Recapitulating endocrine cell clustering in culture promotes maturation of human stem-cell-derived β cells

Despite advances in the differentiation of insulin-producing cells from human embryonic stem cells, the generation of mature functional beta cells in vitro has remained elusive. To accomplish this goal, we have developed cell culture conditions to closely mimic events occurring during pancreatic islet organogenesis and beta cell maturation. In particular, we have focused on recapitulating endocrine cell clustering by isolating and reaggregating immature beta-like cells to form islet-sized enriched beta-clusters (eBCs). eBCs display physiological properties analogous to primary human beta cells, including robust dynamic insulin secretion, increased calcium signalling in response to secretagogues, and improved mitochondrial energization. Notably, endocrine cell clustering induces metabolic maturation by driving mitochondrial oxidative respiration, a process central to stimulus-secretion coupling in mature beta cells. eBCs display glucose-stimulated insulin secretion as early as three days after transplantation in mice. In summary, replicating aspects of endocrine cell clustering permits the generation of stem-cell-derived beta cells that resemble their endogenous counterparts.