Differentiation of embryonic stem cells into neural lineages in an alginate encapsulation microenvironment.

摘要

Cell replacement therapies, using renewable stem cell sources, hold tremendous potential to treat a wide range of degenerative diseases. Although many studies have established techniques to successfully differentiate stem cells into different mature cell lineages using growth factors or extracellular matrix protein supplementation in both two and three-dimensional configurations, their practicality is limited by lack of control, low yields of differentiated cells and oftentimes, heterogeneous cell population outcomes. In order to address these issues, we have previously established a murine embryonic stem cell alginate-poly-l-lysine microencapsulation differentiation system. The three-dimensional alginate microenvironment maintains cell viability, is conducive to ES cell differentiation to hepatocyte lineage cells, and sustains differentiated cellular function. In addition, hepatocyte function was contingent upon aggregate formation within the alginate microbeads. The present studies were designed to determine the feasibility of adapting the alginate encapsulation technique to neuronal lineage differentiation. The results of our studies indicate that by incorporating the soluble inducer, retinoic acid into the permeable microcapsule system, cell aggregation was decreased and neuronal lineage differentiation enhanced. In conjunction with the mechanical and physical characterization of the alginate crosslinking network, we have determined that 2.2% alginate microencapsulation can be optimally adapted to both hepatocyte and neuronal differentiation from embryonic stem cells. However, differentiation could be directed away from the hepatocyte and towards the neural lineage by lowering initial seeding density and physical cell-cell aggregation blocking, even in the absence of RA. This study promises to offer insights into targeting cellular differentiation towards both endodermal and ectodermal cell lineages, and could potentially be generalizable and adaptable to the differentiation of other stem cell types given the correct inducible factors and material properties.