Three-dimensional cell culture systems: useful models for studying host-bacteria interactions

Listeria monocytogenes is a highly invasive foodborne bacterium, and its pathogenesis is not completely understood1. To better comprehend this topic, we used a three dimensional (3D) eukaryotic cell culture model. Epithelial intestinal Caco-2 cells (BCRJ 0059) were grown on collagen beads (Cytodex® 3, Sigma- Aldrich, USA) with a microgravity environment in a rotating wall vessel bioreactor (Synthecon, USA), kept at 37°C in humidified 5% CO2 atmosphere2. For starting up the 55 ml capacity bioreactor, it was filled with DMEM medium added 250 mg of beads and 107 Caco-2 cells. After 28 days of incubation, the 3D cells were collected and an aliquot was used for counting, with a total recovery calculated as 108 cells. The organoid-like structures were used for adhesion and invasion assays with L. monocytogenes, with rates of ca. 49% adhesion and ca. 33% invasion, indicating this system permits a confident in vitro evaluation of host-microbe interactions.