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1.
JCI Insight ; 3(11)2018 06 07.
Artigo em Inglês | MEDLINE | ID: mdl-29875311

RESUMO

Pulmonary arterial hypertension (PAH) remains a disease with limited therapeutic options and dismal prognosis. Despite its etiologic heterogeneity, the underlying unifying pathophysiology is characterized by increased vascular tone and adverse remodeling of the pulmonary circulation. Myeloperoxidase (MPO), an enzyme abundantly expressed in neutrophils, has potent vasoconstrictive and profibrotic properties, thus qualifying as a potential contributor to this disease. Here, we sought to investigate whether MPO is causally linked to the pathophysiology of PAH. Investigation of 2 independent clinical cohorts revealed that MPO plasma levels were elevated in subjects with PAH and predicted adverse outcome. Experimental analyses showed that, upon hypoxia, right ventricular pressure was less increased in Mpo-/- than in WT mice. The hypoxia-induced activation of the Rho-kinase pathway, a critical subcellular signaling pathway yielding vasoconstriction and structural vascular remodeling, was blunted in Mpo-/- mice. Mice subjected to i.v. infusion of MPO revealed activation of Rho-kinase and increased right ventricular pressure, which was prevented by coinfusion of the Rho-kinase inhibitor Y-27632. In the Sugen5416/hypoxia rat model, PAH was attenuated by the MPO inhibitor AZM198. The current data demonstrate a tight mechanistic link between MPO, the activation of Rho-kinase, and adverse pulmonary vascular function, thus pointing toward a potentially novel avenue of treatment.


Assuntos
Hipertensão Pulmonar/patologia , Hipóxia/patologia , Peroxidase/metabolismo , Artéria Pulmonar/patologia , Quinases Associadas a rho/metabolismo , Adulto , Amidas/administração & dosagem , Animais , Estudos de Coortes , Modelos Animais de Doenças , Feminino , Humanos , Hipertensão Pulmonar/sangue , Hipertensão Pulmonar/mortalidade , Hipertensão Pulmonar/fisiopatologia , Hipóxia/sangue , Hipóxia/etiologia , Infusões Intravenosas , Estimativa de Kaplan-Meier , Pulmão/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Pessoa de Meia-Idade , Peroxidase/administração & dosagem , Peroxidase/sangue , Artéria Pulmonar/fisiopatologia , Piridinas/administração & dosagem , Ratos , Ratos Sprague-Dawley , Proteínas Recombinantes/administração & dosagem , Proteínas Recombinantes/sangue , Proteínas Recombinantes/metabolismo , Remodelação Vascular/efeitos dos fármacos , Remodelação Vascular/fisiologia , Vasoconstrição/efeitos dos fármacos , Vasoconstrição/fisiologia , Quinases Associadas a rho/antagonistas & inibidores
2.
Circ Res ; 121(1): 56-70, 2017 Jun 23.
Artigo em Inglês | MEDLINE | ID: mdl-28404615

RESUMO

RATIONALE: Ventricular arrhythmias remain the leading cause of death in patients suffering myocardial ischemia. Myeloperoxidase, a heme enzyme released by polymorphonuclear neutrophils, accumulates within ischemic myocardium and has been linked to adverse left ventricular remodeling. OBJECTIVE: To reveal the role of myeloperoxidase for the development of ventricular arrhythmias. METHODS AND RESULTS: In different murine models of myocardial ischemia, myeloperoxidase deficiency profoundly decreased vulnerability for ventricular tachycardia on programmed right ventricular and burst stimulation and spontaneously as assessed by ECG telemetry after isoproterenol injection. Experiments using CD11b/CD18 integrin-deficient (CD11b-/-) mice and intravenous myeloperoxidase infusion revealed that neutrophil infiltration is a prerequisite for myocardial myeloperoxidase accumulation. Ventricles from myeloperoxidase-deficient (Mpo-/-) mice showed less pronounced slowing and decreased heterogeneity of electric conduction in the peri-infarct zone than wild-type mice. Expression of the redox-sensitive gap junctional protein Cx43 (Connexin 43) was reduced in the peri-infarct area of wild-type compared with Mpo-/- mice. In isolated wild-type cardiomyocytes, Cx43 protein content decreased on myeloperoxidase/H2O2 incubation. Mapping of induced pluripotent stem cell-derived cardiomyocyte networks and in vivo investigations linked Cx43 breakdown to myeloperoxidase-dependent activation of matrix metalloproteinase 7. Moreover, Mpo-/- mice showed decreased ventricular postischemic fibrosis reflecting reduced accumulation of myofibroblasts. Ex vivo, myeloperoxidase was demonstrated to induce fibroblast-to-myofibroblast transdifferentiation by activation of p38 mitogen-activated protein kinases resulting in upregulated collagen generation. In support of our experimental findings, baseline myeloperoxidase plasma levels were independently associated with a history of ventricular arrhythmias, sudden cardiac death, or implantable cardioverter-defibrillator implantation in a cohort of 2622 stable patients with an ejection fraction >35% undergoing elective diagnostic cardiac evaluation. CONCLUSIONS: Myeloperoxidase emerges as a crucial mediator of postischemic myocardial remodeling and may evolve as a novel pharmacological target for secondary disease prevention after myocardial ischemia.


Assuntos
Arritmias Cardíacas/metabolismo , Isquemia Miocárdica/metabolismo , Miócitos Cardíacos/metabolismo , Peroxidase/deficiência , Remodelação Ventricular/fisiologia , Animais , Arritmias Cardíacas/patologia , Células Cultivadas , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Isquemia Miocárdica/patologia , Miócitos Cardíacos/patologia , Técnicas de Cultura de Órgãos
3.
Cardiovasc Res ; 109(1): 174-84, 2016 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-26598510

RESUMO

AIM: Atrial fibrosis, one of the most striking features in the pathology of atrial fibrillation (AF), is promoted by local and systemic inflammation. Electrophilic fatty acid nitroalkenes, endogenously generated by both metabolic and inflammatory reactions, are anti-inflammatory mediators that in synthetic form may be useful as drug candidates. Herein we investigate whether an exemplary nitro-fatty acid can limit atrial fibrosis and AF. METHODS AND RESULTS: Wild-type C57BL6/J mice were treated for 2 weeks with angiotensin II (AngII) and vehicle or nitro-oleic acid (10-nitro-octadec-9-enoic acid, OA-NO2, 6 mg/kg body weight) via subcutaneous osmotic minipumps. OA-NO2 significantly inhibited atrial fibrosis and depressed vulnerability for AF during right atrial electrophysiological stimulation to levels observed for AngII-naive animals. Left atrial epicardial mapping studies demonstrated preservation of conduction homogeneity by OA-NO2. The protection from fibrotic remodelling was mediated by suppression of Smad2-dependent myofibroblast transdifferentiation and inhibition of Nox2-dependent atrial superoxide formation. CONCLUSION: OA-NO2 potently inhibits atrial fibrosis and subsequent AF. Nitro-fatty acids and possibly other lipid electrophiles thus emerge as potential therapeutic agents for AF, either by increasing endogenous levels through dietary modulation or by administration as synthetic drugs.


Assuntos
Angiotensina II/farmacologia , Fibrilação Atrial/prevenção & controle , Remodelamento Atrial/efeitos dos fármacos , Ácidos Linoleicos/farmacologia , Nitrocompostos/farmacologia , Potenciais de Ação/efeitos dos fármacos , Animais , Transdiferenciação Celular/efeitos dos fármacos , Células Cultivadas , Conexina 43/análise , Fibrose , Átrios do Coração/patologia , Camundongos , Camundongos Endogâmicos C57BL , Proteína Smad2/antagonistas & inibidores
4.
Free Radic Biol Med ; 90: 252-260, 2016 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-26620549

RESUMO

Inflammation is an immune response triggered by microbial invasion and/or tissue injury. While acute inflammation is directed toward invading pathogens and injured cells, thus enabling tissue regeneration, chronic inflammation can lead to severe pathologies and tissue dysfunction. These processes are linked with macrophage polarization into specific inflammatory "M1-like" or regulatory "M2-like" subsets. Nitro-fatty acids (NO2-FAs), produced endogenously as byproducts of metabolism and oxidative inflammatory conditions, may be useful for treating diseases associated with dysregulated immune homeostasis. The goal of this study was to characterize the role of nitro-oleic acid (OA-NO2) in regulating the functional specialization of macrophages induced by bacterial lipopolysaccharide or interleukin-4, and to reveal specific signaling mechanisms which can account for OA-NO2-dependent modulation of inflammation and fibrotic responses. Our results show that OA-NO2 inhibits lipopolysaccharide-stimulated production of both pro-inflammatory and immunoregulatory cytokines (including transforming growth factor-ß) and inhibits nitric oxide and superoxide anion production. OA-NO2 also decreases interleukin-4-induced macrophage responses by inhibiting arginase-I expression and transforming growth factor-ß production. These effects are mediated via downregulation of signal transducers and activators of transcription, mitogen-activated protein kinase and nuclear factor-кB signaling responses. Finally, OA-NO2 inhibits fibrotic processes in an in vivo model of angiotensin II-induced myocardial fibrosis by attenuating expression of α-smooth muscle actin, systemic transforming growth factor-ß levels and infiltration of both "M1-" and "M2-like" macrophage subsets into afflicted tissue. Overall, the electrophilic fatty acid derivative OA-NO2 modulates a broad range of "M1-" and "M2-like" macrophage functions and represents a potential therapeutic approach to target diseases associated with dysregulated macrophage subsets.


Assuntos
Ativação de Macrófagos/efeitos dos fármacos , Ácidos Oleicos/farmacologia , Animais , Células Cultivadas , Fibrose , Interleucina-4/farmacologia , Lipopolissacarídeos/farmacologia , Camundongos , Camundongos Endogâmicos C57BL , Miocárdio/patologia , Óxido Nítrico/biossíntese , PPAR gama/fisiologia , Fator de Transcrição STAT3/fisiologia , Superóxidos/metabolismo
5.
J Mol Cell Cardiol ; 74: 353-63, 2014 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-24976018

RESUMO

Myeloperoxidase (MPO) is a heme enzyme abundantly expressed in polymorphonuclear neutrophils. MPO is enzymatically capable of catalyzing the generation of reactive oxygen species (ROS) and the consumption of nitric oxide (NO). Thus MPO has both potent microbicidal and, upon binding to the vessel wall, pro-inflammatory properties. Interestingly, MPO - a highly cationic protein - has been shown to bind to both endothelial cells and leukocyte membranes. Given the anionic surface charge of red blood cells, we investigated binding of MPO to erythrocytes. Red blood cells (RBCs) derived from patients with elevated MPO plasma levels showed significantly higher amounts of MPO by flow cytometry and ELISA than healthy controls. Heparin-induced MPO-release from patient-derived RBCs was significantly increased compared to controls. Ex vivo experiments revealed dose and time dependency for MPO-RBC binding, and immunofluorescence staining as well as confocal microscopy localized MPO-RBC interaction to the erythrocyte plasma membrane. NO-consumption by RBC-membrane fragments (erythrocyte "ghosts") increased with incrementally greater concentrations of MPO during incubation, indicating preserved catalytic MPO activity. In vivo infusion of MPO-loaded RBCs into C57BL/6J mice increased local MPO tissue concentrations in liver, spleen, lung, and heart tissue as well as within the cardiac vasculature. Further, NO-dependent relaxation of aortic rings was altered by RBC bound-MPO and systemic vascular resistance significantly increased after infusion of MPO-loaded RBCs into mice. In summary, we find that MPO binds to RBC membranes in vitro and in vivo, is transported by RBCs to remote sites in mice, and affects endothelial function as well as systemic vascular resistance. RBCs may avidly bind circulating MPO, and act as carriers of this leukocyte-derived enzyme.


Assuntos
Síndrome Coronariana Aguda/sangue , Eritrócitos/metabolismo , Insuficiência Cardíaca/sangue , Peroxidase/sangue , Síndrome Coronariana Aguda/patologia , Animais , Aorta/efeitos dos fármacos , Transporte Biológico , Células Cultivadas , Endotélio Vascular/efeitos dos fármacos , Eritrócitos/patologia , Coração/efeitos dos fármacos , Insuficiência Cardíaca/patologia , Heparina/química , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Óxido Nítrico/metabolismo , Óxido Nítrico/farmacologia , Técnicas de Cultura de Órgãos , Peroxidase/farmacologia , Ligação Proteica , Técnicas de Cultura de Tecidos , Resistência Vascular/efeitos dos fármacos
6.
PLoS One ; 9(2): e89307, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-24558493

RESUMO

BACKGROUND: Recent observational clinical and ex-vivo studies suggest that inflammation and in particular leukocyte activation predisposes to atrial fibrillation (AF). However, whether local binding and extravasation of leukocytes into atrial myocardium is an essential prerequisite for the initiation and propagation of AF remains elusive. Here we investigated the role of atrial CD11b/CD18 mediated infiltration of polymorphonuclear neutrophils (PMN) for the susceptibility to AF. METHODS AND RESULTS: C57bl/6J wildtype (WT) and CD11b/CD18 knock-out (CD11b(-/-)) mice were treated for 14 days with subcutaneous infusion of angiotensin II (Ang II), a known stimulus for PMN activation. Atria of Ang II-treated WT mice were characterized by increased PMN infiltration assessed in immunohistochemically stained sections. In contrast, atrial sections of CD11b(-/-) mice lacked a significant increase in PMN infiltration upon Ang II infusion. PMN infiltration was accompanied by profoundly enhanced atrial fibrosis in Ang II treated WT as compared to CD11b(-/-) mice. Upon in-vivo electrophysiological investigation, Ang II treatment significantly elevated the susceptibility for AF in WT mice if compared to vehicle treated animals given an increased number and increased duration of AF episodes. In contrast, animals deficient of CD11b/CD18 were entirely protected from AF induction. Likewise, epicardial activation mapping revealed decreased electrical conduction velocity in atria of Ang II treated WT mice, which was preserved in CD11b(-/-) mice. In addition, atrial PMN infiltration was enhanced in atrial appendage sections of patients with persistent AF as compared to patients without AF. CONCLUSIONS: The current data critically link CD11b-integrin mediated atrial PMN infiltration to the formation of fibrosis, which promotes the initiation and propagation of AF. These findings not only reveal a mechanistic role of leukocytes in AF but also point towards a potential novel avenue of treatment in AF.


Assuntos
Angiotensina II/farmacologia , Fibrilação Atrial/etiologia , Fibrilação Atrial/metabolismo , Antígeno CD11b/metabolismo , Antígenos CD18/metabolismo , Neutrófilos/metabolismo , Análise de Variância , Angiotensina II/administração & dosagem , Animais , Antígeno CD11b/genética , Antígenos CD18/genética , Técnicas Eletrofisiológicas Cardíacas/métodos , Imunofluorescência , Átrios do Coração/metabolismo , Immunoblotting , Imuno-Histoquímica , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Neutrófilos/efeitos dos fármacos
7.
Am J Respir Cell Mol Biol ; 51(1): 155-62, 2014 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-24521348

RESUMO

Pulmonary arterial hypertension (PAH) is characterized by adverse remodeling of pulmonary arteries. Although the origin of the disease and its underlying pathophysiology remain incompletely understood, inflammation has been identified as a central mediator of disease progression. Oxidative inflammatory conditions support the formation of electrophilic fatty acid nitroalkene derivatives, which exert potent anti-inflammatory effects. The current study investigated the role of 10-nitro-oleic acid (OA-NO2) in modulating the pathophysiology of PAH in mice. Mice were kept for 28 days under normoxic or hypoxic conditions, and OA-NO2 was infused subcutaneously. Right ventricular systolic pressure (RVPsys) was determined, and right ventricular and lung tissue was analyzed. The effect of OA-NO2 on cultured pulmonary artery smooth muscle cells (PASMCs) and macrophages was also investigated. Changes in RVPsys revealed increased pulmonary hypertension in mice on hypoxia, which was significantly decreased by OA-NO2 administration. Right ventricular hypertrophy and fibrosis were also attenuated by OA-NO2 treatment. The infiltration of macrophages and the generation of reactive oxygen species were elevated in lung tissue of mice on hypoxia and were diminished by OA-NO2 treatment. Moreover, OA-NO2 decreased superoxide production of activated macrophages and PASMCs in vitro. Vascular structural remodeling was also limited by OA-NO2. In support of these findings, proliferation and activation of extracellular signal-regulated kinases 1/2 in cultured PASMCs was less pronounced on application of OA-NO2.Our results show that the oleic acid nitroalkene derivative OA-NO2 attenuates hypoxia-induced pulmonary hypertension in mice. Thus, OA-NO2 represents a potential therapeutic agent for the treatment of PAH.


Assuntos
Modelos Animais de Doenças , Hipertensão Pulmonar/prevenção & controle , Hipertrofia Ventricular Direita/prevenção & controle , Hipóxia/fisiopatologia , Inflamação/prevenção & controle , Ácidos Oleicos/uso terapêutico , Animais , Western Blotting , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Cromatografia Líquida de Alta Pressão , MAP Quinases Reguladas por Sinal Extracelular/genética , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Hipertensão Pulmonar Primária Familiar , Hipertensão Pulmonar/etiologia , Hipertensão Pulmonar/patologia , Hipertrofia Ventricular Direita/etiologia , Hipertrofia Ventricular Direita/patologia , Inflamação/etiologia , Inflamação/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Miócitos de Músculo Liso/efeitos dos fármacos , Miócitos de Músculo Liso/metabolismo , Miócitos de Músculo Liso/patologia , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Superóxidos/metabolismo
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