Culture of Mesenchymal Stem Cells in a Hydrogel Model of Vocal Fold Lamina Propria

Stem cell injection has been proposed as an alternative approach for the restoration of vocal fold (VF) function in patients with VF scarring. To assess the therapeutic efficacy of this treatment strategy, we evaluated the behaviors of human mesenchymal stem cells (hMSCs) in hydrogels derived from thiolated hyaluronic acid (HA-SH) and poly(ethylene glycol) diacrylate (PEG-DA) entrapping assembled collagen fibrils (abbreviated as HPC gels). Three hydrogel formulations with varying amounts of collagen (0, 1 and 2 mg/mL) but a fixed HA-SH (5 mg/mL) and PEG-DA (2 mg/mL) concentration, designated as HPC0, HPC1 and HPC2, were investigated. The HPC gels exhibit similar pore sizes (35-50 nm) and AFM indentation moduli (~175 Pa), although the elastic shear modulus for HPC1 (~32 Pa) is lower than HPC0 and HPC2 (~55 Pa). Although HPC1 and HPC2 gels both promoted the development of an elongated cell morphology, greater cell spreading was observed in HPC2 than in HPC1 by day 7. At the transcript level, cells cultured in HPC1 and HPC2 gels had an increased expression of fibronectin and integrin β1, but a decreased expression of tissue inhibitor of metalloproteinase-1, collagen types I/III and HA synthase-1 when compared to cells cultured in HPC0 gels. Cellular expression of connective tissue growth factor was also elevated in HPC1 and HPC2 cultures. Importantly, the HPC2 hydrogels promoted a signficant up-regulation of matrix metalloproteinase 1, transforming growth factor β1, and epithelial growth factor receptor, indicating an increased tissue turnover. Overall, hMSCs cultured in HPC2 gels adopt a phenotype reminiscent of cells involved in the wound healing process, providing a platform to study the effectiveness of therapeutic stem cell treatments for VF scarring.