Human embryonic stem cells (hESCs) hold great promises as sources of therapeutics due to their self-renewal and multilineage differentiation. Human ESCs can serve as a source of insulin-producing cells for diabetes patients. Current protocols for hESC differentiation to insulin-producing cells are characterized by low efficiency. This implicates that processes for the expansion and directed differentiation of hESCs in large quantities is highly desirable.

Here, we explored the use of microcarrier suspension culture for the propagation of hESCs and their subsequent differentiation towards islet cells. Microcarriers provide a high surface area-to-volume ratio, which is easily adjustable. Pluripotent hESCs seeded on microcarriers were able to attach and grow on the beads with viability of at least 90% for 10 days. A doubling time of ~48 hrs was calculated for hESCs in suspension culture compared to ~32 hrs for hESCs cultured on dishes. The proliferation of hESCs on microbeads depended on culture conditions including the agitation rate. In addition, the cells maintained high expression levels of pluripotency markers OCT4, NANOG, REX1, TRA-1-81 and SSEA4 as assessed by quantitative PCR, immunostaining and flow cytometry. These results support the use of microcarrier suspension systems for hESC expansion.

The observed high expression of pluripotency markers and low or absent expression of germ layer genes suggests that hESCs propagated on microbeads may retain their potential for further multi-lineage differentiation. Current work focuses on directing hESCs cultured in suspension towards endoderm progeny, especially pancreatic islet cells.