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VMAT2 identified as a regulator of late-stage β-cell differentiation

Nature Chemical Biology volume 10, pages 141–148 (2014)Cite this article

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Abstract

Cell replacement therapy for diabetes mellitus requires cost-effective generation of high-quality, insulin-producing, pancreatic β 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. 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 β cells for cell therapy.

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Figure 1: Reserpine identified as a chemical that enhanced ES cell differentiation into pancreatic β cells.
Figure 2: VMAT2- and monoamine-dependent suppression of pancreatic β-cell differentiation.
Figure 3: VMAT2 inhibition increased differentiation into Ngn3-GFP+ cells.
Figure 4: TBZ and dBu-cAMP additively potentiated differentiation into β cells.
Figure 5: Characterization of the purified ES cell–derived Ins+ and Pdx-GFP+ cells.
Figure 6: Transplanted cells reversed hyperglycemia and glucose tolerance.

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Acknowledgements

We thank members of the Gene Technology Center and Center for Animal Resources and Development at Kumamoto University for technical assistance. This work was supported by the Funding Program for Next Generation World-Leading Researchers (to S.K. (no. LS099) and M.U.); the Japan Society for the Promotion of Science, the Realization of Regenerative Medicine (to S.K. and M.U.); the Program for Leading Graduate Schools 'HIGO' (awarded to S.K.); Grants-in-Aid from the Ministry of Education, Culture, Sports, Science and Technology (MEXT), Japan (no. 21390280 to S.K. and no. 22790653 to D. Sakano); and the Collaborative Research Program of Institute for Chemical Research, Kyoto University (grant no. 2010-44). The iCeMS is supported by World Premier International Research Center Initiative, MEXT, Japan.

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  1. Kazuhiko Kume

    Present address: Present address: Department of Neuropharmacology, Graduate School of Pharmaceutical Sciences, Nagoya City University, Nagoya, Japan.,

Authors and Affiliations

  1. Department of Stem Cell Biology, Institute of Molecular Embryology and Genetics, Kumamoto University, Kumamoto, Japan

    Daisuke Sakano, Nobuaki Shiraki, Kazuhide Kikawa, Taiji Yamazoe, Masateru Kataoka, Kahoko Umeda, Kazuhiko Kume & Shoen Kume

  2. Department of Pediatrics, Graduate School of Medical Sciences, Kumamoto University, Kumamoto, Japan

    Kazuhide Kikawa, Shirou Matsumoto & Fumio Endo

  3. Laboratory of Developmental Genetics, Institute of Resource Development and Analysis, Kumamoto University, Kumamoto, Japan

    Kimi Araki

  4. Institute for Chemical Research and Institute for Integrated Cell-Material Sciences, (WPI-iCeMS), Kyoto University, Kyoto, Japan

    Di Mao & Motonari Uesugi

  5. Division of Reproductive Engineering, Center for Animal Resources and Development, Kumamoto University, Kumamoto, Japan

    Naomi Nakagata

  6. Department of Cell and Molecular Biology, Karolinska Institute, Stockholm, Sweden

    Olov Andersson & Didier Stainier

  7. Department of Biochemistry and Biophysics, University of California–San Francisco, San Francisco, California, USA

    Olov Andersson

  8. Department of Developmental Genetics, Max Planck Institute for Heart and Lung Research, Bad Nauheim, Germany

    Didier Stainier

  9. Program for Leading Graduate Schools, Health Life Science Interdisciplinary and Global Oriented (HIGO) Program, Kumamoto University, Kumamoto, Japan

    Shoen Kume

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Contributions

D. Sakano performed chemical screening, cellular and biochemical analyses; D. Sakano, N.S. and K.U. established the ES cell differentiation system; D. Sakano, K. Kikawa, M.K. and T.Y. performed transplantation assays; K.A. established the ES cell line; S.M., F.E. and N.N. helped maintain AKITA mice; D.M. and M.U. provided and analyzed the chemical library; O.A. and D. Stainier provided chemicals; K. Kume and S.K. provided technical advices, S.K. designed the experiments and wrote the paper. All of the authors discussed the results and commented on the manuscript.

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Correspondence to Shoen Kume.

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Sakano, D., Shiraki, N., Kikawa, K. et al. VMAT2 identified as a regulator of late-stage β-cell differentiation. Nat Chem Biol 10, 141–148 (2014). https://doi.org/10.1038/nchembio.1410

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