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1.
PLoS Negl Trop Dis ; 13(7): e0007602, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31365537

RESUMO

TGF-ß involvement in Chagas disease cardiomyopathy has been clearly demonstrated. The TGF-ß signaling pathway is activated in the cardiac tissue of chronic phase patients and is associated with an increase in extracellular matrix protein expression. The aim of this study was to investigate the effect of GW788388, a selective inhibitor of TßR1/ALK5, on cardiac function in an experimental model of chronic Chagas' heart disease. To this end, C57BL/6 mice were infected with Trypanosoma cruzi (102 parasites from the Colombian strain) and treated orally with 3mg/kg GW788388 starting at 120 days post-infection (dpi), when 100% of the infected mice show cardiac damage, and following three distinct treatment schedules: i) single dose; ii) one dose per week; or iii) three doses per week during 30 days. The treatment with GW788388 improved several cardiac parameters: reduced the prolonged PR and QTc intervals, increased heart rate, and reversed sinus arrhythmia, and atrial and atrioventricular conduction disorders. At 180 dpi, 30 days after treatment interruption, the GW3x-treated group remained in a better cardiac functional condition. Further, GW788388 treatment reversed the loss of connexin-43 enriched intercellular plaques and reduced fibrosis of the cardiac tissue. Inhibition of the TGF-ß signaling pathway reduced TGF-ß/pSmad2/3, increased MMP-9 and Sca-1, reduced TIMP-1/TIMP-2/TIMP-4, and partially restored GATA-6 and Tbox-5 transcription, supporting cardiac recovery. Moreover, GW788388 administration did not modify cardiac parasite load during the infection but reduced the migration of CD3+ cells to the heart tissue. Altogether, our data suggested that the single dose schedule was not as effective as the others and treatment three times per week during 30 days seems to be the most effective strategy. The therapeutic effects of GW788388 are promising and suggest a new possibility to treat cardiac fibrosis in the chronic phase of Chagas' heart disease by TGF-ß inhibitors.


Assuntos
Benzamidas/uso terapêutico , Cardiomiopatia Chagásica/tratamento farmacológico , Coração/efeitos dos fármacos , Pirazóis/uso terapêutico , Fator de Crescimento Transformador beta/antagonistas & inibidores , Tripanossomicidas/uso terapêutico , Animais , Cardiomiopatia Chagásica/parasitologia , Cardiomiopatia Chagásica/patologia , Doença Crônica , Conexina 43/metabolismo , Modelos Animais de Doenças , Feminino , Fibrose/tratamento farmacológico , Coração/parasitologia , Sistema de Condução Cardíaco/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos C57BL , Carga Parasitária , Trypanosoma cruzi/efeitos dos fármacos
2.
PLoS Negl Trop Dis ; 13(7): e0007597, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31356587

RESUMO

CD8+ T lymphocytes play an important role in controlling infections by intracellular pathogens. Chemokines and their receptors are crucial for the migration of CD8+ T-lymphocytes, which are the main IFNγ producers and cytotoxic effectors cells. Although the participation of chemokine ligands and receptors has been largely explored in viral infection, much less is known in infection by Trypanosoma cruzi, the causative agent of Chagas disease. After T. cruzi infection, CXCR3 chemokine receptor is highly expressed on the surface of CD8+ T-lymphocytes. Here, we hypothesized that CXCR3 is a key molecule for migration of parasite-specific CD8+ T-cells towards infected tissues, where they may play their effector activities. Using a model of induction of resistance to highly susceptible A/Sn mice using an ASP2-carrying DNA/adenovirus prime-boost strategy, we showed that CXCR3 expression was upregulated on CD8+ T-cells, which selectively migrated towards its ligands CXCL9 and CXCL10. Anti-CXCR3 administration reversed the vaccine-induced resistance to T. cruzi infection in a way associated with hampered cytotoxic activity and increased proapoptotic markers on the H2KK-restricted TEWETGQI-specific CD8+ T-cells. Furthermore, CXCR3 receptor critically guided TEWETGQI-specific effector CD8+ T-cells to the infected heart tissue that express CXCL9 and CXCL10. Overall, our study pointed CXCR3 and its ligands as key molecules to drive T. cruzi-specific effector CD8+ T-cells into the infected heart tissue. The unveiling of the process driving cell migration and colonization of infected tissues by pathogen-specific effector T-cells is a crucial requirement to the development of vaccine strategies.


Assuntos
Vacinas contra Adenovirus/imunologia , Linfócitos T CD8-Positivos/imunologia , Cardiomiopatia Chagásica/imunologia , Quimiotaxia de Leucócito , Miocárdio/metabolismo , Receptores CXCR3/metabolismo , Trypanosoma cruzi/imunologia , Animais , Apoptose , Cardiomiopatia Chagásica/parasitologia , Cardiomiopatia Chagásica/prevenção & controle , Feminino , Coração/parasitologia , Ligantes , Camundongos , Camundongos Endogâmicos C57BL , Miocárdio/imunologia , Receptores CCR2/metabolismo , Baço/imunologia , Regulação para Cima , Vacinas de DNA/imunologia
3.
PLoS Negl Trop Dis ; 13(5): e0007413, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-31145733

RESUMO

Chagas disease, caused by the parasite Trypanosoma cruzi, develops into chronic Chagas' cardiomyopathy in ~30% of infected individuals, characterized by conduction disorders, arrhythmias, heart failure, and even sudden cardiac death. Current anti-parasitic treatments are plagued by significant side effects and poor efficacy in the chronic phase of disease; thus, there is a pressing need for new treatment options. A therapeutic vaccine could bolster the protective TH1-mediated immune response, thereby slowing or halting the progression of chronic Chagas' cardiomyopathy. Prior work in mice has demonstrated therapeutic efficacy of a Tc24 recombinant protein vaccine in the acute phase of Chagas disease. However, it is anticipated that humans will be vaccinated therapeutically when in the chronic phase of disease. This study investigates the therapeutic efficacy of a vaccine prototype containing recombinant protein Tc24, formulated with an emulsion containing the Toll-like receptor 4 agonist E6020 as an immunomodulatory adjuvant in a mouse model of chronic T. cruzi infection. Among outbred ICR mice vaccinated during chronic T. cruzi infection, there is a significant increase in the number of animals with undetectable systemic parasitemia (60% of vaccinated mice compared to 0% in the sham vaccine control group), and a two-fold reduction in cardiac fibrosis over the control group. The vaccinated mice produce a robust protective TH1-biased immune response to the vaccine, as demonstrated by a significant increase in antigen-specific IFNγ-production, the number of antigen-specific IFNγ-producing cells, and IgG2a antibody titers. Importantly, therapeutic vaccination significantly reduced cardiac fibrosis in chronically infected mice. This is a first study demonstrating therapeutic efficacy of the prototype Tc24 recombinant protein and E6020 stable emulsion vaccine against cardiac fibrosis in a mouse model of chronic T. cruzi infection.


Assuntos
Anticorpos Antiprotozoários/imunologia , Cardiomiopatia Chagásica/imunologia , Vacinas Protozoárias/administração & dosagem , Animais , Cardiomiopatia Chagásica/parasitologia , Cardiomiopatia Chagásica/patologia , Cardiomiopatia Chagásica/prevenção & controle , Modelos Animais de Doenças , Feminino , Fibrose , Humanos , Interferon gama/genética , Interferon gama/imunologia , Camundongos , Camundongos Endogâmicos ICR , Miocárdio/patologia , Parasitemia/imunologia , Parasitemia/parasitologia , Parasitemia/patologia , Parasitemia/prevenção & controle , Vacinas Protozoárias/imunologia , Células Th1/imunologia , Trypanosoma cruzi/imunologia , Trypanosoma cruzi/fisiologia , Vacinação
4.
Infect Immun ; 87(6)2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-30936158

RESUMO

In chronic Trypanosoma cruzi infection, the cause of Chagas disease, life-threatening inflammatory diseases develop over time in the heart, esophagus, and colon of some patients. C57BL/6 mice infected with the myotropic Colombiana strain of T. cruzi model many of the immunological and parasitological features of human infection but succumb to chronic paralyzing myositis and skeletal muscle vasculitis, not cardiomyopathy or gastrointestinal disease. Here we show that T cell depletion in the chronic phase of this model increased tissue parasitism to acute-phase levels and induced neutrophilic skeletal muscle inflammation. Conversely, after daily treatment with the trypanocide benznidazole for 8 weeks during the chronic phase, viable parasites were no longer detectable, myositis completely resolved, vasculitis was ∼80% reduced, fibrosis was reduced, and myofiber morphology normalized. After the drug was discontinued, parasitism rebounded, and immunopathology recurred. The parasite load was statistically strongly correlated with the severity of inflammation. Thus, both T cell immunity and trypanocidal pharmacotherapy suppress to very low levels, but do not cure, T. cruzi infection, which is necessary and possibly sufficient to induce crippling chronic skeletal muscle myositis and vasculitis in the model.


Assuntos
Cardiomiopatia Chagásica/parasitologia , Músculo Esquelético/parasitologia , Miosite/parasitologia , Trypanosoma cruzi/fisiologia , Vasculite/parasitologia , Animais , Cardiomiopatia Chagásica/imunologia , Modelos Animais de Doenças , Humanos , Imunidade , Camundongos , Camundongos Endogâmicos C57BL , Miosite/imunologia , Linfócitos T/imunologia , Vasculite/imunologia
5.
Acta Trop ; 189: 30-38, 2019 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-30290285

RESUMO

It is still unclear whether the progression of acute to chronic Chagas cardiomyopathy is predominantly associated with the limited efficacy of aetiological chemotherapy, or with the pharmacological resistance profiles and pathogenicity of specific Trypanosoma cruzi strains. Thus, we tested the hypothesis that parasitic load could be a limited target of aetiological chemotherapy to prevent chronic cardiomyopathy in dogs infected by different T. cruzi strains. Animals were infected with benznidazole-susceptible (Berenice-78) and -resistant (VL-10 and AAS) strains of T. cruzi. A quantitative real-time PCR strategy was developed to comparatively quantify the parasite load of the three different strains using a single standard curve. For dogs infected with the VL-10 strain, benznidazole treatment reduced cardiac parasitism during the acute phase of infection. However, similar parasite load and collagen deposition were detected in the myocardium of treated and untreated animals in the chronic phase of the infection. In animals infected with the AAS strain, benznidazole reduced parasite load, myocarditis and type III collagen deposition in the acute phase. However, increased type III collagen deposition was verified in the chronic phase. Dogs infected with the Berenice-78 strain showed a parasitological cure and no evidence of myocardial fibrosis. Parasitic load and cardiac fibrosis presented no correlation in acute or chronic phases of T. cruzi infection. Our findings in a canine model of Chagas disease suggest that parasite burden is a limited predictor for disease progression after treatment and show that benznidazole, although not inducing parasitological cure, is able to prevent total fibrosis in the early stages of infection, as well as complete prevention of cardiac damage when it eliminates parasites at the onset of infection.


Assuntos
Cardiomiopatia Chagásica/tratamento farmacológico , Cardiomiopatia Chagásica/prevenção & controle , Miocárdio/patologia , Carga Parasitária , Parasitemia/tratamento farmacológico , Trypanosoma cruzi , Doença Aguda , Animais , Cardiomiopatia Chagásica/parasitologia , Doença Crônica , Colágeno Tipo III/metabolismo , DNA de Protozoário/sangue , Modelos Animais de Doenças , Progressão da Doença , Cães , Resistência a Medicamentos , Fibrose , Coração/parasitologia , Miocárdio/metabolismo , Nitroimidazóis/uso terapêutico , Parasitemia/sangue , Parasitemia/parasitologia , Tripanossomicidas/uso terapêutico , Trypanosoma cruzi/genética
6.
Parasit Vectors ; 11(1): 611, 2018 Nov 29.
Artigo em Inglês | MEDLINE | ID: mdl-30497493

RESUMO

BACKGROUND: Trypanosoma cruzi, a hemoflagellate protozoan parasite and the etiological agent of Chagas disease (CD), exhibits great genetic and biological diversity. Infected individuals may present clinical manifestations with different levels of severity. Several hypotheses have been proposed to attempt to correlate the diversity of clinical signs and symptoms to the genetic variability of T. cruzi. This work aimed to investigate the differential expression of proteins from two distinct genetic groups of T. cruzi (discrete typing units TcI and TcII), isolated from chronically infected individuals displaying the cardiac form of CD. For this purpose, epimastigote forms of the two isolates were cultured in vitro and the cells recovered for protein extraction. Comparative two-dimensional (2D) gel electrophoreses were performed and differentially expressed spots selected for identification by mass spectrometry, followed by database searching and protein categorization. RESULTS: The 2D electrophoretic profiles revealed the complex composition of the T. cruzi extracted proteome. Protein spots were distributed along the entire pH and molecular mass ranges attesting for the integrity of the protein preparations. In total, 46 differentially expressed proteins were identified present in 40 distinct spots found in the comparative gel analyses. Of these, 16 displayed upregulation in the gel from TcI-typed parasites and 24 appeared overexpressed in the gel from TcII-typed parasites. Functional characterization of differentially expressed proteins revealed major alterations associated with stress response, lipid and amino acid metabolism in parasites of the TcII isolate, whilst those proteins upregulated in the TcI sample were primarily linked to central metabolic pathways. CONCLUSIONS: The comparative 2D-gel electrophoresis allowed detection of major differences in protein expression between two T. cruzi isolates, belonging to the TcI and TcII genotypes. Our findings suggest that patients displaying the cardiac form of the disease harbor parasites capable of exhibiting distinct proteomic profiles. This should be of relevance to disease prognosis and treatment.


Assuntos
Cardiomiopatia Chagásica/parasitologia , Proteínas de Protozoários/genética , Trypanosoma cruzi/genética , Eletroforese em Gel Bidimensional , Feminino , Variação Genética , Genótipo , Humanos , Proteômica , Proteínas de Protozoários/química , Proteínas de Protozoários/metabolismo , Trypanosoma cruzi/química , Trypanosoma cruzi/isolamento & purificação , Trypanosoma cruzi/metabolismo
7.
Circulation ; 138(12): e169-e209, 2018 09 18.
Artigo em Inglês | MEDLINE | ID: mdl-30354432

RESUMO

BACKGROUND: Chagas disease, resulting from the protozoan Trypanosoma cruzi, is an important cause of heart failure, stroke, arrhythmia, and sudden death. Traditionally regarded as a tropical disease found only in Central America and South America, Chagas disease now affects at least 300 000 residents of the United States and is growing in prevalence in other traditionally nonendemic areas. Healthcare providers and health systems outside of Latin America need to be equipped to recognize, diagnose, and treat Chagas disease and to prevent further disease transmission. METHODS AND RESULTS: The American Heart Association and the Inter-American Society of Cardiology commissioned this statement to increase global awareness among providers who may encounter patients with Chagas disease outside of traditionally endemic environments. In this document, we summarize the most updated information on diagnosis, screening, and treatment of T cruzi infection, focusing primarily on its cardiovascular aspects. This document also provides quick reference tables, highlighting salient considerations for a patient with suspected or confirmed Chagas disease. CONCLUSIONS: This statement provides a broad summary of current knowledge and practice in the diagnosis and management of Chagas cardiomyopathy. It is our intent that this document will serve to increase the recognition of Chagas cardiomyopathy in low-prevalence areas and to improve care for patients with Chagas heart disease around the world.


Assuntos
Cardiomiopatia Chagásica/terapia , Tripanossomicidas/uso terapêutico , Trypanosoma cruzi/efeitos dos fármacos , American Heart Association , Cardiomiopatia Chagásica/diagnóstico , Cardiomiopatia Chagásica/epidemiologia , Cardiomiopatia Chagásica/parasitologia , Humanos , Valor Preditivo dos Testes , Prevalência , Fatores de Risco , Resultado do Tratamento , Tripanossomicidas/efeitos adversos , Trypanosoma cruzi/isolamento & purificação , Estados Unidos
8.
Transpl Infect Dis ; 20(6): e12996, 2018 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30204269

RESUMO

BACKGROUND: Heart transplantation has been shown to be a safe and effective intervention for progressive cardiomyopathy from chronic Chagas disease. However, in the presence of the immunosuppression required for heart transplantation, the likelihood of Chagas disease reactivation is significant. Reactivation may cause myocarditis resulting in allograft dysfunction and the rapid onset of congestive heart failure. Reactivation rates have been well documented in Latin America; however, there is a paucity of data regarding the risk in non-endemic countries. METHODS: We present our experience with 31 patients with chronic Chagas disease who underwent orthotopic heart transplantation in the United States from 2012 to 2016. Patients were monitored following a standard schedule. RESULTS: Of the 31 patients, 19 (61%) developed evidence of reactivation. Among the 19 patients, a majority (95%) were identified by laboratory monitoring using polymerase chain reaction testing. One patient was identified after the onset of clinical symptoms of reactivation. All subjects with evidence of reactivation were alive at follow-up (median: 60 weeks). CONCLUSIONS: Transplant programs in the United States are encouraged to implement a monitoring program for heart transplant recipients with Chagas disease. Our experience using a preemptive approach of monitoring for Chagas disease reactivation was effective at identifying reactivation before symptoms developed.


Assuntos
Cardiomiopatia Chagásica/cirurgia , Insuficiência Cardíaca/cirurgia , Transplante de Coração/efeitos adversos , Imunossupressão/efeitos adversos , Trypanosoma cruzi/isolamento & purificação , Adulto , Idoso , Aloenxertos/parasitologia , Aloenxertos/patologia , Cardiomiopatia Chagásica/epidemiologia , Cardiomiopatia Chagásica/parasitologia , Cardiomiopatia Chagásica/patologia , Feminino , Seguimentos , Coração/parasitologia , Insuficiência Cardíaca/epidemiologia , Insuficiência Cardíaca/parasitologia , Insuficiência Cardíaca/patologia , Humanos , Imunossupressão/métodos , Masculino , Pessoa de Meia-Idade , Miocárdio/patologia , Recidiva , Fatores de Risco , Estados Unidos/epidemiologia
9.
Trials ; 19(1): 507, 2018 Sep 19.
Artigo em Inglês | MEDLINE | ID: mdl-30231899

RESUMO

Several studies evaluating clinical forms of chronic Chagas disease show that about one-third of patients present cardiac involvement. Heart failure, sudden death and cardioembolic stroke are the main mechanisms of death in Chagas heart disease. The impact of specific etiologic treatment on the prognosis of patients with chronic Chagas heart disease is very limited regardless of the presence or absence of heart failure. Patients with symptomatic Chagas heart disease present serum selenium (Se) levels lower than patients without Chagas heart disease. Moreover, Se supplementation in animal models showed promising results. The aim of this trial is to estimate the effect of Se treatment on prevention of heart disease progression in patients with Chagas cardiomyopathy. However, we had to introduce some protocol modifications in order to keep trial feasibility, as follows: the primary outcome was restricted to left ventricular ejection fraction as a continuous variable, excluding disease progression; the follow-up period was decreased from 5 years to 1 year, an adjustment that might increase the participation rate of our study; the superior age limit was increased from 65 to 75 years; and diabetes mellitus was no longer considered an exclusion criterion. All of these protocol modifications were extensively debated by the research team enrolled in the design, recruitment and conduction of the clinical trial to guarantee a high scientific quality. TRIAL REGISTRATION: Clinical Trials.gov, NCT00875173 . Registered on 20 October 2008.


Assuntos
Cardiomiopatia Chagásica/tratamento farmacológico , Suplementos Nutricionais , Selenito de Sódio/uso terapêutico , Adolescente , Adulto , Idoso , Cardiomiopatia Chagásica/diagnóstico , Cardiomiopatia Chagásica/parasitologia , Cardiomiopatia Chagásica/fisiopatologia , Doença Crônica , Suplementos Nutricionais/efeitos adversos , Progressão da Doença , Método Duplo-Cego , Determinação de Ponto Final , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Seleção de Pacientes , Ensaios Clínicos Controlados Aleatórios como Assunto , Selenito de Sódio/efeitos adversos , Volume Sistólico/efeitos dos fármacos , Fatores de Tempo , Resultado do Tratamento , Função Ventricular Esquerda/efeitos dos fármacos , Adulto Jovem
10.
Free Radic Biol Med ; 129: 227-236, 2018 12.
Artigo em Inglês | MEDLINE | ID: mdl-30248443

RESUMO

Elderly organisms are more susceptible to infectious diseases. However, the impact of aging on antiparasitic mechanisms, especially the nitric oxide pathway, is poorly understood. Using an integrated in vivo and in vitro model, we compared the severity of Trypanosoma cruzi infection in young and elderly (8 or 72 weeks old) mice. Forty C57BL/6 mice were randomized into four groups: Y-inf, young infected; Yn-inf, young uninfected; A-inf, aged infected; An-inf, aged uninfected. Parasitemia was measured daily, and animals were euthanized after 15 days of infection. Trypanosoma cruzi-induced inflammatory processes were analyzed in blood and heart samples, as well as in bone marrow-derived macrophages (BMDMs) co-cultured with splenocytes isolated from young or elderly mice. Our results indicated upregulated IgG2b and IL-17 production in elderly animals, which was not sufficient to reduce parasitemia, parasitic load and myocarditis to levels observed in young animals. The higher susceptibility of elderly mice to T. cruzi infection was accompanied by reduced cardiac inducible nitric oxide synthase (iNOS) gene expression, nitric oxide (NO) and IFN-γ levels, as well as an antagonistic upregulation of arginase-1 expression and arginase activity. The same responses were observed when BMDMs co-cultured with splenocytes from elderly mice were stimulated with T. cruzi antigens. Our findings indicate that elderly mice were more susceptible to T. cruzi infection, which was potentially related to an attenuated response to antigenic stimulation, inhibition of iNOS gene expression and NO production, and antagonistic upregulation of arginase gene expression and activity, which created favorable conditions for heart parasitism and myocarditis development.


Assuntos
Envelhecimento/genética , Arginase/genética , Cardiomiopatia Chagásica/genética , Doença de Chagas/genética , Óxido Nítrico Sintase Tipo II/genética , Parasitemia/genética , Trypanosoma cruzi/patogenicidade , Envelhecimento/imunologia , Animais , Antígenos de Protozoários/farmacologia , Arginase/sangue , Cardiomiopatia Chagásica/imunologia , Cardiomiopatia Chagásica/parasitologia , Doença de Chagas/imunologia , Doença de Chagas/parasitologia , Técnicas de Cocultura , Regulação da Expressão Gênica , Coração/parasitologia , Interações Hospedeiro-Patógeno/genética , Interações Hospedeiro-Patógeno/imunologia , Imunoglobulina G/sangue , Imunoglobulina G/genética , Interferon gama/sangue , Interferon gama/genética , Interleucina-10/sangue , Interleucina-10/genética , Interleucina-17/sangue , Interleucina-17/genética , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Macrófagos/parasitologia , Camundongos , Camundongos Endogâmicos C57BL , Óxido Nítrico Sintase Tipo II/sangue , Parasitemia/imunologia , Índice de Gravidade de Doença , Transdução de Sinais , Linfócitos T/efeitos dos fármacos , Linfócitos T/imunologia , Linfócitos T/parasitologia , Trypanosoma cruzi/imunologia
12.
PLoS Negl Trop Dis ; 12(7): e0006687, 2018 07.
Artigo em Inglês | MEDLINE | ID: mdl-30044789

RESUMO

In this study, we have investigated the effects of manganese superoxide dismutase (SOD2 or MnSOD) deficiency on mitochondrial function and oxidative stress during Chagas disease. For this, C57BL/6 wild type (WT) and MnSOD+/- mice were infected with Trypanosoma cruzi (Tc), and evaluated at 150 days' post-infection that corresponded to chronic disease phase. Genetic deletion of SOD2 decreased the expression and activity of MnSOD, but it had no effect on the expression of other members of the SOD family. The myocardial expression and activity of MnSOD were significantly decreased in chronically infected WT mice, and it was further worsened in MnSOD+/- mice. Chronic T. cruzi infection led to a decline in mitochondrial complex I and complex II driven, ADP-coupled respiration and ATP synthesis in the myocardium of WT mice. The baseline oxidative phosphorylation (OXPHOS) capacity in MnSOD+/- mice was decreased, and it had an additive effect on mitochondrial dysregulation of ATP synthesis capacity in chagasic myocardium. Further, MnSOD deficiency exacerbated the mitochondrial rate of reactive oxygen species (ROS) production and myocardial oxidative stress (H2O2, protein carbonyls, malondialdehyde, and 4-hydroxynonenal) in Chagas disease. Peripheral and myocardial parasite burden and inflammatory response (myeloperoxidase, IL-6, lactate dehydrogenase, inflammatory infiltrate) were increased in all chagasic WT and MnSOD+/- mice. We conclude that MnSOD deficiency exacerbates the loss in mitochondrial function and OXPHOS capacity and enhances the myocardial oxidative damage in chagasic cardiomyopathy. Mitochondria targeted, small molecule mitigators of MnSOD deficiency will offer potential benefits in averting the mitochondrial dysfunction and chronic oxidative stress in Chagas disease.


Assuntos
Cardiomiopatia Chagásica/enzimologia , Mitocôndrias/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Superóxido Dismutase/deficiência , Animais , Cardiomiopatia Chagásica/genética , Cardiomiopatia Chagásica/metabolismo , Cardiomiopatia Chagásica/parasitologia , Feminino , Humanos , Peróxido de Hidrogênio/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mitocôndrias/genética , Miocárdio/metabolismo , Fosforilação Oxidativa , Estresse Oxidativo , Superóxido Dismutase/genética , Trypanosoma cruzi/fisiologia
13.
PLoS Negl Trop Dis ; 12(7): e0006589, 2018 07.
Artigo em Inglês | MEDLINE | ID: mdl-30044791

RESUMO

Chronic chagasic cardiomyopathy (CCC) is observed in 30% to 50% of the individuals infected by Trypanosoma cruzi and heart failure is the important cause of death among patients in the chronic phase of Chagas disease. Although some studies have elucidated the role of adaptive immune responses involving T and B lymphocytes in cardiac pathogenesis, the role of innate immunity receptors such as Toll-like receptors (TLRs) and Nod-like receptors (NLRs) in CCC pathophysiology has not yet been determined. In this study, we evaluated the association among innate immune receptors (TLR1-9 and nucleotide-binding domain-like receptor protein 3/NLRP3), its adapter molecules (Myd88, TRIF, ASC and caspase-1) and cytokines (IL-1ß, IL-6, IL-12, IL-18, IL-23, TNF-α, and IFN-ß) with clinical manifestation, digestive and cardiac function in patients with different clinical forms of chronic Chagas disease. The TLR8 mRNA expression levels were enhanced in the peripheral blood mononuclear cells (PBMC) from digestive and cardiodigestive patients compared to indeterminate and cardiac patients. Furthermore, mRNA expression of IFN-ß (cytokine produced after TLR8 activation) was higher in digestive and cardiodigestive patients when compared to indeterminate. Moreover, there was a positive correlation between TLR8 and IFN-ß mRNA expression with sigmoid and rectum size. Cardiac and cardiodigestive patients presented higher TLR2, IL-12 and TNF-α mRNA expression than indeterminate and digestive patients. Moreover, cardiac patients also expressed higher levels of NLRP3, ASC and IL-1ß mRNAs than indeterminate patients. In addition, we showed a negative correlation among TLR2, IL-1ß, IL-12 and TNF-α levels with left ventricular ejection fraction, and positive correlation between NLRP3 with cardiothoracic index, and TLR2, IL-1ß and IL-12 with left ventricular mass index. Together, our data suggest that high expression of innate immune receptors in cardiac and digestive patients may induce an enhancement of cytokine expression and participate of cardiac and digestive dysfunction.


Assuntos
Cardiomiopatia Chagásica/imunologia , Doenças do Sistema Digestório/imunologia , Proteína 3 que Contém Domínio de Pirina da Família NLR/imunologia , Proteínas NLR/imunologia , Adulto , Idoso , Caspase 1/genética , Caspase 1/imunologia , Cardiomiopatia Chagásica/genética , Cardiomiopatia Chagásica/parasitologia , Doenças do Sistema Digestório/genética , Doenças do Sistema Digestório/parasitologia , Feminino , Humanos , Interleucina-12/genética , Interleucina-12/imunologia , Interleucina-1beta/genética , Interleucina-1beta/imunologia , Masculino , Pessoa de Meia-Idade , Proteína 3 que Contém Domínio de Pirina da Família NLR/genética , Proteínas NLR/genética , Trypanosoma cruzi/fisiologia
14.
Mem Inst Oswaldo Cruz ; 113(9): e180171, 2018 Jul 16.
Artigo em Inglês | MEDLINE | ID: mdl-30020318

RESUMO

BACKGROUND: The anti-inflammatory and cardioprotective properties of curcumin (Cur), a natural polyphenolic flavonoid isolated from the rhizomes of Curcuma longa, are increasingly considered to have beneficial effects on the progression of Chagas heart disease, caused by the protozoan parasite Trypanosoma cruzi. OBJECTIVE: To evaluate the effects of oral therapy with Cur on T. cruzi-mediated cardiovasculopathy in acutely infected mice and analyse the in vitro response of parasite-infected human microvascular endothelial cells treated with this phytochemical. METHODS: Inflammation of heart vessels from Cur-treated and untreated infected mice were analysed by histology, with benznidazole (Bz) as the reference compound. Parasitaemia was monitored by the direct method. Capillary permeability was visualised by Evans-blue assay. Myocardial ET-1, IL-6, and TNF-α mRNA expressions were measured by quantitative reverse transcription polymerase chain reaction (qRT-PCR). Microvascular endothelial HMEC-1 cells were infected in vitro with or without addition of Cur or Bz. Induction of the Ca2+/NFAT pathway was assessed by fluorometry, immunoblotting, and reporter assay. FINDINGS: Oral Cur therapy of recently infected mice reduced inflammatory cell infiltration of myocardial arteries without lowering parasite levels. Compared to that of the phosphate-buffered saline-receiving group, hearts from Cur-treated mice showed significantly decreased vessel inflammation scores (p < 0.001), vascular permeabilities (p < 0.001), and levels of IL-6/TNF-α (p < 0.01) and ET-1 (p < 0.05) mRNA. Moreover, Cur significantly (p < 0.05 for transcript; p < 0.01 for peptide) downregulated ET-1 secretion from infected HMEC-1 cells. Remarkably, Cur addition significantly (p < 0.05 at 27.0 µM) interfered with T. cruzi-dependent activation of the Ca2+/NFATc1 signalling pathway that promotes generation of inflammatory agents in HMEC-1 cells. CONCLUSIONS: Oral treatment with Cur dampens cardiovasculopathy in acute Chagas mice. Cur impairs the Ca2+/NFATc1-regulated release of ET-1 from T. cruzi-infected vascular endothelium. These findings identify new perspectives for exploring the potential of Cur-based interventions to ameliorate Chagas heart disease.


Assuntos
Anti-Inflamatórios não Esteroides/farmacologia , Cardiomiopatia Chagásica/tratamento farmacológico , Curcumina/farmacologia , Endotelina-1/efeitos dos fármacos , Fatores de Transcrição NFATC/efeitos dos fármacos , Doença Aguda , Animais , Western Blotting , Permeabilidade Capilar/efeitos dos fármacos , Células Cultivadas , Cardiomiopatia Chagásica/metabolismo , Cardiomiopatia Chagásica/parasitologia , Progressão da Doença , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/parasitologia , Endotelina-1/análise , Endotelina-1/metabolismo , Endotélio Vascular/efeitos dos fármacos , Endotélio Vascular/parasitologia , Ensaio de Imunoadsorção Enzimática , Corantes Fluorescentes , Interleucina-6/sangue , Masculino , Camundongos Endogâmicos C57BL , Fatores de Transcrição NFATC/análise , Fatores de Transcrição NFATC/metabolismo , Reprodutibilidade dos Testes , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fatores de Tempo , Trypanosoma cruzi/efeitos dos fármacos , Fator de Necrose Tumoral alfa/sangue
15.
PLoS Negl Trop Dis ; 12(6): e0006567, 2018 06.
Artigo em Inglês | MEDLINE | ID: mdl-29897909

RESUMO

BACKGROUND: Chagas disease (CD) is a major public health concern in Latin America and a potentially serious emerging threat in non-endemic countries. Although the association between CD and cardiac abnormalities is widely reported, study design diversity, sample size and quality challenge the information, calling for its update and synthesis, which would be very useful and relevant for physicians in non-endemic countries where health care implications of CD are real and neglected. We performed to systematically review and meta-analyze population-based studies that compared prevalence of overall and specific ECG abnormalities between CD and non-CD participants in the general population. METHODS: Six databases (EMBASE, Ovid Medline, Web of Science, Cochrane Central, Google Scholar and Lilacs) were searched systematically. Observational studies were included. Odds ratios (OR) were computed using random-effects model. RESULTS: Forty-nine studies were selected, including 34,023(12,276 CD and 21,747 non-CD). Prevalence of overall ECG abnormalities was higher in participants with CD (40.1%; 95%CIs=39.2-41.0) compared to non-CD (24.1%; 95%CIs=23.5-24.7) (OR=2.78; 95%CIs=2.37-3.26). Among specific ECG abnormalities, prevalence of complete right bundle branch block (RBBB) (OR=4.60; 95%CIs=2.97-7.11), left anterior fascicular block (LAFB) (OR=1.60; 95%CIs=1.21-2.13), combination of complete RBBB/LAFB (OR=3.34; 95%CIs=1.76-6.35), first-degree atrioventricular block (A-V B) (OR=1.71; 95%CIs=1.25-2.33), atrial fibrillation (AF) or flutter (OR=2.11; 95%CIs=1.40-3.19) and ventricular extrasystoles (VE) (OR=1.62; 95%CIs=1.14-2.30) was higher in CD compared to non-CD participants. CONCLUSIONS: This systematic review and meta-analysis provides an update and synthesis in this field. This research of observational studies indicates a significant excess in prevalence of ECG abnormalities (40.1%) related to T. cruzi infection in the general population from Chagas endemic regions, being the most common ventricular (RBBB and LAFB), and A-V B (first-degree) node conduction abnormalities as well as arrhythmias (AF or flutter and VE). Also, prevalence of ECG alterations in children was similar to that in adults and suggests earlier onset of cardiac disease.


Assuntos
Cardiomiopatia Chagásica/fisiopatologia , Eletrocardiografia , Sistema de Condução Cardíaco/fisiopatologia , Adulto , Arritmias Cardíacas/parasitologia , Arritmias Cardíacas/fisiopatologia , Cardiomiopatia Chagásica/parasitologia , Doença de Chagas/complicações , Doença de Chagas/parasitologia , Criança , Feminino , Humanos , Masculino , Estudos Observacionais como Assunto , Razão de Chances , Prevalência
16.
Vector Borne Zoonotic Dis ; 18(8): 417-423, 2018 08.
Artigo em Inglês | MEDLINE | ID: mdl-29768103

RESUMO

Chagas disease is a lingering Public Health problem in Latin America with ∼5.7 million people infected with Trypanosoma cruzi. Transmission is still taking place in most countries of the Americas, including the United States. Dogs are frequently infected with T. cruzi and its high infection prevalence is associated with increased risk of Chagas disease in humans. The city of Mérida in the Yucatan peninsula is endemic for Chagas disease and canines are frequently infected with T. cruzi. The objective of this study was to evaluate the performance of a qualitative point of care (POC) molecular test (RPA-LF, recombinase polymerase amplification-lateral flow) developed in our laboratory for identifying infected dogs. We used retrospective samples of dogs that came for consultation because of cardiac alterations and proved to be infected with T. cruzi as determined by enzyme-linked immunosorbent assay (ELISA), Western blot, and quantitative PCR (qPCR). The analytical sensitivity indicated that RPA-LF amplified T. cruzi DNA in samples containing almost equal to one to two parasites per reaction. Serial twofold dilutions of T. cruzi epimastigotes showed that the test had 95% (19/20) repeatability at concentrations of two parasites per reaction. The test showed no cross reactivity with human DNA or other protozoan parasites (Trypanosoma rangeli, Leishmania spp., and Plasmodium spp.). RPA-LF had the capacity to amplify all discrete typing units (DTUs I-VI) of T. cruzi that circulate in domestic or extradomestic environments. The RPA-LF had 93.2% (95% confidence interval 87.2-98.1) sensitivity and excellent agreement with qPCR used as gold standard (Cohen's Kappa test = 0.963). ELISA was positive in 96.6% (85/88) of dogs, which together with the molecular tests confirmed the frequent contact with infected triatomine bugs in the city of Mérida. These preliminary results on the diagnostic efficacy of the RPA-LF deserve further large-scale field testing of this POC test for T. cruzi infection in endemic areas.


Assuntos
Cardiomiopatia Chagásica/veterinária , Doenças do Cão/parasitologia , Reação em Cadeia da Polimerase/veterinária , Trypanosoma cruzi/genética , Animais , Cardiomiopatia Chagásica/epidemiologia , Cardiomiopatia Chagásica/parasitologia , DNA de Cinetoplasto/genética , DNA de Protozoário/genética , Doenças do Cão/patologia , Cães , Ensaio de Imunoadsorção Enzimática/veterinária , México/epidemiologia , Reação em Cadeia da Polimerase/métodos , Reação em Cadeia da Polimerase em Tempo Real , Estudos Retrospectivos
17.
Acta Trop ; 185: 280-284, 2018 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-29746871

RESUMO

Chagas disease is a major public health problem in Latin America and has spread to other countries due to immigration of infected persons. 10-30% of patients with chronic Chagas disease will develop cardiomyopathy. Chagas cardiomyopathy is the worst form of the disease, due to its high morbidity and mortality. Because of its prognostic value and adequate medical monitoring, it is very important to identify infected people who could develop Chagas cardiomyopathy. The aim of this study was to determine if discrete typing units (DTUs) of Trypanosoma cruzi are related to the presence of heart disease in patients with chronic Chagas disease. A total of 86 untreated patients, 41 with cardiomyopathy and 45 without heart involvement were submitted to clinical study. Electrocardiograms and echocardiograms were performed on the group of cardiopaths, in which all important known causes of cardiomyopathy were discarded. Sinus bradycardia and prolonged QTc interval were the most frequent electrocardiographic alterations and patients were classified in group I (46%) and group II (54%) of New York Hearth Association. In all cases real-time PCR genotyping assays were performed. In the group with cardiomyopathy, the most frequent DTU was TcI (56.1%), followed by TcII (19.5%). Mixed infections TcI + TcII were observed in 7.3% of the patients. In the group without cardiac pathologies, TcI and TcII were found at similar rates (28.9 and 31.1%, respectively) and mixed infections TcI + TcII in 17.8% of the cases. TcIII and TcIV were not detected in any sample. Taken together, our data indicate that chronic Chagas cardiomyopathy in Chile can be caused by strains belonging to TcI and TcII.


Assuntos
Cardiomiopatia Chagásica/parasitologia , Genótipo , Tipagem Molecular , Trypanosoma cruzi/genética , Adulto , Idoso , Doença de Chagas/parasitologia , Chile , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase em Tempo Real , Adulto Jovem
18.
Nat Commun ; 9(1): 1513, 2018 04 17.
Artigo em Inglês | MEDLINE | ID: mdl-29666415

RESUMO

Chagas disease is caused by infection with the protozoan Trypanosoma cruzi (T. cruzi) and is an important cause of severe inflammatory heart disease. However, the mechanisms driving Chagas disease cardiomyopathy have not been completely elucidated. Here, we show that the canonical PI3Kγ pathway is upregulated in both human chagasic hearts and hearts of acutely infected mice. PI3Kγ-deficient mice and mutant mice carrying catalytically inactive PI3Kγ are more susceptible to T. cruzi infection. The canonical PI3Kγ signaling in myeloid cells is essential to restrict T. cruzi heart parasitism and ultimately to avoid myocarditis, heart damage, and death of mice. Furthermore, high PIK3CG expression correlates with low parasitism in human Chagas' hearts. In conclusion, these results indicate an essential role of the canonical PI3Kγ signaling pathway in the control of T. cruzi infection, providing further insight into the molecular mechanisms involved in the pathophysiology of chagasic heart disease.


Assuntos
Cardiomiopatia Chagásica/imunologia , Classe Ib de Fosfatidilinositol 3-Quinase/metabolismo , Transdução de Sinais/imunologia , Trypanosoma cruzi/imunologia , Adulto , Animais , Biópsia , Linhagem Celular , Cardiomiopatia Chagásica/parasitologia , Cardiomiopatia Chagásica/patologia , Classe Ib de Fosfatidilinositol 3-Quinase/genética , Modelos Animais de Doenças , Feminino , Coração/parasitologia , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Pessoa de Meia-Idade , Células Mieloides/imunologia , Células Mieloides/metabolismo , Miocárdio/imunologia , Miocárdio/patologia , Quinoxalinas/farmacologia , Tiazolidinedionas/farmacologia , Trypanosoma cruzi/patogenicidade , Regulação para Cima
19.
Front Immunol ; 9: 202, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29503646

RESUMO

Metabolism provides substrates for reactive oxygen species (ROS) and nitric oxide (NO) generation, which are a part of the macrophage (Mφ) anti-microbial response. Mφs infected with Trypanosoma cruzi (Tc) produce insufficient levels of oxidative species and lower levels of glycolysis compared to classical Mφs. How Mφs fail to elicit a potent ROS/NO response during infection and its link to glycolysis is unknown. Herein, we evaluated for ROS, NO, and cytokine production in the presence of metabolic modulators of glycolysis and the Krebs cycle. Metabolic status was analyzed by Seahorse Flux Analyzer and mass spectrometry and validated by RNAi. Tc infection of RAW264.7 or bone marrow-derived Mφs elicited a substantial increase in peroxisome proliferator-activated receptor (PPAR)-α expression and pro-inflammatory cytokine release, and moderate levels of ROS/NO by 18 h. Interferon (IFN)-γ addition enhanced the Tc-induced ROS/NO release and shut down mitochondrial respiration to the levels noted in classical Mφs. Inhibition of PPAR-α attenuated the ROS/NO response and was insufficient for complete metabolic shift. Deprivation of glucose and inhibition of pyruvate transport showed that Krebs cycle and glycolysis support ROS/NO generation in Tc + IFN-γ stimulated Mφs. Metabolic profiling and RNAi studies showed that glycolysis-pentose phosphate pathway (PPP) at 6-phosphogluconate dehydrogenase was essential for ROS/NO response and control of parasite replication in Mφ. We conclude that IFN-γ, but not inhibition of PPAR-α, supports metabolic upregulation of glycolytic-PPP for eliciting potent ROS/NO response in Tc-infected Mφs. Chemical analogs enhancing the glucose-PPP will be beneficial in controlling Tc replication and dissemination by Mφs.


Assuntos
Cardiomiopatia Chagásica/imunologia , Interações Hospedeiro-Parasita/imunologia , Macrófagos/imunologia , Via de Pentose Fosfato/imunologia , Trypanosoma cruzi/imunologia , Animais , Cardiomiopatia Chagásica/parasitologia , Modelos Animais de Doenças , Humanos , Interferon gama/imunologia , Macrófagos/parasitologia , Camundongos , Camundongos Knockout , Óxido Nítrico/imunologia , Óxido Nítrico/metabolismo , PPAR alfa/genética , PPAR alfa/imunologia , Cultura Primária de Células , Células RAW 264.7 , Espécies Reativas de Oxigênio/imunologia , Espécies Reativas de Oxigênio/metabolismo , Regulação para Cima
20.
Parasit Vectors ; 11(1): 72, 2018 01 30.
Artigo em Inglês | MEDLINE | ID: mdl-29382361

RESUMO

BACKGROUND: Cardiac fibrosis is a consequence of chronic chagasic cardiomyopathy (CCC). In other cardiovascular diseases, the protagonist role of fibroblasts in cardiac fibrosis is well established. However, the role of cardiac fibroblasts (CFs) in fibrosis during the CCC is not clear. Here, our aim was to investigate the effect of Trypanosoma cruzi, the etiological agent of Chagas disease on CFs activation. METHODS: Cardiac fibroblasts were purified from primary cultures of mouse embryo cardiac cells. After two passages, cells were infected with T. cruzi (Y strain) and analyzed at different times for determination of infectivity, activation and production of extracellular matrix components (fibronectin, laminin and collagen IV) by immunofluorescence and western blot. RESULTS: At second passage, cultures were enriched in CFs (95% of fibroblasts and 5% of cardiomyocytes), as revealed by presence of alpha-smooth muscle actin (α-SMA) and discoidin domain receptor 2 (DDR2) and absence of sarcomeric tropomyosin (ST) protein expression. Trypanosoma cruzi infection induced fibroblast-myofibroblast transition, with increased expression of α-SMA after 6 and 24 h post-infection (hpi). Fibronectin was increased at 6, 24 and 48 hpi, laminin was increased at 6 and 24 hpi and collagen IV was increased at 6 hpi. CONCLUSIONS: Our results showed that T. cruzi activates CFs, inducing activation and exacerbates ECM production. Furthermore, our data raise the possibility of the involvement of CFs in heart fibrosis during Chagas disease.


Assuntos
Proteínas da Matriz Extracelular/genética , Fibroblastos/parasitologia , Miofibroblastos/parasitologia , Trypanosoma cruzi/fisiologia , Animais , Western Blotting , Células Cultivadas , Cardiomiopatia Chagásica/parasitologia , Cardiomiopatia Chagásica/fisiopatologia , Colágeno/genética , Fibroblastos/fisiologia , Fibronectinas/genética , Imunofluorescência , Laminina/genética , Camundongos , Miofibroblastos/fisiologia
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