Platform for Induction and Maintenance of Transgene-free HiPSCs Resembling Ground State Pluripotent Stem Cells

Overview

Journal Stem Cell Reports

Publisher Cell Press

Specialty Cell Biology

Date 2014 Mar 28

PMID 24672758

Citations 87

Authors

Bahram Valamehr

Megan Robinson

Ramzey Abujarour

Betsy Rezner

Florin Vranceanu

Thuy Le

Amanda Medcalf

Tom Tong Lee

Michael Fitch

David Robbins

Peter Flynn

Affiliations

Soon will be listed here.

Abstract

Cell banking, disease modeling, and cell therapy applications have placed increasing demands on hiPSC technology. Specifically, the high-throughput derivation of footprint-free hiPSCs and their expansion in systems that allow scaled production remains technically challenging. Here, we describe a platform for the rapid, parallel generation, selection, and expansion of hiPSCs using small molecule pathway inhibitors in stage-specific media compositions. The platform supported efficient and expedited episomal reprogramming using just OCT4/SOX2/SV40LT combination (0.5%-4.0%, between days 12 and 16) in a completely feeder-free environment. The resulting hiPSCs are transgene-free, readily cultured, and expanded as single cells while maintaining a homogeneous and genomically stable pluripotent population. hiPSCs generated or maintained in the media compositions described exhibit properties associated with the ground state of pluripotency. The simplicity and robustness of the system allow for the high-throughput generation and rapid expansion of a uniform hiPSC product that is applicable to industrial and clinical-grade use.

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References

Xu Y, Zhu X, Hahm H, Wei W, Hao E, Hayek A . Revealing a core signaling regulatory mechanism for pluripotent stem cell survival and self-renewal by small molecules. Proc Natl Acad Sci U S A. 2010; 107(18):8129-34. PMC: 2889586. DOI: 10.1073/pnas.1002024107. View

Nakagawa M, Koyanagi M, Tanabe K, Takahashi K, Ichisaka T, Aoi T . Generation of induced pluripotent stem cells without Myc from mouse and human fibroblasts. Nat Biotechnol. 2007; 26(1):101-6. DOI: 10.1038/nbt1374. View

Chan Y, Goke J, Ng J, Lu X, Gonzales K, Tan C . Induction of a human pluripotent state with distinct regulatory circuitry that resembles preimplantation epiblast. Cell Stem Cell. 2013; 13(6):663-75. DOI: 10.1016/j.stem.2013.11.015. View

Pera M, Trounson A . Human embryonic stem cells: prospects for development. Development. 2004; 131(22):5515-25. DOI: 10.1242/dev.01451. View

Valamehr B, Abujarour R, Robinson M, Le T, Robbins D, Shoemaker D . A novel platform to enable the high-throughput derivation and characterization of feeder-free human iPSCs. Sci Rep. 2012; 2:213. PMC: 3252544. DOI: 10.1038/srep00213. View

Beattie G, Lopez A, Bucay N, Hinton A, Firpo M, King C . Activin A maintains pluripotency of human embryonic stem cells in the absence of feeder layers. Stem Cells. 2005; 23(4):489-95. DOI: 10.1634/stemcells.2004-0279. View

ODoherty R, Greiser U, Wang W . Nonviral methods for inducing pluripotency to cells. Biomed Res Int. 2013; 2013:705902. PMC: 3693118. DOI: 10.1155/2013/705902. View

Abujarour R, Valamehr B, Robinson M, Rezner B, Vranceanu F, Flynn P . Optimized surface markers for the prospective isolation of high-quality hiPSCs using flow cytometry selection. Sci Rep. 2013; 3:1179. PMC: 3560358. DOI: 10.1038/srep01179. View

Shu J, Wu C, Wu Y, Li Z, Shao S, Zhao W . Induction of pluripotency in mouse somatic cells with lineage specifiers. Cell. 2013; 153(5):963-75. PMC: 4640445. DOI: 10.1016/j.cell.2013.05.001. View

10.

Rao M . iPSC crowdsourcing: a model for obtaining large panels of stem cell lines for screening. Cell Stem Cell. 2013; 13(4):389-91. DOI: 10.1016/j.stem.2013.09.005. View

11.

Marks H, Kalkan T, Menafra R, Denissov S, Jones K, Hofemeister H . The transcriptional and epigenomic foundations of ground state pluripotency. Cell. 2012; 149(3):590-604. PMC: 3398752. DOI: 10.1016/j.cell.2012.03.026. View

12.

Warren L, Manos P, Ahfeldt T, Loh Y, Li H, Lau F . Highly efficient reprogramming to pluripotency and directed differentiation of human cells with synthetic modified mRNA. Cell Stem Cell. 2010; 7(5):618-30. PMC: 3656821. DOI: 10.1016/j.stem.2010.08.012. View

13.

Zhou H, Li W, Zhu S, Joo J, Do J, Xiong W . Conversion of mouse epiblast stem cells to an earlier pluripotency state by small molecules. J Biol Chem. 2010; 285(39):29676-80. PMC: 2943300. DOI: 10.1074/jbc.C110.150599. View

14.

Wang W, Yang J, Liu H, Lu D, Chen X, Zenonos Z . Rapid and efficient reprogramming of somatic cells to induced pluripotent stem cells by retinoic acid receptor gamma and liver receptor homolog 1. Proc Natl Acad Sci U S A. 2011; 108(45):18283-8. PMC: 3215025. DOI: 10.1073/pnas.1100893108. View

15.

Gafni O, Weinberger L, AlFatah Mansour A, Manor Y, Chomsky E, Ben-Yosef D . Derivation of novel human ground state naive pluripotent stem cells. Nature. 2013; 504(7479):282-6. DOI: 10.1038/nature12745. View

16.

Li R, Liang J, Ni S, Zhou T, Qing X, Li H . A mesenchymal-to-epithelial transition initiates and is required for the nuclear reprogramming of mouse fibroblasts. Cell Stem Cell. 2010; 7(1):51-63. DOI: 10.1016/j.stem.2010.04.014. View

17.

Nichols J, Smith A . Pluripotency in the embryo and in culture. Cold Spring Harb Perspect Biol. 2012; 4(8):a008128. PMC: 3405859. DOI: 10.1101/cshperspect.a008128. View

18.

Polo J, Anderssen E, Walsh R, Schwarz B, Nefzger C, Lim S . A molecular roadmap of reprogramming somatic cells into iPS cells. Cell. 2012; 151(7):1617-32. PMC: 3608203. DOI: 10.1016/j.cell.2012.11.039. View

19.

Rais Y, Zviran A, Geula S, Gafni O, Chomsky E, Viukov S . Deterministic direct reprogramming of somatic cells to pluripotency. Nature. 2013; 502(7469):65-70. DOI: 10.1038/nature12587. View

20.

Thomson J, Itskovitz-Eldor J, Shapiro S, Waknitz M, Swiergiel J, Marshall V . Embryonic stem cell lines derived from human blastocysts. Science. 1998; 282(5391):1145-7. DOI: 10.1126/science.282.5391.1145. View