FIGURE 2.

Methods used to generate MSC spheroids ex vivo. (A) In hanging drop technique, MSCs are aggregated by gravitational force. (B) Forced aggregation technique (or pellet culture) is used to generate MSC aggregates by gravitational force that are further induced toward 3D differentiation protocols. (C) Low-attachment surfaces allow the self-organization of cells into suspended spheroids. (D) Magnetic levitation diminish gravitational force, promotes cell–cell contact and induces cell aggregation in vitro. (E) In a spinner flask bioreactor system, cells are continuously mixed by stirring. (F) Rotating wall vessel technique simulates microgravity by constant circular rotation and, therefore, cells are continuously in suspension. Both dynamic culturing techniques (spinner flask and rotating wall vessel) form viable compact MSC spheroids but with altered cell size, altered phenotypic and molecular profiles, and enhanced differentiation potential compared to 2D conventional MSC cultures.