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Stem Cells Int. 2019 May 5;2019:8469012. doi: 10.1155/2019/8469012. eCollection 2019.

Spontaneously Formed Spheroids from Mouse Compact Bone-Derived Cells Retain Highly Potent Stem Cells with Enhanced Differentiation Capability.

Chen K1, Li X1,2,3, Li N1, Dong H1, Zhang Y4, Yoshizawa M1,2, Kagami H1,2,3,5.

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Department of Hard Tissue Research, Graduate School of Oral Medicine, Matsumoto Dental University, Shiojiri 399-0781, Japan.
Department of Oral and Maxillofacial Surgery, School of Dentistry, Matsumoto Dental University, Shiojiri 399-0781, Japan.
Institute for Oral Science, Matsumoto Dental University, Shiojiri 399-0781, Japan.
Department of Stomatology, Shanghai Tenth People's Hospital, Tongji University School of Medicine, Shanghai 200072, China.
Department of General Medicine, IMSUT Hospital, Institute of Medical Science, University of Tokyo, Tokyo 108-8639, Japan.


The results from our recent study showed the presence of two distinct spheroid-forming mechanisms, i.e., spontaneous and mechanical. In this study, we focused on the spontaneously formed spheroids, and the character of spontaneously formed spheroids from mouse compact bone-derived cells (CBDCs) was explored. Cells from (C57BL/6J) mouse leg bones were isolated, and compact bone-derived cells were cultured after enzymatic digestion. Spontaneous spheroid formation was achieved on a culture plate with specific water contact angle as reported. The expression levels of embryonic stem cell markers were analyzed using immunofluorescence and quantitative reverse transcription polymerase chain reaction. Then, the cells from spheroids were induced into osteogenic and neurogenic lineages. The spontaneously formed spheroids from CBDCs were positive for ES cell markers such as SSEA1, Sox2, Oct4, and Nanog. Additionally, the expressions of fucosyltransferase 4/FUT4 (SSEA1), Sox2, and Nanog were significantly higher than those in monolayer cultured cells. The gene expression of mesenchymal stem cell markers was almost identical in both spheroids and monolayer-cultured cells, but the expression of Sca-1 was higher in spheroids. Spheroid-derived cells showed significantly higher osteogenic and neurogenic marker expression than monolayer-cultured cells after induction. Spontaneously formed spheroids expressed stem cell markers and showed enhanced osteogenic and neurogenic differentiation capabilities than cells from the conventional monolayer culture, which supports the superior stemness.

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