Trypanosoma cruzi activates mouse cardiac fibroblasts in vitro leading to fibroblast-myofibroblast transition and increase in expression of extracellular matrix proteins

Abstract 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

Trypanosoma cruzi infection results in the reduced expression of caveolin-3 in the heart

Caveolae are motile, membrane-bound compartments that contain a number of molecules that participate in cell signaling. Caveolins are protein markers of caveolae and function in a variety of biological processes. Caveolin-3 (Cav-3) is expressed in muscle cells and Cav-3 null mice display a cardiomyopathic phenotype. Ultrastructural cytochemistry, confocal microscopy and immunoblotting revealed a reduction in Cav-3 expression and an activation of ERK (extracellular-signal-regulated kinase) 48 hours after Trypanosoma cruzi infection of cultured cardiac myocytes. CD-1 mice infected with the Brazil strain of T. cruzi displayed reduced expression of Cav-3 and activation of ERK 66 days post infection (dpi). By 180 dpi there was a normalization of these values. These data suggest that the reduction in Cav-3 expression and the activation of ERK during the early phase of infection may contribute to the pathogenesis of chagasic cardiomyopathy

Acute Chagas Disease Induces Cerebral Microvasculopathy in Mice

Made available in DSpace on 2015-04-06T17:18:20Z (GMT). No. of bitstreams: 2 lindice_nisimuraetal_IOC_2014.pdf: 732522 bytes, checksum: c1a2d01674a5e613bc1ec25d3bd17e11 (MD5) license.txt: 1914 bytes, checksum: 7d48279ffeed55da8dfe2f8e81f3b81f (MD5) Previous issue date: 2014Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Investigação Cardiovascular. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Investigação Cardiovascular. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório Morfologia e Morfogênese Vital Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Imunofarmacologia. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Investigação Cardiovascular. Rio de janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Imunofarmacologia. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Ultra-estrutura Celular. Rio de Janeiro, Rj, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Investigação Cardiovascular. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Investigação Cardiovascular. Rio de Janeiro, RJ, Brasil.Cardiomyopathy is the main clinical form of Chagas dise ase (CD); however, cerebra l manifestations, such as meningoencephalitis, ischemic stroke and cognitive impairment, can also occur. The aim of the present study was to investigate functional microvascular alterations and oxidative stress in the brain of mice in acute CD. Acute CD was induced in Swiss Webster mice (SWM) with the Y strain of Trypanosoma cruzi ( T. cruzi ). Cerebral functional capillary density (the number of spontaneously perfused capillaries), leukocyte rolling and adhesion and the microvascular endothelial- dependent response were analyzed over a period of fifteen days using intravital video-microscopy. We also evaluated cerebral oxidative stress with the thiobarbituric acid reactive species TBARS method. Compared with the non-infected group, acute CD significantly induced cerebral functional microvascular alterations, including (i) functional capillary rarefaction, (ii) increased leukocyte rolling and adhesion, (iii) the formation of microvascular platelet-leukocyte aggregates, and (iv) alteration of the endothelial response to acetylcholine. Moreover, cerebral oxidative stress increased in infected animals. We concluded that acute CD in mice induced cerebral microvasculopathy, characterized by a reduced incidence of perfused capillaries, a high number of microvascular platelet-leukocyte aggregates, a marked increase in leukocyte- endothelium interactions and brain arteriolar endothelial dysfunction associated with oxidative stress. These results suggest the involvement of cerebral microcirculation alterations in the neurological manifestations of CD.Cardiomyopathy is the main clinical form of Chagas disease (CD); however, cerebral manifestations, such as meningoencephalitis, ischemic stroke and cognitive impairment, can also occur. The aim of the present study was to investigate functional microvascular alterations and oxidative stress in the brain of mice in acute CD. Acute CD was induced in Swiss Webster mice (SWM) with the Y strain of Trypanosoma cruzi (T. cruzi). Cerebral functional capillary density (the number of spontaneously perfused capillaries), leukocyte rolling and adhesion and the microvascular endothelial-dependent response were analyzed over a period of fifteen days using intravital video-microscopy. We also evaluated cerebral oxidative stress with the thiobarbituric acid reactive species TBARS method. Compared with the non-infected group, acute CD significantly induced cerebral functional microvascular alterations, including (i) functional capillary rarefaction, (ii) increased leukocyte rolling and adhesion, (iii) the formation of microvascular platelet-leukocyte aggregates, and (iv) alteration of the endothelial response to acetylcholine. Moreover, cerebral oxidative stress increased in infected animals. We concluded that acute CD in mice induced cerebral microvasculopathy, characterized by a reduced incidence of perfused capillaries, a high number of microvascular platelet-leukocyte aggregates, a marked increase in leukocyte-endothelium interactions and brain arteriolar endothelial dysfunction associated with oxidative stress. These results suggest the involvement of cerebral microcirculation alterations in the neurological manifestations of CD

Data from: Cruzipain activates latent TGF-β from host cells during T. cruzi invasion

Several studies indicate that the activity of cruzipain, the main lysosomal cysteine peptidase of Trypanosoma cruzi, contributes to parasite infectivity. In addition, the parasitic invasion process of mammalian host cells is described to be dependent on the activation of the host TGF-β signaling pathway by T. cruzi. Here, we tested the hypothesis that cruzipain could be an important activator of latent TGF-β and thereby trigger TGF-β-mediated events crucial for the development of Chagas disease. We found that live epimastigotes of T. cruzi, parasite lysates and purified cruzipain were able to activate latent TGF-β in vitro. This activation could be inhibited by the cysteine peptidase inhibitor Z-Phe-Ala-FMK. Moreover, transfected parasites overexpressing chagasin, a potent endogenous cruzipain inhibitor, prevented latent TGF-β activation. We also observed that T. cruzi invasion, as well as parasite intracellular growth, were inhibited by the administration of Z-Phe-Ala-FMK or anti-TGF-β neutralizing antibody to Vero cell cultures. We further demonstrated that addition of purified cruzipain enhanced the invasive activity of trypomastigotes and that this effect could be completely inhibited by addition of a neutralizing anti-TGF-β antibody. Taken together, these results demonstrate that the activities of cruzipain and TGF-β in the process of cell invasion are functionally linked. Our data suggest that cruzipain inhibition is an interesting chemotherapeutic approach for Chagas disease not only because of its trypanocidal activity, but also due to the inhibitory effect on TGF-β activation

Selenium, TGF-Beta and Infectious Endemic Cardiopathy: Lessons from Benchwork to Clinical Application in Chagas Disease

For over 60 years, selenium (Se) has been known as an essential microelement to many biological functions, including cardiovascular homeostasis. This review presents a compilation of studies conducted in the past 20 years related to chronic Chagas disease cardiomyopathy (CCC), caused by Trypanosoma cruzi infection, a neglected disease that represents a global burden, especially in Latin America. Experimental and clinical data indicate that Se may be used as a complementary therapy to prevent heart failure and improve heart function. Starting from the main questions “Is Se deficiency related to heart inflammation and arrhythmogenesis in CCC?” and “Could Se be recommended as a therapeutic strategy for CCC?”, we show evidence implicating the complex and multidetermined CCC physiopathology, discussing its possible interplays with the multifunctional cytokine TGF-β as regulators of immune response and fibrosis. We present two new proposals to face this global public health challenge in vulnerable populations affected by this parasitic disease: fibrosis modulation mediated by TGF-β pathways and the possible use of selenoproteins as antioxidants regulating the increased reactive oxygen stress present in CCC inflammatory environments. We assess the opportunity to consider the beneficial effects of Se in preventing heart failure as a concept to be applied for CCC patients.SCOPUS: re.jinfo:eu-repo/semantics/publishe

Acute CD causes cerebral functional capillary rarefaction.

<p>Perfused cerebral arterioles and capillaries (arrows) can be observed by the fluorescence of FITC-dextran in the non-infected (A) and <i>T. cruzi</i>-infected animals at 8 (B) and 15 (C) dpi. A collapse in the microcirculation can be observed at 15 dpi (C). In (D), the graph shows a significant reduction in the number of perfused blood vessels (capillary density) in the infected animals at 15 dpi (405±31.4 capillaries/mm²) compared with the non-infected controls (514±1 capillaries/mm²) and with the <i>T. cruzi</i>-infected mice at 8 dpi (535±31.2 capillaries/mm²). Quantitative data are expressed as means ± SEM (n = 5–8/group). One-way ANOVA test; bar = 100 µm; dpi: days post-infection; <i>p</i><0.05: * comparing the infected animals at 15 dpi with NI group; # comparing 15 to 8 dpi. dpi: days post infection. NI: non-infected.</p

Rhodamine-labeled leukocytes in cerebral venules in acute CD.

<p>The images show venules of the non-infected (A) and <i>T. cruzi</i>-infected (B) mice at 8 and 15 dpi (C and D). The leukocyte-endothelium interaction (arrows) in the venules of the infected animals can be observed. Note the microvascular platelet-leukocyte aggregates (arrowhead) at 15 dpi in the infected animals (C and D). dpi: days post-infection; NI: non-infected.</p

Unraveling Chagas disease transmission through the oral route: Gateways to Trypanosoma cruzi infection and target tissues

Submitted by Sandra Infurna ([email protected]) on 2017-07-20T11:15:17Z No. of bitstreams: 1 dea_Villaverde_etal_IOC_2017.pdf: 4095153 bytes, checksum: 9d778615a28eca5e7d9aecc418adb476 (MD5)Approved for entry into archive by Sandra Infurna ([email protected]) on 2017-07-20T11:38:43Z (GMT) No. of bitstreams: 1 dea_Villaverde_etal_IOC_2017.pdf: 4095153 bytes, checksum: 9d778615a28eca5e7d9aecc418adb476 (MD5)Made available in DSpace on 2017-07-20T11:38:43Z (GMT). No. of bitstreams: 1 dea_Villaverde_etal_IOC_2017.pdf: 4095153 bytes, checksum: 9d778615a28eca5e7d9aecc418adb476 (MD5) Previous issue date: 2017Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Pesquisa sobre o Timo. Rio de Janeiro, RJ. Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Pesquisa sobre o Timo. Rio de Janeiro, RJ. Brasil.Universidade Federal do Rio de Janeiro. Centro Nacional de Biologia Estrutural e Bioimagem. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Biologia Molecular e Doenças Endêmicas. Rio de Janeiro, RJ. Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Pesquisa sobre o Timo. Rio de Janeiro, RJ. Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Pesquisa sobre o Timo. Rio de Janeiro, RJ. Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Pesquisa sobre o Timo. Rio de Janeiro, RJ. Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Biologia Molecular e Doenças Endêmicas. Rio de Janeiro, RJ. Brasil.Universidade Federal do Rio de Janeiro. Instituto de Microbiologia Paulo de Góes. Departamento de Imunologia. Rio de Janeiro, RJ, BrasilFundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Pesquisa sobre o Timo. Rio de Janeiro, RJ. Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Inovações em Terapias, Ensino e Bioprodutos. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Pesquisa sobre o Timo. Rio de Janeiro, RJ. Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Pesquisa sobre o Timo. Rio de Janeiro, RJ. Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Pesquisa sobre o Timo. Rio de Janeiro, RJ. Brasil.Oral transmission of Trypanosoma cruzi, the causative agent of Chagas disease, is the most important route of infection in Brazilian Amazon and Venezuela. Other South American countries have also reported outbreaks associated with food consumption. A recent study showed the importance of parasite contact with oral cavity to induce a highly severe acute disease in mice. However, it remains uncertain the primary site of parasite entry and multiplication due to an oral infection. Here, we evaluated the presence of T. cruzi Dm28c luciferase (Dm28c-luc) parasites in orally infected mice, by bioluminescence and quantitative real-time PCR. In vivo bioluminescent images indicated the nasomaxillary region as the site of parasite invasion in the host, becoming consistently infected throughout the acute phase. At later moments, 7 and 21 days post-infection (dpi), luminescent signal is denser in the thorax, abdomen and genital region, because of parasite dissemination in different tissues. Ex vivo analysis demonstrated that the nasomaxillary region, heart, mandibular lymph nodes, liver, spleen, brain, epididymal fat associated to male sex organs, salivary glands, cheek muscle, mesenteric fat and lymph nodes, stomach, esophagus, small and large intestine are target tissues at latter moments of infection. In the same line, amastigote nests of Dm28c GFP T. cruzi were detected in the nasal cavity of 6 dpi mice. Parasite quantification by real-time qPCR at 7 and 21 dpi showed predominant T. cruzi detection and expansion in mouse nasal cavity. Moreover, T. cruzi DNA was also observed in the mandibular lymph nodes, pituitary gland, heart, liver, small intestine and spleen at 7 dpi, and further, disseminated to other tissues, such as the brain, stomach, esophagus and large intestine at 21 dpi. Our results clearly demonstrated that oral cavity and adjacent compartments is the main target region in oral T. cruzi infection leading to parasite multiplication at the nasal cavity