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Cell and Tissue Research

, Volume 366, Issue 1, pp 113–127 | Cite as

Characteristics of three-dimensional prospectively isolated mouse bone marrow mesenchymal stem/stromal cell aggregates on nanoculture plates

  • Chizuka ObaraEmail author
  • Ken-ichi Tomiyama
  • Kazuya Takizawa
  • Rafiqul Islam
  • Takeshi Yasuda
  • Takaya Gotoh
  • Katsushi TajimaEmail author
Regular Article

Abstract

Three-dimensional (3-D) aggregate culturing is useful for investigating the functional properties of mesenchymal stem/stromal cells (MSCs). For 3-D MSC analysis, however, pre-expansion of MSCs with two-dimensional (2-D) monolayer culturing must first be performed, which might abolish their endogenous properties. To avoid the need for 2-D expansion, we used prospectively isolated mouse bone marrow (BM)-MSCs and examined the differences in the biological properties of 2-D and 3-D MSC cultures. The BM-MSCs self-assembled into aggregates on nanoculture plates (NCP) that have nanoimprinted patterns with a low-cellular binding texture. The 3-D MSCs proliferated at the same rate as 2-D-cultured cells by only diffusion culture and secreted higher levels of pro-angiogenic factors such as vascular endothelial growth factor and hepatocyte growth factor (HGF). Conditioned medium from 3-D MSC cultures promoted more capillary formation than that of 2-D MSCs in an in vitro tube formation assay. Matrigel-implanted 3-D MSC aggregates tended to induce angiogenesis in host mice. The 3-D culturing on NCP induced alpha-fetoprotein (AFP) expression in MSCs without the application of AFP- or endodermal-inducible factors, possibly via an HGF-autocrine mechanism, and maintained their differentiation ability for adipocytes, osteocytes, and chondrocytes. Prospectively isolated mouse BM-MSCs expressed low/negative stemness-related genes including Oct3/4, Nanog, and Sox2, which were not enhanced by NCP-based 3-D culturing, suggesting that some of these cells differentiate into meso-endodermal layer cells. Culturing of prospectively isolated MSCs on NCP in 3-D allows the analysis of the biological properties of more closely endogenous BM-MSCs and might contribute to tissue engineering and repair.

Keywords

Nanoculture plate Mouse bone marrow mesenchymal stem/stromal cells 3-D aggregate culture Alpha-fetoprotein Tissue engineering 

Notes

Acknowledgments

We thank Ms. S. Fukuzaki, N. Gotoh, K. Noshiro, and Mr. M. Hama for expert technical assistance, and Drs. I. Tanaka, H. Ishihara, H. Yakumaru, M. Hazawa, and Y. Michikawa for helpful suggestions. We are also grateful to the FACS support team of the National Institute of Radiological Sciences for their technical support regarding the flow cytometry experiments.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Supplementary material

441_2016_2405_MOESM1_ESM.pdf (2.7 mb)
ESM 1 (PDF 2,735 kb)

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Copyright information

© Springer-Verlag Berlin Heidelberg 2016

Authors and Affiliations

  • Chizuka Obara
    • 1
    Email author
  • Ken-ichi Tomiyama
    • 1
  • Kazuya Takizawa
    • 1
  • Rafiqul Islam
    • 1
  • Takeshi Yasuda
    • 1
  • Takaya Gotoh
    • 1
  • Katsushi Tajima
    • 1
    Email author
  1. 1.Research Center for Radiation Emergency MedicineNational Institute of Radiological SciencesChiba-cityJapan

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