Microscale multilayer cocultures for biomimetic blood vessels

<jats:title>Abstract</jats:title><jats:p>Living tissues have complex and well‐organized microstructures. Although microtechnology has been used to create <jats:italic>in vivo</jats:italic>‐like cell microstructures <jats:italic>in vitro</jats:italic>, 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 <jats:italic>in vivo</jats:italic>‐like function. This approach provides a method to fabricate microscale hierarchical “neotissues” with 3D configurations of matrix materials and multiple cell types, and an <jats:italic>in vitro</jats:italic> cell coculture model to understand 3D processes of cell–cell and cell–matrix interactions. © 2004 Wiley Periodicals, Inc. J Biomed Mater Res 72A: 146–160, 2005</jats:p>