Skip to main content

Generation of rat-induced pluripotent stem cells: Reprogramming and culture medium

Abstract

The rat represents an animal model highly attractive for studying pharmacology, physiology, aging, cardiovascular diseases, etc., that in many aspects is more adequate than the mouse model. Derivation of induced pluripotent stem cells from rats (riPS) opens the opportunity for gene targeting in specific rat strains, as well as for the development of new protocols for the treatment of different degenerative diseases. Here we report an improved protocol for riPS cell generation, which is based on lentivirus delivery of reprogramming factors with their subsequent excision from the genome, application of serum-free media and chemical inhibitors MEK and GSK. We compared various conditions for riPS cell derivation, analyzed the cell karyotype, and assessed the pluripotency of the established cells. These data may prompt further iPS cell-based gene targeting in rat, as well as the development of iPS-based cell therapy, using this animal model.

This is a preview of subscription content, access via your institution.

Abbreviations

iPS cells:

induced pluripotent stem cells

MEF:

mouse embryonic fibroblasts

ES cells:

embryonic stem cells

EGFP:

enhanced green fluorescent protein

GSK:

glycogen synthase kinase

LIF:

leukemia inhibitory factor

PBS:

phosphate buffer saline

References

  1. Aoi, T., Yae, K., Nakagawa, M., Ichisaka, T., Okita, K., Takahashi, K., Chiba, T., and Yamanaka, S., Generation of Pluripotent Stem Cells from Adult Mouse Liver and Stomach Cells, Science, 2008, vol. 321, pp. 699–702.

    PubMed  Article  CAS  Google Scholar 

  2. Brambrink, T., Foreman, R., Welstead, G.G., Lengner, C.J., Wernig, M., Suh, H., and Jaenisch, R., Sequential Expression of Pluripotency Markers during Direct Reprogramming of Mouse Somatic Cells, Cell Stem Cell, 2008, vol. 2, pp. 151–159.

    PubMed  Article  CAS  Google Scholar 

  3. Buehr, M., Meek, S., Blair, K., Yang, J., Ure, J., Silva, J., McLay, R., Hall, J., Ying, Q.L., and Smith, A., Capture of Authentic Embryonic Stem Cells from Rat Blastocysts, Cell, 2008, vol. 135, pp. 1287–1298.

    PubMed  Article  CAS  Google Scholar 

  4. Evans, M.J. and Kaufman, M.H., Establishment in Culture of Pluripotential Cells from Mouse Embryos, Nature, 1981, vol. 292, pp. 154–156.

    PubMed  Article  CAS  Google Scholar 

  5. Hanna, J., Markoulaki, S., Schorderet, P., Carey, B.W., Beard, C., et al., Direct Reprogramming of Terminally Differentiated Mature B Lymphocytes to Pluripotency, Cell, 2008, vol. 133, pp. 250–264.

    PubMed  Article  CAS  Google Scholar 

  6. Holubcová, Z., Matula, P., Sedláčková, M., Vinarský, V., Doležalová, D., Bárta, T., Dvořák, P., and Hampl, A., Human Embryonic Stem Cells Suffer from Centrosomal Amplification, Stem Cells, 2010, vol. 29, pp. 46–56.

    Article  Google Scholar 

  7. Li, W., Wei, W., Zhu, S., Zhu, J., Shi, Y., Lin, T., Hao, E., Hayek, A., Deng, H., and Ding, S., Generation of Rat and Human Induced Pluripotent Stem Cells by Combining Genetic Reprogramming and Chemical Inhibitors, Cell Stem Cell, 2009, vol. 4, pp. 16–19.

    PubMed  Article  Google Scholar 

  8. Liao, J., Cui, C., Chen, S., Ren, J., Chen, J., Gao, Y., Li, H., Jia, N., Cheng, L., Xiao, H., and Xiao, L., Generation of Induced Pluripotent Stem Cell Lines from Adult Rat Cells, Cell. Stem Cell, 2009, vol. 4, pp. 11–5.

    PubMed  Article  CAS  Google Scholar 

  9. Minina, Iu.M., Zhdanova, N.S., Shilov, A.G., Tolkunova, E.N., Liskovykh, M.A., and Tomilin, A.N., Chromosomal Instability of in vitro Cultured Mouse Embryonic Stem Cells and Induced Pluripotent Stem Cells, Tsitologiya, 2010, vol. 52, pp. 420–425.

    Google Scholar 

  10. O’Brien, S.J., Menninger, J.C., and Nash, W.G., Atlas of Mammalian Chromosomes, Hoboken, NJ: Wiley-Liss. 2006, vol. XLII.

    Book  Google Scholar 

  11. Okita, K., Ichisaka, T., and Yamanaka, S., Generation of Germline-Competent Induced Pluripotent Stem Cells, Nature, 2007, vol. 448, pp. 313–317.

    PubMed  Article  CAS  Google Scholar 

  12. Silva, J., Barrandon, O., Nichols, J., Kawaguchi, J., Theunissen, T.W., and Smith, A., Promotion of Reprogramming to Ground State Pluripotency by Signal Inhibition, PLoS Biol., 2008, vol. 6, p. e253.

  13. Stadtfeld, M., Brennand, K., and Hochedlinger, K., Reprogramming of Pancreatic β Cells into Induced Pluripotent Stem Cells, Curr. Biol., 2008, vol. 18, pp. 890–894.

    PubMed  Article  CAS  Google Scholar 

  14. Sugawara, A., Goto, K., Sotomaru, Y., Sofuni, T., and Ito, T., Current Status of Chromosomal Abnormalities in Mouse Embryonic Stem Cell Lines Used in Japan, Comp. Med., 2006, vol. 56, pp. 31–34.

    PubMed  CAS  Google Scholar 

  15. Takahashi, K. and Yamanaka, S., Induction of Pluripotent Stem Cells from Mouse Embryonic and Adult Fibroblast Cultures by Defined Factors, Cell, 2006, vol. 126, pp. 663–676.

    PubMed  Article  CAS  Google Scholar 

  16. Wiznerowicz, M. and Trono, D., Conditional Suppression of Cellular Genes: Lentivirus Vector-Mediated Drug-Inducible RNA Interference, J. Virol., 2003, vol. 77, pp. 8957–8961.

    PubMed  Article  CAS  Google Scholar 

  17. Ying, Q.L., Wray, J., Nichols, J., Batlle-Morera, L., Doble, B., Woodgett, J., Cohen, P., and Smith, A., The Ground State of Embryonic Stem Cell Self-Renewal, Nature, 2008, vol. 453, pp. 519–523.

    PubMed  Article  CAS  Google Scholar 

  18. Yu, J., Vodyanik, M.A., Smuga-Otto, K., Antosiewicz-Bourget, J., Frane, J.L., Tian, S., Nie, J., Jonsdottir, G.A., Ruotti, V., Stewart, R., Slukvin, I.I., and Thomson, J.A., Induced Pluripotent Stem Cell Lines Derived from Human Somatic Cells, Science, 2007, vol. 318, pp. 1917–1920.

    PubMed  Article  CAS  Google Scholar 

Download references

Author information

Affiliations

Authors

Corresponding author

Correspondence to M. A. Liskovykh.

Additional information

Original Russian Text © M.A. Liskovykh, I.A. Chuykin, A. Ranjan, D.A. Safina, E.N. Tolkunova, Yu.M. Minina, N.S. Zhdanova, P.A. Dyban, John J. Mullins, E.I. Kostyleva, E.V. Chikhirzhina, M. Bader, N. Alenina, A.N. Tomilin, 2011, published in Tsitologiya, 2011, Vol. 53, No. 12, pp. 939–945.

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Liskovykh, M.A., Chuykin, I.A., Ranjan, A. et al. Generation of rat-induced pluripotent stem cells: Reprogramming and culture medium. Cell Tiss. Biol. 6, 115–121 (2012). https://doi.org/10.1134/S1990519X12020095

Download citation

Keywords

  • induced pluripotent stem cells
  • rat
  • reprogramming