Cell-surface markers for the isolation of pancreatic cell types derived from human embryonic stem cells

Abstract

Using a flow cytometry–based screen of commercial antibodies, we have identified cell-surface markers for the separation of pancreatic cell types derived from human embryonic stem (hES) cells. We show enrichment of pancreatic endoderm cells using CD142 and of endocrine cells using CD200 and CD318. After transplantation into mice, enriched pancreatic endoderm cells give rise to all the pancreatic lineages, including functional insulin-producing cells, demonstrating that they are pancreatic progenitors. In contrast, implanted, enriched polyhormonal endocrine cells principally give rise to glucagon cells. These antibodies will aid investigations that use pancreatic cells generated from pluripotent stem cells to study diabetes and pancreas biology.

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Figure 1: Cell-surface markers for endocrine and PE cells in hES cell–derived pancreatic cultures.
Figure 2: FACS isolation of CD142 and CD200 cell subsets from hES cell–derived pancreatic cultures.
Figure 3: CD142, CD200 and CD318 immuno-magnetic cell separations from hES cell–derived pancreatic cultures.
Figure 4: Immunofluorescence analyses of transplants of unenriched or CD318-enriched endocrine cells.
Figure 5: Immunofluorescence and functional analyses of transplants of unenriched or CD142-enriched PE cells.

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Acknowledgements

We thank O. Madsen (Hagedorn Research Institute), C. Wright (Vanderbilt University), J. Johnson (UT Southwestern Medical Center) and BD Biosciences for providing antibodies and A. Elefanty and E. Stanley (Monash University) for providing the MEL1 hES cell line. The CyT203 and CyT49 hES cell lines were derived with partial funding from the Juvenile Diabetes Research Foundation.

Author information

O.G.K. and A.G.B. wrote the paper. O.G.K. and A.G.B. designed, directed and interpreted experiments with intellectual contributions from E.E.B., M.M., L.A.M., E.K., K.A.D., K.K. and M.K.C. The antibody screen was proposed by A.G.B. and carried out by O.G.K., M.Y.C. and M.M. K.A.D. suggested the Y-27632 compound. M.M. developed and performed the flow cytometry assays and analyses with assistance from M.Y.C. and K.G.R. O.G.K., A.G.B., M.Y.C. and T.M.O. performed the cell culture experiments and immuno-magnetic cell separations. M.Y.C. and O.G.K. performed qPCR and immunofluorescence analyses of in vitro material. L.A.M., E.K. and M.R. executed the in vivo experiments, including transplantations and C-peptide assays. K.K. performed the histological and immunofluorescence analyses of implanted grafts.

Correspondence to Olivia G Kelly.

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The authors are employees or former employees of ViaCyte (formerly Novocell).

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Supplementary Table 1 and Supplementary Figures 1–9 (PDF 2938 kb)

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Kelly, O., Chan, M., Martinson, L. et al. Cell-surface markers for the isolation of pancreatic cell types derived from human embryonic stem cells. Nat Biotechnol 29, 750–756 (2011). https://doi.org/10.1038/nbt.1931

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