Labeled probes inserted in the macrophage membrane are transferred to the parasite surface and internalized during cell invasion by Toxoplasma gondii.
Tachyzoites of Toxoplasma gondii attach to the macrophage surface and are internalized either by a phagocytic process, which can be inhibited by cytochalasin D, or by an active process, independent of host cell actin. Previous studies have shown that parasite attachment induces the secretion of macromolecules found in the apical organelles (micronemes and rhoptries) and subsequent/concomitant parasite internalization with the formation of a membrane-bound vacuole known as the parasitophorous vacuole. In the present study we labeled the macrophage surface with fluorescent probes that bind to proteins (DiIC16) and lipids (DTAF) and then allowed control or cytochalasin-D-treated cells to interact with untreated or antibody-coated tachyzoites of T. gondii. The interaction was interrupted at different time points by fixation and the distribution of the probes was analyzed by confocal laser scanning microscopy. Following attachment of the parasites to the macrophage surface, intense labeling of the parasite surface was observed, suggesting transfer of components of the macrophage surface to the parasite surface. Nonadherent parasites were not labeled. Immediately after attachment, most of the parasites were internalized and labeling of the internalized parasites as well as of the parasitophorous vacuole, probably of its membrane, was evident, indicating that surface components of the macrophage are involved in the formation of the parasitophorous vacuole.
Pacheco-Soares
C
C
Laboratório de Biologia Celular e Tecidual, Centro de Biociências e Biotecnologia, Universidade Estadual do Norte Fluminense, Rio de Janeiro, RJ, Brasil.
De Souza
W
W
D000818:Animals; D002232:Carbocyanines / Q000378:metabolism; D015638:Cytochalasin D / Q000494:pharmacology; D005452:Fluoresceins / Q000378:metabolism; D005456:Fluorescent Dyes / Q000378:metabolism; D017737:Macrophages, Peritoneal / Q000378:metabolism / Q000469:parasitology*; D008563:Membrane Lipids / Q000378:metabolism*; D008565:Membrane Proteins / Q000378:metabolism*; D051379:Mice; D018613:Microscopy, Confocal; D014122:Toxoplasma / Q000378:metabolism / Q000472:pathogenicity*
C035005:3,3'-dihexadecylindocarbocyanine; D002232:Carbocyanines; D005452:Fluoresceins; D005456:Fluorescent Dyes; D008563:Membrane Lipids; D008565:Membrane Proteins; D015638:Cytochalasin D; C039083:5-((4,6-dichloro-1,3,5-triazin-2-yl)amino)fluorescein