摘要
目的 研究人MSC诱导为成骨细胞后 ,其在脱钙骨上的粘附特性。方法 采用密度梯度法分离人骨髓中骨髓基质干细胞 ,条件培养液诱导培养至第 3代细胞 ,经相差显微镜观察 ,钙结节茜素红染色 ,免疫组化检测Ⅰ型胶原和骨钙蛋白 ,RT -PCR检测Ⅰ型胶原和骨钙蛋白mRNA的表达。不同浓度细胞悬液接种脱钙骨 ,MTT计数未粘附细胞量 ,计算不同浓度下MSC的粘附率 ,单位重量脱钙骨上MSC最多能粘附的细胞数量。结果 MSC经体外诱导形成钙结节 ,Ⅰ型胶原和骨钙蛋白免疫组化结果阳性 ,RT -PCR证实Ⅰ型胶原和骨钙蛋白mRNA的表达。脱钙骨上接种MSC达到饱和后 ,MSC的粘附率随接种浓度上升而下降。单位重量脱钙骨最多粘附的MSC量为 3 .5×1 0 7/ g。 结论 人MSC在体外经条件培养液诱导可表达成骨细胞表型 ,在脱钙骨上有良好的粘附能力 。
Objective To study the feasibility of osteoblast phenotype differentiation of bone marrow MSC and its adhersion rate on decalcified bone. Methods Human MSC were isolated from bone marrow, which were aspirated from iliac crest of volunteers and cultured in defined medium in vitro. The mineralized node was stained by Alizarin-Red. The expression of typeⅠ collagen and osteocalcin in 3rd generation MSC were detected by immunohistological staining technique, and their mRNA expression by RT-PCR. With different densities, cell suspension were seeded on decalcified bone, MTT automated colormetric microassay was adapted to count the number of cells adhered to the decalcified bone. Results The differentiation of MSC to osteoblastic phenotype was verified by the positive staining of mineralized node and ColⅠand OCN by immmunohistochemistry and RT-PCR. Conclusion Bone marrow mesenchymal stem cell can differentiate to osteoblast phenotype. The maximal number which MSC could adhere to decalcified bone is 35×10 7/g.
出处
《实用美容整形外科杂志》
2003年第2期66-69,共4页
Journal of Practical Aesthetic and Plastic Surgery
基金
国家重点基础研究发展规划 ( 973)资助项目( 19990 54 30 5)
上海市科委软骨组织工程的临床应用资助项目( 0 0DJ14 0 0 1- 1)
