Characteristics of three-dimensional prospectively isolated mouse bone marrow mesenchymal stem/stromal cell aggregates on nanoculture plates.
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.
Research Center for Radiation Emergency Medicine, National Institute of Radiological Sciences, 4-9-1, Anagawa, Inage-ku, Chiba-city, Chiba, 263-8555, Japan. obara.chizuka@qst.go.jp.
Chizuka
C
Obara
Research Center for Radiation Emergency Medicine, National Institute of Radiological Sciences, 4-9-1, Anagawa, Inage-ku, Chiba-city, Chiba, 263-8555, Japan.
Ken-Ichi
K
Tomiyama
Research Center for Radiation Emergency Medicine, National Institute of Radiological Sciences, 4-9-1, Anagawa, Inage-ku, Chiba-city, Chiba, 263-8555, Japan.
Kazuya
K
Takizawa
Research Center for Radiation Emergency Medicine, National Institute of Radiological Sciences, 4-9-1, Anagawa, Inage-ku, Chiba-city, Chiba, 263-8555, Japan.
Rafiqul
R
Islam
Research Center for Radiation Emergency Medicine, National Institute of Radiological Sciences, 4-9-1, Anagawa, Inage-ku, Chiba-city, Chiba, 263-8555, Japan.
Takeshi
T
Yasuda
Research Center for Radiation Emergency Medicine, National Institute of Radiological Sciences, 4-9-1, Anagawa, Inage-ku, Chiba-city, Chiba, 263-8555, Japan.
Takaya
T
Gotoh
Research Center for Radiation Emergency Medicine, National Institute of Radiological Sciences, 4-9-1, Anagawa, Inage-ku, Chiba-city, Chiba, 263-8555, Japan. tajima@qst.go.jp.
Katsushi
K
Tajima
D000818:Animals; D015415:Biomarkers / Q000378:metabolism; D001854:Bone Marrow Cells / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D002449:Cell Aggregation / Q000187:drug effects; D018929:Cell Culture Techniques / Q000379:methods*; D019070:Cell Lineage / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D002469:Cell Separation / Q000379:methods*; D048430:Cell Shape / Q000187:drug effects; D048429:Cell Size / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D058948:Cell Tracking; D002478:Cells, Cultured; D017077:Culture Media, Conditioned / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000187:drug effects; D021621:Imaging, Three-Dimensional; D008297:Male; D059630:Mesenchymal Stem Cells / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D008810:Mice, Inbred C57BL; D036103:Nanotechnology / Q000379:methods*; D018919:Neovascularization, Physiologic / Q000187:drug effects
D015415:Biomarkers; D017077:Culture Media, Conditioned
3-D aggregate culture; Alpha-fetoprotein; Mouse bone marrow mesenchymal stem/stromal cells; Nanoculture plate; Tissue engineering