Ping Zhou1, Jia-Min Shi2, Jing-E Song1, Yu Han1, Hong-Jiao Li1, Ya-Meng Song1, Fang Feng1, Jian-Lin Wang2, Rui Zhang3,4, Feng Lan5. 1. School and Hospital of Stomatology, Lanzhou University, No.222 Tianshui South Road, Chengguan District, Lanzhou, 730000, Gansu Province, People's Republic of China. 2. College of Life Sciences, Lanzhou University, No.222 Tianshui South Road, Chengguan District, Lanzhou, 730000, Gansu Province, People's Republic of China. 3. School and Hospital of Stomatology, Lanzhou University, No.222 Tianshui South Road, Chengguan District, Lanzhou, 730000, Gansu Province, People's Republic of China. zhangrui@lzu.edu.cn. 4. College of Life Sciences, Lanzhou University, No.222 Tianshui South Road, Chengguan District, Lanzhou, 730000, Gansu Province, People's Republic of China. zhangrui@lzu.edu.cn. 5. National Center for Cardiovascular Diseases, Fuwai Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, 100037, People's Republic of China. fenglan@ccmu.edu.cn.
Abstract
BACKGROUND: Derivation of osteoblast-like cells from human pluripotent stem cells (hPSCs) is a popular topic in bone tissue engineering. Although many improvements have been achieved, the low induction efficiency because of spontaneous differentiation hampers their applications. To solve this problem, a detailed understanding of the osteogenic differentiation process of hPSCs is urgently needed. METHODS: Monolayer cultured human embryonic stem cells and human-induced pluripotent stem cells were differentiated in commonly applied serum-containing osteogenic medium for 35 days. In addition to traditional assays such as cell viability detection, reverse transcription-polymerase chain reaction, immunofluorescence, and alizarin red staining, we also applied studies of cell counting, cell telomerase activity, and flow cytometry as essential indicators to analyse the cell type changes in each week. RESULTS: The population of differentiated cells was quite heterogeneous throughout the 35 days of induction. Then, cell telomerase activity and cell cycle analyses have value in evaluating the cell type and tumourigenicity of the obtained cells. Finally, a dynamic map was made to integrate the analysis of these results during osteogenic differentiation of hPSCs, and the cell types at defined stages were concluded. CONCLUSIONS: Our results lay the foundation to improve the in vitro osteogenic differentiation efficiency of hPSCs by supplementing with functional compounds at the desired stage, and then establishing a stepwise induction system in the future.
BACKGROUND: Derivation of osteoblast-like cells from human pluripotent stem cells (hPSCs) is a popular topic in bone tissue engineering. Although many improvements have been achieved, the low induction efficiency because of spontaneous differentiation hampers their applications. To solve this problem, a detailed understanding of the osteogenic differentiation process of hPSCs is urgently needed. METHODS: Monolayer cultured human embryonic stem cells and human-induced pluripotent stem cells were differentiated in commonly applied serum-containing osteogenic medium for 35 days. In addition to traditional assays such as cell viability detection, reverse transcription-polymerase chain reaction, immunofluorescence, and alizarin red staining, we also applied studies of cell counting, cell telomerase activity, and flow cytometry as essential indicators to analyse the cell type changes in each week. RESULTS: The population of differentiated cells was quite heterogeneous throughout the 35 days of induction. Then, cell telomerase activity and cell cycle analyses have value in evaluating the cell type and tumourigenicity of the obtained cells. Finally, a dynamic map was made to integrate the analysis of these results during osteogenic differentiation of hPSCs, and the cell types at defined stages were concluded. CONCLUSIONS: Our results lay the foundation to improve the in vitro osteogenic differentiation efficiency of hPSCs by supplementing with functional compounds at the desired stage, and then establishing a stepwise induction system in the future.
Authors: Nico Heins; Mikael C O Englund; Cecilia Sjöblom; Ulf Dahl; Anna Tonning; Christina Bergh; Anders Lindahl; Charles Hanson; Henrik Semb Journal: Stem Cells Date: 2004 Impact factor: 6.277
Authors: Jeffrey M Karp; Lino S Ferreira; Ali Khademhosseini; Albert H Kwon; Judy Yeh; Robert S Langer Journal: Stem Cells Date: 2005-10-27 Impact factor: 6.277
Authors: W Liu; S Toyosawa; T Furuichi; N Kanatani; C Yoshida; Y Liu; M Himeno; S Narai; A Yamaguchi; T Komori Journal: J Cell Biol Date: 2001-10-01 Impact factor: 10.539