茜素红半定量测定细胞基质钙含量的实验研究  

The Research of Extracellular Matrix Calcium Mineralization using Alizarin Red S Assay

作　　者：李宇琳[1] 周恒[1] 刘广鹏[1] 崔磊[1] 

机构地区：[1]上海组织工程研究与开发中心,上海市200235

出　　处：《组织工程与重建外科杂志》2008年第2期61-64,共4页Journal of Tissue Engineering and Reconstructive Surgery

基　　金：国家高技术发展计划(863)项目(2006AA02A123)

摘　　要：目的使用茜素红检测的方法,对细胞在成骨诱导和未诱导状态下,细胞基质中钙盐沉积情况进行半定量分析,研究细胞基质中钙盐沉积情况变化趋势规律,为临床实践提供理论参考。方法对4例正常人骨髓间充质干细胞(Bone marrow mesenchymal stem cells,BMSCs)及4例正常人脂肪间充质干细胞(Adipose-derived mesenchymal stem cells,ASCs),使用茜素红半定量法检测成骨诱导及未诱导细胞基质中钙盐沉积情况进行半定量分析,研究细胞基质中钙盐沉积情况变化趋势规律。结果对于相同代次的间充质干细胞,未诱导组的钙盐沉积情况,在7d后没有显著的改变;而诱导组钙盐的沉积在10～14d明显增多。实验结果同时显示,同一样本的间充质干细胞,随着代次增加,未诱导组钙盐的沉积时间延后,且沉积量逐渐减少。诱导组钙盐的沉积情况,7d时没有显著的改变;14d时,随代次增加,钙盐沉积量逐渐减少。结论体外成骨诱导的第3～4代的间充质干细胞在第10～14天已经具有较强的成骨活性,可能比较适宜体内回植。本实验为组织工程骨的体外构建时间提供了理论依据。Objective To investigate the extracellular matrix （ECM） Calcium mineralization using the method of halfquantitative Alizarin Red S assay, determining the trend of ECM Calcium mineralization in vitro and providing reference to clinical application. Methods The ECM Calcium mineralization of human bone marrow mesenehymal stem cells （BMSCs） and adipose-derived mesenchymal stem cells （ASCs） was determined using the method of half-quantitative Alizarin Red S assay, under the condition of osteogenic induced or non-induced. Results In the non-induced group of the same passage stem cells, after 7 days of culture, there was no significant change of the ECM Calcium mineralization in vitro. For the osteogenic-induced group, the Alizarin Red S results showed a trend of continuously increased mineralization after 10 days of culture. The results also illustrated that along with passaging, ECM Calcium mineralization in vitro of the non-induced group would take a longer time with fewer amount of calcium. Similar trend also occurred to the induced group. Conclusion Based on the results, after being osteogenically induced for 10 to 14 days in vitro, the stem cells of P3 to P4 may be suitable for in vivo implantation.

关 键 词：茜素红 细胞基质 钙离子 骨髓基质干细胞 脂肪间充质干细胞 种子细胞 

分 类 号：Q813.11[生物学—生物工程]