Transforming growth factor-ß as a therapeutic target for the cardiac damage of Chagas disease.

Transforming growth factor beta (TGF-ß) 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-ß; (ii) the potential involvement of TGF-ß pathway on T. cruzi invasion of host cells; (iii) association of TGF-ß 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-ß to treat the cardiac alterations of Chagas disease-affected patients.

In Chagas disease, transforming growth factor beta neutralization reduces Trypanosoma cruzi infection and improves cardiac performance.

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-ß (TGF-ß), a key cytokine controlling extracellular matrix synthesis and degradation. TGF-ß 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-ß 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-ß 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-ß 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-ß neutralization.

CD8low T cells expanded following acute Trypanosoma cruzi infection and benznidazole treatment are a relevant subset of IFN-γ producers.

CD8 T cells are regarded as pivotal players in both immunoprotection and immunopathology following Trypanosoma cruzi infection. Previously, we demonstrated the expansion of CD8+ T lymphocytes in the spleen of T. cruzi-infected mice under treatment with benznidazole (N-benzyl-2-nitroimidazole acetamide; Bz), a drug available for clinical therapy. This finding underlies the concept that the beneficial effects of Bz on controlling acute T. cruzi infection are related to a synergistic process between intrinsic trypanocidal effect and indirect triggering of the active immune response. In the present study, we particularly investigated the effect of Bz treatment on the CD8+ T cell subset following T. cruzi infection. Herein we demonstrated that, during acute T. cruzi infection, Bz treatment reduces and abbreviates the parasitemia, but maintains elevated expansion of CD8+ T cells. Within this subset, a remarkable group of CD8low cells was found in both Bz-treated and non-treated infected mice. In Bz-treated mice, early pathogen control paralleled the lower frequency of recently activated CD8low cells, as ascertained by CD69 expression. However, the CD8low subset sustains significant levels of CD44highCD62Llow and CD62LlowT-bethigh effector memory T cells, in both Bz-treated and non-treated infected mice. These CD8low cells also comprise the main group of spontaneous interferon (IFN)-γ-producing CD8+ T cells. Interestingly, following in vitro anti-CD3/CD28 stimulation, CD8+ T cells from Bz-treated T. cruzi-infected mice exhibited higher frequency of IFN-γ+ cells, which bear mostly a CD8low phenotype. Altogether, our results point to the marked presence of CD8low T cells that arise during acute T. cruzi infection, with Bz treatment promoting their significant expansion along with a potential effector program for high IFN-γ production.

TGF-ß inhibitor therapy decreases fibrosis and stimulates cardiac improvement in a pre-clinical study of chronic Chagas' heart disease.

TGF-ß involvement in Chagas disease cardiomyopathy has been clearly demonstrated. The TGF-ß 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 TßR1/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-ß signaling pathway reduced TGF-ß/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-ß inhibitors.

Transforming growth factor-ß as a therapeutic target for the cardiac damage of Chagas disease

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.

Experimental infection with Trypanosoma cruzi increases the population of CD8(+), but not CD4(+), immunoglobulin G Fc receptor-positive T lymphocytes.

It is well established that activating-type Fc receptors for immunoglobulin G (FcgammaR), such as FcgammaRI and FcgammaRIII, are essential for inducing inflammatory responses. On the other hand, a unique inhibitory FcgammaR, FcgammaRIIB, inhibits intracellular signaling upon engagement of immunoglobulin G-immune complexes, suppressing inflammation and autoimmunity. The expression of FcgammaRIIB on B lymphocytes, natural killer cells, macrophages, mast cells, and a number of other cell types has been demonstrated for many years. However, the expression on T lymphocytes is probably restricted to activated cells in a narrow window of time. The controversy regarding the FcgammaR expression on T lymphocytes is attributable to considerable heterogeneity of cellular subpopulations and activation stages during immune responses in vivo. We addressed here this question by using mice experimentally infected with Trypanosoma cruzi, and we found an increase in the CD8(+) FcgammaR(+) population but not in the CD4(+) FcgammaR(+) population. Moreover, CD8(+) FcgammaR(+) T cells predominantly composed the cardiac inflammatory infiltration induced by the infection. These results indicate a novel pattern of FcgammaR expression on T cells in a pathological situation, and possible functional roles of this phenomenon are discussed.