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Cell replacement therapy for diabetes mellitus requires cost-effective generation of high-quality, insulin-producing, pancreatic b cells from pluripotent stem cells. Development of this technique has been hampered by a lack of knowledge of the molecular mechanisms underlying β -cell differentiation. The present study identified reserpine and tetrabenazine (TBZ), both vesicular monoamine transporter 2 (VMAT2) inhibitors, as promoters of late-stage differentiation of Pdx1 -positive pancreatic progenitor cells into Neurog3 (referred to henceforth as Ngn3) -positive endocrine precursors (Figure) . VMAT2-controlled monoamines, such as dopamine, histamine and serotonin, negatively regulated β -cell differentiation. Reserpine or TBZ acted additively with dibutyryl adenosine 3′,5′-cyclic AMP, a cell-permeable cAMP analog, to potentiate differentiation of embryonic stem (ES) cells into β cells that exhibited glucose-stimulated insulin secretion. When ES cell–derived β cells were transplanted into AKITA diabetic mice, the cells reversed hyperglycemia. Our protocol provides a basis for the understanding of β -cell differentiation and its application to a cost-effective production of functional b cells for cell therapy.

 

ES cells differentiate through the mesendoderm, the definitive endoderm, pancreatic progenitor s and the endocrine progenitor stage, until mature β-cells. The cytosolic monoamine storage decreases when VMAT2 is inhibited. It leads to the increasing of the β-cell induction rate . This is through promoting differentiation of endocrine progenitors from pancreatic progenitors. Moreover, cell permeable cAMP renders the glucose-dependent insulin secreting abilities. It is possible to induce mature β -cell s by using VMAT2 inhibitor and cAMP.