3D printing technologies have great potential in the production of scaffolds to be applied in the design of tissue models for personalised medicine, drug development and toxicological testing. The most common 3D printing technology is Fused Deposition Modelling (FDM). In FDM a thermoplastic polymer is extruded above its melting temperature to create a 3D object. Among thermoplastic polymers, poly(ester-urethane)s (PUs) represent interesting materials due to their high chemical versatility, which can be exploited to synthesize a plethora of polymers matching the requirements of different applications. For instance, degradable PUs have been extensively used in cardiac tissue engineering because of their elastomeric properties. In this study we engineered biphasic scaffolds consisting of 3D printed PU scaffolds filled with GelMA hydrogels, mimicking the young and aged cardiac tissue.

PU was synthesized using poly(ε-caprolactone) diol, 2000Da, 1,4-diisocyanatobutane and L-lysine ethyl ester. The PU was characterized by FT-IR, SEC and tensile tests. Thermal characterization was carried out by TGA, DSC and rheology, to assess polymer suitability for processing in the melt state. The PU was then microfabricated into scaffolds by melt-extrusion additive-manufacturing (fig. 1) and the extruded polymer was characterized to evaluate its stability during the printing process. The scaffolds were surface plasma-treated in the presence of acrylic acid vapor and then grafted with fibronectin through the carbodiimide chemistry. GelMAs with two different degrees of methacryloylation were successfully synthesized as assessed by the colorimetric Ninhydrin assay and FT-IR and NMR analyses. Then, GelMA hydrogels were designed by solubilizing the polymer in a watery medium added with a catalytic amount of a water-soluble photoinitiator. Different GelMA concentrations were exploited as tuning parameters to modulate the mechanical performances of photo-cured gels, obtained by irradiating GelMA aqueous solutions under cell-friendly conditions. Post-curing storage modulus values measured through photo-rheological time sweep tests varied between few and tens kPa. These tunable properties can mimic the cardiac aging process. Lastly, GelMA gels incubated at 25 °C under dynamic flow conditions exhibited stability up to 10 days of observation.

GelMA was loaded with hiPSC-CMs, embedded in the 3D scaffolds and photo-cured, to establish cardiac tissue models. Cells showed viability and contraction ability.