Abstract
Ecto-Nucleoside Triphosphate Diphosphohydrolases (Ecto-NTPDases) are enzymes that hydrolyze tri- and/or di-phosphate nucleotides. Evidences point to their participation in Trypanosoma cruzi virulence and infectivity. In this work, we evaluate TcNTPDase-1 gene expression in comparison with ecto-NTPDase activity, in order to study the role of TcNTPDase-1 in parasite virulence, infectivity and adaptation to heat shock. Comparison between distinct T. cruzi isolates (Y, 3663 and 4167 strains, and Dm28c, LL014 and CL-14 clones) showed that TcNTPDase-1 expression was 7.2 ± 1.5 times higher in the Dm28c than the CL-14 avirulent clone. A remarkable expression increase was also observed in the trypomastigote and amastigote forms (22.5 ± 5.6 and 16.3 ± 3.8 times higher than epimastigotes, respectively), indicating that TcNTPDase-1 is overexpressed in T. cruzi infective forms. Moreover, heat shock and long-term cultivation also induced a significant increment on TcNTPDase-1 expression. Our results suggest that TcNTPDase-1 plays an important role on T. cruzi infectivity and adaptation to stress conditions, such as long-term cultivation and heat shock.
Highlights
Ecto-Nucleoside Triphosphate Diphosphohydrolases (Ecto-NTPDases) are enzymes that hydrolyze tri- and/or di-phosphate nucleotides
Our results suggest that TcNTPDase-1 plays an important role on T. cruzi infectivity and adaptation to stress conditions, such as long-term cultivation and heat shock
Ethical approval In order to perform the experimental infections with Trypanosoma cruzi, swiss mice obtained from the animal facilities of the Oswaldo Cruz Foundation (CECAL/Fiocruz, Rio de Janeiro, Brazil) were housed under specific pathogen free conditions in a 12-hour light-dark cycle with access to food and water ad libitum
Summary
Chagas disease is a neglected illness caused by the protozoan parasite Trypanosoma cruzi, which affects 8 million people in endemic areas of Latin America [1,2]. We have developed a quantitative Real-Time RT-PCR assay to quantify TcNTPDase-1 mRNA levels, using TcGAPDH and TcCalmoduline as housekeeping genes (Additional file 1: Figure S1), aiming to contribute to the knowledge about the role of this NTPDase to T. cruzi infectivity and virulence. In this sense, we evaluated the TcNTPDase-1. The epimastigote and metacyclic trypomastigote forms interact with the triatomine insect vector at 28°C and the amastigote and trypomastigote forms interact with the mammalian host at 37°C This heat shock may induce a response from the parasite, promoting the modulation of ecto-ATPase activity [14]. Considering the importance of the ecto-NTPDase activity on the parasite’s purine salvage pathway [15] and heat shock adaptation, the expression of TcNTPDase-1 was analyzed during T. cruzi epimastigote cultivation, at different temperatures, to investigate the expression regulation in response to long-term cultivation or induced by heat shock
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