Regulating myogenic differentiation of mesenchymal stem cells using thermosensitive hydrogels

Stem cell therapy has potential to regenerate skeletal muscle tissue in ischemic limb. However, the delivered stem cells experience low rate of myogenic differentiation. Employing injectable hydrogels as stem cell carriers may enhance the myogenic differentiation as their modulus may be tailored to induce the differentiation. Yet current approaches used to manipulate hydrogel modulus often simultaneously vary other properties that also affect stem cell differentiation, such as chemical structure, composition and water content. Thus it is challenging to demonstrate the decoupled effect of hydrogel modulus on stem cell differentiation. In this report, we decoupled the hydrogel modulus from chemical structure, composition, and water content using injectable and thermosensitive hydrogels. The hydrogels were synthesized from N-isopropylacrylamide (NIPAAm), acrylic acid (AAc), and degradable macromer 2-hydroxyethyl methacrylate-oligomer [oligolatide, oligohydroxybutyrate, or oligo(trimethylene carbonate)]. We found that using the same monomer composition and oligomer chemical structure but different oligomer length can independently vary hydrogel modulus. Rat bone marrow mesenchymal stem cells (MSCs) were encapsulated in the hydrogels with elastic expansion moduli of 11, 20, and 40 kPa, respectively. After 14 days of culture, significant myogenic differentiation was achieved for the hydrogel with elastic expansion modulus of 20 kPa, as judged from both the gene and protein expression. In addition, MSCs exhibited an elastic expansion modulus-dependent proliferation rate. The most significant proliferation was observed in the hydrogel with elastic expansion modulus of 40 kPa. These results demonstrate that the developed injectable and thermosensitive hydrogels with suitable modulus has the potential to deliver stem cells into ischemic limb for enhanced myogenic differentiation and muscle regeneration.

Statement of significance: Stem cell therapy for skeletal muscle regeneration in ischemic limb experiences low rate of myogenic differentiation. Employing injectable hydrogels as stem cell carriers may enhance the myogenic differentiation as hydrogel modulus may be modulated to induce the differentiation. Yet current approaches used to modulate hydrogel modulus may simultaneously vary other properties that also affect stem cell myogenic differentiation, such as chemistry, composition and water content. In this report, we decoupled the hydrogel modulus from chemistry, composition, and water content using injectable and thermosensitive hydrogels. We found that mesenchymal stem cells best differentiated into myogenic lineage in the hydrogel with elastic modulus of 20 kPa.