Chinese Journal of Tissue Engineering Research ›› 2012, Vol. 16 ›› Issue (32): 5967-5972.doi: 10.3969/j.issn.2095-4344.2012.32.015

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Time-dependent morphological characteristics of mouse embryonic stem cells differentiating into osteoblasts

Shi Xiao-lu1, Li Hua-jing1, Ma Yan2, Bi Xiao-juan2, Xie Feng-lian3, Wang Lu-xiang4，Liu Li-zhong1   

  1. 1Otorhinolaryngologic Department,
    2Medical Research Center, First Affiliated Hospital of Xinjiang Medical University, Urumqi 830054, Xinjiang Uyghur Autonomous Region, China;
    3Xinjiang Medical University, Urumqi 830054, Xinjiang Uyghur Autonomous Region, China;
    4Xinjiang University, Urumqi 830000, Xinjiang Uyghur Autonomous Region, China
  • Received:2012-01-30 Revised:2012-04-11 Online:2012-08-05 Published:2012-08-05
  • Contact: Liu Li-zhong, Otorhinolaryngologic Department, First Affiliated Hospital of Xinjiang Medical University, Urumqi 830054, Xinjiang Uyghur Autonomous Region, China liulizhongno.1@163.com
  • About author:Shi Xiao-yu★, Master, Otorhinolaryngologic Department, First Affiliated Hospital of Xinjiang Medical University, Urumqi 830054, Xinjiang Uyghur Autonomous Region, China lauren84103@sina.com

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Abstract:

BACKGROUND: Nano-hydroxyapatite has good biocompatibility and bioactivity. It can tightly connect with in vitro tissues within a short time. Therefore, nano-hydroxyapatite has been widely used for bone tissue engineering.
OBJECTIVE: To investigate the time-dependent morphological characteristics of mouse embryonic stem cells differentiating into osteoblasts.
METHODS: P3 embryonic stem cells from C57 mouse were prepared into embryoid bodies. Primary embryonic stem cells were identified by alkaline phosphatase staining and immunohistochemical analysis (oct 4, sox2, nanog). After osteogenic induction by L-vitamin C, β-phosphoglycerol, and dexamethasone, cells were identified by alizarin red S, collagen Ⅰ, and von Kossa staining. The morphology and proliferation of cells when cultured with nano-hydroxyapatite were observed using scanning electron microscope.
RESULTS AND CONCLUSION: The expression of alkaline phosphatase showed no obvious change in each group within 7 days after adherence of embryoid bodies, but increased gradually after 14 days. Green nodules of varying size with clear border could be seen under optical microscope. After Alizarin red S staining, orange red nodules of varying size with clear border could be observed. Black precipitates appeared in the cell mass after von Kossa staining for 14 days, and amplified after 21 days. After co-culture with nano-hydroxyapatite for 1, 3, 5, 7, 10 days, the majority of embryonic stem cells grew in cell mass. These findings suggest that co-application of vitamin C, β-phosphoglycerol and dexamethasone effectively promote embryonic stem cells to differentiate into osteoblasts; embryonic stem cells composited onto scaffold material can be used in bone tissue engineering.

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