Abstract
Successful engineering of functional salivary glands necessitates the creation of cell-instructive environments for ex vivo expansion and lineage specification of primary human salivary gland stem cells (hS/PCs). Herein, basement membrane mimetic hydrogels are prepared using hyaluronic acid, cell adhesive peptides, and hyperbranched polyglycerol (HPG), with or without sulfate groups, to produce "hyperGel+" or "hyperGel", respectively. Differential scanning fluorescence experiments confirm the ability of the sulfated HPG precursor to stabilize fibroblast growth factor 10. The hydrogels are nanoporous, cytocompatible, and cell-permissive, enabling the development of multicellular hS/PC spheroids in 14 days. The incorporation of sulfated HPG species in the hydrogel enhances cell proliferation. Culture of hS/PCs in hyperGel+ in the presence of a Rho kinase inhibitor Y-27632 (Y-27) leads to the development of spheroids with a central lumen, increases the expression of acinar marker aquaporin-3 at the transcript level (AQP3), and decreases the expression of ductal marker keratin 7 at both the transcript (KRT7) and the protein levels (K7). Reduced expression of transforming growth factor beta (TGF-beta) targets SMAD2/3 is also observed in Y27-treated cultures, suggesting attenuation of TGF-beta signaling. Thus, hyperGel+ cooperates with the Rho-associated protein kinase inhibitor to promote the development of lumened spheroids with enhanced expression of acinar markers.
Basement membrane mimetic hydrogels (hyperGel+) are prepared using thiolated hyaluronic acid and hyperbranched polyglycerol with acrylate and sulfate groups. When combined with a soluble Rho kinase inhibitor, the cell-permissive nanoporous hydrogel supports the development of secretory spheroids with a central lumen from dispersed human primary salivary gland stem/progenitor cells.image (c) 2023 WILEY-VCH GmbH