Embedded Bioprinting of Dense Cellular Constructs in Bone Allograft-Enhanced Hydrogel Matrices for Bone Tissue Engineering

Abstract

Bone tissue engineering aims to address critical-sized defects by developing biomimetic scaffolds that promote repair and regeneration. This study introduces a material extrusion-based embedded bioprinting approach to fabricate dense cellular constructs within methacrylated hyaluronic acid (MeHA) hydrogels enhanced with bioactive microparticles. Composite matrices containing human bone allograft or tricalcium phosphate (TCP) particles were evaluated for their rheological, mechanical, and osteoinductive properties. High cell viability (>95%) and uniform strand dimensions were achieved across all bioprinting conditions, demonstrating the method’s ability to preserve cellular integrity and structural fidelity. The inclusion of bone or TCP particles did not significantly alter the viscosity, crosslinking kinetics, or compressive modulus of the MeHA hydrogels, ensuring robust mechanical stability and shape retention. However, bone allograft particles significantly enhanced osteogenic differentiation of human mesenchymal stem cells (hMSCs), as evidenced by increased alkaline phosphatase (ALP) activity and calcium deposition. Notably, osteogenesis was observed even in basal media, with a dose-dependent response to bone particle concentration, highlighting the intrinsic bioactivity of allograft particles. This study demonstrates the potential of combining embedded bioprinting with bioactive matrices to create dense, osteoinductive cellular constructs. The ability to induce osteogenesis without external growth factors positions this platform as a scalable and clinically relevant solution for bone repair and regeneration.