Microfluidic fabrication of bioactive microgels for rapid formation and enhanced differentiation of stem cell spheroids

A major challenge in tissue engineering is to develop robust protocols for differentiating ES and iPS cells to functional adult tissues at a clinically relevant scale. The goal of this study is to develop a high throughput platform for generating bioactive, stem cell-laden microgels to direct differentiation in a well-defined microenvironment. We describe a droplet microfluidics system for fabricating microgels composed of polyethylene glycol and heparin, with tunable geometric, mechanical, and chemical properties, at kHz rates. Heparin-containing hydrogel particles sequestered growth factors Nodal and FGF-2, which are implicated in specifying pluripotent cells to definitive endoderm. Mouse ESCs were encapsulated into heparin microgels with a single dose of Nodal and FGF-2, and expressed high levels of endoderm markers Sox17 and FoxA2 after 5 days. These results highlight the use of microencapsulation for tailoring the stem cell microenvironment to promote directed differentiation, and may provide a straightforward path to large scale bioprocessing in the future. Statement of Significance Multicellular spheroids and microtissues are valuable for tissue engineering, but fabrication approaches typically sacrifice either precision or throughput. Microfluidic encapsulation in polymeric biomaterials is a promising technique for rapidly generating cell aggregates with excellent control of microenvironmental parameters. Here we describe the microfluidic fabrication of bioactive, heparin-based microgels, and demonstrate the adsorption of heparin-binding growth factors for enhancing directed differentiation of embryonic stem cells toward endoderm. This approach also facilitated a ∼90-fold decrease in consumption of exogenous growth factors compared to conventional differentiation protocols.