MEM INST OSWALDO CRUZ, RIO DE JANEIRO, 94 (Suppl.I) September 1999
PAGES: 153-154 DOI: Full paper
Why Studies on Invasion of Host Cell by Trypanosoma cruzi Using Stablished Cell Lines or Primary Cell Cultures Give Conflicting Results?

Helene S Barbosa

Laboratório de Ultra-estrutura Celular, Departamento de Ultra-estrutura e Biologia Celular, Instituto Oswaldo Cruz, Av. Brasil 4365, 21045-900 Rio de Janeiro, RJ, Brasil

Studies in vitro of the interaction of Trypanosoma cruzi with host cells did not always involve cells that are in vivotargets of infection. Tumoral cells and cells lines of different origins, which are commonly used, do not represent the real possibility of the interaction of the parasite within mammal host. In the last 15 years our group has been using primary cultures of heart and skeletal muscular cells, the main target cells during evolution of Chagas disease, to approach the biology of the T. cruzi, the molecular events of the parasite interaction, the formation of the parasitophorous vacuole and the intracellular fate of the parasites (Meirelles et al. 1986, Araújo-Jorge et al. 1992, Barbosa & Meirelles 1992, 1993). One of the points that remains under discussion, mainly in the last 10 years, has been the mechanism of invasion of phagocytic and non-professional phagocytic cellsby T. cruzi. A series of papers have reported that cytochalasins B (CB) and D (CD) block the entry of epimastigotes and trypomastigotes into macrophages, Vero cells and fibroblasts (Alexander 1975, Nogueira & Cohn 1976, Ebert & Barbosa 1981, Henriquez et al. 1981, Meirelles et al. 1982, Zenian & Kierszenbaum 1983), while others have reported active penetration of trypomastigote forms into CB-treated fibroblasts, MDCK and HeLa cells (Schenkman et al. 1991, Schenkman & Mortara 1992). Amastigote forms, on the other hand, invade HeLa cells after association with surface microvilli and mobilization of actin microfilaments (Mortara 1991). CD treatment has been also shown to enhance T. cruzi invasion of rat kidney epithelial cells, and the with disruption of cell microfilaments facilitates the access of lysosomes to the adhesion site (Tardieux et al. 1992). Our results obtained with cardiomyocytes showed that the infection rate ranges from 65 to 75% when CB and CD are used. Ultrastructural analysis on the first 30 min of interaction showed that pseudopodia-like expansions of the host cell membranes occur in the adhesion step of the parasite, which are later enclosed by projections of the host cell membrane. Infected cells treated with Triton X-100 demonstrated active mobilization of cytoskeleton filaments at the site of parasite invasion and "sleeve-like" membrane extensions around the parasites. Fixed parasites were never seen inside cardio-myocytes, neither live parasites did invade fixed cells. Our data do not preclude the possibility of additional mechanism(s) of penetration that might require more active participation of the parasites for complete invasion to occur, but indicate that endocytosis is the main process involved in the uptake of metacyclic forms of T. cruzi by cardiomyocytes (Barbosa & Meirelles 1995). In a recent paper, De Souza et al. (1998) suggested that both active penetration and typical phagocytosis can be used by the parasites to invade macrophages and Vero cells, and that both process can occur in the same cell.

Does the use of different host cells, as well as different strains and evolutive forms of the parasites increase the knowledge on the biology of the parasite or does it amplify the differences in results?

This question has been partially answered during Dr Mortara presentation in this round-table: the distribution of different host cell components during the parasite invasion is dependent on the infective forms and also on the host cells, which has been demonstrated by the recruitment of extracellular matrix components, integrin receptors and cytoskeleton elements of HeLa and Vero cells (Procópio et al. 1998).

From the above remarks, a question still unsolved: Can the mechanisms of T. cruzi invasion described with cell lines and tumor cells be considered as universal, despite the fact that these cells are not involved in the in vivo system during the Chagas disease?



Alexander J 1975. Effect of the antiphagocytic agent Cytochalasin B on macrophages invasion by Leishmania mexicana promastigotes and Trypanosoma cruzi epimastigotes. J Protozool 22: 237-240.

Araújo-Jorge TC, Barbosa HS, Meirelles MNL 1992. Trypanosoma cruzi recognition by macrophages and muscle cells: perspectives after a 15-year study. Mem Inst Oswaldo Cruz 87: 43-56.

Barbosa HS, Meirelles, MNL 1992. Ultrastrucutural detection in vitro of WGA-RCAI and Con A-binding sites involved in the invasion of heart muscle cells by Trypanosoma cruzi. Parasitol Res 78: 404-409.

Barbosa HS, Meirelles, MNL 1993. The role of RCA-binding sites in the adhesion of Trypanosoma cruzi to heart muscle cells, as revealed by electron spectroscopic imaging. J Submicroc Pathol 25: 47-51.

Barbosa HS, Meirelles, MNL 1995. Evidence of participation of cytoskeleton of heart muscle cells during the invasion of Trypanosoma cruzi. Cell Struct Funct 20: 275-284.

De Souza W, Carvalho TU, De Melo, ET, Coimbra, ES, Rosestolato CT, Ferreira SR, Vieira M 1998. The use of confocal laser scanning microscopy to analyse the process of parasitic protozoan-host cell interaction. Braz J Med Biol Res 31: 1459-1470.

Ebert F, Barbosa HS 1981. The influence of Cytochalasin B on the interaction of T. cruzi and mouse peritoneal macrophages. Rev Inst Med Trop São Paulo 23: 61-67.

Henriquez D, Piras R, Piras MM 1981. The effect of surface membrane modification of fibroblastic cells on the entry process of Trypanosoma cruzi trypomastigotes. Mol Biochem Parasitol 2: 359-366.

Meirelles MNL, Araújo-Jorge TC, De Souza W 1982. Interaction of Trypanosoma cruzi with macrophages in vitro: Dissociation of the attachment and internalization phases by low temperature and cytochalasin B. Z Parasitenkd68: 7-14.

Meirelles MNL, Araújo-Jorge TC, Miranda CF, De Souza W, Barbosa HS 1986. Interaction of Trypanosoma cruziwith heart muscle cells: ultrastructural and cytochemical analysis of endocytic vacuole formation and effect upon myogenesis in vitro. Eur J Cell Biol 41: 198-206.

Mortara R 1991. Trypanosoma cruzi: Amastigotes and trypomastigotes interact with different structures on the surface of HeLa cells. Exp Parasitol 73: 1-14.

Nogueira N, Cohn Z 1976. Trypanosoma cruzi: Mechanism of entry and intracellular fate in mammalian cells. J Exp Med 143: 1402-1420.

Procópio DO, Silva S, Cunningham CC, Mortara RA 1998. Trypanosoma cruzi: Effect of protein kinase inhibitors and cytoskeleton protein organization and expression on host cell invasion by amastigotes and metacyclic trypomastigotes. Exp Parasitol 90: 1-13.

Schenkman S, Mortara RA 1992. HeLa cells extend and internalize pseudopodia during active invasion byTrypanosoma cruzi trypomastigotes. J Cell Sci 101: 895-905.

Schenkman S, Robbins ES, Nussenzweig V 1991. Attachment of Trypanosoma cruzi to mammalian cells requires parasite energy, and invasion can be independent of the target cell cytoskeleton. Infect Immun 59: 645-654.

Tardieux I, Webster P, Ravesloot J, Boron W, Lunn JA, Heuser JE, Andrews NW 1992. Lysosome recruitment and fusion are early events required for trypanosome invasion of mammalian cells. Cell 71: 1117-1130.

Zenian A, Kierszenbaum F 1983. Trypanosoma cruzi: differences in cell surface interaction of circulating (trypomastigote) and culture (epimastigote) forms with macrophages. J Parasitol 69: 660-665.

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Received 9 June 1999

Accepetd August 1999


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