摘要
目的观察脉冲电磁场对小鼠胚胎干细胞向成骨细胞分化的影响。方法将小鼠4d龄胚胎干细胞(embryonic stem cells,ESCs)衍生的胚胎体单细胞按5×104ml-1接种到细胞培养板,在DMEM基础培养液中使胚胎体单细胞贴壁培养。于第14～21天时,分别给予脉冲电磁场、化学诱导剂以及联合刺激。采用茜素红染色、骨钙素免疫组织化学染色成骨分化关键因子Osterix和Cbfa-1/Runx-2的相对定量RT-PCR检测等方法观测成骨分化情况。结果茜素红染色和RT-PCR结果均显示:脉冲电磁场组、化学诱导组以及联合培养组的成骨诱导效果均比空白对照组显著增加,其中又以联合培养组成骨诱导效果最明显。骨钙素免疫组化染色也显示:联合培养组中骨钙素分泌最多。结论脉冲电磁场可以促进小鼠胚胎干细胞向成骨细胞分化,而且脉冲电磁场和常规化学诱导剂联合应用会产生协同效应,其成骨诱导效果优于两者单独应用。
Objective To investigate the effect of pulsed electromagnetic fields （PEMFs） on the osteoblast differentiation of mouse embryonic stem cells. Methods The ceils isolated from 4-day embryoid bodies （EBs） derived from mouse embryonic stem ceils （mESCs） were inoculated into each well of cell culture plate at 5 ×10^4ml-1 in DMEM and then cultured. During the period from 14 to 21 d after the EBs-derived single ceils attachment, the cells was divided into 4 groups： control （group A）, PEMFs （group B, 50 Hz, 1.1 roT, twice per day, 30 rain per time, in an interval of 12 h）, chemical inductor （group C, 50 μg/ml Vitamin C, 10 mmol/L β-glycerophosphate and 1 μ moL/L dexamethasone in the culture medium）, B ＋ C （group D）. Histochemical staining for Alizarin Red, immunocytochemical staining for osteoclacin expression and PCR for Osterix and Cbfa-l/Runx-2 were applied to measure the osteoblast differentiation. Results Alizarin Red staining revealed that compared with group A, the surface area of bone nodule colonies of group B, C and D were increased respectively by 123% , 229% and 262% （P 〈0.01 ）. The surface area of bone nodule colonies of group D was raised by 62% compared with that of group B （P 〈 0.01 ） and by 10% with that of group C （P 〈 0.05）. Immunocytochemical staining for osteoclacin revealed that the expression of osteoclacin were the most in group D. PCR revealed that the expression of Osterix and Cbfa-1/Runx-2 of group B, C and D were more than that of group A. Conclusion PEMF accelerates the differentiation of mESCs to osteoblasts. Combining PEMFs and chemical inducers together have synergistic effect.
出处
《第三军医大学学报》
CAS
CSCD
北大核心
2009年第9期772-775,共4页
Journal of Third Military Medical University
基金
国家高技术研究发展计划(863计划)(2006AA02Z4E3)~~
关键词
脉冲电磁场
胚胎干细胞
成骨细胞
定向诱导
骨组织工程
pulsed electromagnetic fields
embryonic stem cells
osteoblast
directed differentiation
bone tissue engeering
