Assembly of Microgel Building Blocks into Cell-instructive Microporous Annealed Particle Hydrogels for Tissue Engineering

Abstract

Therapeutic cell delivery is a major strategy to stimulate the repair in tissue engineering and regenerative medicine. Efficacious cell therapies require a biomaterial carrier to retain injected cells in degenerated sites and guide cell function. Hydrogels, consisting of water-swollen polymeric network, are particularly of interest as cell delivery vehicles due to their structural similarity to native extracellular matrix. The common approach is to inject hydrogel precursor solutions with cells and cured in situ for encapsulation, which myriad evidences have suggested to improve the cell retention and viability. However, the resulting polymeric network are nanoporous, which form physical constrains for cell behaviors. Microporous annealed particle (MAP) hydrogels are an emerging class of biomaterials to overcome this challenge by providing permissive environment to cells. The overall goal of this work was to develop cellinstructive MAP hydrogels as a therapeutic cell delivery platform to improve regeneration outcomes. Norbornene-bearing poly(ethylene glycol) microgels were assembled into MAP hydrogels via thiol-ene click chemistry with the addition of bis-thiol linker and human mesenchymal stem cells were incorporated during microgel assembly. Since cells interact with microgel surface within MAP hydrogels, we demonstrated the possibility of leveraging the rich body of knowledge regarding cell behavior in 2D environments to direct cellular behavior within these 3D scaffolds. We revealed cells responded to stiffness by activating mechanosensing pathways. In addition, long-term cell spreading, proliferation, and microenvironmental remodeling were dependent critically on the susceptibility of the MAP hydrogels to degradation. The paracrine secretion of cells was influenced by both signaling from specific integrin-binding peptides and degradability of the MAP hydrogels. The healing outcomes of an optimized formulation was evaluated in a mouse femoral defect model, and significant new bone formation was seen using these MAP hydrogels. We further developed a novel microfluidic method to create physicochemical gradients in MAP hydrogels for screening cell-material interactions. Last, we showed the use of microgels as biomaterial inks to construct MAP hydrogels with anatomically relevant structures in 3D printing.