Protein-based conductive, injectable, biodegradable hydrogels - Project Summary/Abstract Many cells are responsive to electrically conductive materials; however, to date electrical conductivity is mostly achieved through graphene or synthetic polymers. These materials have limited translational use due to a lack of biodegradability and rigid mechanical properties. To overcome these challenges, we propose the design of a recombinant engineered, conductive, injectable, and biodegradable hydrogel that has the potential to induce regeneration across a wide range of tissues. We have recently pioneered the synthesis of a fully recombinant gel that incorporates electrically-conductive protein nanowires (ePN), an engineered matrix-like protein, and the polysaccharide hyaluronic acid (HA). While the ePN provides conductivity, the engineered matrix-like protein and HA provide biochemical ligands that promote cell adhesion. The hydrogel material is crosslinked through dynamic covalent chemistry, allowing for tunable viscoelastic properties and injectability. The resulting gel supports three-dimensional cell culture and biodegrades in response to cell-secreted enzymes. As the spinal cord is an electrically conductive tissue, we will demonstrate the efficacy of our technology in a cell-based therapy for spinal cord injury (SCI). Less than 1% of SCI patients have full neurological recovery by the time of hospital discharge. We previously demonstrated with non-conductive hydrogels that intraspinal transplantation of neural progenitor cells (NPCs) can significantly improve function in a rodent SCI model, but only when they are sufficiently matured into a neuronal phenotype. We have also demonstrated that NPCs enhance their neuronal maturation in vitro when grown on conductive biomaterials that were rigid and non-biodegradable. Thus, we hypothesize that our new hydrogel will facilitate the intraspinal injection of NPCs and significantly promote their neuronal maturation, thus resulting in significant functional and histological improvements. In Aim 1, we identify the gel formulation that best promotes neuronal differentiation and maturation of human induced pluripotent stem cell-derived NPCs in vitro. Specifically, we will tune the bulk conductivity of the fabricated gels through altering the ePN concentration and amino acid sequence. Recombinant engineering of ePN allows for tunability of the electrical conductivity along a single protein wire. The cell morphology, gene expression, and protein expression of encapsulated NPCs in the gels without and with varying levels of conductivity will be quantified. In Aim 2, we will select the gel variant that provides the best in vitro results for assessment in a preclinical, rat model of cervical SCI. NPCs will be transplanted within the conductive, biodegradable gel and evaluated for functional behavior over 6 weeks. Histological outcomes include transplanted cell survival and neurite outgrowth. Controls include conductive gels without cells and non-conductive gels with cells. This study would represent the first use of conductive, biodegradable, recombinant nanowires in tissue engineering, which can have broad application in conductive tissues including brain, cardiac muscle, skeletal muscle, and skin.
