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Introdução/Objetivo: As infecções causadas por agentes multirresistentes são um problema de saúde mundial, com altas taxas de mortalidade. A identificação rápida dessas infecções é essencial, devido à sua natureza contagiosa, dificuldade de tratamento e custos hospitalares. Para abordar esse desafio, o desenvolvimento de métodos de detecção rápidos e acessíveis, bem como terapias alternativas, são necessários. Uma solução promissora é o uso de aptâmeros, que são moléculas capazes de interagir com bactérias. Esses aptâmeros têm potencial para reconhecer agentes infecciosos e/ou inibir suas funções. O objetivo desse estudo foi selecionar e identificar aptâmeros capazes de se ligar a células de A. baumannii multirresistentes. Métodos: Uma cepa de A. baumannii isolada de amostra clínica foi submetida ao protocolo de cell-SELEX. O sucesso das rodadas de seleção foi acompanhado por RT-PCR (curva de melting comparada com controle positivo) e citometria de fluxo com aptâmeros marcados com FAM (deslocamento de pico de fluorescência indicando ligação dos aptâmeros às células). Os aptâmeros foram identificados por sequenciamento utilizando a plataforma illumina. Resultados: Primeiramente, otimizamos uma metodologia interna, previamente descrita, baseada em cell-SELEX, para identificação de aptâmeros com execução rápida e baixo custo. Foram realizadas 15 rodadas de cell-SELEX, sendo duas negativas utilizando células de K. pneumoniae. Os ensaios de citometria de fluxo revelaram que após a 15ª rodada além dos aptâmeros selecionados se ligarem à célula alvo, eles também apresentaram preferência de ligação quando comparados com o controle negativo. O sequenciamento revelou as 10 sequencias mais frequentemente encontradas após a seleção na 13ª e 15ª rodada, sugerindo os aptâmeros mais selecionados para ligação às células alvo e provavelmente os melhores candidatos. Os aptâmeros que se mantiveram nas duas rodadas e que aumentaram o número de cópias na 15ª com relação à 13ª foram selecionados. A avaliação inicial da estrutura tridimensional revelou que 3 aptâmeros apresentavam estruturas similares, indicando uma possível convergência na seleção. Testes de confirmação da especificidade e sensibilidade para cada novo aptâmero identificado estão sendo realizados. Conclusão: Esses dados indicam que em breve a tecnologia baseada em aptâmero pode se tornar uma alternativa tangível às abordagens tradicionais ao diagnóstico e terapia de bactérias multirresistentes como A. baumanni.

Chronic Chagasic cardiomyopathy (CCC), a progressive inflammatory and fibrosing disease, is the most prominent clinical form of Chagas disease, a neglected tropical disease caused by Trypanosoma cruzi infection. During CCC, the parasite remains inside the cardiac cells, leading to tissue damage, involving extensive inflammatory response and irregular fibrosis. Among the fibrogenic factors is transforming growth factor-β (TGF-β), a key cytokine controlling extracellular matrix synthesis and degradation. TGF-β is involved in CCC onset and progression, with increased serum levels and activation of its signaling pathways in the cardiac tissue, which crucially contributes to fibrosis. Inhibition of the TGF-β signaling pathway attenuates T. cruzi infection and prevents cardiac damage in an experimental model of acute Chagas disease. The aim of this study was to investigate the effect of TGF-β neutralization on T. cruzi infection in both in vitro and in vivo pre-clinical models, using the 1D11 monoclonal antibody. To this end, primary cultures of cardiac cells were infected with T. cruzi trypomastigote forms and treated with 1D11. For in vivo studies, 1D11 was administered in different schemes for acute and chronic phase models (Swiss mice infected with 104 parasites from the Y strain and C57BL/6 mice infected with 102 parasites from the Colombian strain, respectively). Here we show that the addition of 1D11 to cardiac cells greatly reduces cardiomyocyte invasion by T. cruzi and the number of parasites per infected cell. In both acute and chronic experimental models, T. cruzi infection altered the electrical conduction, decreasing the heart rate, increasing the PR interval and the P wave duration. The treatment with 1D11 reduced cardiac fibrosis and reversed electrical abnormalities improving cardiac performance. Taken together, these data further support the major role of the TGF-β signaling pathways in T. cruzi-infection and their biological consequences on parasite/host interactions. The therapeutic effects of the 1D11 antibody are promising and suggest a new possibility to treat cardiac fibrosis in the chronic phase of Chagas’ heart disease by TGF-β neutralization.

Transforming growth factor beta (TGF-β) is deeply involved on the pathogenesis of Chagas disease. Our group has been investigating the participation of this pleiotropic cytokine in different aspects of Chagas disease over the last 20 years. Important observations have been made, such as: (i) the ability of Trypanosoma cruzi in activating latent TGF-β; (ii) the potential involvement of TGF-β pathway on T. cruzi invasion of host cells; (iii) association of TGF-β with parasite intracellular replication; (iv) cardiac fibrosis development and maintenance; (v) disruption of Connexin-43 plaque structures and (vi) inflammation and immune response. In this perspective article we intend to discuss the advances of the potential use of new therapies targeting TGF-β to treat the cardiac alterations of Chagas disease-affected patients.

TGF-β involvement in Chagas disease cardiomyopathy has been clearly demonstrated. The TGF-β 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 TβR1/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-β signaling pathway reduced TGF-β/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-β inhibitors.