Regulation of Stem Cell Function in an Engineered Vocal Fold-Mimetic Environment

Human mesenchymal stem cells (hMSCs) have been proposed as therapeutic cells for the treatment of vocal fold (VF) scarring. Although functional recovery was observed in animal models after stem cell injection, it is not clear how injected stem cells interact locally with the extracellular matrix (ECM) of the lamina propria (LP) and how such interactions affect stem cell behaviors to improve function. Herein, we developed an in vitro cell culture platform where hMSCs were encapsulated in a LP-mimetic matrix, derived from hyaluronic acid (HA), poly(ethylene glycol) (PEG) and collagen, and cultured dynamically in a custom-designed VF bioreactor. The cell culture system was characterized by oscillatory shear rheology, laser doppler vibrometry (LDV), and digital image correlation (DIC). A constitutive finite element analysis (FEA) model was further developed to predict vibratory responses of the hydrogel. LDV analysis demonstrated an average displacement of 47 μm in the center of the hydrogel construct at 200 Hz applied frequency without any harmonics. The predicted strains throughout the hydrogel ranged from 0 to 0.03, in good agreement with reported values for the VF. The 3D cellular construct was subjected to vibrational stimulations at 200 Hz for an optimized duration of 1 h, as confirmed by a maximal c-Fos upregulation at the transcript level. Vibrational culture over a 3-day period with a 1h-on/1h-off pattern did not compromise the overall cell viability, but resulted in a significant downregulation of fibrogenic markers and diminished staining for alpha smooth muscle actin (αSMA). Collectively, high frequency mechanical loading resulted in the loss of myofibrogenic potential and a shift away from a fibrotic phenotype.