Cellularized Thermoformed Scaffolds with Human Septal Chondrocytes Support In Vivo Cartilage Maturation for Auricular Reconstruction

This study developed a tissue-engineered auricular cartilage platform combining a 3D-printed and thermoformed poly(lactide-b-ethylene glycol) scaffold with clinical-grade alginate and human nasal septal chondrocytes. Initial in vitro experiments compared nasal septum and costal cartilage chondrocytes and found that both maintained high viability after encapsulation, but nasal septal chondrocytes showed greater extracellular matrix deposition and were selected for subsequent studies. Simplified 1 x 1 cm cellularized PLA-PEG/alginate constructs and full ear-shaped thermoformed constructs were then implanted subcutaneously in immunodeficient mice. After 12 weeks, cellularized constructs showed cartilage-like extracellular matrix formation, glycosaminoglycan deposition, collagen type II expression, limited collagen type X expression, and human Ku80 staining demonstrating that newly formed cartilage was predominantly produced by the implanted human cells. Transcriptomic analysis of the ear-shaped constructs showed strong upregulation of cartilage matrix and maturation genes including COL2A1, ACAN, PRG4, COMP, MATN4, and ELN, together with activation of extracellular matrix organization and chondrogenic pathways. UniVert compression measurements showed that cellularization and subsequent tissue formation changed the mechanical response of the simplified constructs, particularly at higher deformation, with cellularized constructs exhibiting lower Young’s modulus and a more sustained, tissue-like mechanical response compared with acellular controls. Although the constructs showed robust cartilage maturation, some fine auricular anatomical detail was lost as alginate degraded. Overall, the study demonstrates the potential of human nasal septal chondrocytes combined with PLA-PEG and alginate scaffolds for clinically relevant engineered auricular cartilage reconstruction.