GelMA hydrogel modulates macrophage polarization to promote tissue regeneration

Objectives: This study aimed to investigate how the biophysical and biochemical properties of gelatin methacrylate (GelMA) hydrogels modulate macrophage polarization and, subsequently, influence dental pulp stem cell (DPSC) behavior for dental pulp regeneration. Methods: GelMA hydrogels were synthesized and characterized using scanning electron microscopy (SEM) and Fourier transform infrared (FTIR) spectroscopy. The stiffness of 5%, 10%, and 20% GelMA formulations was quantified after light curing. THP-1 monocytes were seeded on GelMA-coated plates (2 × 10 6 cells/well) and differentiated into macrophages (M0) using 320 nM phorbol 12-myristate 13-acetate (PMA) for 24 hours. M0 macrophages were maintained for 48 hours to generate conditioned media (CM). Cell viability was assessed using the CCK8 assay. The expression of pro-inflammatory (IL-12) and pro-regenerative (FN-1) markers in macrophages and odontogenic differentiation markers ( DMP1, DSPP ) in DPSCs cultured with macrophage CM were determined by RT-qPCR. Results: SEM and FTIR confirmed successful GelMA synthesis, with characteristic peaks at 3300, 1625, and 1500 cm −1 . Mechanical testing revealed stiffness values of 22.24 ± 4.64 kPa (5%), 76.31 ± 20.16 kPa (10%), and 282.2 ± 27.11 kPa (20%) (p < 0.05). No significant cytotoxicity was observed across all groups. FN-1 expression was significantly upregulated in macrophages cultured on all GelMA substrates (p < 0.05), while IL-12 expression was downregulated exclusively in the 20% GelMA group (p < 0.05), indicating a shift toward a pro-regenerative M2-like phenotype. Correspondingly, DPSCs treated with CM from the 20% GelMA macrophage group showed significantly higher DMP1 and DSPP expression (p < 0.05) compared with controls. Conclusions: The 20% GelMA hydrogel demonstrated superior mechanical strength and enhanced macrophage polarization toward a regenerative phenotype, which subsequently promoted odontogenic differentiation in DPSCs. These findings suggest that GelMA’s mechanical tuning can strategically regulate immune–stem cell crosstalk to enhance dental pulp tissue regeneration.