Arterial thrombosis is the main instigating factor of heart attacks and strokes, which result in over 14 million deaths worldwide every year. The mechanism of thrombosis involves factors from the blood and the vessel wall, and it also relies strongly on 3D vessel geometry and local blood flow patterns. Microfluidic chipbased vascular models allow controlled in vitro studies of the interaction between vessel wall and blood in thrombosis, but until now, they could not fully recapitulate the 3D geometry and blood flow patterns of real-life healthy or diseased arteries. Here we present a method for fabricating microfluidic chips containing miniaturized vascular structures that closely mimic architectures found in both healthy and stenotic blood vessels. By applying stereolithography (SLA) 3D printing of computed tomography angiography (CTA) data, 3D vessel constructs were produced with diameters of 400 μm, and resolution as low as 25 μm. The 3D-printed templates in turn were used as moulds for polydimethylsiloxane (PDMS)-based soft lithography to create microfluidic chips containing miniaturized replicates of in vivo vessel geometries. By applying computational fluid dynamics (CFD) modeling a correlation in terms of flow fields and local wall shear rate was found between the original and miniaturized artery. The walls of the microfluidic chips were coated with human umbilical vein endothelial cells (HUVECs) which formed a confluent monolayer as confirmed by confocal fluorescence microscopy. The endothelialised microfluidic devices, with healthy and stenotic geometries, were perfused with human whole blood with fluorescently labeled platelets at physiologically relevant shear rates. After 15 minutes of perfusion the healthy geometries showed no sign of thrombosis, while the stenotic geometries did induce thrombosis at and downstream of the stenotic area. Overall, the novel methodology reported here, overcomes important design limitations found in typical 2Dwafer-based soft lithography microfabrication techniques and shows great potential for controlled studies of the role of 3D vessel geometries and blood flow patterns in arterial thrombosis.

Biofabrication of reinforced 3D-scaff olds using two-component hydrogels Two strategies were developed to 3D print two-component hydrogels by exploiting polymers with thermogelling and chemoselective crosslinking properties. Pre-crosslinking by oxo-ester mediated native chemical ligation of the two components results in well-defi ned 3D printed constructs. Alternatively, mechanical reinforcement of these hydrogels through covalent cross-linking with a functionalized thermoplastic polymer can be achieved by the same chemoselective chemistry. Good cell compatibility was observed for hyaluronic acid containing scaff olds indicating their potential as new materials for biofabrication applications. precursors proceeded after extrusion to form mechanically stable hydrogels that exhibited a storage modulus of 9 kPa after 3 hours. Flow and elastic properties of the polymer solutions and hydrogels were analyzed under similar conditions to those used during the 3D-printing process. These experiments showed the ability to extrude the hydrogels, as well as their rapid recovery after applied shear forces. Hydrogels were printed in grid-like structures, hollow cones and a model representing a femoral condyle, with a porosity of 48 AE 2%. Furthermore, an N-hydroxysuccinimide functionalized thermoplastic poly-e-caprolactone (PCL) derivative was successfully synthesized and 3D-printed. We demonstrated that covalent grafting of the developed hydrogel to the thermoplastic reinforced network resulted in improved mechanical properties and yielded high construct integrity. Reinforced constructs also containing hyaluronic acid showed high cell viability of chondrocytes, underlining their potential for further use in regenerative medicine applications.