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Cultivation of Human Bone-Like Tissue from Pluripotent Stem Cell-Derived Osteogenic Progenitors in Perfusion Bioreactors

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Book cover Biomimetics and Stem Cells

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1202))

Abstract

Human pluripotent stem cells represent an unlimited source of skeletal tissue progenitors for studies of bone biology, pathogenesis, and the development of new approaches for bone reconstruction and therapies. In order to construct in vitro models of bone tissue development and to grow functional, clinical-size bone substitutes for transplantation, cell cultivation in three-dimensional environments composed of porous osteoconductive scaffolds and dynamic culture systems—bioreactors—has been studied. Here, we describe a stepwise procedure for the induction of human embryonic and induced pluripotent stem cells (collectively termed PSCs) into mesenchymal-like progenitors, and their subsequent cultivation on decellularized bovine bone scaffolds in perfusion bioreactors, to support the development of viable, stable bone-like tissue in defined geometries.

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References

  1. de Peppo GM, Marolt D (2013) Modulating the biochemical and biophysical culture environment to enhance osteogenic differentiation and maturation of human pluripotent stem cell-derived mesenchymal progenitors. Stem Cell Res Ther 5:106

    Google Scholar 

  2. Tandon N, Marolt D, Cimetta E, Vunjak-Novakovic G (2013) Bioreactor engineering of stem cell environments. Biotechnol Adv. 7:1020–1031

    Google Scholar 

  3. Marolt D, Knezevic M, Vunjak-Novakovic G (2010) Bone tissue engineering with human stem cells. Stem Cell Res Ther 2:10

    Article  Google Scholar 

  4. Tachibana M, Amato P, Sparman M, Gutierrez NM, Tippner-Hedgesm R, Ma H, Kang E, Fulati A, Lee HS, Sritanaudomchai H, Masterson K, Larson J, Eaton D, Sadler-Fredd K, Battaglia D, Lee D, Wu D, Jensen J, Patton P, Gokhale S, Stouffer RL, Wolf D, Mitalipov S (2013) Human embryonic stem cells derived by somatic cell nuclear transfer. Cell 6:1228–1238

    Article  Google Scholar 

  5. Takahashi K, Tanabe K, Ohnuki M, Narita M, Ichisaka T, Tomoda K, Yamanaka S (2007) Induction of pluripotent stem cells from adult human fibroblasts by defined factors. Cell 5:861–872

    Article  Google Scholar 

  6. Murry CE, Keller G (2008) Differentiation of embryonic stem cells to clinically relevant populations: lessons from embryonic development. Cell 4:661–680

    Article  Google Scholar 

  7. Kuznetsov SA, Cherman N, Robey PG (2011) In vivo bone formation by progeny of human embryonic stem cells. Stem Cells Dev 2:269–287

    Article  Google Scholar 

  8. Levi B, Hyun JS, Montoro DT, Lo DD, Chan CK, Hu S, Sun N, Lee M, Grova M, Connolly AJ, Wu JC, Gurtner GC, Weissman IL, Wan DC, Longaker MT (2012) In vivo directed differentiation of pluripotent stem cells for skeletal regeneration. Proc Natl Acad Sci U S A 50:20379–20384

    Article  Google Scholar 

  9. Fröhlich M, Grayson WL, Wan LQ, Marolt D, Drobnic M, Vunjak-Novakovic G (2008) Tissue engineered bone grafts: biological requirements, tissue culture and clinical relevance. Curr Stem Cell Res Ther 4:254–264

    Article  Google Scholar 

  10. Grayson WL, Fröhlich M, Yeager K, Bhumiratana S, Chan ME, Cannizzaro C, Wan LQ, Liu XS, Guo XE, Vunjak-Novakovic G (2010) Engineering anatomically shaped human bone grafts. Proc Natl Acad Sci U S A 8:3299–3304

    Article  Google Scholar 

  11. Grayson WL, Bhumiratana S, Cannizzaro C, Chao PH, Lennon DP, Caplan AI, Vunjak-Novakovic G (2008) Effects of initial seeding density and fluid perfusion rate on formation of tissue-engineered bone. Tissue Eng Part A 11:1809–1820

    Article  Google Scholar 

  12. Grayson WL, Marolt D, Bhumiratana S, Fröhlich M, Guo XE, Vunjak-Novakovic G (2011) Optimizing the medium perfusion rate in bone tissue engineering bioreactors. Biotechnol Bioeng 5:1159–1170

    Article  Google Scholar 

  13. Marcos-Campos I, Marolt D, Petridis P, Bhumiratana S, Schmidt D, Vunjak-Novakovic G (2012) Bone scaffold architecture modulates the development of mineralized bone matrix by human embryonic stem cells. Biomaterials 33:8329–8342

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  14. de Peppo GM, Marcos-Campos I, Kahler DJ, Alsalman D, Shang L, Vunjak-Novakovic G, Marolt D (2013) Engineering bone tissue substitutes from human induced pluripotent stem cells. Proc Natl Acad Sci U S A 21:8680–8685

    Article  Google Scholar 

  15. Marolt D, Campos IM, Bhumiratana S, Koren A, Petridis P, Zhang G, Spitalnik PF, Grayson WL, Vunjak-Novakovic G (2012) Engineering bone tissue from human embryonic stem cells. Proc Natl Acad Sci U S A 22:8705–8709

    Article  Google Scholar 

  16. de Peppo GM, Sjovall P, Lennerås M, Strehl R, Hyllner J, Thomsen P, Karlsson C (2010) Osteogenic potential of human mesenchymal stem cells and human embryonic stem cell-derived mesodermal progenitors: a tissue engineering perspective. Tissue Eng Part A 11:3413–3426

    Article  Google Scholar 

  17. de Peppo GM, Svensson S, Lennerås M, Synnergren J, Stenberg J, Strehl R, Hyllner J, Thomsen P, Karlsson C (2010) Human embryonic mesodermal progenitors highly resemble human mesenchymal stem cells and display high potential for tissue engineering applications. Tissue Eng Part A 7:2161–2182

    Article  Google Scholar 

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Acknowledgements

This work was supported by the New York Stem Cell Foundation-Helmsley Investigator Award (to D.M.); the Leona M. and Harry B. Helmsley Charitable Trust; Robin Chemers Neustein; Goldman Sachs Gives, at the recommendation of Alan and Deborah Cohen; New York State Stem Cell Science Shared Facility, Grant C024179; National Institutes of Health Grants DE016525 and EB002520, (to G.V.-N.); and the New York Stem Cell Foundation.

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Correspondence to Darja Marolt .

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© 2013 Springer Science+Business Media New York

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de Peppo, G.M., Vunjak-Novakovic, G., Marolt, D. (2013). Cultivation of Human Bone-Like Tissue from Pluripotent Stem Cell-Derived Osteogenic Progenitors in Perfusion Bioreactors. In: Vunjak-Novakovic, G., Turksen, K. (eds) Biomimetics and Stem Cells. Methods in Molecular Biology, vol 1202. Humana Press, New York, NY. https://doi.org/10.1007/7651_2013_52

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  • DOI: https://doi.org/10.1007/7651_2013_52

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-1331-2

  • Online ISBN: 978-1-4939-1332-9

  • eBook Packages: Springer Protocols

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