RESUMO
Macrophage plasticity is critical for normal tissue repair to ensure transition from the inflammatory to the proliferative phase of healing. We examined macrophages isolated from wounds of patients afflicted with diabetes and of healthy controls and found differential expression of the methyltransferase Setdb2. Myeloid-specific deletion of Setdb2 impaired the transition of macrophages from an inflammatory phenotype to a reparative one in normal wound healing. Mechanistically, Setdb2 trimethylated histone 3 at NF-κB binding sites on inflammatory cytokine gene promoters to suppress transcription. Setdb2 expression in wound macrophages was regulated by interferon (IFN) ß, and under diabetic conditions, this IFNß-Setdb2 axis was impaired, leading to a persistent inflammatory macrophage phenotype in diabetic wounds. Setdb2 regulated the expression of xanthine oxidase and thereby the uric acid (UA) pathway of purine catabolism in macrophages, and pharmacologic targeting of Setdb2 or the UA pathway improved healing. Thus, Setdb2 regulates macrophage plasticity during normal and pathologic wound repair and is a target for therapeutic manipulation.
Assuntos
Proteínas de Transporte/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Histona-Lisina N-Metiltransferase/metabolismo , Macrófagos/fisiologia , Proteínas Nucleares/metabolismo , Idoso , Animais , Proteínas de Transporte/genética , Diferenciação Celular , Células Cultivadas , Modelos Animais de Doenças , Feminino , Histona-Lisina N-Metiltransferase/genética , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , Pessoa de Meia-Idade , Proteínas Nucleares/genética , Fenótipo , Ácido Úrico/metabolismo , CicatrizaçãoRESUMO
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/metabolismoRESUMO
Macrophages play a critical role in the establishment of a regulated inflammatory response following tissue injury. Following injury, CCR2+ monocytes are recruited from peripheral blood to wound tissue, and direct the initiation and resolution of inflammation that is essential for tissue repair. In pathologic states where chronic inflammation prevents healing, macrophages fail to transition to a reparative phenotype. Using a murine model of cutaneous wound healing, we found that CCR2-deficient mice (CCR2-/- ) demonstrate significantly impaired wound healing at all time points postinjury. Flow cytometry analysis of wounds from CCR2-/- and WT mice revealed a significant decrease in inflammatory, Ly6CHi recruited monocyte/macrophages in CCR2-/- wounds. We further show that wound macrophage inflammatory cytokine production is decreased in CCR2-/- wounds. Adoptive transfer of mT/mG monocyte/macrophages into CCR2+/+ and CCR2-/- mice demonstrated that labeled cells on days 2 and 4 traveled to wounds in both CCR2+/+ and CCR2-/- mice. Further, adoptive transfer of monocyte/macrophages from WT mice restored normal healing, likely through a restored inflammatory response in the CCR2-deficient mice. Taken together, these data suggest that CCR2 plays a critical role in the recruitment and inflammatory response following injury, and that wound repair may be therapeutically manipulated through modulation of CCR2.
Assuntos
Macrófagos/transplante , Receptores CCR2/genética , Cicatrização/genética , Cicatrização/fisiologia , Animais , Inflamação/imunologia , Macrófagos/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptores CCR2/metabolismoRESUMO
The healing of cutaneous wounds is dependent on the progression through distinct, yet overlapping phases of wound healing, including hemostasis, inflammation, proliferation, and resolution/remodeling. The failure of these phases to occur in a timely, progressive fashion promotes pathologic wound healing. The macrophage (MΦ) has been demonstrated to play a critical role in the inflammatory phase of tissue repair, where its dynamic plasticity allows this cell to mediate both tissue-destructive and -reparative functions. The ability to understand and control both the initiation and the resolution of inflammation is critical for treating pathologic wound healing. There are now a host of studies demonstrating that metabolic and epigenetic regulation of gene transcription can influence MΦ plasticity in wounds. In this review, we highlight the molecular and epigenetic factors that influence MΦ polarization in both physiologic and pathologic wound healing, with particular attention to diabetic wounds.
Assuntos
Diabetes Mellitus/imunologia , Inflamação/imunologia , Macrófagos/imunologia , Cicatrização/imunologia , Animais , Diferenciação Celular , Complicações do Diabetes/imunologia , Epigênese Genética , Regulação da Expressão Gênica , Humanos , Mediadores da Inflamação/imunologia , CamundongosRESUMO
Deep venous thrombosis and residual thrombus burden correlates with circulating IL-6 levels in humans. To investigate the cellular source and role of IL-6 in thrombus resolution, Wild type C57BL/6J (WT), and IL-6-/- mice underwent induction of VT via inferior vena cava (IVC) stenosis or stasis. Vein wall (VW) and thrombus were analyzed by western blot, immunohistochemistry, and flow cytometry. Adoptive transfer of WT bone marrow derived monocytes was performed into IL6-/- mice to assess for rescue. Cultured BMDMs from WT and IL-6-/- mice underwent quantitative real time PCR and immunoblotting for fibrinolytic factors and matrix metalloproteinase activity. No differences in baseline coagulation function or platelet function were found between WT and IL-6-/- mice. VW and thrombus IL-6 and IL-6 leukocyte-specific receptor CD126 were elevated in a time-dependent fashion in both VT models. Ly6Clo Mo/MØ were the predominant leukocyte source of IL-6. IL-6-/- mice demonstrated larger, non-resolving stasis thrombi with less neovascularization, despite a similar number of monocytes/macrophages (Mo/MØ). Adoptive transfer of WT BMDM into IL-6-/- mice undergoing stasis VT resulted in phenotype rescue. Human specimens of endophlebectomized tissue showed co-staining of Monocyte and IL-6 receptor. Thrombosis matrix analysis revealed significantly increased thrombus fibronectin and collagen in IL-6-/- mice. MMP9 activity in vitro depended on endogenous IL-6 expression in Mo/MØ, and IL-6-/- mice exhibited stunted matrix metalloproteinase activity. Lack of IL-6 signaling impairs thrombus resolution potentially via dysregulation of MMP-9 leading to impaired thrombus recanalization and resolution. Restoring or augmenting monocyte-mediated IL-6 signaling in IL-6 deficient or normal subjects, respectively, may represent a non-anticoagulant target to improve thrombus resolution.
Assuntos
Trombose , Doenças Vasculares , Trombose Venosa , Animais , Humanos , Camundongos , Modelos Animais de Doenças , Interleucina-6/metabolismo , Camundongos Endogâmicos C57BL , Monócitos/metabolismo , Trombose/metabolismo , Doenças Vasculares/metabolismo , Veia Cava Inferior/metabolismo , Trombose Venosa/genéticaRESUMO
Symptomatic or ruptured thoracoabdominal aortic aneurysms (TAAA) carry a high morbidity and mortality. Modern fenestrated and/or branched endovascular devices (B/FEVAR) have improved the immediate peri-operative mortality of TAAA and have increased the number of people that can undergo repair - in those who might otherwise be prohibitively high risk for surgery. Most modern B/FEVAR are custom made devices that require 6-12 weeks to assemble and ship to the site of implantation. Thus, patients who require more urgent repair due to symptomatic or ruptured aneurysms may not have access to this potentially life saving technology. Physician-modified endografts (PMEGs), or traditional endografts that have been back-table modified to have fenestrations or branches, have partially fixed this problem as they can be constructed in less than an hour and can provide similar results to modern custom made devices. Here we review the existing data behind the use of PMEGs in urgent and emergent aortic pathology and summarize a case describing one methodology for PMEG construction that has been standardized at our institution.
Assuntos
Aneurisma da Aorta Torácica , Implante de Prótese Vascular , Procedimentos Endovasculares , Médicos , Aneurisma da Aorta Torácica/diagnóstico por imagem , Aneurisma da Aorta Torácica/cirurgia , Prótese Vascular , Implante de Prótese Vascular/efeitos adversos , Procedimentos Endovasculares/efeitos adversos , Humanos , Complicações Pós-Operatórias , Desenho de Prótese , Estudos Retrospectivos , Fatores de Risco , Fatores de Tempo , Resultado do TratamentoRESUMO
Control of inflammation is critical for the treatment of nonhealing wounds, but a delicate balance exists between early inflammation that is essential for normal tissue repair and the pathologic inflammation that can occur later in the repair process. This necessitates the development of novel therapies that can target inflammation at the appropriate time during repair. Here, we found that SIRT3 is essential for normal healing and regulates inflammation in wound macrophages after injury. Under prediabetic conditions, SIRT3 was decreased in wound macrophages and resulted in dysregulated inflammation. In addition, we found that FABP4 regulates SIRT3 in human blood monocytes, and inhibition of FABP4 in wound macrophages decreases inflammatory cytokine expression, making FABP4 a viable target for the regulation of excess inflammation and wound repair in diabetes. Using a series of ex vivo and in vivo studies with genetically engineered mouse models and diabetic human monocytes, we showed that FABP4 expression is epigenetically upregulated in diabetic wound macrophages and, in turn, diminishes SIRT3 expression, thereby promoting inflammation. These findings have significant implications for controlling inflammation and promoting tissue repair in diabetic wounds.
Assuntos
Diabetes Mellitus Experimental/tratamento farmacológico , Sirtuína 3/farmacologia , Cicatrização/efeitos dos fármacos , Animais , Células Cultivadas , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Experimental/patologia , Macrófagos/metabolismo , Macrófagos/patologia , Camundongos , Camundongos Endogâmicos C57BLRESUMO
BACKGROUND: Whereas chronic venous insufficiency and varicose veins (VVs) are a universally recognized problem, they are frequently underappreciated as major contributors to long-term morbidity in the elderly despite the increasing prevalence with age. Previous studies have demonstrated that chronic venous insufficiency and VV treatments in patients ≥65 years old yield an overall benefit; however, there have been few data as to whether octogenarians are undergoing these procedures and with what success. As such, our objectives were to investigate the procedures selected, to examine clinical outcomes after VV procedures in elderly patients ≥80 years old, and to explore complication rates (both systemic and leg specific) after VV procedures in patients ≥80 years old. METHODS: We performed a retrospective review using the Vascular Quality Initiative Varicose Vein Registry of all VV procedures performed for ≥C2 disease from January 2015 to February 2017. We divided all procedures into three age groups: patients <65 years, patients ≥65 to 79 years, and patients ≥80 years. Statistical testing included χ2 test for categorical variables and Student t-test for continuous variables. Two comparisons were performed: first, comparing patients <65 years old with patients ≥65 to 79 years old; and second, comparing patients ≥65 to 79 years old with patients ≥80 years old. RESULTS: There were a total of 12,262 procedures performed, with 8608 procedures in the patients <65 years, 3226 in patients 65 to 79 years, and 428 procedures in patients ≥80 years. A total of 22,050 veins were treated during the 12,262 procedures. Almost half of procedures (46.51%; n = 5703) had only one vein treated during a single procedure. Between age groups, the percentage of one vein treated increased as the patient's age increased, ranging from 45.39% (n = 3875) for patients <65 years to 48.55% (n = 1555) for patients between 65 and 79 years and 64.08% (n = 273) for patients ≥80 years. Patients in the group ≥80 years had an overall lower average body mass index and were more likely to be receiving anticoagulation and to undergo truncal procedures alone compared with the other groups. The group ≥80 years had a significant improvement in both Venous Clinical Severity Score (4.37 ± 4.16; P < .001) and patient-reported outcomes (8.79 ± 7.27; P < .001) from before to after the procedure. Overall complications were low in all age groups. The octogenarians had no higher risk of systemic complications. CONCLUSIONS: Vascular specialists are performing VV procedures in octogenarians and are more likely to perform truncal only therapy. In addition, octogenarians have statistically significant improvement of Venous Clinical Severity Score and patient-reported outcomes with a low risk of complications despite more advanced venous disease at presentation.
Assuntos
Varizes/cirurgia , Procedimentos Cirúrgicos Vasculares , Insuficiência Venosa/cirurgia , Fatores Etários , Idoso , Idoso de 80 Anos ou mais , Distribuição de Qui-Quadrado , Doença Crônica , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Medidas de Resultados Relatados pelo Paciente , Recuperação de Função Fisiológica , Sistema de Registros , Estudos Retrospectivos , Fatores de Risco , Índice de Gravidade de Doença , Resultado do Tratamento , Varizes/diagnóstico por imagem , Varizes/fisiopatologia , Procedimentos Cirúrgicos Vasculares/efeitos adversos , Insuficiência Venosa/diagnóstico por imagem , Insuficiência Venosa/fisiopatologiaRESUMO
Macrophages are essential immune cells necessary for regulated inflammation during wound healing. Recent studies have identified that Notch plays a role in macrophage-mediated inflammation. Thus, we investigated the role of Notch signaling on wound macrophage phenotype and function during normal and diabetic wound healing. We found that Notch receptor and ligand expression are dynamic in wound macrophages during normal healing. Mice with a myeloid-specific Notch signaling defect (DNMAMLfloxedLyz2Cre+ ) demonstrated delayed early healing (days 1-3) and wound macrophages had decreased inflammatory gene expression. In our physiologic murine model of type 2 diabetes (T2D), Notch receptor expression was significantly increased in wound macrophages on day 6, following the initial inflammatory phase of wound healing, corresponding to increased inflammatory cytokine expression. This increase in Notch1 and Notch2 was also observed in human monocytes from patients with T2D. Further, in prediabetic mice with a genetic Notch signaling defect (DNMAMLfloxedLyz2Cre+ on a high-fat diet), improved wound healing was seen at late time points (days 6-7). These findings suggest that Notch is critical for the early inflammatory phase of wound healing and directs production of macrophage-dependent inflammatory mediators. These results identify that canonical Notch signaling is important in directing macrophage function in wound repair and define a translational target for the treatment of non-healing diabetic wounds.
RESUMO
Macrophages are critical for the initiation and resolution of the inflammatory phase of wound repair. In diabetes, macrophages display a prolonged inflammatory phenotype in late wound healing. Mixed-lineage leukemia-1 (MLL1) has been shown to direct gene expression by regulating nuclear factor-κB (NF-κB)-mediated inflammatory gene transcription. Thus, we hypothesized that MLL1 influences macrophage-mediated inflammation in wound repair. We used a myeloid-specific Mll1 knockout (Mll1f/fLyz2Cre+ ) to determine the function of MLL1 in wound healing. Mll1f/fLyz2Cre+ mice display delayed wound healing and decreased wound macrophage inflammatory cytokine production compared with control animals. Furthermore, wound macrophages from Mll1f/fLyz2Cre+ mice demonstrated decreased histone H3 lysine 4 trimethylation (H3K4me3) (activation mark) at NF-κB binding sites on inflammatory gene promoters. Of note, early wound macrophages from prediabetic mice displayed similarly decreased MLL1, H3K4me3 at inflammatory gene promoters, and inflammatory cytokines compared with controls. Late wound macrophages from prediabetic mice demonstrated an increase in MLL1, H3K4me3 at inflammatory gene promoters, and inflammatory cytokines. Prediabetic macrophages treated with an MLL1 inhibitor demonstrated reduced inflammation. Finally, monocytes from patients with type 2 diabetes had increased Mll1 compared with control subjects without diabetes. These results define an important role for MLL1 in regulating macrophage-mediated inflammation in wound repair and identify a potential target for the treatment of chronic inflammation in diabetic wounds.
Assuntos
Diabetes Mellitus Tipo 2/metabolismo , Histona-Lisina N-Metiltransferase/metabolismo , Macrófagos/fisiologia , Proteína de Leucina Linfoide-Mieloide/metabolismo , Obesidade/metabolismo , Cicatrização/fisiologia , Animais , Células Cultivadas , Dieta Hiperlipídica , Histona-Lisina N-Metiltransferase/genética , Humanos , Inflamação/metabolismo , Camundongos , Camundongos Knockout , Proteína de Leucina Linfoide-Mieloide/genética , Estado Pré-Diabético/metabolismoRESUMO
Venous thrombosis (VT), a leading cause of morbidity and mortality worldwide, has recently been linked to neutrophil activation and release of neutrophil extracellular traps (NETs) via a process called NETosis. The use of various in vivo thrombosis models and genetically modified mice has more precisely defined the exact role of NETosis in the pathogenesis of VT. Translational large animal VT models and human studies have confirmed the presence of NETs in pathologic VT. Activation of neutrophils, with subsequent NETosis, has also been linked to acute infection. This innate immune response, while effective for bacterial clearance from the host by formation of an intravascular bactericidal "net," also triggers thrombosis. Intravascular thrombosis related to such innate immune mechanisms has been coined immunothrombosis. Dysregulated immunothrombosis has been proposed as a mechanism of pathologic micro- and macrovascular thrombosis in sepsis and autoimmune disease. In this focused review, we will address the dual role of NETs in the pathogenesis of VT and immunothrombosis.