(195a) Characterizing the hMSC-Mediated Remodeling of Polymer-Peptide Hydrogels on Multiple Length Scales Using Bi-Disperse Multiple Particle Tracking Microrheology

Human mesenchymal stem cells (hMSCs) migrate out of the bone marrow to wounds to coordinate the healing process. hMSCs use matrix metalloproteinases (MMPs) and cytoskeletal tension to degrade and remodel their microenvironment, enabling migration. To study this cell-mediated remodeling, we encapsulate hMSCs in a well-defined hydrogel, cross-linking norbornene-functionalized 4-arm poly(ethylene glycol) with an MMP-cleavable peptide cross-linker. We also embed fluorescently labeled probe particles to enable multiple particle tracking microrheology (MPT) measurements. MPT is used to quantify local rheology around a migrating hMSC by measuring the Brownian motion of the particles in the pericellular region. hMSC remodeling impacts the material on several length scales. In order for a cell to migrate, it must use MMPs to break individual cross-links on the nanometer scale. The cell then pulls itself forward using its cellular extensions which are on the order of 1 µm in diameter. Finally, the collective remodeling activity and migration of all encapsulated cells results in bulk degradation of the entire hydrogel. In order to characterize rheological changes on several length scales, we extend traditional MPT by using multiple particle sizes simultaneously in a technique called bi-disperse MPT. We characterize cellular remodeling by measuring the motion of 0.5 and 2.0 µm particles simultaneously. As a control, we degrade the material using collagenase in the absence of cells and measure the same degradation on each scale. Cell-mediated degradation remodels the material more on the 2.0 µm length scale than on the 0.5 µm scale. We then inhibit cytoskeletal tension and measure the same remodeling on both the 0.5 and 2.0 µm length scales. This demonstrates that cytoskeletal tension is responsible for the increased remodeling of the larger length scale. In separate experiments we encapsulate cells with 4.5 µm particles alone to measure a larger length scale. We measure that cell speed correlates differently with material rheology on each length scale, emphasizing the importance of length scale dependent remodeling during hMSC migration. These findings can be used to enhance the design of implantable materials for hMSC delivery by providing a better understanding of how length scale can be incorporated into material structure to instruct cell behavior.