Precise Preparation of a Multilayer Tubular Cell Sheet with Well-Aligned Cells in Different Layers to Simulate Native Arteries

Compared with artificial vascular grafts, bottom-up tubular cell sheets (TCSs) without scaffolds have shown promise for patients with cardiovascular disease. However, TCS therapy also faces the challenges of lengthy maturation time, elaborate operation, and weak mechanical strength. In this work, a structured small-diameter vascular graft (SDVG), consisting of three layers of TCSs, with different cell types and arrangements, was fabricated using layer-by-layer assembly of naturally formed TCSs and further cell culture. To this end, a surface-patterned collagen-coated cylindrical substrate was designed for the efficient harvesting of naturally formed and well-aligned TCSs. The patterned collagen (type I) layer facilitated the adhesion and orientation of cells, and a continuous tubular cell monolayer was naturally formed after approximately 4 days in cell culture. Biocompatible near-infrared (NIR) light was used to trigger the photothermal phase transition of the collagen coated on the cylindrical substrate to dissociate the collagen layer. As a result, an intact TCS could be harvested within a few minutes. These naturally formed and well-aligned TCSs exhibited outstanding free-standing performance without rugosity, facilitating their operability and practical application. A ring tensile test showed that orientation was critical for improving the mechanical properties of TCSs. The layer-by-layer assembly of SDVGs not only is easy to manipulate and has a short preparation time but also overcomes the bottleneck of forming a hierarchically structured vascular graft. This approach shows promise for repairing damaged blood vessels.