Abstract
Toxoplasma gondii multiplies within parasitophorous vacuole that is not recognized by the primary no oxidative defense of host cells, mainly represented by the fusion with acidic organelles. Recent studies have already shown that hydroxyurea arrested the intracellular parasites leading to its destruction. In the present work we investigated the cellular mechanism involved in the destruction of intracellular Toxoplasma gondii. Fluorescent vital stains were used in order to observe possible acidification of parasitophorous vacuole-containing Toxoplasma gondii in presence of hydroxyurea. Vero cells infected with tachyzoites were treated with hydroxyurea for 12, 24 or 48 hours. Fluorescence, indicative of acidification, was observed in the parasitophorous vacuole when the cultures were incubated in presence of acridine orange. LysoTracker red was used in order to determine whether lysosomes were involved in the acidification process. An intense fluorescence was observed after 12 and 24 hours of incubation with hydroxyurea, achieving it is highly intensity after 48 hours of treatment. Ultrastructural cytochemistry for localization of the acid phosphatase lysosomal enzyme was realized. Treated infected cultures showed reaction product in vesicles fusing with vacuole or associated with intravacuolar parasites. These results suggest that fusion with lysosomes and acidification of parasitophorous vacuole leads to parasites destruction in the presence pf hydroxyurea.
Highlights
Toxoplasma gondii is a cosmopolitan protozoan parasite able to invade and replicate within nucleated cells of vertebrates (Tenter et al 2000, Hill and Dubey 2002)
Following invasion of host cells the infective form of Toxoplasma gondii – tachyzoites – resides within membrane-bound vacuoles known as parasitophorous vacuole (PV) (Sibley et al 1985, Sinai and Joiner 1997, Beyer et al 2002)
Studies showed that intravacuolar parasites as T. gondii are eliminated of host cells in presence of hydroxyurea (Melo et al 2000, Melo and Beiral 2003)
Summary
Toxoplasma gondii is a cosmopolitan protozoan parasite able to invade and replicate within nucleated cells of vertebrates (Tenter et al 2000, Hill and Dubey 2002). Intracellular vacuoles usually undergo fusion with acidic organelles as lysosomes and endosomes, leading to acidification and digestion of intravacuolar materials (Tjeiie et al 2000). The lysosome fusion with vacuoles is the primary no oxidative defense of the host cells against intracellular parasites (Kornfeld and Mellman 1989). PV containing T. gondii avoids fusion with acidic organelles as lysosomes and endosomes (Hirsch and Jones 1972, Finlay and Falkow 1989, Joiner et al 1990, Sibley 1993) and this characteristic is responsible for the intracellular survival of the parasite. Studies showed that intravacuolar parasites as T. gondii are eliminated of host cells in presence of hydroxyurea (Melo et al 2000, Melo and Beiral 2003). In the present study we showed the cellular mechanism involved in the elimination of intracellular T. gondii. Understanding the interactions of this vacuole with the host cell is important, since they are one of the keys to successful intracellular parasitism
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