The lamina propria (LP) of the vocal fold (VF), the major vibratory tissue during phonation, is a laminated structure with increasing stiffness from superficial layer (SLP) to deep layer (DLP). Tetrazine ligation is one of the fast bioorthogonal reactions based on the inverse-electron-demand Diels-Alder cycloaddition between s-tetrazines (Tz) and trans-cyclooctenes (TCO). We are developing a hyaluronic acid (HA) based multilayer hydrogel capable of mimicing the extracellular environment with a spatial gradient of matrix stiffness. Tetrazine modified hyaluronic acid (HA-Tz), norbornene (Nb) modified matrix metalloproteinase (MMP)-sensitive peptide (bis-Nb-SMR), as well as TCO-tagged cell adhesive peptide (RGD-TCO), were synthesized and used to construct soft homogeneous hydrogel (soft-gel) based on relatively slow Tz-Nb ligation with excess amount of tetrazine. The residual Tz was used to stiffen the matrix while TCO modified HA (HA-TCO) diffused from the top, resulting in multilayer structure (multilayer gel) with spatial stiffness difference, similar to the modulus range in LP. Human mesenchymal stem cells (hMSCs) were encapsulated in the hydrogel and the culture was maintained for up to 14 days. Compare to the control soft gel, multilayer gel showed spatial difference of cell morphology, hMSCs kept in a rounded phenotype in stiff part, meanwhile transitioned to an extended phenotype in soft part, with an increasing secreted MMP level. Our results suggest that the spatial stiffness difference we created in the multilayer hydrogel system is capable to direct stem cell fate, providing a potential system to study VF tissue regeneration and response to various chemical physical and biomechanical cues.

Constructing vocal fold-mimetic synthetic extracellular matrices (sECMs) with a multilayered hydrogel based on bioorthogonal reactions

Macromolecular Chemistry: The Second Century

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ACS Spring 2020 National Meeting & Expo

The official archive and repository for all ACS Meetings & Expositions abstracts and select posters or presentations.