phenotype and is critical for normal tissue repair following injury. Evidence suggests that epigenetic alterations play a critical role in establishing macrophage phenotype and function during normal and pathologic wound repair. Here, we find in human and murine wound macrophages that cyclooxygenase 2/prostaglandin E2 (COX-2/PGE2) is elevated in diabetes and regulates downstream macrophage-mediated inflammation and host defense. Using single-cell RNA sequencing of human wound tissue, we identify increased NF-κB-mediated inflammation in diabetic wounds and show increased COX-2/PGE2 in diabetic macrophages. Further, we identify that COX-2/PGE2 production in wound macrophages requires epigenetic regulation of 2 key enzymes in the cytosolic phospholipase A2/COX-2/PGE2 (cPLA2/COX-2/PGE2) pathway. We demonstrate that TGF-ß-induced miRNA29b increases COX-2/PGE2 production via inhibition of DNA methyltransferase 3b-mediated hypermethylation of the Cox-2 promoter. Further, we find mixed-lineage leukemia 1 (MLL1) upregulates cPLA2 expression and drives COX-2/PGE2. Inhibition of the COX-2/PGE2 pathway genetically (Cox2fl/fl Lyz2Cre+) or with a macrophage-specific nanotherapy targeting COX-2 in tissue macrophages reverses the inflammatory macrophage phenotype and improves diabetic tissue repair. Our results indicate the epigenetically regulated PGE2 pathway controls wound macrophage function, and cell-targeted manipulation of this pathway is feasible to improve diabetic wound repair.


Assuntos
Diabetes Mellitus Experimental/fisiopatologia , Diabetes Mellitus/fisiopatologia , Dinoprostona/farmacologia , Epigênese Genética , Regulação da Expressão Gênica/efeitos dos fármacos , Inflamação/prevenção & controle , Macrófagos/efeitos dos fármacos , Cicatrização , Idoso , Animais , Ciclo-Oxigenase 2/metabolismo , Humanos , Inflamação/genética , Inflamação/imunologia , Inflamação/patologia , Macrófagos/metabolismo , Macrófagos/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , NF-kappa B/metabolismo , Ocitócicos/farmacologia , Fenótipo , Pseudomonas aeruginosa/efeitos dos fármacos , Transdução de Sinais
6.
Tuberculosis (Edinb) ; 124: 101980, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32801053

RESUMO

Tuberculosis (TB) is the leading cause of mortality among infectious diseases worldwide. The study of molecular targets for therapy and diagnosis suggested that Notch signaling is an important pathway for the maintenance of the immune response during Mycobacterium tuberculosis (Mtb) infection. We evaluated the participation of the Notch pathway in the modulation of immune response during Mtb infection, and observed that patients with active TB had increased DLL4 expression in intermediate and non-classic monocytes. Further, patients with moderate and advanced lung injury have higher Notch1 expression in CD4+ T cells when compared to patients with a minimal lung injury. When we considered the severity of disease in active TB patients, the expression of the DLL4 in intermediate monocytes and the expression of Notch1 in CD4+ T cells are positively correlated with the degree of lung injury. In vitro, PBMCs treated with the Notch pharmacological inhibitor reduced the production of IL-17A and IL-2, whereas anti-hDLL4 treatment promoted a significant increase in TNF-α and phagocytosis. We suggest that Notch1 and DLL4 are associated with immune response activation in human tuberculosis, and can be a novel target to be exploited in the future in the searching of biomarkers.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Linfócitos T CD4-Positivos/metabolismo , Proteínas de Ligação ao Cálcio/metabolismo , Pulmão/metabolismo , Mycobacterium tuberculosis/imunologia , Receptor Notch1/metabolismo , Tuberculose Pulmonar/metabolismo , Adulto , Biomarcadores/metabolismo , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/microbiologia , Estudos de Casos e Controles , Células Cultivadas , Citocinas/metabolismo , Progressão da Doença , Feminino , Interações Hospedeiro-Patógeno , Humanos , Pulmão/imunologia , Pulmão/microbiologia , Masculino , Pessoa de Meia-Idade , Fagocitose , Índice de Gravidade de Doença , Transdução de Sinais , Tuberculose Pulmonar/imunologia , Tuberculose Pulmonar/microbiologia , Adulto Jovem
7.
Eur J Immunol ; 50(12): 1929-1940, 2020 12.
Artigo em Inglês | MEDLINE | ID: mdl-32662520

RESUMO

Chronic macrophage inflammation is a hallmark of type 2 diabetes (T2D) and linked to the development of secondary diabetic complications. T2D is characterized by excess concentrations of saturated fatty acids (SFA) that activate innate immune inflammatory responses, however, mechanism(s) by which SFAs control inflammation is unknown. Using monocyte-macrophages isolated from human blood and murine models, we demonstrate that palmitate (C16:0), the most abundant circulating SFA in T2D, increases expression of the histone demethylase, Jmjd3. Upregulation of Jmjd3 results in removal of the repressive histone methylation (H3K27me3) mark on NFκB-mediated inflammatory gene promoters driving macrophage-mediated inflammation. We identify that the effects of palmitate are fatty acid specific, as laurate (C12:0) does not regulate Jmjd3 and the associated inflammatory profile. Further, palmitate-induced Jmjd3 expression is controlled via TLR4/MyD88-dependent signaling mechanism, where genetic depletion of TLR4 (Tlr4-/- ) or MyD88 (MyD88-/- ) negated the palmitate-induced changes in Jmjd3 and downstream NFκB-induced inflammation. Pharmacological inhibition of Jmjd3 using a small molecule inhibitor (GSK-J4) reduced macrophage inflammation and improved diabetic wound healing. Together, we conclude that palmitate contributes to the chronic Jmjd3-mediated activation of macrophages in diabetic peripheral tissue and a histone demethylase inhibitor-based therapy may represent a novel treatment for nonhealing diabetic wounds.


Assuntos
Histona Desmetilases/metabolismo , Histona Desmetilases com o Domínio Jumonji/metabolismo , Macrófagos/metabolismo , Palmitatos/metabolismo , Receptor 4 Toll-Like/metabolismo , Cicatrização/fisiologia , Animais , Diabetes Mellitus Tipo 2 , Humanos , Inflamação/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Monócitos/metabolismo , NF-kappa B/metabolismo , Transdução de Sinais/fisiologia , Regulação para Cima/fisiologia
8.
J Immunol ; 204(9): 2503-2513, 2020 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-32205424

RESUMO

Macrophages are critical for the initiation and resolution of the inflammatory phase of wound healing. In diabetes, macrophages display a prolonged inflammatory phenotype preventing tissue repair. TLRs, particularly TLR4, have been shown to regulate myeloid-mediated inflammation in wounds. We examined macrophages isolated from wounds of patients afflicted with diabetes and healthy controls as well as a murine diabetic model demonstrating dynamic expression of TLR4 results in altered metabolic pathways in diabetic macrophages. Further, using a myeloid-specific mixed-lineage leukemia 1 (MLL1) knockout (Mll1f/fLyz2Cre+ ), we determined that MLL1 drives Tlr4 expression in diabetic macrophages by regulating levels of histone H3 lysine 4 trimethylation on the Tlr4 promoter. Mechanistically, MLL1-mediated epigenetic alterations influence diabetic macrophage responsiveness to TLR4 stimulation and inhibit tissue repair. Pharmacological inhibition of the TLR4 pathway using a small molecule inhibitor (TAK-242) as well as genetic depletion of either Tlr4 (Tlr4-/- ) or myeloid-specific Tlr4 (Tlr4f/fLyz2Cre+) resulted in improved diabetic wound healing. These results define an important role for MLL1-mediated epigenetic regulation of TLR4 in pathologic diabetic wound repair and suggest a target for therapeutic manipulation.


Assuntos
Diabetes Mellitus Tipo 2/genética , Diabetes Mellitus Tipo 2/imunologia , Epigênese Genética/genética , Macrófagos/fisiologia , Receptor 4 Toll-Like/genética , Cicatrização/genética , Idoso , Animais , Epigênese Genética/imunologia , Feminino , Histonas/genética , Histonas/imunologia , Humanos , Inflamação/genética , Inflamação/imunologia , Mediadores da Inflamação/imunologia , Macrófagos/imunologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Pessoa de Meia-Idade , Proteína de Leucina Linfoide-Mieloide/genética , Proteína de Leucina Linfoide-Mieloide/imunologia , Regiões Promotoras Genéticas/genética , Regiões Promotoras Genéticas/imunologia , Receptor 4 Toll-Like/imunologia , Cicatrização/imunologia
9.
JCI Insight ; 5(5)2020 03 12.
Artigo em Inglês | MEDLINE | ID: mdl-32069267

RESUMO

A critical component of wound healing is the transition from the inflammatory phase to the proliferation phase to initiate healing and remodeling of the wound. Macrophages are critical for the initiation and resolution of the inflammatory phase during wound repair. In diabetes, macrophages display a sustained inflammatory phenotype in late wound healing characterized by elevated production of inflammatory cytokines, such as TNF-α. Previous studies have shown that an altered epigenetic program directs diabetic macrophages toward a proinflammatory phenotype, contributing to a sustained inflammatory phase. Males absent on the first (MOF) is a histone acetyltransferase (HAT) that has been shown be a coactivator of TNF-α signaling and promote NF-κB-mediated gene transcription in prostate cancer cell lines. Based on MOF's role in TNF-α/NF-κB-mediated gene expression, we hypothesized that MOF influences macrophage-mediated inflammation during wound repair. We used myeloid-specific Mof-knockout (Lyz2Cre Moffl/fl) and diet-induced obese (DIO) mice to determine the function of MOF in diabetic wound healing. MOF-deficient mice exhibited reduced inflammatory cytokine gene expression. Furthermore, we found that wound macrophages from DIO mice had elevated MOF levels and higher levels of acetylated histone H4K16, MOF's primary substrate of HAT activity, on the promoters of inflammatory genes. We further identified that MOF expression could be stimulated by TNF-α and that treatment with etanercept, an FDA-approved TNF-α inhibitor, reduced MOF levels and improved wound healing in DIO mice. This report is the first to our knowledge to define an important role for MOF in regulating macrophage-mediated inflammation in wound repair and identifies TNF-α inhibition as a potential therapy for the treatment of chronic inflammation in diabetic wounds.


Assuntos
Diabetes Mellitus Experimental/imunologia , Histona Acetiltransferases/metabolismo , Macrófagos/imunologia , Fator de Necrose Tumoral alfa/fisiologia , Animais , Diabetes Mellitus Experimental/fisiopatologia , Etanercepte/farmacologia , Inflamação/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fator de Necrose Tumoral alfa/antagonistas & inibidores , Cicatrização/fisiologia
10.
Microbes Infect ; 22(8): 312-321, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-31958572

RESUMO

Pneumococcal conjugate vaccination (PCV) may prevent influenza-related pneumonia, including Streptococcus pneumoniae pneumonia. To investigate PCV efficacy against secondary pneumococcal pneumonia following influenza, PCV was administered intramuscularly 2 and 5 weeks before S. pneumoniae serotype-3 colonization of murine nasopharynges followed by intranasal challenge with a sublethal dose of influenza A virus. Bacterial and viral loads, including innate immune responses were compared across conditions. PCV vaccination improved the survival of mice with secondary pneumococcal pneumonia and significantly reduced the pulmonary bacterial burden. Increased monocyte/macrophage influx into the lungs, alleviated loss of alveolar macrophages and decreased neutrophil influx into the lungs occurred in PCV-treated mice irrespective of pneumococcal colonization. Higher monocyte chemoattractant protein 1 levels and lower levels of CXCL1, interferon-γ, interleukin-17A, and IL-10, were detected in PCV-treated mice. Additionally, PCV treatment activated the macrophage intracellular killing of S. pneumoniae. Collectively, PCV potentially modulates the host's innate immunity and specific antibodies induction. Macrophage-related innate immunity should be further explored to elucidate the efficacy and mechanisms of PCV versus influenza-related life-threatening diseases.


Assuntos
Coinfecção/imunologia , Imunidade Inata , Macrófagos/imunologia , Infecções por Orthomyxoviridae/imunologia , Vacinas Pneumocócicas/imunologia , Pneumonia Pneumocócica/imunologia , Animais , Anticorpos Antibacterianos/sangue , Antígeno B7-2/metabolismo , Carga Bacteriana , Coinfecção/microbiologia , Coinfecção/mortalidade , Coinfecção/virologia , Citocinas/metabolismo , Modelos Animais de Doenças , Vírus da Influenza A , Pulmão/imunologia , Pulmão/microbiologia , Pulmão/virologia , Macrófagos/microbiologia , Camundongos , Neutrófilos/imunologia , Infecções por Orthomyxoviridae/microbiologia , Infecções por Orthomyxoviridae/mortalidade , Infecções por Orthomyxoviridae/virologia , Fagocitose , Vacinas Pneumocócicas/administração & dosagem , Pneumonia Pneumocócica/microbiologia , Pneumonia Pneumocócica/mortalidade , Pneumonia Pneumocócica/virologia , Streptococcus pneumoniae , Taxa de Sobrevida , Vacinação , Vacinas Conjugadas/administração & dosagem , Vacinas Conjugadas/imunologia
11.
Am J Physiol Lung Cell Mol Physiol ; 318(1): L200-L211, 2020 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-31747308

RESUMO

Stem cell factor (SCF) and its receptor c-kit have been implicated in inflammation, tissue remodeling, and fibrosis. Ingenuity Integrated Pathway Analysis of gene expression array data sets showed an upregulation of SCF transcripts in idiopathic pulmonary fibrosis (IPF) lung biopsies compared with tissue from nonfibrotic lungs that are further increased in rapid progressive disease. SCF248, a cleavable isoform of SCF, was abundantly and preferentially expressed in human lung fibroblasts and fibrotic mouse lungs relative to the SCF220 isoform. In fibroblast-mast cell coculture studies, blockade of SCF248 using a novel isoform-specific anti-SCF248 monoclonal antibody (anti-SCF248), attenuated the expression of COL1A1, COL3A1, and FN1 transcripts in cocultured IPF but not normal lung fibroblasts. Administration of anti-SCF248 on days 8 and 12 after bleomycin instillation in mice significantly reduced fibrotic lung remodeling and col1al, fn1, acta2, tgfb, and ccl2 transcript expression. In addition, bleomycin increased numbers of c-kit+ mast cells, eosinophils, and ILC2 in lungs of mice, whereas they were not significantly increased in anti-SCF248-treated animals. Finally, mesenchymal cell-specific deletion of SCF significantly attenuated bleomycin-mediated lung fibrosis and associated fibrotic gene expression. Collectively, these data demonstrate that SCF is upregulated in diseased IPF lungs and blocking SCF248 isoform significantly ameliorates fibrotic lung remodeling in vivo suggesting that it may be a therapeutic target for fibrotic lung diseases.


Assuntos
Fibrose Pulmonar Idiopática/metabolismo , Pulmão/metabolismo , Isoformas de Proteínas/metabolismo , Fator de Células-Tronco/metabolismo , Animais , Bleomicina/farmacologia , Contagem de Células/métodos , Células Cultivadas , Técnicas de Cocultura/métodos , Feminino , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Fibrose/tratamento farmacológico , Fibrose/metabolismo , Humanos , Fibrose Pulmonar Idiopática/tratamento farmacológico , Pulmão/efeitos dos fármacos , Mastócitos/efeitos dos fármacos , Mastócitos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Transdução de Sinais/efeitos dos fármacos , Regulação para Cima/efeitos dos fármacos
12.
Thromb Haemost ; 120(2): 289-299, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31887775

RESUMO

Venous thrombosis (VT) resolution is a complex process, resembling sterile wound healing. Infiltrating blood-derived monocyte/macrophages (Mo/MΦs) are essential for the regulation of inflammation in tissue repair. These cells differentiate into inflammatory (CD11b+Ly6CHi) or proreparative (CD11b+Ly6CLo) subtypes. Previous studies have shown that infiltrating Mo/MΦs are important for VT resolution, but the precise roles of different Mo/MΦs subsets are not well understood. Utilizing murine models of stasis and stenosis inferior vena cava thrombosis in concert with a Mo/MΦ depletion model (CD11b-diphtheria toxin receptor [DTR]-expressing mice), we examined the effect of Mo/MΦ depletion on thrombogenesis and VT resolution. In the setting of an 80 to 90% reduction in circulating CD11b+Mo/MΦs, we demonstrated that Mo/MΦs are not essential for thrombogenesis, with no difference in thrombus size, neutrophil recruitment, or neutrophil extracellular traps found. Conversely, CD11b+Mo/MΦ are essential for VT resolution. Diphtheria toxoid (DTx)-mediated depletion after thrombus creation depleted primarily CD11b+Ly6CLo Mo/MΦs and resulted in larger thrombi. DTx-mediated depletion did not alter CD11b+Ly6CHi Mo/MΦ recruitment, suggesting a protective effect of CD11b+Ly6CLo Mo/MΦs in VT resolution. Confirmatory Mo/MΦ depletion with clodronate lysosomes showed a similar phenotype, with failure to resolve VT. Adoptive transfer of CD11b+Ly6CLo Mo/MΦs into Mo/MΦ-depleted mice reversed the phenotype, restoring normal thrombus resolution. These findings suggest that CD11b+Ly6CLo Mo/MΦs are essential for normal VT resolution, consistent with the known proreparative function of this subset, and that further study of Mo/MΦ subsets may identify targets for immunomodulation to accelerate and improve thrombosis resolution.


Assuntos
Lisossomos/metabolismo , Macrófagos/citologia , Monócitos/citologia , Trombose/sangue , Trombose Venosa/sangue , Transferência Adotiva , Animais , Antígenos Ly/metabolismo , Antígenos CD11/metabolismo , Separação Celular , Toxina Diftérica/farmacologia , Ensaio de Imunoadsorção Enzimática , Inflamação , Leucócitos , Camundongos , Camundongos Endogâmicos C57BL , Neutrófilos/citologia , Fenótipo
13.
Front Immunol ; 10: 2148, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31708911

RESUMO

Inflammation after trauma is both critical to normal wound healing and may be highly detrimental when prolonged or unchecked with the potential to impair physiologic healing and promote de novo pathology. Mechanical strain after trauma is associated with impaired wound healing and increased inflammation. The exact mechanisms behind this are not fully elucidated. Neutrophil extracellular traps (NETs), a component of the neutrophil response to trauma, are implicated in a range of pro-inflammatory conditions. In the current study, we evaluated their role in linking movement and inflammation. We found that a link exists between the disruption and amplification of NETs which harbors the potential to regulate the wound's response to mechanical strain, while leaving the initial inflammatory signal necessary for physiologic wound healing intact.


Assuntos
Movimento Celular/imunologia , Armadilhas Extracelulares/imunologia , Neutrófilos/imunologia , Estresse Mecânico , Ferimentos e Lesões/imunologia , Animais , Humanos , Inflamação/imunologia , Inflamação/patologia , Camundongos , Neutrófilos/patologia , Ferimentos e Lesões/patologia
14.
Arterioscler Thromb Vasc Biol ; 39(11): 2353-2366, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31644352

RESUMO

OBJECTIVE: Sepsis represents an acute life-threatening disorder resulting from a dysregulated host response. For patients who survive sepsis, there remains long-term consequences, including impaired inflammation, as a result of profound immunosuppression. The mechanisms involved in this long-lasting deficient immune response are poorly defined. Approach and Results: Sepsis was induced using the murine model of cecal ligation and puncture. Following a full recovery period from sepsis physiology, mice were subjected to our wound healing model and wound macrophages (CD11b+, CD3-, CD19-, Ly6G-) were sorted. Post-sepsis mice demonstrated impaired wound healing and decreased reepithelization in comparison to controls. Further, post-sepsis bone marrow-derived macrophages and wound macrophages exhibited decreased expression of inflammatory cytokines vital for wound repair (IL [interleukin]-1ß, IL-12, and IL-23). To evaluate if decreased inflammatory gene expression was secondary to epigenetic modification, we conducted chromatin immunoprecipitation on post-sepsis bone marrow-derived macrophages and wound macrophages. This demonstrated decreased expression of Mll1, an epigenetic enzyme, and impaired histone 3 lysine 4 trimethylation (activation mark) at NFκB (nuclear factor kappa-light-chain-enhancer of activated B cells)-binding sites on inflammatory gene promoters in bone marrow-derived macrophages and wound macrophages from postcecal ligation and puncture mice. Bone marrow transplantation studies demonstrated epigenetic modifications initiate in bone marrow progenitor/stem cells following sepsis resulting in lasting impairment in peripheral macrophage function. Importantly, human peripheral blood leukocytes from post-septic patients demonstrate a significant reduction in MLL1 compared with nonseptic controls. CONCLUSIONS: These data demonstrate that severe sepsis induces stable mixed-lineage leukemia 1-mediated epigenetic modifications in the bone marrow, which are passed to peripheral macrophages resulting in impaired macrophage function and deficient wound healing persisting long after sepsis recovery.


Assuntos
Epigênese Genética , Inflamação/fisiopatologia , Macrófagos/fisiologia , Sepse/genética , Sepse/fisiopatologia , Cicatrização/fisiologia , Animais , Células da Medula Óssea/fisiologia , Células Cultivadas , Citocinas/metabolismo , Modelos Animais de Doenças , Feminino , Histona-Lisina N-Metiltransferase/genética , Histonas/genética , Humanos , Tolerância Imunológica , Masculino , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos , Proteína de Leucina Linfoide-Mieloide/genética , NF-kappa B/genética , Regiões Promotoras Genéticas , Sepse/metabolismo
15.
J Immunol ; 202(6): 1777-1785, 2019 03 15.
Artigo em Inglês | MEDLINE | ID: mdl-30710046

RESUMO

Myeloid cells are critical for orchestrating regulated inflammation during wound healing. TLRs, particularly TLR4, and its downstream-signaling MyD88 pathway play an important role in regulating myeloid-mediated inflammation. Because an initial inflammatory phase is vital for tissue repair, we investigated the role of TLR4-regulated, myeloid-mediated inflammation in wound healing. In a cutaneous tissue injury murine model, we found that TLR4 expression is dynamic in wound myeloid cells during the course of normal wound healing. We identified that changes in myeloid TLR4 during tissue repair correlated with increased expression of the histone methyltransferase, mixed-lineage leukemia 1 (MLL1), which specifically trimethylates the histone 3 lysine 4 (H3K4me3) position of the TLR4 promoter. Furthermore, we used a myeloid-specific Mll1 knockout (Mll1f/fLyz2Cre+ ) to determine MLL1 drives Tlr4 expression during wound healing. To understand the critical role of myeloid-specific TLR4 signaling, we used mice deficient in Tlr4 (Tlr4-/- ), Myd88 (Myd88 -/-), and myeloid-specific Tlr4 (Tlr4f/fLyz2Cre+) to demonstrate delayed wound healing at early time points postinjury. Furthermore, in vivo wound myeloid cells isolated from Tlr4-/- and Myd88 -/- wounds demonstrated decreased inflammatory cytokine production. Importantly, adoptive transfer of monocyte/macrophages from wild-type mice trafficked to wounds with restoration of normal healing and myeloid cell function in Tlr4-deficient mice. These results define a role for myeloid-specific, MyD88-dependent TLR4 signaling in the inflammatory response following cutaneous tissue injury and suggest that MLL1 regulates TLR4 expression in wound myeloid cells.


Assuntos
Histona-Lisina N-Metiltransferase/metabolismo , Histonas/metabolismo , Proteína de Leucina Linfoide-Mieloide/metabolismo , Pele/metabolismo , Receptor 4 Toll-Like/biossíntese , Cicatrização/fisiologia , Animais , Metilação de DNA/fisiologia , Feminino , Regulação da Expressão Gênica/fisiologia , Inflamação/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Células Mieloides/metabolismo , Fator 88 de Diferenciação Mieloide/metabolismo , Transdução de Sinais/fisiologia , Pele/lesões
16.
J Immunol ; 202(3): 931-942, 2019 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-30578307

RESUMO

Sepsis is the leading cause of death in the intensive care unit with an overall mortality rate of 20%. Individuals who are obese and have type 2 diabetes have increased recurrent, chronic, nosocomial infections that worsen the long-term morbidity and mortality from sepsis. Additionally, animal models of sepsis have shown that obese, diabetic mice have lower survival rates compared with nondiabetic mice. Neutrophils are essential for eradication of bacteria, prevention of infectious complications, and sepsis survival. In diabetic states, there is a reduction in neutrophil chemotaxis, phagocytosis, and reactive oxygen species (ROS) generation; however, few studies have investigated the extent to which these deficits compromise infection eradication and mortality. Using a cecal ligation and puncture model of sepsis in lean and in diet-induced obese mice, we demonstrate that obese diabetic mice have decreased "emergency hematopoiesis" after an acute infection. Additionally, both neutrophils and monocytes in obese, diabetic mice have functional defects, with decreased phagocytic ability and a decreased capacity to generate ROS. Neutrophils isolated from obese diabetic mice have decreased transcripts of Axl and Mertk, which partially explains the phagocytic dysfunction. Furthermore, we found that exogenous GM-CSF administration improves sepsis survival through enhanced neutrophil and monocytes phagocytosis and ROS generation abilities in obese, diabetic mice with sepsis.


Assuntos
Diabetes Mellitus Experimental/imunologia , Fator Estimulador de Colônias de Granulócitos e Macrófagos/farmacologia , Imunidade Inata/efeitos dos fármacos , Obesidade/imunologia , Sepse/imunologia , Animais , Bactérias , Citocinas/genética , Citocinas/imunologia , Diabetes Mellitus Experimental/microbiologia , Modelos Animais de Doenças , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Monócitos/imunologia , Monócitos/patologia , Neutrófilos/imunologia , Neutrófilos/patologia , Obesidade/microbiologia , Fagocitose , Sepse/tratamento farmacológico , Sepse/microbiologia
18.
J Invest Dermatol ; 139(5): 1161-1170, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30465800

RESUMO

Wound healing is a well-coordinated process that involves inflammatory mediators and cellular responses; however, if any disturbances are present during this process, tissue repair is impaired. Chronic wounds are one of the serious long-term complications associated with diabetes mellitus. The chemokine receptor CCR4 and its respective ligands, CCL17 and CCL22, are involved in regulatory T cell recruitment and activation in inflamed skin; however, the role of regulatory T cells in wounds is still not clear. Our aim was to investigate the role of CCR4 and regulatory T cells in cutaneous wound healing in diabetic mice. Alloxan-induced diabetic wild- type mice (diabetic) developed wounds that were difficult to heal, differently from CCR4-/- diabetic mice (CCR4-/- diabetic), and also from anti-CCL17/22 or anti-CD25-injected diabetic mice that presented with accelerated wound healing and fewer regulatory T cells in the wound bed. Consequently, CCR4-/- diabetic mice also presented with alteration on T cells population in the wound and draining lymph nodes; on day 14, these mice also displayed an increase of collagen fiber deposition. Still, cytokine levels were decreased in the wounds of CCR4-/- diabetic mice on day 2. Our data suggest that the receptor CCR4 and regulatory T cells negatively affect wound healing in diabetic mice.


Assuntos
Quimiocina CCL17/antagonistas & inibidores , Quimiocina CCL22/antagonistas & inibidores , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Tipo 1/metabolismo , Receptores CCR4/metabolismo , Cicatrização/efeitos dos fármacos , Aloxano/farmacologia , Análise de Variância , Animais , Biópsia por Agulha , Quimiocina CCL17/farmacologia , Quimiocina CCL22/farmacologia , Quimiocinas/metabolismo , Diabetes Mellitus Experimental/patologia , Diabetes Mellitus Tipo 1/tratamento farmacológico , Humanos , Imuno-Histoquímica , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Reação em Cadeia da Polimerase em Tempo Real/métodos , Cicatrização/fisiologia
19.
Sci Rep ; 8(1): 188, 2018 01 09.
Artigo em Inglês | MEDLINE | ID: mdl-29317674

RESUMO

Acute liver injury (ALI) is characterized by hepatocyte damage and inflammation. In the present study, we examined whether the absence of Sprouty-related EVH1-domain-containing protein 2 (Spred2), a negative regulator of the Ras/Raf/ERK/MAPK pathway, influences ALI induced by D-galactosamine (D-GalN) and lipopolysaccharide (LPS). Compared to wild-type mice, Spred2-/- mice developed exacerbated liver injury represented by enhanced hepatocyte damage and inflammation. Enhanced ERK activation was observed in Spred2-/--livers, and the MEK/ERK inhibitor U0126 ameliorated ALI. Hepatic tumour necrosis factor α (TNFα) and interleukin (IL)-1ß levels were increased in Spred-2-/--livers, and the neutralization of TNFα dramatically ameliorated ALI, which was associated with decreased levels of endogenous TNFα and IL-1ß. When mice were challenged with D-GalN and TNFα, much severer ALI was observed in Spred2-/- mice with significant increases in endogenous TNFα and IL-1ß in the livers. Immunohistochemically, Kupffer cells were found to produce TNFα, and isolated Kupffer cells from Spred2-/- mice produced significantly higher levels of TNFα than those from wild-type mice after LPS stimulation, which was significantly decreased by U0126. These results suggest that Spred2 negatively regulates D-GalN/LPS-induced ALI under the control of TNFα in Kupffer cells. Spred2 may present a therapeutic target for the treatment of ALI.


Assuntos
Falência Hepática/metabolismo , Proteínas Repressoras/genética , Animais , Células Cultivadas , Galactosamina/toxicidade , Hepatócitos/metabolismo , Interleucina-1beta/genética , Interleucina-1beta/metabolismo , Macrófagos do Fígado/metabolismo , Lipopolissacarídeos/toxicidade , Falência Hepática/etiologia , Falência Hepática/genética , Sistema de Sinalização das MAP Quinases , Camundongos , Camundongos Endogâmicos C57BL , Proteínas Repressoras/deficiência , Proteínas Repressoras/metabolismo , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/metabolismo
20.
J Immunol ; 199(7): 2555-2569, 2017 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-28827285

RESUMO

Although most novel tuberculosis (TB) vaccines are designed for delivery via the muscle or skin for enhanced protection in the lung, it has remained poorly understood whether systemic vaccine-induced memory T cells can readily home to the lung mucosa prior to and shortly after pathogen exposure. We have investigated this issue by using a model of parenteral TB immunization and intravascular immunostaining. We find that systemically induced memory T cells are restricted to the blood vessels in the lung, unable to populate either the lung parenchymal tissue or the airway under homeostatic conditions. We further find that after pulmonary TB infection, it still takes many days before such T cells can enter the lung parenchymal tissue and airway. We have identified the acquisition of CXCR3 expression by circulating T cells to be critical for their entry to these lung mucosal compartments. Our findings offer new insights into mucosal T cell biology and have important implications in vaccine strategies against pulmonary TB and other intracellular infections in the lung.


Assuntos
Pulmão/imunologia , Mycobacterium tuberculosis/imunologia , Receptores CXCR3/metabolismo , Vacinas contra a Tuberculose/imunologia , Tuberculose Pulmonar/imunologia , Transferência Adotiva , Animais , Antígenos de Bactérias/imunologia , Linfócitos T CD8-Positivos/imunologia , Movimento Celular , Imunização , Memória Imunológica , Leucócitos/imunologia , Pulmão/citologia , Pulmão/microbiologia , Camundongos , Receptores CXCR3/genética , Receptores CXCR3/imunologia , Mucosa Respiratória/imunologia , Mucosa Respiratória/microbiologia , Transdução de Sinais , Vacinas contra a Tuberculose/administração & dosagem , Tuberculose Pulmonar/microbiologia
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