Neuronal Outgrowth and Differentiation on Poly(glycerol sebacate)

ABSTRACTThe importance of the extracellular mechanical environment in stem cell differentiation has been extensively studied over the last decade. In neuronal cell differentiation, matrix stiffness and neurite outgrowth have been correlated, highlighting the impact of matrix effects on neuronal cell morphology. In addition, on materials that approach the physiological mechanical properties of brain tissue, neurons from mixed phenotype primary cultures will prevail. However, if the same mixed culture is grown on polystyrene, glial populations are more prevalent. Enhancing the understanding of these differentiation processes will further expand the ability to design materials for neuronal implants that are conducive to neuronal survival, resist glial scarring and promote neurite outgrowth and cell connectivity. Specifically, elastomers such as poly(glycerol sebacate) (PGS) hold promise in neuronal tissue engineering, due to their mechanical tunability. PGS is biocompatible, biodegradable and possesses mechanical properties similar to that of living tissue. Neuronal cell differentiation was studied on PGS, using P19 embryonic carcinoma cells, which can be differentiated into a neuronal phenotype using retinoic acid. Varying cure temperatures of PGS including 120°, 140° and 165°C were selected, which equate to an elastic modulus of 0.07, 0.43 and 2.30 MPa respectively. Cells were characterized via immunocytochemistry. A primarily astrocytic population, with limited neuronal differentiation and neurite outgrowth were observed on the PGS 120°C. Cells grown on PGS 140°C demonstrated marked neurite outgrowth, with an increase in neuronal cells. Cells grown on the PGS 165°C exhibited the largest population of neurons, with significant neurite outgrowth. These results indicate that substrate mechanical properties do impact neuronal differentiation, but that a material with a Young’s modulus similar to that of neuronal tissue (PGS 120°C) may not necessarily be the most conducive to in vitro differentiation.