Three-Dimensional Differentiation of Embryonic Stem Cells into Islet-like Insulin-Producing Clusters

Abstract

The production of mature pancreatic cells that function similarly to primary islets is the premise of cell therapy for diabetes. Here, we describe a novel approach to generating more mature insulin-producing cells from embryonic stem (ES) cells. A three-dimensional (3D) ES cell pancreatic differentiation system was developed and used to direct the ES cell differentiation into glucose-responsive, insulin-secreting cells. Using mouse ES cells as a model, we demonstrate that more mature insulin-producing cells can be generated from ES cells in 3D cultures. The 3D differentiated pancreatic endocrine cells can assemble into an islet-like tissue structure that displays greater similarities in phenotype and gene expression profile to adult mouse pancreatic islets, that is, with beta cells in the core and non-beta cells forming the mantel, leading to a significant improvement of the maturity of the insulin-producing cells. Our findings show that nearly 50-60% of the cells in 3D formed cell clusters express insulin. More importantly, those cells exhibit a high level of glucose-responsive insulin and C-peptide syntheses and release. A high level of expression of glucose transporter-2 was also detected in these cells. Compared to two-dimensional ES cell-derived insulin-producing cells, the insulin release from 3D ES cell-derived insulin-producing cells showed a nearly fivefold (p<0.05) increase when exposed to a high glucose (27.7 mM) medium. This 3D culture model provides an excellent system to study pancreatic endocrine morphogenesis and tissue organization. This study also demonstrates the feasibility of producing clinically relevant beta cells from ES cells in a 3D environment.