Controlling size, shape and homogeneity of embryoid bodies using poly(ethylene glycol) microwells

TLDR

Directed differentiation of embryonic stem cells is valuable for disease modeling and therapeutic cell generation, yet conventional suspension cultures produce embryoid bodies with heterogeneous sizes and shapes that can affect differentiation outcomes. The authors employ microfabricated poly(ethylene glycol) wells to template the formation of homogeneous embryoid bodies. Embryoid bodies formed in PEG microwells displayed controlled, uniform sizes and shapes, maintained high viability (>95 %), and could be readily retrieved, demonstrating the method’s potential for generating homogeneous embryoid bodies for in‑vitro differentiation studies.

Abstract

Directed differentiation of embryonic stem (ES) cells is useful for creating models of human disease and could potentially generate a wide array of functional cell types for therapeutic applications. Methods to differentiate ES cells often involve the formation of cell aggregates called embryoid bodies (EBs), which recapitulate early stages of embryonic development. EBs are typically made from suspension cultures, resulting in heterogeneous structures with a wide range of sizes and shapes, which may influence differentiation. Here, we use microfabricated cell-repellant poly(ethylene glycol) (PEG) wells as templates to initiate the formation of homogenous EBs. ES cell aggregates were formed with controlled sizes and shapes defined by the geometry of the microwells. EBs generated in this manner remained viable and maintained their size and shape within the microwells relative to their suspension counterparts. Intact EBs could be easily retrieved from the microwells with high viability (>95%). These results suggest that the microwell technique could be a useful approach for in vitro studies involving ES cells and, more specifically, for initiating the differentiation of EBs of greater uniformity based on controlled microenvironments.