Development of a Synthetic Extracellular Matrix for Augmentation of Placental-derived Stem Cell Central Nervous System Therapy

Introduction: The prevalence of spinal cord injury (SCI) in the United States is estimated between 250,000-363,000, with an estimated 17,000 new cases each year. Following injury, inflammation of central nervous system tissue leads to neuronal and glial cell loss, axonal degeneration, extracellular matrix inhibition, and glial scar formation, all of which pose an impediment to central nervous system regeneration after injury. Our goal was to develop an injectable synthetic extracellular matrix (sECM) that augments stem cell therapy and provides a microenvironment that favors neural tissue regeneration over glial scar formation.

Methods: Hydrogels were formed using Michael addition of acrylate-functionalized hyaluronic acid (HA-AC) with a linker peptide and pro-neurogenic and angiogenic peptides. Rheology was conducted to determine the stiffness of the sECM. H1 NMR was performed to analyze each step of the chemical formation of the hydrogel to confirm the presence of appropriate functional groups. Neurons and placental-derived stem cells (PSC) were co-cultured in a non-adherent 96-well plate both with and without the synthetic extracellular matrix. Cells in sECM were incubated for 7 days before fixing and immunostaining for further analysis.

Results: This novel sECM composition transitions from a liquid to a hydrogel at 37°C (body temperature), has a stiffness of 350-400 Pa which emulates the softness of the brain and spinal cord, and forms a hydrogel at 37°C with cells present within the initial liquid solution. Neurons co-cultured with PSCs in the sECM formed tissue in vitro in which neurons spread, formed neurites and neuronal networks.

Conclusion : This novel sECM is injectable, is soft (350-400 Pa), and supports in vitro tissue formation with viable neurons, neurite formation, and most importantly the formation of neuronal networks. Overall, the biomechanical properties of the sECM are unique and distinctively engineered for the treatment of injured central nervous system tissue.