Multicellular organotypic model of human intestinal mucosa

A method of preparing a multi-cellular three-dimensional tissue construct, comprising

• growing human intestinal epithelial, primary human umbilical vein endothelial cells (HUVEC), and CCD-18Co primary human fibroblast cells separately as two-dimensional (2-D) confluent monolayers, wherein the human intestinal epithelial cells comprises a HCT-8 human intestinal epithelial cell line;
• embedding the fibroblast cells and the endothelial cells, in a collagen-I extracellular matrix mixture enriched with gut basement membrane proteins comprising laminin, collagen IV, fibronectin and heparin sulfate proteoglycan, at a concentration of 1.0-1.2×106 and 1.5-2.0×106 cells/mL for HUVEC and CCD-18Co cells, respectively, to form a cell-containing gel;
• with a specific gravity similar to culture medium;
• adding the cell-containing gel to a rotating wall vessel bioreactor and suspending said HCT-8 human intestinal epithelial cells in medium and then adding the suspended cells at a concentration of about 107 cells/mL to the rotating wall vessel to form a cell culture;
• and
• applying microgravity conditions that produce laminar flow and minimize shear force inside the culture, allowing the cells to culture, while adding primary human lymphocytes to the cell culture, to form a multi-cellular three-dimensional tissue construct;
• wherein the epithelial cells form a simple columnar epithelium and differentiate into multiple lineages.