摘要
目的:研究人脐带间充质干细胞(hUCMSCs)的成骨分化及其在成骨诱导过程中成骨相关标记物的变化情况。方法:通过原代培养的方法获得hUCMSCs;通过流式细胞技术对获得的细胞进行鉴定;通过茜素红染色和细胞免疫荧光技术对成骨诱导后的细胞进行鉴定;在成骨诱导过程中设立时间点(0d、7d、14d、21d),对成骨相关标记物(碱性磷酸酶、骨桥蛋白、Runx2)进行检测。结果:通过原代培养成功获得hUCMSCs;流式分析结果显示其表达间充质干细胞标记物;进行成骨诱导21d后茜素红染色阳性,免疫荧光染色检测胞浆内大量表达骨桥蛋白;碱性磷酸酶活性和骨桥蛋白(mRNA水平)在第7天时即表现出具有统计学差异的升高,Runx2(mRNA水平)在第7天时达到峰值,随后下降。结论:hUCMSCs的成骨分化过程是一个连续的过程;体外诱导7d后成骨分化进行已经全面启动;经过诱导的hUCMSCs比未诱导的细胞更适合用于骨缺损修复。
Objective： To investigate the change of osteogenic related markers in human umbilical cord mesenchymal stem cells （hUCMSCs） during osteogenic differentiation. Methods： The hUCMSCs were obtained by primary culture and identified using flow cytometry. Alizarin red staining and Immunofluorescence were performed to assess hUCMSC proliferation and differentiation potential in vitro. Alkaline phosphatase activity （ALP）, mRNA level of osteopontin （OPN） and Runx2 were measured at 0, 7, 14, 21d. Results： The hUCMSCs could be obtained by primary culture of human umbilical cords. The flow cytometry results showed that the cells expressed numerous mesenchymal stem cell markers. After osteogenic induction for 21 days in vitro, Alizarin red stain was positive, and immunofluorescence staining showed that OPN was greatly expressed in the cytoplasm. The ALP activity and the mRNA expression levels of OPN increased at day 7 and kept a high level during osteogenic differentiation. The mRNA expression levels of Runx2 had peak value at day 7 and then declined gradually. Conclusion： The osteogenic differentiation of hUCMSCs is a continuous process which started completely after 7 days induction. The predifferentiated hUCMSCs may be superior to the undifferentiated cells in bone defect repair.
出处
《口腔医学研究》
CAS
CSCD
2014年第8期709-712,共4页
Journal of Oral Science Research
基金
国家自然科学基金(编号:81100792)
关键词
间充质干细胞
脐带
细胞培养
成骨分化
Mesenchymal stem cells Umbilical cord Cell culture Osteogenic differentiation
