Effect of suramin on trypomastigote forms of Trypanosoma cruzi: changes on cell motility and on the ultrastructure of the flagellum-cell body attachment region.

Suramin has been previously reported to inhibit distinct cell enzymes and to affect the synthesis and distribution of cytoskeleton proteins. Our study indicates that prolonged incubation of Trypanosoma cruzi infected-LLC-MK2 cells in the presence of 500 microM suramin during the intracellular development of the parasite caused morphological changes on trypomastigote forms characterized by a partial or complete detachment of the flagellum from the cell body, besides an accentuated decrease on parasite motility. Immunofluorescence analysis of the region of adhesion between the cell body and the flagellum on trypomastigotes obtained from suramin-treated host cells after the completion of cell cycle did not show any difference in the localization of FAZ antigens recognized by 4D9 and L3B2 monoclonal antibodies despite the presence of a detached flagellum. On the other hand, suramin caused a significant increase on the phenotypic expression of FRA antigen, which was observed throughout the surface of trypomastigotes. Cytochemical localization of cationized ferritin in trypomastigotes obtained from suramin-treated host cells showed that anionic particles gained access to the space between the cell and flagellar membranes, as well as to the flagellar pocket, indicating an alteration on extracellular components of the region of adhesion between the cell body and the flagellum.

A Mg-dependent ecto-ATPase is increased in the infective stages of Trypanosoma cruzi.

In this work, we describe the ability of living epimastigotes of Trypanosoma cruzi to hydrolyze extracellular ATP. In these intact parasites, there was a low level of ATP hydrolysis in the absence of any divalent metal (2.42 +/- 0.31 nmol Pi/h x 10(8) cells). ATP hydrolysis was stimulated by MgCl2, and the Mg-dependent ecto-ATPase activity was 27.15 +/- 2.91 nmol Pi/h x 10(8) cells. The addition of MgCl2 to the extracellular medium increased the ecto-ATPase activity in a dose-dependent manner. This stimulatory activity was also observed when MgCl2 was replaced by MnCl2, but not by CaCl2 or SrCl2. The apparent Km for Mg-ATP2- was 0.61 mM, and free Mg2+ did not increase the ecto-ATPase activity. This ecto-ATPase activity was insensitive to the inhibitors of other ATPase and phosphatase activities. To confirm that this Mg-dependent ATPase was an ecto-ATPase, we used an impermeant inhibitor, DIDS (4, 4'.diisothiocyanostylbene 2'-2'-disulfonic acid) as well as suramin, an antagonist of P2 purinoreceptors and inhibitor of some ecto-ATPases. These two reagents inhibited the Mg2+-dependent ATPase activity in a dose-dependent manner. A comparison among the Mg2+-ecto-ATPase activities of the three forms of T. cruzi showed that the noninfective epimastigotes were less efficient at hydrolyzing ATP than the infective trypomastigote and amastigote stages.

Ecto-ATPase activity on the surface of Trypanosoma cruzi and its possible role in the parasite-host cell interaction.

This study describes the possible role of Mg(2+)-dependent ecto-ATPase activity on the Trypanosoma cruzi-host cell interaction. Mg(2+)-dependent ecto-ATPase activity is observed on the cell body and flagellar membranes of the parasite and is about 20 times greater in trypomastigotes, as compared with epimastigotes. Suramin (a competitive antagonist of P2 receptors) and the impermeant agent 4,4'-diisothiocyanostylbene 2',2'-disulfonic acid (DIDS), both inhibitors of ecto-ATPases, strongly inhibited ATPase activity and the adhesion and internalization of both evolutive forms by mouse resident macrophages. Suramin inhibited the growth of epimastigotes, suggesting a direct participation of ecto-ATPase activity in this process. To overcome the presence of suramin in the culture medium during the time of growth, Mg(2+) ecto-ATPase activity was enhanced 4-fold, as compared with control parasites. The over-expression in enzyme activity was followed by a dramatic increase in the adhesion of epimastigotes to resident macrophages above the level observed for non-treated parasites.