Mechanotransductive Engineering of Neural Stem Cell Behavior

Abstract

Neural stem cells (NSCs) play important roles in learning and memory in the adult mammalian brain and may also serve as a source of cells in cell replacement therapies to treat neurodegenerative diseases. Therefore, investigating how NSC behavior is regulated is crucial to understanding the fundamental biology of the brain as well as in engineering biomedical therapies. Towards these ends, an increasing wealth of knowledge in the NSC field describes a complex picture of biochemical and genetic regulation of NSC self-renewal and differentiation. However, little is known about the biophysical control of NSC behavior by the extracellular matrix (ECM). Here we demonstrate that ECM-derived mechanical signals can act with Rho GTPases to regulate NSC stiffness and differentiation. Culturing NSCs on increasingly stiff ECMs suppresses neurogenesis and enhances gliogenesis, even in the absence of exogenous differentiating agents. This shift is accompanied by enhanced RhoA and Cdc42 activation and increased cellular stiffness. Direct manipulation of RhoA and Cdc42 activity disrupts the ability of NSCs to sense ECM stiffness and tips the balance between neurogenesis and gliogenesis in the presence and absence of exogenous differentiation cues. Inhibitors of a downstream effector of RhoA, Rho kinase, as well as inhibition of myosin II contractility rescues neuronal differentiation of NSCs cultured on stiff substrates as well as for NSCs expressing CA RhoA and CA Cdc42, suggesting that NSC stiffness/contractility regulates NSC differentiation. These results establish Rho GTPase-based mechanotransduction and cellular stiffness as novel regulators of NSC behavior.