Bioengineered artificial blood vessels have been a major area of interest over the last decade. Of particular interest are small diameter vessels, as surgical options are currently limited. This study aimed to fabricate a small diameter, heterogeneous bilayer blood vessel-like construct in a single step with gelatin methacryloyl (GelMA) bioink using a 3D micro-extrusion bioprinter on a solid platform. GelMA was supplemented with Hyaluronic acid (HA), glycerol and gelatin to form a GelMA bioink with good printability, mechanical strength, and biocompatibility. Two separate concentrations of GelMA bioink with unique pore sizes were selected to fabricate a heterogeneous bilayer. A higher concentration of GelMA bioink (6% w/v GelMA, 2% gelatin, 0.3% w/v HA, 10% v/v glycerol) was used to load human umbilical vein endothelial cells (HUVECs) and form an inner, endothelial tissue layer. A lower concentration of GelMA bioink (4% w/v GelMA, 4% gelatin, 0.3% w/v HA, 10% v/v glycerol) was used to load smooth muscle cells (SMCs) and form an outer, muscular tissue layer. Bioprinted blood vessel-like grafts were then assessed for mechanical properties with Instron mechanical testing, and suture-ability, and for biological properties including viability, proliferation, and histological analysis. The resulting 20 mm long, 4.0 mm diameter lumen heterogeneous bilayer blood vessel-like construct closely mimics a native blood vessel and maintains high cell viability and proliferation. Our results represent a novel strategy for small diameter blood vessel biofabrication.

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