Differentiation of Embryonic Stem Cells into Neural Cells on 3D Poly (D,L-Lactic Acid) Scaffolds versus 2D Cultures

Abstract

In this study, a highly porous poly (D, L-lactic acid) (PDLLA) scaffold was designed and fabricated using dioxane and thermal-induced phase separation (TIPS) methods (liquid-liquid and solid-liquid). Additionally, we characterized the ability of mouse embryonic stem cells (ESCs) to differentiate into neural cells in PDLLA scaffold with uniform porosity, interconnectivity, and high porosity, and then compared them with cells seeded under conventional two-dimensional (2D) culture conditions. Histochemistry staining showed the migration of differentiated cells through the scaffold. Immunofluorescence analysis of the differentiated cells by counting positive cells revealed that the PDLLA scaffold resulted in a significantly greater number of neural markers, microtubule associated protein-2, β-tubulin III, neurofilament protein, and glial fibrillary acidic protein (the astrocyte marker) when compared to those in 2D culture condition. Moreover, the expression of Nestin, Mash1, Pax6, and HB9 increased significantly in 3D scaffolds when compared with 2D cultures as detected by semi-quantitative RT-PCR. Scanning electron microscopy of differentiated neurons on scaffolds also demonstrated favorable results for neurite outgrowth. The results of this study demonstrated a promising effect of 3D scaffold culture for neural cell differentiation from ESCs in prospective tissue engineering applications.