Dataset Information

Differential Role of TGF-β in Extracellular Matrix Regulation During Trypanosoma cruzi-Host Cell Interaction.

ABSTRACT: Transforming growth factor beta (TGF-β) is a determinant for inflammation and fibrosis in cardiac and skeletal muscle in Chagas disease. To determine its regulatory mechanisms, we investigated the response of Trypanosoma cruzi-infected cardiomyocytes (CM), cardiac fibroblasts (CF), and L6E9 skeletal myoblasts to TGF-β. Cultures of CM, CF, and L6E9 were infected with T. cruzi (Y strain) and treated with TGF-β (1-10 ng/mL, 1 h or 48 h). Fibronectin (FN) distribution was analyzed by immunofluorescence and Western blot (WB). Phosphorylated SMAD2 (PS2), phospho-p38 (p-p38), and phospho-c-Jun (p-c-Jun) signaling were evaluated by WB. CF and L6E9 showed an increase in FN from 1 ng/mL of TGF-β, while CM displayed FN modulation only after 10 ng/mL treatment. CF and L6E9 showed higher PS2 levels than CM, while p38 was less stimulated in CF than CM and L6E9. T. cruzi infection resulted in localized FN disorganization in CF and L6E9. T. cruzi induced an increase in FN in CF cultures, mainly in uninfected cells. Infected CF cultures treated with TGF-β showed a reduction in PS2 and an increase in p-p38 and p-c-Jun levels. Our data suggest that p38 and c-Jun pathways may be participating in the fibrosis regulatory process mediated by TGF-β after T. cruzi infection.

SUBMITTER: Silva TA

PROVIDER: S-EPMC6801917 | biostudies-literature | 2019 Sep

REPOSITORIES: biostudies-literature

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Differential Role of TGF-β in Extracellular Matrix Regulation During Trypanosoma cruzi-Host Cell Interaction.

Silva Tatiana Araújo TA Ferreira Luis Felipe de Carvalho LFC Pereira Mirian Claudia de Souza MCS Calvet Claudia Magalhães CM

International journal of molecular sciences 20190929 19

Transforming growth factor beta (TGF-β) is a determinant for inflammation and fibrosis in cardiac and skeletal muscle in Chagas disease. To determine its regulatory mechanisms, we investigated the response of Trypanosoma cruzi-infected cardiomyocytes (CM), cardiac fibroblasts (CF), and L6E9 skeletal myoblasts to TGF-β. Cultures of CM, CF, and L6E9 were infected with T. cruzi (Y strain) and treated with TGF-β (1-10 ng/mL, 1 h or 48 h). Fibronectin (FN) distribution was analyzed by i ...[more]

PMID: 31569452

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Project description:Trypanosoma cruzi is the protozoan that causes Chagas disease (CD), an endemic parasitosis in Latin America distributed around the globe. If CD is not treated in acute phase, the parasite remains silent for years in the host's tissues in a chronic form, which may progress to cardiac, digestive or neurological manifestations. Recently, studies indicated that the gastrointestinal tract represents an important reservoir for T. cruzi in the chronic phase. During interaction T. cruzi and host cells release extracellular vesicles (EVs) that modulates the immune system and infection, but the dynamics of secretion of host and parasite molecules through these EVs is not understood. Now, we used two cell lines: mouse myoblast cell line C2C12, and human intestinal epithelial cell line Caco-2to simulate the environments found by the parasite in the host. We isolated large EVs (LEVs) from the interaction of T. cruzi CL Brener and Dm28c/C2C12 and Caco-2 cells upon 2 and 24 h of infection. Our data showed that at two hours there is a strong cellular response mediated by EVs, both in the number, variety and enrichment/targeting of proteins found in LEVs for diverse functions. Qualitative and quantitative analysis showed that proteins exported in LEVs of C2C12 and Caco-2 have different patterns. We found a predominance of host proteins at early infection. The parasite-host cell interaction induces a switch in the functionality of proteins carried by LEVs and a heterogeneous response depending on the tissues analyzed. Protein-protein interaction analysis showed that cytoplasmic and mitochondrial homologues of the same parasite protein, tryparedoxin peroxidase, were differentially packaged in LEVs, also impacting the interacting molecule of this protein in the host. These data provide new evidence that the interaction with T. cruzi leads to a rapid tissue response through the release of LEVs, reflecting the enrichment of some proteins that could modulate the infection environment.

Project description:Trypanosoma cruzi, the etiological agent of Chagas disease in humans, infects a wide variety of vertebrates. Trypomastigotes, the parasite infective forms, invade mammalian cells by a still poorly understood mechanism. Adhesion of tissue culture- derived trypomastigotes to the extracellular matrix (ECM) prior to cell invasion has been shown to be a relevant part of the process. Changes in phosphorylation, S-nitrosylation, and nitration levels of proteins, in the late phase of the interaction (2 h), leading to the reprogramming of both trypomastigotes metabolism and the DNA binding profile of modified histones, were described by our group. Here, the involvement of calcium signaling at a very early phase of parasite interaction with ECM is described. Increments in the intracellular calcium concentrations during trypomastigotes-ECM interaction depends on the Ca2+ uptake from the extracellular medium, since it is inhibited by EGTA or Nifedipine, an inhibitor of the L-type voltage gated Ca2+ channels and sphingosine-dependent plasma membrane Ca2+ channel, but not by Vanadate, an inhibitor of the plasma membrane Ca2+-ATPase. Furthermore, Nifedipine inhibits the invasion of host cells by tissue culture- derived trypomastigotes in a dose-dependent manner, reaching 95% inhibition at 100 µM Nifedipine. These data indicate the importance of both Ca2+ uptake from the medium and parasite-ECM interaction for host-cell invasion. Previous treatment of ECM with protease abolishes the Ca2+ uptake, further reinforcing the possibility that these events may be connected. The mitochondrion plays a relevant role in Ca2+ homeostasis in trypomastigotes during their interaction with ECM, as shown by the increment of the intracellular Ca2+ concentration in the presence of Antimycin A, in contrast to other calcium homeostasis disruptors, such as Cyclopiazonic acid for endoplasmic reticulum and Bafilomycin A for acidocalcisome. Total phosphatase activity in the parasite decreases in the presence of Nifedipine, EGTA, and Okadaic acid, implying a role of calcium in the phosphorylation level of proteins that are interacting with the ECM in tissue culture- derived trypomastigotes. In summary, we describe here the increment of Ca2+ at an early phase of the trypomastigotes interaction with ECM, implicating both nifedipine-sensitive Ca2+ channels in the influx of Ca2+ and the mitochondrion as the relevant organelle in Ca2+ homeostasis. The data unravel a complex sequence of events prior to host cell invasion itself.

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Dataset Information

Differential Role of TGF-β in Extracellular Matrix Regulation During Trypanosoma cruzi-Host Cell Interaction.

Publications

Differential Role of TGF-β in Extracellular Matrix Regulation During <i>Trypanosoma cruzi</i>-Host Cell Interaction.

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