3d Bioprinting of Graphene Oxide-Incorporated Cell-Laden Bone Mimicking Scaffolds for Promoting Scaffold Fidelity, Osteogenic Differentiation and Mineralization

Abstract

Bioprinting is a promising technique for facilitating the fabrication of engineered bone tissues for patient-specific defect repair and for developing in vitro tissue/organ models for ex vivo tests. However, polymer-based ink materials often result in insufficient mechanical strength, low scaffold fidelity and loss of osteogenesis induction because of the intrinsic swelling/shrinking and bioinert properties of most polymeric hydrogels. In this work, we developed a novel human mesenchymal stem cell (hMSC)-laden graphene oxide (GO)/alginate/gelatin composite bioink and investigated the influence of GO incorporation on bioprintability, scaffold fidelity, osteogenic differentiation and extracellular matrix (ECM) mineralization. Our results showed that the GO composite bioinks (0.5GO, 1GO, 2GO) with higher GO concentrations (0.5, 1 and 2 mg/ml) improved the bioprintability with better shear thinning and shear recovery properties. The scaffold fidelity, compressive modulus and cell viability of the 3D bioprinted cell-laden scaffolds were improved with higher GO incorporation at day 1. After culturing in the osteogenic media for 42 days, the 2GO group swelled and had the lowest compressive modulus. The higher GO concentration increased the cell body size and DNA content. The 1GO group had the highest osteogenic differentiation of hMSC with the upregulation of osteogenic-related gene (ALPL, BGLAP, PHEX) expression. To mimic critical-sized calvarial bone defects in mice and prove scaffold fidelity, 3D cell-laden GO defect scaffolds with complex geometries were successfully bioprinted, and the influence of GO incorporation on mineral formation and scaffold fidelity was investigated. 1GO maintained the best scaffold fidelity and had the highest mineral volume after culturing in the bioreactor for 42 days. In conclusion, cell-laden GO composite bioinks had better bioprintability, scaffold fidelity, cell proliferation, osteogenic differentiation and ECM mineralization than the pure alginate/gelatin system. The optimal GO incorporation was 1 mg/ml, which demonstrated great potential for 3D bioprinting of bone tissue model and tissue engineering applications.