Impaired frequencies and function of platelets and tissue remodeling in chronic Chagas disease

RESEARCH ARTICLE Impaired frequencies and function of platelets and tissue remodeling in chronic Chagas disease Claudia Pengue1☯, Gonzalo Cesar2☯, Marı́a Gabriela Alvarez1, Graciela Bertocchi1, Bruno Lococo1, Rodolfo Viotti1, Marı́a Ailén Natale2, Melisa D. Castro Eiro2, Silvia S. Cambiazzo3, Nancy Perroni1, Myriam Nuñez4, Marı́a Cecilia Albareda2*, Susana A. Laucella ID1,2* a1111111111 a1111111111 a1111111111 a1111111111 a1111111111 1 Hospital Interzonal General de Agudos Eva Perón, Buenos Aires, Argentina, 2 Instituto Nacional de Parasitologı́a Dr. M. Fatala Chaben, Buenos Aires, Argentina, 3 Hospital General de Agudos Dr. Teodoro Álvarez, Buenos Aires, Argentina, 4 Departamento de Matemática y Fı́sica, Facultad Farmacia y Bioquı́mica, Universidad de Buenos Aires, Argentina ☯ These authors contributed equally to this work. * (SAL); (MCA) Abstract OPEN ACCESS Citation: Pengue C, Cesar G, Alvarez MG, Bertocchi G, Lococo B, Viotti R, et al. (2019) Impaired frequencies and function of platelets and tissue remodeling in chronic Chagas disease. PLoS ONE 14(6): e0218260. https://doi.org/10.1371/journal. pone.0218260 Editor: Pablo Garcia de Frutos, Institut d’Investigacions Biomediques de Barcelona, SPAIN Received: November 23, 2018 Accepted: May 29, 2019 Published: June 14, 2019 Copyright: © 2019 Pengue et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Chronic inflammation, as a consequence of the persistent infection with Trypanosoma cruzi, leads to continuous activation of the immune system in patients with chronic Chagas disease. We have previously shown that increased sera levels of soluble P-selectin are associated with the severity of the cardiomyopathy distinctive of chronic Chagas disease. In this study, we explored the expression of biomarkers of platelet and endothelial activation, tissue remodeling, and mediators of the coagulation cascade in patients at different clinical stages of chronic Chagas heart disease. The frequencies of activated platelets, measured by the expression of CD41a and CD62P were decreased in patients with chronic Chagas disease compared with those in uninfected subjects, with an inverse association with disease severity. Platelet activation in response to adenosine diphosphate was also decreased in T. cruziinfected subjects. A major proportion of T. cruzi infected subjects showed increased serum levels of fibrinogen. Patients with severe cardiac dysfunction showed increased levels of endothelin-1 and normal values of procollagen I. In conclusion, chronic infection with T. cruzi induced hemostatic alterations, even in those patients who do not yet present cardiac symptoms. Data Availability Statement: All relevant data are within the manuscript and its Supporting Information files. Introduction Funding: This work was supported by the National Scientific and Technical Research Council, Argentina (CONICET PIP 2013 N˚037 to SAL), the Ministry of Health of the Province of Buenos Aires and the National Ministry of Health of Argentina. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. SAL and MCA are Trypanosoma cruzi, the causative agent of Chagas disease, infects 6–7 million people in Central and South America, as well as in countries historically nonendemic for T. cruzi infection [1]. The acute phase is characterized by the presence of a large number of parasites in the circulation and even though the immune response is able to control the infection, the parasite can survive establishing a chronic infection. Chronic inflammation, as a consequence of the persistent infection with T. cruzi, leads to continuous activation of the immune system in chronic Chagas disease patients [2–5]. PLOS ONE | https://doi.org/10.1371/journal.pone.0218260 June 14, 2019 1 / 15 Biomarkers of altered microcirculation in chronic Chagas disease members of The National Scientific and Technical Research Council, Argentina. MAN and MDCE are CONICET Ph.D. fellows. GC is a Ph.D. fellow of the Scientific and Technological Research Fund (FONCyT), Argentina. Competing interests: The authors have declared that no competing interests exist. Inflammatory mediators regulate the expression of different adhesion molecules that participate in the recruitment of leukocytes and monocytes to sites of infection [6]. Among the latter group, platelet selectin (P-selectin) redistributes to the surface of platelets and endothelial cells within minutes after activation [7–9], and the P-selectin glycoprotein ligand-1 (PSGL-1) is constitutively expressed and participates in both leukocyte recruitment and the formation of platelet thrombi [10]. We have previously shown that increased serum levels of soluble P-selectin are associated with the severity of the cardiomyopathy that is distinctive of chronic Chagas disease [3]. However, children in early stages of T. cruzi infection also displayed high s-P-selectin titers, and these levels decreased following treatment with benznidazole [11]. Although increased s-Pselectin levels in chronic T. cruzi infection probably reflect alterations in the microcirculation that might eventually result in a pathogenic mechanism, it can also be reflective of an ongoing immune response to keep the parasite under control [3,12–15]. Of note, it is becoming more evident that platelets have inflammatory functions and can influence both innate and adaptive immune responses [16–18]. Here, we explored the expression of platelet and endothelial activation, tissue remodeling, and mediators of the coagulation cascade in patients at different clinical stages of chronic Chagas heart disease. The findings reported in the present work show platelet dysfunction and alterations in hemostatic factors in chronic Chagas disease. Materials and methods Selection of the study population Subjects were recruited at the Chagas disease Section, Cardiology Department, Hospital Interzonal General de Agudos “Eva Perón”, Buenos Aires, Argentina. A positive T. cruzi infection was determined by indirect immunofluorescence assay, hemagglutination, and enzyme-linked immunoassay techniques [19]. Subjects testing positive in at least two of these tests were considered to be infected. Chronically T. cruzi-infected subjects were evaluated clinically and stratified according to a modified version of the Kuschnir grading system, as follows [20,21]. Group 0 (G0) had normal electrocardiograph (ECG), chest radiograph, and echocardiograph findings; Group 1 (G1) had normal chest radiograph and echocardiograph findings but abnormal electrocardiograph findings; Group 2 (G2) had ECG abnormalities and heart enlargement as determined by chest X-ray; and Group 3 (G3) had ECG abnormalities, heart enlargement and clinical or radio (...truncated)