(170e) Mesenchymal Stem Cell Motility in 3D Hydrogel Scaffolds

Currently, a large amount of effort is being put forth on combining marrow progenitor cells, or mesenchymal stem cells (MSCs) with 3D scaffolds to direct bone regeneration in critical size bone defects. Consequently, many people are investigating how the bio-physical and ?chemical properties of 3D scaffolds can facilitate MSC survival, proliferation, and differentiation into appropriate lineages. However, little is known about how implanted MSCs migrate through these scaffolds. Here, we present a synthetic poly(ethylene glycol) (PEG) system in which adhesivity, matrix stiffness, and porosity can be independently tuned. We are using this highly tunable system to analyze distinct biophysical features of 3D scaffolds on MSC migration. Combined with existing data on MSC proliferation, survival, and differentiation in 3D scaffolds by our lab and others, this information on MSC motility could be very powerful for future intelligent scaffold design for MSC-directed bone regeneration in vivo.

To create hydrogels with a macroporous internal structure, we have adapted and modified a previously published method. Briefly, poly(methyl methacrylate) (PMMA) microparticles are forced into an ordered array and dried. A polymer solution based on PEG dimethacrylate (PEGDMA) is then poured around the ordered PMMA beads and polymerized via UV-light. Post-polymerization, the PMMA beads are leached away from the hydrogels, leaving a PEG-based hydrogel with internal macropores. The concentration of the crosslinker PEGDMA (from 10-34%) and a hetero-bifunctional PEG-methacrylate (PEGMA) (from 1-12.5 mM) are varied to control the bulk mechanical properties and the cell-adhesive ligand density, respectively. Immortalized marrow derived stem cells (hTERT MSCs) are seeded into the porous hydrogels, and their migration tracked via confocal microscopy.

In these 3D scaffold-like hydrogels, the interconnected pore sizes can be tightly controlled from 7-17um independently of stiffness, the bulk mechanical properties range from 10 to over 1000kPa independent of pore size (following a predictive model), and the concentration of adhesive ligand can be tuned independent of stiffness. Thus far, we have seen that pore size is the dominating biophysical cue in controlling MSC migration speed in these gels, with average cell speeds hovering around 10um/hr in gels with 7um pores, to over 30um/hr in gels with 17um pores. The effects of adhesive ligand density and stiffness on MSC migration in these 3D scaffolds (in comparison to 2D analogous systems) will also be discussed.