Induced Pluripotent Stem (iPS) Cell Culture Methods and Induction of Differentiation into Endothelial Cells

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Induced Pluripotent Stem (iPS) Cells

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1357))

Abstract

The study of stem cell behavior and differentiation in a developmental context is complex, time-consuming, and expensive, and for this reason, cell culture remains a method of choice for developmental and regenerative biology and mechanistic studies. Similar to ES cells, iPS cells have the ability to differentiate into endothelial cells (ECs), and the route for differentiation appears to mimic the developmental process that occurs during the formation of an embryo. Traditional EC induction methods from embryonic stem (ES) cells rely mostly on the formation of embryoid body (EB), which employs feeder or feeder-free conditions in the presence or absence of supporting cells. Similar to ES cells, iPS cells can be cultured in feeder layer or feeder-free conditions. Here, we describe the iPS cell culture methods and induction differentiation of these cells into ECs. We use anti-mouse Flk1 and anti-mouse VE-cadherin to isolate and characterize mouse ECs, because these antibodies are commercially available and their use has been described in the literature, including by our group. The ECs produced by this method have been used by our laboratory, and we have demonstrated their in vivo potential. We also discuss how iPS cells differ in their ability to differentiate into endothelial cells in culture.

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References

  1. Evans MJ, Kaufman MH (1981) Establishment in culture of pluripotential cells from mouse embryos. Nature 292:154–156

    Article  PubMed  CAS  Google Scholar 

  2. Bradley A, Robertson E (1986) Embryo-derived stem cells: a tool for elucidating the developmental genetics of the mouse. Curr Top Dev Biol 20:357–371

    Article  PubMed  CAS  Google Scholar 

  3. Nagy A, Gócza E, Diaz EM, Prideaux VR, Iványi E, Markkula M, Rossant J (1990) Embryonic stem cells alone are able to support fetal development in the mouse. Development 110(3):815–821

    PubMed  CAS  Google Scholar 

  4. Nagy A, Rossant J, Nagy R, Abramow-Newerly W, Roder JC (1993) Derivation of completely cell culture-derived mice from early-passage embryonic stem cells. Proc Natl Acad Sci U S A 90(18):8424–8428

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  5. Wilmut I, Schnieke AE, McWhir J, Kind AJ, Campbell KH (1997) Viable offspring derived from fetal and adult mammalian cells. Nature 385:810–813

    Article  PubMed  CAS  Google Scholar 

  6. Thomson JA, Itskovitz-Eldor J, Shapiro SS, Waknitz MA, Swiergiel JJ, Marshall VS, Jones JM (1998) Embryonic stem cell lines derived from human blastocysts. Science 282(5391):1145–1147

    Article  PubMed  CAS  Google Scholar 

  7. Amit M, Carpenter MK, Inokuma MS, Chiu CP, Harris CP, Waknitz MA, Itskovitz-Eldor J, Thomson JA (2000) Clonally derived human embryonic stem cell lines maintain pluripotency and proliferative potential for prolonged periods of culture. Dev Biol 227(2):271–278

    Article  PubMed  CAS  Google Scholar 

  8. Waddington CH (1957) The strategy of the genes. Geo Allen and Unwin, London

    Google Scholar 

  9. Takahashi K, Yamanaka S (2006) Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors. Cell 126(4):663–676

    Article  PubMed  CAS  Google Scholar 

  10. Okita K, Ichisaka T, Yamanaka S (2007) Generation of germline-competent induced pluripotent stem cells. Nature 448(7151):313–317

    Article  PubMed  CAS  Google Scholar 

  11. Yu J, Vodyanik MA, Smuga-Otto K, Antosiewicz-Bourget J, Frane JL, Tian S, Nie J, Jonsdottir GA, Ruotti V, Stewart R, Slukvin II, Thomson JA (2007) Induced pluripotent stem cell lines derived from human somatic cells. Science 318(5858):1917–1920

    Article  PubMed  CAS  Google Scholar 

  12. Wernig M, Meissner A, Foreman R, Brambrink T, Ku M, Hochedlinger K, Bernstein BE, Jaenisch R (2007) In vitro reprogramming of fibroblasts into a pluripotent ES-cell-like state. Nature 448(7151):318–324

    Article  PubMed  CAS  Google Scholar 

  13. Wang Y, Baskerville S, Shenoy A, Babiarz JE, Baehner L, Blelloch R (2008) Embryonic stem cell-specific microRNAs regulate the G1-S transition and promote rapid proliferation. Nat Genet 40:1478–1483

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  14. Judson RL, Babiarz JE, Venere M, Blelloch R (2009) Embryonic stem cell-specific microRNAs promote induced pluripotency. Nat Biotechnol 27:459–461

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  15. Subramanyam D, Lamouille S, Judson RL, Liu JY, Bucay N, Derynck R, Blelloch R (2011) Multiple targets of miR-302 and miR-372 promote reprogramming of human fibroblasts to induced pluripotent stem cells. Nat Biotechnol 29(5):443–448

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  16. Gonzalez F, Boue S, Belmonte JC (2011) Methods for making induced pluripotent stem cells: reprogramming a la carte. Nat Rev Genet 12:231–242

    Article  PubMed  CAS  Google Scholar 

  17. Giorgetti A et al (2009) Generation of induced pluripotent stem cells from human cord blood using OCT4 and SOX2. Cell Stem Cell 5:353–357

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  18. Utikal J, Maherali N, Kulalert W, Hochedlinger K (2009) Sox2 is dispensable for the reprogramming of melanocytes and melanoma cells into induced pluripotent stem cells. J Cell Sci 122:3502–3510

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  19. Eminli S, Utikal J, Arnold K, Jaenisch R, Hochedlinger K (2008) Reprogramming of neural progenitor cells into induced pluripotent stem cells in the absence of exogenous Sox2 expression. Stem Cells 26(10):2467–2474

    Article  PubMed  CAS  Google Scholar 

  20. Kim JB, Greber B, Araúzo-Bravo MJ, Meyer J, Park KI, Zaehres H, Schöler HR (2009) Direct reprogramming of human neural stem cells by OCT4. Nature 461:649–653

    Article  PubMed  CAS  Google Scholar 

  21. Li Y, Zhang Q, Yin X, Yang W, Du Y, Hou P, Ge J, Liu C, Zhang W, Zhang X, Wu Y, Li H, Liu K, Wu C, Song Z, Zhao Y, Shi Y, Deng H (2011) Generation of iPSCs from mouse fibroblasts with a single gene, Oct4, and small molecules. Cell Res 21(1):196–204

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  22. Theunissen TW, Jaenisch R (2014) Molecular control of induced pluripotency. Cell Stem Cell 14(6):720–734

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  23. Boland MJ, Hazen JL, Nazor KL, Rodriguez AR, Gifford W, Martin G, Kupriyanov S, Baldwin KK (2009) Adult mice generated from induced pluripotent stem cells. Nature 461:91–94

    Article  PubMed  CAS  Google Scholar 

  24. Woltjen K, Michael IP, Mohseni P, Desai R, Mileikovsky M, Hämäläinen R, Cowling R, Wang W, Liu P, Gertsenstein M, Kaji K, Sung HK, Nagy A (2009) piggyback transposition reprograms fibroblasts to induced pluripotent stem cells. Nature 458(7239):766–770

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  25. Kaji K, Norrby K, Paca A, Mileikovsky M, Mohseni P, Woltjen K (2009) Virus-free induction of pluripotency and subsequent excision of reprogramming factors. Nature 458(7239):771–775

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  26. Woltjen K, Hämäläinen R, Kibschull M, Mileikovsky M, Nagy A (2011) Transgene-free production of pluripotent stem cells using piggyBac transposons. Methods Mol Biol 767:87–103

    Article  PubMed  CAS  Google Scholar 

  27. Papapetrou EP, Sadelain M (2011) Generation of transgene-free human induced pluripotent stem cells with an excisable single polycistronic vector. Nat Protoc 6:1251–1273

    Article  PubMed  CAS  Google Scholar 

  28. Kohler EE, Wary KK, Li F, Chatterjee I, Urao N, Toth PT, Ushio-Fukai M, Rehman J, Park C, Malik AB (2013) Flk1+ and VE-cadherin+ endothelial cells derived from iPSCs recapitulates vascular development during differentiation and display similar angiogenic potential as ESC-derived cells. PLoS One 8(12):e85549

    Article  PubMed  PubMed Central  Google Scholar 

  29. Yamashita J, Itoh H, Hirashima M, Ogawa M, Nishikawa S, Yurugi T, Naito M, Nakao K, Nishikawa S (2000) Flk1-positive cells derived from embryonic stem cells serve as vascular progenitors. Nature 408:92–96

    Article  PubMed  CAS  Google Scholar 

  30. Levenberg S, Golub JS, Amit M, Itskovitz-Eldor J, Langer R (2002) Endothelial cells derived from human embryonic stem cells. Proc Natl Acad Sci U S A 99:4391–4396

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  31. Choi KD, Yu J, Smuga-Otto K, Salvagiotto G, Rehrauer W, Vodyanik M, Thomson J, Slukvin I (2009) Hematopoietic and endothelial differentiation of human induced pluripotent stem cells. Stem Cells 27(3):559–567

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  32. Yin L, Ohanyan V, Pung YF, Delucia A, Bailey E, Enrick M, Stevanov K, Kolz CL, Guarini G, Chilian WM (2012) Induction of vascular progenitor cells from endothelial cells stimulates coronary collateral growth. Circ Res 110(2):241–252

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  33. Yang HM, Moon SH, Choi YS, Park SJ, Lee YS, Lee HJ, Kim SJ, Chung HM (2013) Therapeutic efficacy of human embryonic stem cell-derived endothelial cells in humanized mouse models harboring a human immune system. Arterioscler Thromb Vasc Biol 33:2839–2849

    Article  PubMed  CAS  Google Scholar 

  34. Adams WJ, Zhang Y, Cloutier J, Kuchimanchi P, Newton G, Sehrawat S, Aird WC, Mayadas TN, Luscinskas FW, García-Cardeña G (2013) Functional vascular endothelium derived from human induced pluripotent stem cells. Stem Cell Reports 1(2):105–113

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  35. Prasain N, Lee MR, Vemula S, Meador JL, Yoshimoto M, Ferkowicz MJ, Fett A, Gupta M, Rapp BM, Saadatzadeh MR, Ginsberg M, Elemento O, Lee Y, Voytik-Harbin SL, Chung HM, Hong KS, Reid E, O’Neill CL, Medina RJ, Stitt AW, Murphy MP, Rafii S, Broxmeyer HE, Yoder MC (2014) Differentiation of human pluripotent stem cells to cells similar to cord-blood endothelial colony-forming cells. Nat Biotechnol 32(11):1151–1157

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  36. Belair DG, Whisler JA, Valdez J, Velazquez J, Molenda JA, Vickerman V, Lewis R, Daigh C, Hansen TD, Mann DA, Thomson JA, Griffith LG, Kamm RD, Schwartz MP, Murphy WL (2014) Human vascular tissue models formed from human induced pluripotent stem cell derived endothelial cells. Stem Cell Rev (in press)

    Google Scholar 

  37. Kusuma S, Facklam A, Gerecht S (2014) Characterizing human pluripotent-stem-cell-derived vascular cells for tissue engineering applications. Stem Cells Dev (in press)

    Google Scholar 

Download references

Acknowledgments

The research in the authors’ laboratory is supported by grants from the National Institutes of Health (NIH) and the American Heart Association (AHA) to KKW.

[According to the PloS One Open Access Journal policy no permission is required to reproduce text and the Figures (Kohler EE et al., PLoS One 8(12):e85549.)]

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Correspondence to Kishore K. Wary .

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Chatterjee, I., Li, F., Kohler, E.E., Rehman, J., Malik, A.B., Wary, K.K. (2015). Induced Pluripotent Stem (iPS) Cell Culture Methods and Induction of Differentiation into Endothelial Cells. In: Turksen, K., Nagy, A. (eds) Induced Pluripotent Stem (iPS) Cells. Methods in Molecular Biology, vol 1357. Humana Press, New York, NY. https://doi.org/10.1007/7651_2015_203

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  • DOI: https://doi.org/10.1007/7651_2015_203

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-3054-8

  • Online ISBN: 978-1-4939-3055-5

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