Neural Stem Cell Differentiation in 2D and 3D Microenvironments

Abstract

Neural Stem Cells (NSCs) have tremendous potential for tissue engineering applications because of their high regenerative capacity to promote functional recovery following disease and injury in the central nervous system. Despite their great potential, current methods to culture NSCs are limited; e.g., adherent 2D cultures are greatly simplified vs. the in vivo microenvironment by imposing altered tissue-specific architecture and mechanical and biochemical cues, and cell morphology. Environmental cues are critical for cellular maturation and function and in vivo these are presented in a 3D environment. Recent studies with non-neuronal cells demonstrate that in a 3D matrix, cells dramatically alter their morphology and signaling pathways, with in vitro 3D environments being a better representation of in vivo systems. The main goal of this study is to define how NSC differentiation and cell-matrix signaling is altered in 2D and 3D systems. We hypothesize that 3D culture imposes changes in matrix-ligand organization and alters NSC behavior by modulating cytoskeletal signaling and differentiation outcome. To test our hypothesis we cultured mouse embryonic NSCs in 2D and 3D biomaterials and observed differences in cell behavior and β1 - integrin signaling with altered culture dimensionality using immunocytochemistry and flow cytometry. In this study we show that NSCs sense the dimensionality of their environment and alter motility: in 3D, individual cells adapt a random migration pattern and extend longer neurites than in 2D where the cells undergo chain migration. In addition, the differentiation of the NSCs into the neuronal phenotype is increased in 2D vs 3D culture. These results confirm our hypothesis and provide a foundation to design optimal biomaterials towards the development of therapeutics for nerve repair and neurodegenerative disorders.