3D bioprinting of graphene oxide-incorporated cell-laden bone mimicking scaffolds for promoting scaffold fidelity, osteogenic differentiation and mineralization

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. Here, we developed a human mesenchymal stem cells (hMSCs)-laden graphene oxide (GO)/alginate/gelatin composite bioink to form 3D bone-mimicking scaffolds using a 3D bioprinting technique. 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, scaffold fidelity, compressive modulus and cell viability at day 1. The higher GO concentration increased the cell body size and DNA content, but the 2GO group swelled and had the lowest compressive modulus at day 42. 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. 1GO maintained the best scaffold fidelity and had the highest mineral volume after culturing in the bioreactor for 42 days. In conclusion, GO composite bioinks had better bioprintability, scaffold fidelity, cell proliferation, osteogenic differentiation and ECM mineralization than the pure alginate/gelatin system. The optimal GO group was 1GO, which demonstrated the potential for 3D bioprinting of bone tissue models and tissue engineering applications. Statement of significance In this work, we developed a novel human mesenchymal stem cell (hMSC)-laden GO/alginate/ gelatin composite bioink to form 3D bone mimicking scaffolds. Our results showed that the GO composite bioinks with the optimal GO incorporation could improve bioprintability, scaffold fidelity, compressive modulus and cell viability. The higher GO concentration increased the cell body size and cell proliferation. The 1GO group (1 mg/ml) had the highest osteogenic differentiation of hMSC with the upregulation of osteogenic-related gene expression (ALPL, BGLAP, PHEX) at day 42. To mimic critical-sized calvarial bone defects in mice, 3D cell-laden GO defect scaffolds with complex geometries were successfully bioprinted. 1GO maintained the best scaffold fidelity and had the highest mineral volume after culturing in the bioreactor for 42 days. (c) 2020 Acta Materialia Inc. Published by Elsevier Ltd. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/)

Automated summary

In this study, a novel human mesenchymal stem cell (hMSC)-laden GO/alginate/gelatin composite bioink was developed for forming 3D bone mimicking scaffolds. The GO composite bioinks showed improved bioprintability, scaffold fidelity, cell proliferation, osteogenic differentiation, and ECM mineralization. The optimal GO group was 1GO, which demonstrated great potential for 3D bioprinting of bone tissue models and tissue engineering applications.