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Medical Molecular Morphology

, Volume 49, Issue 2, pp 119–126 | Cite as

A lower volume culture method for obtaining a larger yield of neuron-like cells from mesenchymal stem cells

  • Atsushi ShimomuraEmail author
  • Akiko Iizuka-Kogo
  • Naoki Yamamoto
  • Ryuji Nomura
Original Paper

Abstract

Mesenchymal stem cells (MSCs) represent a promising cell source for stem cell therapy to replace neurons damaged by neurodegenerative diseases. A system designed for in vitro neuronal differentiation of MSCs is an indispensable technique, which provides MSC-derived functional neurons for cell-replacement therapies and valuable information in pre-clinical research. This study investigated the effects of reducing the volume of neural induction medium on cell viability and neural differentiation of MSCs. When MSCs were differentiated in low volumes of neural induction medium, rather than using the conventional method, the cell density on culture dishes significantly increased. The % cell death, including apoptosis and necrosis, was significantly lower in the lower volume method than in the conventional method. There were no significant differences between the lower volume and conventional methods in the expression levels of the neuronal marker genes. In an analysis of immunostaining for a mature neuronal marker, no significant difference was detected between the media volumes. These findings demonstrate that neuronal induction of MSCs in low volumes of differentiation medium promoted survival during differentiation and resulted in larger numbers of MSC-derived neurons, compared to the conventional method. This novel lower volume method offers both financial and cell-yield advantages over the conventional method.

Keywords

A lower volume culture method In vitro neuronal differentiation Mesenchymal stem cell Neural differentiation 

Notes

Acknowledgments

The authors wish to thank Ms. Kazuko Hikita, Mr. Kazuhiro Yanagisawa, Mr. Yohei Takeuchi, Ms. Eri Nishigaki, and Ms. Mayu Tsunekawa for their helpful support. This research was supported by JSPS KAKENHI Grant Number 24592567 and the Aichi Cancer Research Foundation.

Compliance with ethical standards

Conflict of interest

The authors have declared that no competing interests exist.

References

  1. 1.
    Kondo T, Matsuoka AJ, Shimomura A, Koehler KR, Chan RJ, Miller JM, Srour EF, Hashino E (2011) Wnt signaling promotes neuronal differentiation from mesenchymal stem cells through activation of Tlx3. Stem Cells 29:836–846CrossRefPubMedPubMedCentralGoogle Scholar
  2. 2.
    Kondo T, Johnson SA, Yoder MC, Romand R, Hashino E (2005) Sonic hedgehog and retinoic acid synergistically promote sensory fate specification from bone marrow-derived pluripotent stem cells. Proc Natl Acad Sci USA 102:4789–4794CrossRefPubMedPubMedCentralGoogle Scholar
  3. 3.
    Dezawa M, Kanno H, Hoshino M, Cho H, Matsumoto N, Itokazu Y, Tajima N, Yamada H, Sawada H, Ishikawa H, Mimura T, Kitada M, Suzuki Y, Ide C (2004) Specific induction of neuronal cells from bone marrow stromal cells and application for autologous transplantation. J Clin Invest 113:1701–1710CrossRefPubMedPubMedCentralGoogle Scholar
  4. 4.
    Park HW, Cho JS, Park CK, Jung SJ, Park CH, Lee SJ, Oh SB, Park YS, Chang MS (2012) Directed induction of functional motor neuron-like cells from genetically engineered human mesenchymal stem cells. PLoS ONE 7:e35244CrossRefPubMedPubMedCentralGoogle Scholar
  5. 5.
    Barzilay R, Kan I, Ben-Zur T, Bulvik S, Melamed E, Offen D (2008) Induction of human mesenchymal stem cells into dopamine-producing cells with different differentiation protocols. Stem Cells Dev 17:547–554CrossRefPubMedGoogle Scholar
  6. 6.
    Hass R, Kasper C, Bohm S, Jacobs R (2011) Different populations and sources of human mesenchymal stem cells (MSC): a comparison of adult and neonatal tissue-derived MSC. Cell Commun Signal 9:12CrossRefPubMedPubMedCentralGoogle Scholar
  7. 7.
    Malladi P, Xu Y, Chiou M, Giaccia AJ, Longaker MT (2006) Effect of reduced oxygen tension on chondrogenesis and osteogenesis in adipose-derived mesenchymal cells. Am J Physiol Cell Physiol 290:C1139–C1146CrossRefPubMedGoogle Scholar
  8. 8.
    Wang DW, Fermor B, Gimble JM, Awad HA, Guilak F (2005) Influence of oxygen on the proliferation and metabolism of adipose derived adult stem cells. J Cell Physiol 204:184–191CrossRefPubMedGoogle Scholar
  9. 9.
    Meirelles Lda S, Fontes AM, Covas DT, Caplan AI (2009) Mechanisms involved in the therapeutic properties of mesenchymal stem cells. Cytokine Growth Factor Rev 20:419–427CrossRefPubMedGoogle Scholar
  10. 10.
    Murry CE, Reinecke H, Pabon LM (2006) Regeneration gaps: observations on stem cells and cardiac repair. J Am Coll Cardiol 47:1777–1785CrossRefPubMedGoogle Scholar
  11. 11.
    Shapiro AM, Lakey JR, Ryan EA, Korbutt GS, Toth E, Warnock GL, Kneteman NM, Rajotte RV (2000) Islet transplantation in seven patients with type 1 diabetes mellitus using a glucocorticoid-free immunosuppressive regimen. N Engl J Med 343:230–238CrossRefPubMedGoogle Scholar
  12. 12.
    Zweigerdt R (2009) Large scale production of stem cells and their derivatives. Adv Biochem Eng Biotechnol 114:201–235PubMedGoogle Scholar
  13. 13.
    Kitada M (2012) Mesenchymal cell populations: development of the induction systems for Schwann cells and neuronal cells and finding the unique stem cell population. Anat Sci Int 87:24–44CrossRefPubMedGoogle Scholar
  14. 14.
    Woodbury D, Reynolds K, Black IB (2002) Adult bone marrow stromal stem cells express germline, ectodermal, endodermal, and mesodermal genes prior to neurogenesis. J Neurosci Res 69:908–917CrossRefPubMedGoogle Scholar

Copyright information

© The Japanese Society for Clinical Molecular Morphology 2015

Authors and Affiliations

  • Atsushi Shimomura
    • 1
    • 2
    Email author
  • Akiko Iizuka-Kogo
    • 3
  • Naoki Yamamoto
    • 4
  • Ryuji Nomura
    • 5
  1. 1.Department of Communication DisordersHealth Sciences University of Hokkaido School of Psychological ScienceSapporoJapan
  2. 2.Department of Communication DisordersHealth Sciences University of Hokkaido School of Rehabilitation SciencesTobetsuJapan
  3. 3.Department of Anatomy and Cell BiologyGunma University School of MedicineMaebashiJapan
  4. 4.Laboratory of Molecular Biology and HistochemistryFujita Health University Joint Research LaboratoryToyoakeJapan
  5. 5.Department of Anatomy IFujita Health University School of MedicineToyoakeJapan

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