3D-printed sacrificial molds for high-resolution, patient-specific hydrogel heart valve engineering

Abstract The fabrication of anatomically accurate, cellularized heart valve substitutes remains a significant challenge in tissue engineering, particularly for pediatric and patient-specific applications. While three-dimensional (3D) bioprinting enables the creation of complex geometries, it often compromises cell viability and lacks the precision required for small-scale constructs. In this study, we present a high-fidelity, reproducible molding technique using 3D-printed sugar glass molds to engineer custom, alginate-based hydrogel cellularized heart valves. Human adipose-derived stromal cells (ASCs) were used as the cell source due to their accessibility and regenerative potential. This approach overcomes the limitations of conventional molding and bioprinting by enabling the reproduction of intricate anatomical features, including the sinuses of Valsalva, which are critical for physiological hemodynamics. The molding method maintains high cell viability (>90%) at the time of fabrication and the process supports both scalability and automation. Sugar glass molds for valve sizes from 16 to 26 mm inner diameter were printed with 90% of the mold surface within a ±0.3 mm deviation of the reference computer-aided design model. Cellularized valves cultured in a custom perfusion bioreactor retained structural integrity and cell viability over a 14 d period. This biofabrication strategy offers a promising platform for engineering patient-specific heart valves and also lays the groundwork for in vitro disease modeling, including valve mineralization, using living cells such as ASCs.