The present study was conducted to manipulate various biomaterials to find potential hydrogel formulations through three-dimensional (3D) bioprinting fabrication for tissue repair, reconstruction, or regeneration. The hydrogels were prepared using sodium alginate and gelatin combined with different concentrations of Pluronic F127 (6% (3 g), 8% (4 g), and 10% (5 g)) and were marked as AGF-6%, AGF-8%, and AGF-10%, respectively. The properties of the hydrogels were investigated using a contact angle goniometer, rheometer, and 3D bioprinter. In addition, the osteoblast-like cell line (MG-63) was used to evaluate the cell viability including hydrogels before and after 3D bioprinting. It was found that the ratio of contact angle was lowest at AGF-6%, and the rheological results were higher for all samples of AGF-6%, AGF-8%, and AGF-10% compared with the control sample. The printability indicated that the AGF-6% hydrogel possessed great potential in creating a cell scaffold with shape integrity. Moreover, the live/dead assay also presented the highest numbers of live cells before printing compared with after printing. However, the number of live cells on day 7 was higher than on day 1 before and after printing (** p < 0.01). Therefore, the combination of AGF-6% could be developed as a biofunctional hydrogel formulation for potential tissue regeneration applications.

This study evaluated whether three-dimensional alginate–gelatin hydrogels (AGHs) crosslinked with calcium chloride (CaCl₂) enhance the osteo-odontogenic differentiation of odontoblast-like cells in vitro. Two seeding configurations were compared: inter-hydrogel (INT) surface seeding and intra-hydrogel (INTR) encapsulation. Here, the MDPC-23 cells were cultured in AGHs crosslinked with 70 or 100 mM CaCl₂ and assessed for proliferation, cytoskeletal morphology, alkaline phosphatase (ALPase) activity, osteo-odontogenic gene expression, and mineralized nodule formation. After 7 days, cell proliferation was significantly greater in the alginate–gelatin hydrogel (AGH) groups than in the control group. Cells in the intra alginate–gelatin hydrogel 100 (INTR-AGH100) remained predominantly rounded, whereas those in the inter alginate–gelatin hydrogel 100 (INT-AGH100) formed irregular clusters on the hydrogel surface. ALPase activity was highest in INTR-AGH100 at the early stage of culture. Both INT-AGH100 and INTR-AGH100 showed significantly increased expression of DSPP, DMP-1, BSP, OCN, OPN, and Runx-2, together with enhanced mineralized nodule formation. Although no significant differences were detected between the two seeding strategies in all assays, distinct morphological patterns were observed, and the INTR configuration showed relatively greater early differentiation-related activity. These findings suggest that 100 mM CaCl₂-crosslinked AGHs provide a favorable three-dimensional microenvironment under the present experimental conditions and represent a promising in vitro scaffold platform to support future studies of scaffold-guided dentin regeneration.