摘要
目的研究人骨髓基质干细胞经体外诱导表达成骨细胞表型,作为骨组织工程临床应用种子细胞的可行性。方法人骨髓中密度梯度法分离骨髓基质干细胞,诱导组予条件培养液,对照组予单纯DMEM培养液,分别培养至第3代细胞,计算各代倍增时间和3代总的扩增倍数,经相差显微镜观察,钙结节茜素红染色,四环素荧光,免疫荧光检测骨钙蛋白,成骨细胞转录因子RT-PCR检测Ⅰ型胶原,骨钙蛋白mRNA表达。结果诱导组骨髓基质干细胞(MSC)经体外诱导,三代平均扩增163.4倍后,形成钙结节,骨钙蛋白(OCN),成骨细胞转录因子(cbfa1)免疫荧光结果阳性,RT-PCR证实Ⅰ型胶原,骨钙蛋白mRNA的表达;对照组未能形成钙结节,无成骨细胞特征性蛋白OCN,cbfa1的表达。结论人MSC体外经条件培养液诱导培养三代后仍可表达成骨细胞表型,可以满足骨组织工程临床应用对种子细胞质和量的需要。
Objective To study the feasibility of osteobla st phenotype differentiation of bon e marrow mesenchymal stem cells(MSC),and using them as the stem cells of bone ti ssue engineering for clinical application.Methods Human MSC were isolated from bone marrow,which aspirated from iliac crest and cultu red in defined medium in vitro.The mi neralized node was stained by Alizarin-Red .The expression of typeⅠcollagen,osteocalcin and cbfa1in 3th passage MSC were detected by immun ohistological staining technique,and their mRNA expression by RT-PCR.Results The differentiation of MSC to osteob lastic phenotype was verified by the positive result of mineralized node and ColⅠ,OCN and cbfa1by immmunohistochemistry and RT-PCR.Conclusion Bone marrow mesenchymal stem cells c an differentiate to osteoblast phen o-type.It' s optimistic to use MSC as the stem cell of bone tissue engineering for clin ical application.
出处
《中国临床康复》
CSCD
2002年第22期3340-3341,I001,共3页
Chinese Journal of Clinical Rehabilitation
基金
国家重点基础研究发展规划(973):G1999054305
上海市科委:软骨组织工程的临床应用(00DJ14001-1)
