Engineering Tissues without the Use of a Synthetic Scaffold: A Twenty-Year History of the Self-Assembly Method

Figure 2

Schema for the production of the “flat” model by the self-assembly technique. Upper panel: classic self-assembly technique. (1) Mesenchymal cells are seeded into 3 cell culture dishes and cultivated for 28 days in the presence of ascorbate. (2) The 3 stromal sheets are peeled from the dish and (3) they are stacked to form a thicker tissue. (4) To ensure fusion of the 3 sheets, a mechanical load is applied on the stacked sheets for a variable time. (5) Then, epithelial cells are seeded on the top of the construct and culture continued for 7 days before (6) tissues are placed at the air/liquid interface for 21 days in order to obtain a mature epithelium. Middle panel: reseeding self-assembly technique. Mesenchymal cells are seeded into cell culture dishes and cultivated for 14 days in the presence of ascorbate before (2) a second seeding of mesenchymal cells on the top of the first stromal sheet. After another 14 days, (3) epithelial cells are seeded on the top of the construct and culture continued for 7 days before (6) tissues are placed at the air/liquid interface for 21 days in order to obtain a mature epithelium. Lower panel: tools required to produce tissue by the self-assembly technique. The anchoring paper is used to limit the contraction of the tissue by the mesenchymal cells and to help in manipulation. The metal weights help to stabilize the anchoring paper during the different cell culture steps. The surgical Ligaclips eliminate the movement of stromal sheets during the fusion step. The surgical sponge is needed to avoid the direct contact between the heavy metal weights and the tissue, and these weights are responsible for the mechanical load favoring the stroma sheet fusion. Finally the air/liquid support maintains the epithelium in contact with the air and the stroma in the cell culture medium.