Transmigration of Trypanosoma cruzi Trypomastigotes through 3D Spheroids Mimicking Host Tissues

RODRIGUEZ MATIAS EXQUIEL; MASIP YAMIL; RIZZI MARIANA; SANCHEZ DANIEL; CAEIRO LUCAS; TEKIEL VALERIA

T. cruzi Infection: Methods and Protocols, Methods in Molecular Biology

Humana press

Lugar: New York; Año: 2018; p. 165 - 177

While cellular invasion by T. cruzi trypomastigotes and intracellular amastigote replication are wellcharacterizedevents that have been described by using 2D monolayer cultures, other relevant parasite?hostinteractions, like the dynamics of tissue invasiveness, cannot be captured using monolayer cultures.Spheroids constitute a valuable three-dimensional (3D) culture system because they mimic the microarchitectureof tissues and provide an environment similar to the encountered in natural infections, whichincludes the presence of extracellular matrix as well as 3D cell?cell interactions. In this work, we describe aprotocol for studying transmigration of T. cruzi trypomastigotes into 3D spheroids. In the experimentalsetup, cells and parasites are labelled with two fluorescent dyes, allowing their visualization by confocalmicroscopy.We also describe the general procedure and setting of the confocal microscope and downstreamapplications for acquisition and reconstruction of 3D images. This model was employed to analyze thetransmigration of trypomastigotes from the highly virulent and pantropic RA T. cruzi strain. Of course,other aspects encountered by T. cruzi in the mammalian host environment can be studied with thismethodology.