Engineering Highly Porous, Fiber-Reinforced Granular Hydrogel Scaffolds

Whitewolf, Jack, Biomedical Engineering - School of Engineering and Applied Science, University of Virginia
Highley, Chris, EN-Biomed Engr Dept, University of Virginia

This dissertation spans two separate projects I led during my PhD. The first project investigated the conformal encapsulation of stem cells using modified hyaluronic acids: Microand nanoencapsulation of cells has been studied as a strategy to protect cells from environmental stress and promote survival during delivery. Hydrogels used in encapsulation can be modified to immunoisolate cells as well as to influence cell behaviors and direct where they collect or are assembled in their surroundings. Here, we report a system that conformally encapsulated stem cells using hyaluronic acid (HA). We successfully modified HA with lipid, thiol, and maleimide pendant groups to facilitate a hydrogel system in which HA was deposited onto cell plasma membranes and subsequently crosslinked through thiol-maleimide click chemistry. We demonstrated conformal encapsulation of both neural stem cells (NSCs) and mesenchymal stromal cells (MSCs), with viability of both cell types greater than 90% after encapsulation. Additional material could be added to the conformal hydrogel through alternating addition of thiol-modified and maleimide-modified HA in a layering process. After encapsulation, we tracked egress and viability of the cells over days and observed differential responses of cell types to conformal hydrogels both according to cell type and the amount of material deposited on the cell surfaces. Through the design of the conformal hydrogels, we showed that multicellular assembly could be created in suspension and that encapsulated cells could be immobilized on surfaces. In conjunction with photolithography, conformal hydrogels enabled rapid assembly of encapsulated cells on hydrogel substrates with resolution at the scale of 100 μm.
The second project was aimed to develop a new biomaterial platform using highly porous granular scaffolds: scaffolds composed of crosslinked hydrogel microparticles (HMPs), or granular hydrogel scaffolds, contain pore spaces much greater in size than conventional bulk hydrogels. Packing of HMPs creates interconnected, micron-scaled openings that are preserved upon crosslinking. Packed HMP scaffolds, also known as microporous annealed particle (MAP) scaffolds, greatly improve cell migration and proliferation. However, while higher porosity allows for more cellular infiltration, it also reduces the mechanical integrity of the scaffolds. Here, we address this challenge by demonstrating 1) the fabrication of high-porosity granular scaffolds beyond what is possible from particle packing using sacrificial HMPs; and 2) stabilization of the high-porosity scaffolds by incorporating electrospun hydrogel fibers within the scaffolds. We fabricated granular scaffolds using norbornene-modified hyaluronic acid (norHA) HMPs and gelatin HMPs as the sacrificial population. We created HMP scaffolds with up to 50% porosity by incorporating (0-50 vol.%) sacrificial gelatin HMPs. When electrospun norHA fibers were incorporated in scaffolds at 5-10 vol.%, highly porous scaffolds retained their structure over a period of 28 days. Additionally, HUVECs were mixed with HMPs prior to scaffold formation and exhibited high viability. The HUVEC-HMP mixture could be injected through a needle and then crosslinked to form a scaffold, with post-injection viability >80%. Overall, this study demonstrates the development and characterization of a stable, highly porous granular scaffold system with high processibility and cytocompatibility.

PHD (Doctor of Philosophy)
Biomaterials, Granular hydrogel scaffold, Nanoencapsulation, Hyaluronic acid
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