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1.
Eur Heart J ; 44(44): 4696-4712, 2023 Nov 21.
Artigo em Inglês | MEDLINE | ID: mdl-37944136

RESUMO

BACKGROUND AND AIMS: Developing novel therapies to battle the global public health burden of heart failure remains challenging. This study investigates the underlying mechanisms and potential treatment for 4-hydroxynonenal (4-HNE) deleterious effects in heart failure. METHODS: Biochemical, functional, and histochemical measurements were applied to identify 4-HNE adducts in rat and human failing hearts. In vitro studies were performed to validate 4-HNE targets. RESULTS: 4-HNE, a reactive aldehyde by-product of mitochondrial dysfunction in heart failure, covalently inhibits Dicer, an RNase III endonuclease essential for microRNA (miRNA) biogenesis. 4-HNE inhibition of Dicer impairs miRNA processing. Mechanistically, 4-HNE binds to recombinant human Dicer through an intermolecular interaction that disrupts both activity and stability of Dicer in a concentration- and time-dependent manner. Dithiothreitol neutralization of 4-HNE or replacing 4-HNE-targeted residues in Dicer prevents 4-HNE inhibition of Dicer in vitro. Interestingly, end-stage human failing hearts from three different heart failure aetiologies display defective 4-HNE clearance, decreased Dicer activity, and miRNA biogenesis impairment. Notably, boosting 4-HNE clearance through pharmacological re-activation of mitochondrial aldehyde dehydrogenase 2 (ALDH2) using Alda-1 or its improved orally bioavailable derivative AD-9308 restores Dicer activity. ALDH2 is a major enzyme responsible for 4-HNE removal. Importantly, this response is accompanied by improved miRNA maturation and cardiac function/remodelling in a pre-clinical model of heart failure. CONCLUSIONS: 4-HNE inhibition of Dicer directly impairs miRNA biogenesis in heart failure. Strikingly, decreasing cardiac 4-HNE levels through pharmacological ALDH2 activation is sufficient to re-establish Dicer activity and miRNA biogenesis; thereby representing potential treatment for patients with heart failure.


Assuntos
Insuficiência Cardíaca , MicroRNAs , Humanos , Ratos , Animais , MicroRNAs/metabolismo , Ribonuclease III/genética , Ribonuclease III/metabolismo , Aldeídos/metabolismo , Aldeídos/farmacologia , Processamento de Proteína Pós-Traducional , Aldeído-Desidrogenase Mitocondrial/genética
2.
Int J Mol Sci ; 25(1)2023 Dec 19.
Artigo em Inglês | MEDLINE | ID: mdl-38203212

RESUMO

Parasitemia and inflammatory markers are cross-sectionally associated with chronic Chagas cardiomyopathy (CCC) among patients with Trypanosoma cruzi. However, the prospective association of the parasite load and host immune response-related characteristics with CCC (that is, progressors) among T. cruzi seropositive individuals has only been partially defined. In a cohort of T. cruzi seropositive patients in Montes Claros and São Paulo, Brazil who were followed over 10 years, we identified the association of a baseline T. cruzi parasite load and systemic markers of inflammation with a decline in cardiac function and/or the presence of cardiac congestion 10 years later. The progressors (n = 21) were individuals with a significant decline in the left ventricular ejection fraction and/or elevated markers of cardiac congestion after 10 years. The controls (n = 31) had normal markers of cardiac function and congestion at the baseline and at the follow-up. They were matched with the progressors on age, sex, and genetic ancestry. The progressors had higher mean parasite loads at the baseline than the controls (18.3 vs. 0.605 DNA parasite equivalents/20 mL, p < 0.05). Of the 384 inflammation-related proteins analyzed, 47 differed significantly at a false discovery rate- (FDR-) corrected p < 0.05 between the groups. There were 44 of these 47 proteins that were significantly higher in the controls compared to in the progressors, including the immune activation markers CCL21, CXCL12, and HCLS1 and several of the tumor necrosis factor superfamily of proteins. Among the individuals who were seropositive for T. cruzi at the baseline and who were followed over 10 years, those with incident CCC at the 10-year marker had a comparatively higher baseline of T. cruzi parasitemia and lower baseline markers of immune activation and chemotaxis. These findings generate the hypothesis that the early impairment of pathogen-killing immune responses predisposes individuals to CCC, which merits further study.


Assuntos
Doença de Chagas , Parasitos , Trypanosoma cruzi , Humanos , Animais , Trypanosoma cruzi/genética , Brasil/epidemiologia , Parasitemia , Volume Sistólico , Função Ventricular Esquerda , DNA , Inflamação
3.
Antimicrob Agents Chemother ; 66(11): e0028422, 2022 11 15.
Artigo em Inglês | MEDLINE | ID: mdl-36314800

RESUMO

Drug combinations and drug repurposing have emerged as promising strategies to develop novel treatments for infectious diseases, including Chagas disease. In this study, we aimed to investigate whether the repurposed drugs chloroquine (CQ) and colchicine (COL), known to inhibit Trypanosoma cruzi infection in host cells, could boost the anti-T. cruzi effect of the trypanocidal drug benznidazole (BZN), increasing its therapeutic efficacy while reducing the dose needed to eradicate the parasite. The combination of BZN and COL exhibited cytotoxicity to infected cells and low antiparasitic activity. Conversely, a combination of BZN and CQ significantly reduced T. cruzi infection in vitro, with no apparent cytotoxicity. This effect seemed to be consistent across different cell lines and against both the partially BZN-resistant Y and the highly BZN-resistant Colombiana strains. In vivo experiments in an acute murine model showed that the BZN+CQ combination was eight times more effective in reducing T. cruzi infection in the acute phase than BZN monotherapy. In summary, our results demonstrate that the concomitant administration of CQ and BZN potentiates the trypanocidal activity of BZN, leading to a reduction in the dose needed to achieve an effective response. In a translational context, it could represent a higher efficacy of treatment while also mitigating the adverse effects of high doses of BZN. Our study also reinforces the relevance of drug combination and repurposing approaches in the field of Chagas disease drug discovery.


Assuntos
Doença de Chagas , Nitroimidazóis , Tripanossomicidas , Trypanosoma cruzi , Camundongos , Animais , Reposicionamento de Medicamentos , Cloroquina/farmacologia , Cloroquina/uso terapêutico , Doença de Chagas/tratamento farmacológico , Doença de Chagas/parasitologia , Nitroimidazóis/farmacologia , Nitroimidazóis/uso terapêutico , Tripanossomicidas/farmacologia , Tripanossomicidas/uso terapêutico
4.
Immun Ageing ; 19(1): 57, 2022 Nov 16.
Artigo em Inglês | MEDLINE | ID: mdl-36384671

RESUMO

BACKGROUND: Although older adults are at a high risk of severe or critical Covid-19, there are many cases of unvaccinated centenarians who had a silent infection or recovered from mild or moderate Covid-19. We studied three Brazilian supercentenarians, older than 110 years, who survived Covid-19 in 2020 before being vaccinated. RESULTS: Despite their advanced age, humoral immune response analysis showed that these individuals displayed robust levels of IgG and neutralizing antibodies (NAbs) against SARS-CoV-2. Enrichment of plasma proteins and metabolites related to innate immune response and host defense was also observed. None presented autoantibodies (auto-Abs) to type I interferon (IFN). Furthermore, these supercentenarians do not carry rare variants in genes underlying the known inborn errors of immunity, including particular inborn errors of type I IFN. CONCLUSION: These observations suggest that their Covid-19 resilience might be a combination of their genetic background and their innate and adaptive immunity.

5.
AIDS Res Ther ; 19(1): 2, 2022 01 12.
Artigo em Inglês | MEDLINE | ID: mdl-35022035

RESUMO

BACKGROUND: We developed a personalized Monocyte-Derived Dendritic-cell Therapy (MDDCT) for HIV-infected individuals on suppressive antiretroviral treatment and evaluated HIV-specific T-cell responses. METHODS: PBMCs were obtained from 10 HIV+ individuals enrolled in trial NCT02961829. Monocytes were differentiated into DCs using IFN-α and GM-CSF. After sequencing each patient's HIV-1 Gag and determining HLA profiles, autologous Gag peptides were selected based on the predicted individual immunogenicity and used to pulse MDDCs. Three doses of the MDDCT were administered every 15 days. To assess immunogenicity, patients' cells were stimulated in vitro with autologous peptides, and intracellular IL-2, TNF, and interferon-gamma (IFN-γ) production were measured in CD4+ and CD8+ T-cells. RESULTS: The protocol of ex-vivo treatment with IFN-α and GM-CSF was able to induce maturation of MDDCs, as well as to preserve their viability for reinfusion. MDDCT administration was associated with increased expression of IL-2 in CD4+ and CD8+ T-cells at 15 and/or 30 days after the first MDDCT administration. Moreover, intracellular TNF and IFN-γ expression was significantly increased in CD4+ T-cells. The number of candidates that increased in vitro the cytokine levels in CD4+ and CD8+ T cells upon stimulation with Gag peptides from baseline to day 15 and from baseline to day 30 and day 120 after MDDCT was significant as compared to Gag unstimulated response. This was accompanied by an increasing trend in the frequency of polyfunctional T-cells over time, which was visible when considering both cells expressing two and three out of the three cytokines examined. CONCLUSIONS: MDDC had a mature profile, and this MDDCT promoted in-vitro T-cell immune responses in HIV-infected patients undergoing long-term suppressive antiretroviral treatment. Trial registration NCT02961829: (Multi Interventional Study Exploring HIV-1 Residual Replication: a Step Towards HIV-1 Eradication and Sterilizing Cure, https://www.clinicaltrials.gov/ct2/show/NCT02961829 , posted November 11th, 2016).


Assuntos
Infecções por HIV , HIV-1 , Linfócitos T CD4-Positivos , Linfócitos T CD8-Positivos , Terapia Baseada em Transplante de Células e Tecidos , Células Dendríticas , Infecções por HIV/tratamento farmacológico , Humanos
6.
Clin Infect Dis ; 73(4): 672-679, 2021 08 16.
Artigo em Inglês | MEDLINE | ID: mdl-33539531

RESUMO

BACKGROUND: Chagas disease is an infectious disease caused by the parasite Trypanosoma cruzi and is endemic from Latin American countries. The goal of our study was to identify novel genetic loci associated with chronic Chagas cardiomyopathy development in Chagas disease patients from different Latin American populations. METHODS: We performed a cross-sectional, nested case-control study including 3 sample collections from Colombia, Argentina, and Bolivia. Samples were genotyped to conduct a genome-wide association study (GWAS). These results were meta-analyzed with summary statistic data from Brazil, gathering a total of 3413 Chagas disease patients. To identify the functional impact of the associated variant and its proxies, we performed an in silico analysis of this region. RESULTS: The meta-analysis revealed a novel genome-wide statistically significant association with chronic Chagas cardiomyopathy development in rs2458298 (OR = 0.90, 95%CI = 0.87-0.94, P-value = 3.27 × 10-08), nearby the SAC3D1 gene. In addition, further in silico analyses displayed functional relationships between the associated variant and the SNX15, BAFT2, and FERMT3 genes, related to cardiovascular traits. CONCLUSIONS: Our findings support the role of the host genetic factors in the susceptibility to the development of the chronic cardiac form of this neglected disease.


Assuntos
Cardiomiopatia Chagásica , Doença de Chagas , Trypanosoma cruzi , Estudos de Casos e Controles , Cardiomiopatia Chagásica/genética , Estudos Transversais , Estudo de Associação Genômica Ampla , Humanos , Trypanosoma cruzi/genética
7.
J Clin Immunol ; 41(5): 1048-1063, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-33660144

RESUMO

Cardiomyopathies are an important cause of heart failure and sudden cardiac death. Little is known about the role of rare genetic variants in inflammatory cardiomyopathy. Chronic Chagas disease cardiomyopathy (CCC) is an inflammatory cardiomyopathy prevalent in Latin America, developing in 30% of the 6 million patients chronically infected by the protozoan Trypanosoma cruzi, while 60% remain free of heart disease (asymptomatic (ASY)). The cytokine interferon-γ and mitochondrial dysfunction are known to play a major pathogenetic role. Chagas disease provides a unique model to probe for genetic variants involved in inflammatory cardiomyopathy. METHODS: We used whole exome sequencing to study nuclear families containing multiple cases of Chagas disease. We searched for rare pathogenic variants shared by all family members with CCC but absent in infected ASY siblings and in unrelated ASY. RESULTS: We identified heterozygous, pathogenic variants linked to CCC in all tested families on 22 distinct genes, from which 20 were mitochondrial or inflammation-related - most of the latter involved in proinflammatory cytokine production. Significantly, incubation with IFN-γ on a human cardiomyocyte line treated with an inhibitor of dihydroorotate dehydrogenase brequinar (enzyme showing a loss-of-function variant in one family) markedly reduced mitochondrial membrane potential (ΔψM), indicating mitochondrial dysfunction. CONCLUSION: Mitochondrial dysfunction and inflammation may be genetically determined in CCC, driven by rare genetic variants. We hypothesize that CCC-linked genetic variants increase mitochondrial susceptibility to IFN-γ-induced damage in the myocardium, leading to the cardiomyopathy phenotype in Chagas disease. This mechanism may also be operative in other inflammatory cardiomyopathies.


Assuntos
Cardiomiopatia Chagásica/genética , Inflamação/genética , Mitocôndrias/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Predisposição Genética para Doença , Variação Genética , Humanos , Masculino , Pessoa de Meia-Idade , Sequenciamento do Exoma
8.
Metab Brain Dis ; 36(1): 193-197, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-33170418

RESUMO

Lithium activates Wnt/ß-catenin signaling leading to stabilization of free cytosolic ß-catenin. The aim of the present study is to evaluate the in vivo effect of acute and chronic lithium treatment on the expression of ß-catenin target genes, addressing its transcripts HIG2, Bcl-xL, Cyclin D1, c-myc, in cortical and hippocampal tissue from adult mice. Lithium doses were established to yield therapeutic working concentrations. In acute treatment, mice received a 300µL of a 350 mg/kg solution of LiCl by gavage, and were euthanized after 2 h, 6 h and 12 h. To determine the effect of chronic treatment, animals were continuously fed either with chow supplemented with 2 g/kg Li2CO3, or regular chow (controls), being euthanized after 30 days. All animals had access to drinking water and 0.9% saline ad libitum. After acute and chronic treatments samples of peripheral blood were obtained from the tail vein for each animal, and serum concentrations of lithium were determined. All transcripts were up-regulated in cortical and hippocampal tissues of lithium-treated mice, both under acute and chronic treatments. There was a positive correlation between serum lithium concentrations and the increment in the expression of all transcripts. This effect was observed in all time points of the acute treatment (i.e., 2, 6 and 12 hours) and also after 30 days. We conclude that Wnt/ß-catenin transcriptional response (HIG2, Bcl-xL, Cyclin D1 and c-myc) is up-regulated in the mouse brain in response to acute and chronic lithium treatment at therapeutic concentrations.


Assuntos
Antimaníacos/farmacologia , Córtex Cerebral/efeitos dos fármacos , Hipocampo/efeitos dos fármacos , Cloreto de Lítio/farmacologia , Proteínas Wnt/metabolismo , beta Catenina/metabolismo , Animais , Córtex Cerebral/metabolismo , Hipocampo/metabolismo , Camundongos , Via de Sinalização Wnt/efeitos dos fármacos
9.
Int J Mol Sci ; 22(21)2021 Oct 20.
Artigo em Inglês | MEDLINE | ID: mdl-34768767

RESUMO

Mitochondria are the energy center of the cell. They are found in the cell cytoplasm as dynamic networks where they adapt energy production based on the cell's needs. They are also at the center of the proinflammatory response and have essential roles in the response against pathogenic infections. Mitochondria are a major site for production of Reactive Oxygen Species (ROS; or free radicals), which are essential to fight infection. However, excessive and uncontrolled production can become deleterious to the cell, leading to mitochondrial and tissue damage. Pathogens exploit the role of mitochondria during infection by affecting the oxidative phosphorylation mechanism (OXPHOS), mitochondrial network and disrupting the communication between the nucleus and the mitochondria. The role of mitochondria in these biological processes makes these organelle good targets for the development of therapeutic strategies. In this review, we presented a summary of the endosymbiotic origin of mitochondria and their involvement in the pathogen response, as well as the potential promising mitochondrial targets for the fight against infectious diseases and chronic inflammatory diseases.


Assuntos
Infecções/metabolismo , Inflamação/metabolismo , Mitocôndrias/imunologia , Mitocôndrias/microbiologia , Animais , Metabolismo Energético , Humanos , Infecções/tratamento farmacológico , Inflamação/tratamento farmacológico , Mitocôndrias/metabolismo , Dinâmica Mitocondrial
10.
Genet Mol Biol ; 44(1 Suppl 1): e20200302, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33651876

RESUMO

COVID-19 comprises clinical outcomes of SARS-CoV-2 infection and is highly heterogeneous, ranging from asymptomatic individuals to deceased young adults without comorbidities. There is growing evidence that host genetics play an important role in COVID-19 severity, including inborn errors of immunity, age-related inflammation and immunosenescence. Here we present a brief review on the known order of events from infection to severe system-wide disturbance due to COVID-19 and summarize potential candidate genes and pathways. Finally, we propose a strategy of subject's ascertainment based on phenotypic extremes to take part in genomic studies and elucidate intrinsic risk factors involved in COVID-19 severe outcomes.

11.
J Nucl Cardiol ; 26(5): 1569-1579, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-29392628

RESUMO

BACKGROUND: Myocardial perfusion defects (MPD) due to coronary microvascular dysfunction is frequent in chronic Chagas cardiomyopathy (CCC) and may be involved with development of myocardial damage. We investigated whether MPD precedes left ventricular systolic dysfunction and tested the hypothesis that prolonged use of dipyridamole (DIPY) could reduce MPD in an experimental model of CCC in hamsters. METHODS AND RESULTS: We investigated female hamsters 6-months after T. cruzi infection (baseline condition) and control animals, divided into T. cruzi-infected animals treated with DIPY (CH + DIPY) or placebo (CH + PLB); and uninfected animals treated with DIPY (CO + DIPY) or placebo (CO + PLB). The animals were submitted to echocardiogram and rest SPECT-Sestamibi-Tc99m myocardial perfusion scintigraphy. Next, the animals were treated with DIPY (4 mg/kg bid, intraperitoneal) or saline for 30 days, and reevaluated with the same imaging methods. At baseline, the CH + PLB and CH + DIPY groups showed larger areas of perfusion defect (13.2 ± 13.2% and 17.3 ± 13.2%, respectively) compared with CO + PLB and CO + DIPY (3.8 ± 2.2% e 3.5 ± 2.7%, respectively), P < .05. After treatment, we observed: reduction of perfusion defects only in the CH + DIPY group (17.3 ± 13.2% to 6.8 ± 7.6%, P = .001) and reduction of LVEF in CH + DIPY and CH + PLB groups (from 65.3 ± 9.0% to 53.6 ± 6.9% and from 69.3 ± 5.0% to 54.4 ± 8.6%, respectively, P < .001). Quantitative histology revealed greater extents of inflammation and interstitial fibrosis in both Chagas groups, compared with control group (P < .001), but no difference between Chagas groups (P > .05). CONCLUSIONS: The prolonged use of DIPY in this experimental model of CCC has reduced the rest myocardial perfusion defects, supporting the notion that those areas correspond to viable hypoperfused myocardium.


Assuntos
Cardiomiopatia Chagásica/diagnóstico por imagem , Cardiomiopatia Chagásica/tratamento farmacológico , Dipiridamol/administração & dosagem , Coração/diagnóstico por imagem , Animais , Cricetinae , Modelos Animais de Doenças , Ecocardiografia , Feminino , Ventrículos do Coração/diagnóstico por imagem , Imagem de Perfusão do Miocárdio , Perfusão , Tecnécio Tc 99m Sestamibi , Tomografia Computadorizada de Emissão de Fóton Único , Tomografia Computadorizada por Raios X , Trypanosoma cruzi , Vasodilatadores/administração & dosagem
12.
Eur Arch Otorhinolaryngol ; 276(11): 3247-3249, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31363902

RESUMO

PURPOSE: The pathogenesis of persistent allergic rhinitis with chronic and refractory nasal obstruction is still unknown. Inflammation and tissue remodeling are known to play a role, but this has not been studied thoroughly. The purpose of this study is to identify the profile of gene expression of inflammatory and remodeling markers in nasal mucosa of patients with PAR and chronic obstruction. METHODS: After informed consent, we obtained nasal mucosa tissue from five aeroallergen-sensitized PAR patients undergoing anterior turbinectomy, and control non-sensitized individuals undergoing cerebrospinal fluid fistula repair or rhinoplasty. We assessed the expression of 34 genes related to inflammation and tissue remodeling using the real-time polymerase chain reaction (qPCR) to quantify each mRNA. RESULTS: IL-4 mRNA was upregulated in nasal mucosa of all five patients; CCR3, CCR8 and Eotaxin-2 were upregulated in four out of five patient samples; while IL-5 and IL-13 were upregulated in two of them. TGF-ß1 was not upregulated in PAR samples. mRNA from metalloproteinases MMP-7, MMP13 and MMP15 were upregulated in three out of five samples. Our results indicate a typical mRNA expression profile of the infiltrating inflammatory Th2 cells and eosinophils, combined with altered gene expression of remodeling-related proteins in stromal cells from the mucosa. CONCLUSION: Prolonged allergen challenge can lead to persistent upregulation of genes for inflammatory mediators such as IL-4 Th2/eosinophil cytokines, chemokines and receptors, which may play an important role in maintaining PAR with chronic nasal obstruction. Our findings may have therapeutic implications, including the use of anti-IL4, -CCR3 or -MMP therapy to ameliorate the condition.


Assuntos
Mediadores da Inflamação , Interleucina-4/análise , Metaloproteases/análise , Mucosa Nasal/imunologia , Obstrução Nasal , Receptores CCR3/análise , Rinite Alérgica/imunologia , Adulto , Biomarcadores/análise , Feminino , Perfilação da Expressão Gênica , Humanos , Mediadores da Inflamação/análise , Mediadores da Inflamação/classificação , Masculino , Pessoa de Meia-Idade , Obstrução Nasal/etiologia , Obstrução Nasal/imunologia , Rinite Alérgica/complicações , Rinite Alérgica/patologia , Tempo , Regulação para Cima
13.
PLoS Pathog ; 12(4): e1005593, 2016 04.
Artigo em Inglês | MEDLINE | ID: mdl-27128676

RESUMO

The ß1i, ß2i and ß5i immunoproteasome subunits have an important role in defining the repertoire of MHC class I-restricted epitopes. However, the impact of combined deficiency of the three immunoproteasome subunits in the development of protective immunity to intracellular pathogens has not been investigated. Here, we demonstrate that immunoproteasomes play a key role in host resistance and genetic vaccination-induced protection against the human pathogen Trypanosoma cruzi (the causative agent of Chagas disease), immunity to which is dependent on CD8+ T cells and IFN-γ (the classical immunoproteasome inducer). We observed that infection with T. cruzi triggers the transcription of immunoproteasome genes, both in mice and humans. Importantly, genetically vaccinated or T. cruzi-infected ß1i, ß2i and ß5i triple knockout (TKO) mice presented significantly lower frequencies and numbers of splenic CD8+ effector T cells (CD8+CD44highCD62Llow) specific for the previously characterized immunodominant (VNHRFTLV) H-2Kb-restricted T. cruzi epitope. Not only the quantity, but also the quality of parasite-specific CD8+ T cell responses was altered in TKO mice. Hence, the frequency of double-positive (IFN-γ+/TNF+) or single-positive (IFN-γ+) cells specific for the H-2Kb-restricted immunodominant as well as subdominant T. cruzi epitopes were higher in WT mice, whereas TNF single-positive cells prevailed among CD8+ T cells from TKO mice. Contrasting with their WT counterparts, TKO animals were also lethally susceptible to T. cruzi challenge, even after an otherwise protective vaccination with DNA and adenoviral vectors. We conclude that the immunoproteasome subunits are key determinants in host resistance to T. cruzi infection by influencing both the magnitude and quality of CD8+ T cell responses.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Doença de Chagas/imunologia , Complexo de Endopeptidases do Proteassoma/imunologia , Vacinas Protozoárias/imunologia , Adolescente , Adulto , Animais , Ensaio de Imunoadsorção Enzimática , Feminino , Citometria de Fluxo , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase em Tempo Real , Trypanosoma cruzi , Adulto Jovem
14.
Crit Care ; 22(1): 68, 2018 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-29540208

RESUMO

BACKGROUND: Exosomes isolated from plasma of patients with sepsis may induce vascular apoptosis and myocardial dysfunction by mechanisms related to inflammation and oxidative stress. Despite previous studies demonstrating that these vesicles contain genetic material related to cellular communication, their molecular cargo during sepsis is relatively unknown. In this study, we evaluated the presence of microRNAs (miRNAs) and messenger RNAs (mRNAs) related to inflammatory response and redox metabolism in exosomes of patients with septic shock. METHODS: Blood samples were collected from 24 patients with septic shock at ICU admission and after 7 days of treatment. Twelve healthy volunteers were used as control subjects. Exosomes were isolated by ultracentrifugation, and their miRNA and mRNA content was evaluated by qRT-PCR array. RESULTS: As compared with healthy volunteers, exosomes from patients with sepsis had significant changes in 65 exosomal miRNAs. Twenty-eight miRNAs were differentially expressed, both at enrollment and after 7 days, with similar kinetics (18 miRNAs upregulated and 10 downregulated). At enrollment, 35 differentially expressed miRNAs clustered patients with sepsis according to survival. The pathways enriched by the miRNAs of patients with sepsis compared with control subjects were related mostly to inflammatory response. The comparison of miRNAs from patients with sepsis according to hospital survival demonstrated pathways related mostly to cell cycle regulation. At enrollment, sepsis was associated with significant increases in the expression of mRNAs related to redox metabolism (myeloperoxidase, 64-fold; PRDX3, 2.6-fold; SOD2, 2.2-fold) and redox-responsive genes (FOXM1, 21-fold; SELS, 16-fold; GLRX2, 3.4-fold). The expression of myeloperoxidase mRNA remained elevated after 7 days (65-fold). CONCLUSIONS: Exosomes from patients with septic shock convey miRNAs and mRNAs related to pathogenic pathways, including inflammatory response, oxidative stress, and cell cycle regulation. Exosomes may represent a novel mechanism for intercellular communication during sepsis.


Assuntos
Exossomos/química , MicroRNAs/análise , Choque Séptico/fisiopatologia , Adulto , Idoso , Brasil , Exossomos/metabolismo , Exossomos/patologia , Feminino , Proteína Forkhead Box M1/análise , Proteína Forkhead Box M1/sangue , Glutarredoxinas/análise , Glutarredoxinas/sangue , Humanos , Inflamação/complicações , Inflamação/diagnóstico , Inflamação/metabolismo , Unidades de Terapia Intensiva/organização & administração , Masculino , Proteínas de Membrana/análise , Proteínas de Membrana/sangue , MicroRNAs/sangue , MicroRNAs/metabolismo , Pessoa de Meia-Idade , Estresse Oxidativo , Avaliação de Resultados da Assistência ao Paciente , Peroxidase/análise , Peroxidase/sangue , Peroxirredoxina III/análise , Peroxirredoxina III/sangue , Estudos Prospectivos , RNA Mensageiro/análise , RNA Mensageiro/sangue , RNA Mensageiro/metabolismo , Selenoproteínas/análise , Selenoproteínas/sangue , Choque Séptico/metabolismo , Superóxido Dismutase/análise , Superóxido Dismutase/sangue
15.
J Infect Dis ; 215(3): 387-395, 2017 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-28003350

RESUMO

Chagas disease, caused by the protozoan parasite Trypanosoma cruzi, affects 7 million people in Latin American areas of endemicity. About 30% of infected patients will develop chronic Chagas cardiomyopathy (CCC), an inflammatory cardiomyopathy characterized by hypertrophy, fibrosis, and myocarditis. Further studies are necessary to understand the molecular mechanisms of disease progression. Transcriptome analysis has been increasingly used to identify molecular changes associated with disease outcomes. We thus assessed the whole-blood transcriptome of patients with Chagas disease. Microarray analysis was performed on blood samples from 150 subjects, of whom 30 were uninfected control patients and 120 had Chagas disease (1 group had asymptomatic disease, and 2 groups had CCC with either a preserved or reduced left ventricular ejection fraction [LVEF]). Each Chagas disease group displayed distinct gene expression and functional pathway profiles. The most different expression patterns were between CCC groups with a preserved or reduced LVEF. A more stringent analysis indicated that 27 differentially expressed genes, particularly those related to natural killer (NK)/CD8+ T-cell cytotoxicity, separated the 2 groups. NK/CD8+ T-cell cytotoxicity could play a role in determining Chagas disease progression. Understanding genes associated with disease may lead to improved insight into CCC pathogenesis and the identification of prognostic factors for CCC progression.


Assuntos
Cardiomiopatia Chagásica/genética , Disfunção Ventricular/genética , Linfócitos T CD8-Positivos/imunologia , Cardiomiopatia Chagásica/sangue , Cardiomiopatia Chagásica/fisiopatologia , Citotoxicidade Imunológica/genética , Perfilação da Expressão Gênica , Redes Reguladoras de Genes , Humanos , Células Matadoras Naturais/imunologia , Análise em Microsséries , Pessoa de Meia-Idade , Miocárdio/patologia , Reação em Cadeia da Polimerase em Tempo Real , Disfunção Ventricular/sangue , Disfunção Ventricular/parasitologia
16.
Clin Infect Dis ; 65(7): 1103-1111, 2017 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-28575239

RESUMO

Background: Chagas disease, caused by the protozoan Trypanosoma cruzi, is endemic in Latin America and affects 10 million people worldwide. Approximately 12000 deaths attributable to Chagas disease occur annually due to chronic Chagas disease cardiomyopathy (CCC), an inflammatory cardiomyopathy presenting with heart failure and arrythmia; 30% of infected subjects develop CCC years after infection. Genetic mechanisms play a role in differential progression to CCC, but little is known about the role of epigenetic modifications in pathological gene expression patterns in CCC patients' myocardium. DNA methylation is the most common modification in the mammalian genome. Methods: We investigated the impact of genome-wide cardiac DNA methylation on global gene expression in myocardial samples from end-stage CCC patients, compared to control samples from organ donors. Results: In total, 4720 genes were differentially methylated between CCC patients and controls, of which 399 were also differentially expressed. Several of them were related to heart function or to the immune response and had methylation sites in their promoter region. Reporter gene and in silico transcription factor binding analyses indicated promoter methylation modified expression of key genes. Among those, we found potassium channel genes KCNA4 and KCNIP4, involved in electrical conduction and arrythmia, SMOC2, involved in matrix remodeling, as well as enkephalin and RUNX3, potentially involved in the increased T-helper 1 cytokine-mediated inflammatory damage in heart. Conclusions: Results support that DNA methylation plays a role in the regulation of expression of pathogenically relevant genes in CCC myocardium, and identify novel potential disease pathways and therapeutic targets in CCC.


Assuntos
Cardiomiopatia Chagásica/genética , Doença de Chagas/genética , Metilação de DNA/genética , Adolescente , Adulto , Idoso , Cardiomiopatia Chagásica/parasitologia , Doença de Chagas/parasitologia , Doença Crônica , Impressões Digitais de DNA/métodos , Feminino , Expressão Gênica/genética , Coração/parasitologia , Humanos , Inflamação/genética , Inflamação/parasitologia , Masculino , Pessoa de Meia-Idade , Miocárdio/metabolismo , Canais de Potássio/genética , Regiões Promotoras Genéticas/genética , Trypanosoma cruzi/patogenicidade , Adulto Jovem
18.
J Infect Dis ; 214(1): 161-5, 2016 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-26951817

RESUMO

Long noncoding RNAs (lncRNAs) modulate gene expression at the epigenetic, transcriptional, and posttranscriptional levels. Dysregulation of the lncRNA known as myocardial infarction-associated transcript (MIAT) has been associated with myocardial infarction. Chagas disease causes a severe inflammatory dilated chronic cardiomyopathy (CCC). We investigated the role of MIAT in CCC. A whole-transcriptome analysis of heart biopsy specimens and formalin-fixed, paraffin-embedded samples revealed that MIAT was overexpressed in patients with CCC, compared with subjects with noninflammatory dilated cardiomyopathy and controls. These results were confirmed in a mouse model. Results suggest that MIAT is a specific biomarker of CCC.


Assuntos
Doença de Chagas/complicações , Doença de Chagas/genética , Perfilação da Expressão Gênica , Infarto do Miocárdio/etiologia , Infarto do Miocárdio/genética , RNA Longo não Codificante , Animais , Doença de Chagas/fisiopatologia , Feminino , Humanos , Masculino , Camundongos , Fatores de Transcrição
20.
Immunol Cell Biol ; 93(10): 868-76, 2015 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-25953029

RESUMO

DNA vaccines have failed to induce satisfactory immune responses in humans. Several mechanisms of double-stranded DNA (dsDNA) sensing have been described, and modulate DNA vaccine immunogenicity at many levels. We hypothesized that the immunogenicity of DNA vaccines in humans is suppressed by APOBEC (apolipoprotein B (APOB) mRNA-editing, catalytic polypeptide)-mediated plasmid degradation. We showed that plasmid sensing via STING (stimulator of interferon (IFN) genes) and TBK-1 (TANK-binding kinase 1) leads to IFN-ß induction, which results in APOBEC3A mRNA upregulation through a mechanism involving protein kinase C signaling. We also showed that murine APOBEC2 expression in HEK293T cells led to a 10-fold reduction in intracellular plasmid levels and plasmid-encoded mRNA, and a 2.6-fold reduction in GFP-expressing cells. A bicistronic DNA vaccine expressing an immunogen and an APOBEC2-specific shRNA efficiently silenced APOBEC2 both in vitro and in vivo, increasing the frequency of induced IFN-γ-secreting T cells. Our study brings new insights into the intracellular machinery involved in dsDNA sensing and how to modulate it to improve DNA vaccine immunogenicity in humans.


Assuntos
Apolipoproteínas B/metabolismo , Citidina Desaminase/metabolismo , HIV-1/fisiologia , Proteínas Musculares/metabolismo , Proteínas/metabolismo , Linfócitos T/imunologia , Vacinas de DNA/imunologia , Desaminases APOBEC , Animais , Antígenos Virais/genética , Apolipoproteínas B/genética , Citidina Desaminase/genética , Células HEK293 , Antígenos HLA-DR/genética , Humanos , Imunomodulação , Interferon beta/metabolismo , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Camundongos Transgênicos , Proteínas Musculares/genética , Fragmentos de Peptídeos/genética , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas/genética , Edição de RNA , RNA Interferente Pequeno/genética , Transdução de Sinais/genética , Transgenes/genética , Vacinas de DNA/genética
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