Microscale multilayer cocultures for biomimetic blood vessels

Living tissues have complex and well-organized microstructures. Although microtechnology has been used to create in vivo-like cell microstructures in vitro, most available microscale systems are two-dimensional, and few three-dimensional (3D) systems have been explored. This article demonstrates a 3D hierarchical biomimetic multilayer microsystem created by a generally applicable technique. The technique employs layer-by-layer microfluidics to build layers of cells and biopolymers in microchannels, allowing controlled patterning of cells and their microenvironments in the x, y, and z-dimension. As a prototype, a multilayer system was created using three vascular cell types within heterogeneous types of biopolymers to mimic the structure and composition of a blood vessel wall. The effects of matrix composition and multilayer configurations on 3D cell-cell interactions and cell biology were revealed. Cell migration in the z-dimension, matrix remodeling, intercellular adhesion molecule expression and actin organization were examined under different 3D coculture conditions. A more biomimetic coculture was found to reproduce a more stable structure and in vivo-like function. This approach provides a method to fabricate microscale hierarchical "neotissues" with 3D configurations of matrix materials and multiple cell types, and an in vitro cell coculture model to understand 3D processes of cell-cell and cell-matrix interactions.