Hydrogels are hydrophilic polymeric matrices that can support cell and tissue growth in three-dimension. With careful design, hydrogels serve not only as a 3D platform for studying cell behaviors, but also as scaffolds for culturing and differentiating stem/progenitor cells.
We aim to develop a multifunctional hydrogel system that initially supports cell proliferation, but at a later stage can be "switched" into a microenvironment that promotes cell/tissue differentiation into a specific cell type. Our central hypothesis is that cell expansion and differentiation can be achieved in a single hydrogel matrix incorporated with dynamic biophysical and biochemical cues. We will test this hypothesis by developing a versatile hydrogel system using multiple cytocompatible thiol-ene "click" reactions. In the initial stage, we will design locally degradable thiol-ene gels to promote proliferation of PANC-1 cells (a pancreatic ductal epithelial cell line) (Aim 1). We will incorporate matrix metalloproteinase 2 (MMP-2) specific peptide substrates as hydrogel crosslinkers. This allows the gels to degrade locally (thus creating additional space for cell proliferation) by MMP-2 secreted from PANC-1 cells. The proliferation of encapsulated cells will be promoted by tuning cell-ECM and cell-cell interactions in hydrogels, as well as providing the encapsulated cells with diffusible soluble growth factors. Next, we will "switch" the cell-laden hydrogels from a "pro-proliferation" to a "pro-differentiation" microenvironment (Aim 2a). We will achieve this by performing a second thiol-ene click reaction to introduce pro-differentiation cues within the cell-laden hydrogels, thus promoting the formation of islet-like, insulin secreting cell clusters. The differentiation process will be a combinatorial result of using serum-free culture media, affinity-based recruitment of basic fibroblast growth factor (bFGF), and timely conjugation of glucagon-like peptide 1 (GLP-1). The differentiated cell clusters can be retrieved from the erodible gels (due to specific enzyme activity) for further characterization and biological/clinical applications (Aim 2b). The differentiated clusters are expected to form tight aggregates due to strong cell-cell interactions. Together, our strategies facilitate the formation of islet-like cell clusters with natural cell-cell interactions and normal insulin secretion profiles.
This proposal aims to design biomaterial devices that incorporate signals to promote the proliferation of epithelial cells and to enhance the differentiation of these cells into insulin-producing cells. If successful, this strategy will provide alternative cell sources to benefit type 1 diabetic patients.
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