RAFAEL RIBEIRO ALMEIDA
Projetos de Pesquisa
Unidades Organizacionais
Instituto do Coração, Hospital das Clínicas, Faculdade de Medicina
LIM/19 - Laboratório de Histocompatibilidade e Imunidade Celular, Hospital das Clínicas, Faculdade de Medicina
LIM/19 - Laboratório de Histocompatibilidade e Imunidade Celular, Hospital das Clínicas, Faculdade de Medicina
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- Rare Pathogenic Variants in Mitochondrial and Inflammation-Associated Genes May Lead to Inflammatory Cardiomyopathy in Chagas Disease(2021) OUARHACHE, Maryem; MARQUET, Sandrine; FRADE, Amanda Farage; FERREIRA, Ariela Mota; IANNI, Barbara; ALMEIDA, Rafael Ribeiro; NUNES, Joao Paulo Silva; FERREIRA, Ludmila Rodrigues Pinto; RIGAUD, Vagner Oliveira-Carvalho; CANDIDO, Darlan; MADY, Charles; ZANIRATTO, Ricardo Costa Fernandes; BUCK, Paula; TORRES, Magali; GALLARDO, Frederic; ANDRIEUX, Pauline; BYDLOWSKY, Sergio; LEVY, Debora; ABEL, Laurent; CARDOSO, Clareci Silva; SANTOS-JUNIOR, Omar Ribeiro; OLIVEIRA, Lea Campos; OLIVEIRA, Claudia Di Lorenzo; NUNES, Maria Do Carmo; COBAT, Aurelie; KALIL, Jorge; RIBEIRO, Antonio Luiz; SABINO, Ester Cerdeira; CUNHA-NETO, Edecio; CHEVILLARD, ChristopheCardiomyopathies 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-gamma 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-gamma 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 (Delta psi M), 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-gamma-induced damage in the myocardium, leading to the cardiomyopathy phenotype in Chagas disease. This mechanism may also be operative in other inflammatory cardiomyopathies.
- Co-Exposure of Cardiomyocytes to IFN-gamma and TNF-alpha Induces Mitochondrial Dysfunction and Nitro-Oxidative Stress: Implications for the Pathogenesis of Chronic Chagas Disease Cardiomyopathy(2021) NUNES, Joao Paulo Silva; ANDRIEUX, Pauline; BROCHET, Pauline; ALMEIDA, Rafael Ribeiro; KITANO, Eduardo; HONDA, Andre Kenji; IWAI, Leo Kei; ANDRADE-SILVA, Debora; GOUDENEGE, David; SILVA, Karla Deysiree Alcantara; VIEIRA, Raquel de Souza; LEVY, Debora; BYDLOWSKI, Sergio Paulo; GALLARDO, Frederic; TORRES, Magali; BOCCHI, Edimar Alcides; MANO, Miguel; SANTOS, Ronaldo Honorato Barros; BACAL, Fernando; POMERANTZEFF, Pablo; LAURINDO, Francisco Rafael Martins; TEIXEIRA, Priscila Camillo; NAKAYA, Helder I.; KALIL, Jorge; PROCACCIO, Vincent; CHEVILLARD, Christophe; CUNHA-NETO, EdecioInfection by the protozoan Trypanosoma cruzi causes Chagas disease cardiomyopathy (CCC) and can lead to arrhythmia, heart failure and death. Chagas disease affects 8 million people worldwide, and chronic production of the cytokines IFN-gamma and TNF-alpha by T cells together with mitochondrial dysfunction are important players for the poor prognosis of the disease. Mitochondria occupy 40% of the cardiomyocytes volume and produce 95% of cellular ATP that sustain the life-long cycles of heart contraction. As IFN-gamma and TNF-alpha have been described to affect mitochondrial function, we hypothesized that IFN-gamma and TNF-alpha are involved in the myocardial mitochondrial dysfunction observed in CCC patients. In this study, we quantified markers of mitochondrial dysfunction and nitro-oxidative stress in CCC heart tissue and in IFN-gamma/TNF-alpha-stimulated AC-16 human cardiomyocytes. We found that CCC myocardium displayed increased levels of nitro-oxidative stress and reduced mitochondrial DNA as compared with myocardial tissue from patients with dilated cardiomyopathy (DCM). IFN-gamma/TNF-alpha treatment of AC-16 cardiomyocytes induced increased nitro-oxidative stress and decreased the mitochondrial membrane potential (Delta psi m). We found that the STAT1/NF-kappa B/NOS2 axis is involved in the IFN-gamma/TNF-alpha-induced decrease of Delta psi m in AC-16 cardiomyocytes. Furthermore, treatment with mitochondria-sparing agonists of AMPK, NRF2 and SIRT1 rescues Delta psi m in IFN-gamma/TNF-alpha-stimulated cells. Proteomic and gene expression analyses revealed that IFN-gamma/TNF-alpha-treated cells corroborate mitochondrial dysfunction, transmembrane potential of mitochondria, altered fatty acid metabolism and cardiac necrosis/cell death. Functional assays conducted on Seahorse respirometer showed that cytokine-stimulated cells display decreased glycolytic and mitochondrial ATP production, dependency of fatty acid oxidation as well as increased proton leak and non-mitochondrial oxygen consumption. Together, our results suggest that IFN-gamma and TNF-alpha cause direct damage to cardiomyocytes' mitochondria by promoting oxidative and nitrosative stress and impairing energy production pathways. We hypothesize that treatment with agonists of AMPK, NRF2 and SIRT1 might be an approach to ameliorate the progression of Chagas disease cardiomyopathy.