Synergy of inorganic and organic inks in bioprinted tissue substitutes: Construct stability and cell response during long-term cultivation in vitro

Abstract

Microextrusion bioprinting, within the emerging field of biofabrication, holds great promise for developing in vitro tissue models or implantable constructs that mimic complexity of native tissues. Despite the considerable progress made fabricating cell-laden scaffolds, mono-material approaches often cannot reproduce complex tissue architecture or satisfy biomechanical requirements, a particular challenge for hard, mineralized bone tissue. Thus, in this work, we focus on the fabrication of high-strength, partly-mineralized composite bone substitutes via multi-channel (bio)printing of inorganic and organic (bio)inks – with the aim of synergistically integrating the advantages of both phases. A hydroxyapatite (HAp) matrix-mimicking, clinically approved, ready-to-use calcium phosphate cement (CPC) ink as a stiff, mechanically robust phase is combined with an organic eggwhite-functionalized bioink of alginate-methylcellulose (AlgMC + EW) delivering cells in a spatially defined distribution. As proven in a novel systematic assessment, cell-free CPC ink and cell-laden AlgMC + EW bioinks exhibited excellent co-printability and shape fidelity in simple, more complex and anatomically shaped biphasic designs. The mechanical properties of such biphasic constructs were tailored via adjusting the ratio of inorganic and organic component. The survival and fate of human mesenchymal stem cells (MSC) and primary human pre-osteoblasts (hOB) in bioprinted inorganic/organic biphasic structures in response to CPC and EW functionalization during a long-term cultivation were investigated, revealing a defined, cell-friendly bioink delivery which allows cells to migrate and proliferate, as well as proving great potential for 3D osteogenesis. In line with further suggested technological refinements, this creates a toolbox for targeted design and construction of possibly pre-vascularized bone structures in clinically relevant dimensions.