Quick layer-by-layer assembly of aligned multilayers of vascular smooth muscle cells in deep microchannels

One of the main challenges in vascular tissue engineering has been mimicking the complex native three-dimensional (3D) architecture of smooth muscle cells (SMCs). In the current study, we performed layer-by-layer (LBL) seeding of SMCs in a microchanneled scaffold, with or without interleaving a thin layer of collagen type I hydrogel, toward fabricating the 3D microarchitecture. This LBL process avoids the "steric hindrance'' effect observed in direct 3D culture of SMCs in collagen hydrogel. More importantly, the LBL process enables the building up of multilayers of aligned and confluent SMCs. Within each layer, the SMCs as well as the SMC F-actin and alpha-actin filaments align along the direction of the scaffold microchannels, which would potentially improve the tensile and contractile strength of the tissue engineered construct, desirable properties for an engineered vasculature. In addition, rapid two- dimensional (2D) patterning of SMCs is possible with high seeding density, which makes the LBL method feasible for fabrication of multilayered structures in a short time, rendering it useful in clinical therapeutic applications.