EXPERIMENTAL PARASITOLOGY 90, 1–13 (1998) ARTICLE NO. PR984314 Trypanosoma cruzi: Effect of Protein Kinase Inhibitors and Cytoskeletal Protein Organization and Expression on Host Cell Invasion by Amastigotes and Metacyclic Trypomastigotes Daniela O. Proco ´ pio, 1 Solange da Silva, C. Casey Cunningham, 2 and Renato A. Mortara 3 Departamento de Microbiologia, Imunologia e Parasitologia, Escola Paulista de Medicina, Universidade Federal de Sa ˜ o Paulo, Rua Botucatu, 862, 6 th floor, 04023-062, Sa ˜ o Paulo, SP, Brazil Proco ´ pio, D. O., da Silva, S., Cunningham, C. C., and Mortara, R. INTRODUCTION A. 1998. Trypanosoma cruzi: Effect of protein kinase inhibitors and cytoskeletal protein organization and expression on host cell invasion by amastigotes and metacyclic trypomastigotes. Experimental Parasi- Trypanosoma cruzi is the flagellated protozoan causative tology 90, 1–13. Although trypomastigotes are regarded as the classic infective forms of T. cruzi, amastigotes generated extracellularly or agent of Chagas’ disease. The infection is transmitted to released from infected cells during lysis may circulate and infect other humans by triatomine vectors or accidental causes, affecting cells. We have compared the infectivity of metacyclic trypomastigotes 16 to 18 millions of individuals, and its geographical distri- and extracellular amastigotes toward HeLa and Vero cells and observed bution ranges from northern Mexico to southern parts of that amastigotes were capable of invading both HeLa and Vero cells Argentina (WHO 1996). The infected insect transmits meta- to a much higher degree than the corresponding metacyclic forms. Second, cell microfilament or microtubule disruption inhibited amasti- cyclic trypomastigotes that invade mammalian cells, escape gote but not trypomastigote entry. Third, cells with altered expression the endocytic vacuole, and transform into rounded amasti- in cytoskeletal components (ABP or gelsolin) internalize amastigotes gote forms. Amastigotes replicate in the cytoplasm and dif- and trypomastigotes with highly contrasting fashion. Fourth, protein ferentiate back into trypomastigotes that are released after kinase inhibitors such as genistein and staurosporine affect the internal- cell lysis. An alternative subcycle can occur in the mamma- ization of amastigotes and trypomastigotes in a host-cell-dependent manner. Our results suggest that extracellular amastigotes and meta- lian host if amastigotes, originated from premature lysis of cyclic trypomastigotes utilize mechanisms to invade host cells with infected cells (designated intracellular amastigotes) (Hudson particular features for each T. cruzi form and for each host cell. When et al. 1984; Nogueira and Cohn 1976; Umezawa et al. 1985; internalized, both forms associate to lysosomes of HeLa cells. q 1998 Behbehani 1973; Carvalho et al. 1981; Ulisses de Carvalho Academic Press and De Souza 1986) or by extracellular differentiation of Index Descriptors and Abbreviations: Trypanosoma cruzi; trypomas- tigotes; amastigotes; host cell cytoskeleton; protein phosphorylation trypomastigotes (designated extracellular amastigotes) (An- inhibitors; cellular invasion. drews et al. 1987; Ley et al. 1988; Mortara 1991; Pan 1978), invade professional or nonprofessional phagocytes. Inside these cells the amastigotes can survive and sustain the para- site’s life cycle. Although intracellular amastigotes are larger 1 Present address: Departamento de Bioquı ´ mica e Imunologia, Insti- and slightly more elongated than the extracellular forms tuto de Cie ˆ ncias Biolo ´ gicas, Universidade Federal de Minas Gerais, (Barros et al. 1996), both share immunological, biochemical, Caixa Postal 486, Belo Horizonte, MG, 30161-970 Brasil. and ultrastructural similarities (Villalta and Kierszenbaum 2 Present address: Harvard Medical School, Division of Experimental 1982; Villalta and Kierszenbaum 1984; Andrews et al. 1987; Medicine, 221 Longwood, Boston 02115-5817, MA. Hudson et al. 1984; Ley et al. 1988) and express antigenic 3 To whom all correspondence should be addressed. Fax: 55 11 571- 1095. E-mail: renato.dmip@epm.br. stage-specific markers of amastigotes (Andrews et al. 1987; 1 0014-4894/98 $25.00 Copyright q 1998 by Academic Press All rights of reproduction in any form reserved.