? DESCRIPTION (provided by applicant): Trypanosoma cruzi is an obligate intracellular parasite that infects blood, heart cells and other cells of the body causing debilitating Chagas heart disease. This chronic disease affects several million individuals around the world, including those in the United States; it causes significant morbidity and mortality, yet remains incurable. The long-term goal of this research is to elucidate the early molecular mechanisms that the parasite uses to make the heart fibrotic, leading to heart failure and death. We have shown that invasive T. cruzi trypomastigotes significantly up-regulate the expression of thrombospondin-1 (TSP-1) to accomplish the process of cellular infection. Knockdown of TSP-1 by RNA interference (RNAi) significantly decreased the infection of heart cells by T. cruzi. We also showed that both TSP-1 and the N-terminal domain of TSP-1 (NTSP) were able to interact specifically with the surface of invasive trypomastigotes. In addition, TSP-1 and NTSP pulled down radiolabeled T. cruzi surface calreticulin (TcCRT) in a mini-proteomic assay, and T. cruzi infection is inhibited when parasites are pre-incubated with an anti-TcCRT antibody. This indicates that the TcCRT-TSP-1 interaction is essential for the process of cellular invasion. We also showed that invasive T. cruzi trypomastigotes regulate the expression of profibrotic genes in heart cells; however the exact mechanism remains unknown. Based on our data, we hypothesize that during cellular infection, the interaction of TcCRT and TSP-1 enhances T. cruzi infectivity and induces a fibro genic response via TGF-? dependent and independent pathways. The objective of this application is to delineate a detailed mechanistic understanding of the early molecular interactions between invasive T. cruzi and heart cells leading to the activation of a pro- fibrotic phenotype. This project is therefore significant because it will characterize the lin between TSP-1 up- regulation, the TcCRT-TSP-1 interaction, which enhances T. cruzi infection, TGF-? activation, and the downstream up-regulation of pro-fibrotic genes and proteins in primary human cardiac myocytes and fibroblasts leading to the development of cardiac fibrosis. Additionally, cardiac macrophages purified from infected mice will be characterized to elucidate the immunological arm of T. cruzi induced cardiomyopathy in the presence and absence of endogenous TSP-1. These studies will fill the gap and advance our mechanistic understanding of one of the significant molecular pathologies occurring during cellular infection by T. cruzi. Understanding the mechanism by which this parasite induces heart disease will pave the way for the development of novel molecular intervention strategies to block cardiac pathology thereby making the findings of high significance in the control of Chagas heart disease.

SC1AI127352

2016-05-05

2021-04-30