Primary bone-derived cells induce osteogenic differentiation without exogenous factors in human embryonic stem cells.
We developed a new and efficient method for osteoblastic differentiation of human embryonic stem cells (hESCs) using primary bone-derived cells (PBDs). Three days after embryoid body (hEB) formation, cells were allowed to adhere to culture surface where PBDs were pre-plated and mitomycin C-treated in DMEM/F12 medium supplemented with 5% knockout serum replacement. As early as 14 days, mineralization and formation of nodule-like structures in cocultured hEBs were prominent by von Kossa and Alizarin S staining, and expressions of osteoblast-specific markers including bone sialoprotein, alkaline phosphates, osteocalcin, collagen 1, and core binding factor alpha1 by RT-PCR. In addition, FACS analysis revealed that over 19% of the differentiated cells expressed osteocalcin. These results suggest that PBDs not only have osteogenic effects releasing osteogenic factors as bone morphogenic protein (BMP) 2 and BMP 4 but also have exerted other effects, whether chemical or physical, for the differentiation of hESCs.
Ahn
Seong Eun
SE
Stem Cell Research Laboratory, Cell and Gene Therapy Research Institute, Pochon CHA University, Seoul 135-081, Republic of Korea.
Kim
Sinae
S
Park
Kyu Hyung
KH
Moon
Sung Hwan
SH
Lee
Hae Jin
HJ
Kim
Gi Jin
GJ
Lee
Young Jae
YJ
Park
Keun Hong
KH
Cha
Kwang Yul
KY
Chung
Hyung Min
HM
D000328:Adult; D015415:Biomarkers; D055396:Bone Morphogenetic Protein 2; D055415:Bone Morphogenetic Protein 4; D019485:Bone Morphogenetic Proteins / Q000096:biosynthesis / Q000235:genetics; D001842:Bone and Bones / Q000166:cytology* / Q000378:metabolism / Q000502:physiology; D002454:Cell Differentiation / Q000502:physiology*; D002460:Cell Line; D002478:Cells, Cultured; D018920:Coculture Techniques; D004622:Embryo, Mammalian / Q000166:cytology* / Q000378:metabolism; D006133:Growth Substances; D006801:Humans; D010012:Osteogenesis / Q000235:genetics / Q000502:physiology*; D013234:Stem Cells / Q000166:cytology* / Q000378:metabolism; D016212:Transforming Growth Factor beta / Q000096:biosynthesis / Q000235:genetics
C525718:BMP2 protein, human; C525809:BMP4 protein, human; D015415:Biomarkers; D055396:Bone Morphogenetic Protein 2; D055415:Bone Morphogenetic Protein 4; D019485:Bone Morphogenetic Proteins; D006133:Growth Substances; D016212:Transforming Growth Factor beta