Methacrylated Hyaluronic Acid–Based Hydrogels Maintain Stemness in Human Dental Pulp Stem Cells

Abstract

2-Aminoethyl methacrylate (AEMA)–modified hyaluronic acid (HA) hydrogels with tunable swelling, degradation, and rheological properties were developed to maintain the native spherical and stemness of human dental pulp stem cells (DPSCs). Two molecular weights (MW) of HA, 18 and 270 kDa, and 25% degree of AEMA substitution were used to synthesize the HA macromers and hydrogels. The HA macromers were characterized by 1H NMR, and the AEMA-HA hydrogels were synthesized by UV polymerization. The swelling, hydrolytic degradation, and rheological properties of the hydrogels strongly depended on the MW of HA. The degradation rate decreased and the storage modulus increased with the increase in the MW of HA. The storage modulus of the hydrogels decreased with degradation of the hydrogels. The hydrogels were not cytotoxic to DPSCs evidenced by MTT assay, Hoechst 33342 live cell staining, and propidium iodide dead cell staining. The hydrogels maintained the native spherical morphology and stemness of DPSCs when the hydrogels’ storage moduli were in the ranges of 486–681 Pa. In contrast, DPSCs grown on 2D cell culture substrates showed a flat and elongated shape and could not maintain their native spherical shape. The hydrogels increased the expression of the stemness including NANOG and SOX2 of DPSCs (3 and 24 folds, respectively) when their storage modulus was 681.41 ± 92.81 Pa in comparison with DPSCs grown on 2D cell culture plates. The designed AEMA-HA hydrogels mimic the in vivo 3D environment to grow DPSCs. Thus, they appear to have great potential for clinical applications in regenerative medicine and other therapeutic remedies after further optimization in the future. Regenerative medicine is a translational approach that offers the possibility to replace or heal the diseased or damaged tissues caused by trauma, age, infection, cancer, or inherited defects. Such reinstatement of the functionality or healing of the damaged cells or tissues often includes the combination of numerous strategies including the use of biomaterials/scaffolds, stem cells, growth factors, and small molecules. In this study, we used hyaluronic acid (HA), a naturally occurring polysaccharide biopolymer commonly found in the extracellular matrix of connective tissues, to synthesize 3D hydrogel scaffolds for growing human dental pulp stem cells (DPSCs). We evaluated the effects of the molecular weight of HA, and the swelling, degradation, and rheological properties of the hydrogels on maintaining the native spherical and stemness of DPSCs for future therapeutic applications including regenerative medicine.