Digestion degree is a key factor to regulate the printability of pure tendon decellularized extracellular matrix bio-ink in extrusion-based 3D cell printing

Improving the printability of pure, decellularized extracellular matrix (dECM) bio-ink without altering its physiological components has been a challenge in three-dimensional (3D) cell printing. To improve the printability of the bio-ink, we first investigated the digestion process of the powdered dECM material obtained from porcine tendons. We manifested the digestion process of tendon derived dECM powders, which includes dissolution, gelatinization and solubilization. After a short dissolution period (around 10 min), we observed a ‘High viscosity slurry’ status (3 h) of the dECM precursors, i.e. the gelatinization process, followed by the solubilization processes, i.e. a ‘Medium viscosity slurry’ period (12 h) and a ‘Low viscosity slurry’ period (72 h). The ‘Medium viscosity slurry’ status of the dECM bio-ink was inhomogeneous and could not be extruded out from the barrel after the pH value was neutralized to 7.4. Although the ‘Low viscosity slurry’ status of the dECM bio-ink has been reported to be extrudable, it has poor printability. This study explores the printability of the ‘High viscosity slurry’ status of the dECM bio-ink, which has not been addressed thus far. The results demonstrate that this less digested status of the dECM bio-ink yields higher shape fidelity and stacking accuracy than the traditional over-digested status of the dECM bio-ink; this indicates better printability of this less digested dECM bio-ink. We compared the performance of the two bio-inks using cell viability tests for 3D cell printing. Bone marrow mesenchymal stem cells derived from rats was printed using the ‘High viscosity slurry’ status of the dECM bio-ink, yielding high cellular viability lasting for 7 d after printing. Thus, the ‘High viscosity slurry’ status of tendon dECM bio-ink can be utilized to fabricate complicated 3D organoid structures; it also shows promise for applications such as regenerative medicine and biomimetic tissue engineering.