Multilayered Engineered Tissue Sheets for Vascularized Tissue Regeneration

A major hurdle in engineering thick and laminated tissues such as skin is how to vascularize the tissue. This study introduces a promising strategy for generating multi-layering engineered tissue sheets consisting of fibroblasts and endothelial cells co-seeded on highly micro-fibrous, biodegradable polycaprolactone membrane. Analysis of the conditions for induction of the vessels in vivo showed that addition of endothelial cell sheets into the laminated structure increases the number of incorporated cells and promotes primitive endothelial vessel growth. In vivo analysis of 11-layered constructs showed that seeding a high number of endothelial cells resulted in better cell survival and vascularization 4 weeks after implantation. Within one week after implantation in vivo, red blood cells were detected in the middle section of three-layered engineered tissue sheets composed of polycaprolactone/collagen membranes. Our engineered tissue sheets have several advantages, such as easy handling for cell seeding, manipulation by stacking each layer, a flexible number of cells for next-step applications and versatile tissue regeneration, and automated thick tissue generation with proper vascularization.