Functionalized Biomimetic Hydrogels Enhance Salivary Stem/Progenitor Cell Organization

Abstract

Complex branched salivary structures remain challenging to replicate within implant ready hydrogels. We showed previously that hyaluronic acid (HA)-based hydrogels enable growth and organization of primary salivary-derived human stem/progenitor cells (hS/PCs) into multicellular spheroids. Here, we systematically functionalized three components of migration-permissive hydrogels to foster salivary tissue morphogenesis. We separately analyzed contributions of an enzymatically degradable crosslinker, a pendant integrin-binding site, and hydrogel porosity to best support high viability, integrin-dependent cell adhesion and migration. Structure size, frequency, and morphology were all affected by hydrogel crosslink density and integration of biofunctional peptides. Viability and proliferation data suggested that integration of integrin binding sites had the greatest effect on hS/PCs behavior. A larger internal matrix space, created by increasing both crosslinker length and PEG content, was needed to form large multicellular hS/PC structures. Peptide-modified hydrogels with more internal space shifted hS/PC organization from spheroidal, surrounded by thick basement membrane, to an asymmetric arrangement with punctate matrix proteins defining a “wrinkled” perimeter. Integrin-binding peptides activated integrin β1, with highest activation observed in hydrogels having both cleavable peptide and integrin ligand. The design parameters we prescribe allowed us to encapsulate hS/PCs in a humanized biomimetic hydrogel matrix able to support morphogenesis needed for salivary restoration.

Hyaluronan Hydrogel “Safety Nets” for 3D Cell Culture Applications

“…The MMP degradation data in Figure 3c is an interesting contrast to one of our prior systems (Martinez et al 2021), which compared similar HA hydrogels, formed with either 100% PQ crosslinker, 100% PQ*, or 100% PEGDA. In that work by Martinez et al, gel pucks expanded in size when incubated with MMP-2, while the present work showed that susceptible gel pucks contracted.…”

Bone Marrow Endothelial Cells Increase Prostate Cancer Cell Apoptosis in 3D Triculture Model of Reactive Stroma

“…Cells were resuspended in HA-SH solution, and then acrylate-PEG-GRGDS (73.7 mg/mL in PBS), acrylate-PEG-YIGSR (73.7 mg/mL), and acrylate-PEG-PQ-PEG-acrylate (37.0 mg/mL in PBS) were added at a volume ratio of 4:0.5:0.5:0.2:0.8 (HA-SH:acrylate-PEG-GRGDS:acrylate-PEG-YIGSR:acrylate-PEG-PQ-PEG-acrylate:PBS) for all experiments. The solution was mixed well, and then 42 µL of gel solution was dispensed into each well cavity of the PDMS mold, as described previously [ 17 ]. Filled molds were placed into the cell culture incubator at 37 °C for 45 min to allow gelation to occur; then, one drop of cell culture media was added to the top of each gel to prevent dehydration, and the molds were placed back into the incubator for 1–2 h. This allowed low-crosslinking-density hydrogels to crosslink more completely for ease of handling before the gels were scored around their circumference with the tip of a sterile needle.…”

Hydrogels for Salivary Gland Tissue Engineering

“…ECM-derived hydrogels contain these motifs, such as RGD (Arginine-Glycine-Aspartic Acid), DGEA (Aspartic acid-Glycine-Glutamic acid-Alanine), YIGSR (Tyrosine-Isoleucine-Glycine-Serine-Arginine) [ 30 ], and heparan sulfate binding domains, which inherently support cell attachment, as opposed to natural bioinert materials such as alginate. So far, limited studies have evaluated the specific benefits of modifying and integrating binding ligands for SG three-dimensional culture [ 31 , 32 ].…”