Influence of In Vitro and In Vivo Oxygen Modulation on β Cell Differentiation From Human Embryonic Stem Cells

The possibility of using human embryonic stem (hES) cell-derived beta cells as an alternative to cadaveric islets for the treatment of type 1 diabetes is now widely acknowledged. However, current differentiation methods consistently fail to generate meaningful numbers of mature, functional beta cells. In order to address this issue, we set out to explore the role of oxygen modulation in the maturation of pancreatic progenitor (PP) cells differentiated from hES cells. We have previously determined that oxygenation is a powerful driver of murine PP differentiation along the endocrine lineage of the pancreas. We hypothesized that targeting physiological oxygen partial pressure (pO(2)) levels seen in mature islets would help the differentiation of PP cells along the beta-cell lineage. This hypothesis was tested both in vivo (by exposing PP-transplanted immunodeficient mice to a daily hyperbaric oxygen regimen) and in vitro (by allowing PP cells to mature in a pet-Fluorocarbon-based culture device designed to carefully adjust pO(2) to a desired range). Our results show that oxygen modulation does indeed contribute to enhanced maturation of PP cells, as evidenced by improved engraftment, segregation of alpha and beta cells, body weight maintenance, and rate of diabetes reversal in vivo, and by elevated expression of pancreatic endocrine makers, beta-cell differentiation yield, and insulin production in vitro. Our studies confirm the importance of oxygen modulation as a key variable to consider in the design of beta-cell differentiation protocols and open the door to future strategies for the transplantation of fully mature beta cells.