A microcarrier-based cultivation system for expansion of primary mesenchymal stem cells
A microcarrier-based cultivation system for expansion of primary mesenchymal stem cells
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DOI：
10.1021/bp060155w
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发表时间：
2007-01-01
影响因子：
2.9
通讯作者：
Ringe, Jochen
中科院分区：
文献类型：
--
作者：
Frauenschuh, Simone;Reichmann, Elisabeth;Ringe, Jochen
微载体培养已被证明允许组织工程相关细胞(如软骨细胞)的广泛细胞扩增，同时保持其表型。我们的目的是研究猪骨髓来源的原代间充质干细胞(MSC)使用商业上可用的Cytodex 1型、2型和3型微载体进行体外三维扩增。经泊松分布分析，Cytodex 1型微载体的贴壁效果最好，3h后贴壁细胞达80%以上。起始细胞密度从1到3×10(6)个/100 cm(2)的不同起始细胞密度对粘附力的影响很小。培养28d后，在Cytodex 1型微载体上检测细胞增殖情况，发现细胞生长情况，并证明细胞可以重新克隆新加入的微载体。扫描电子显微镜显示适当的细胞形态，并确认细胞增殖。从微载体上酶解后，诱导细胞向成骨和向软骨方向分化，并进行相应的组织化学，如von Kossa和阿尔辛蓝染色。综上所述，我们的研究结果表明，微载体上MSC的三维扩展是传统二维单层培养方法的有益替代。
Microcarrier cultures have been shown to allow extensive cell expansion of tissue engineering relevant cells, such as chondrocytes, while maintaining their phenotype. Our aim was to investigate the in vitro three-dimensional expansion of porcine bone-marrow-derived primary mesenchymal stem cells (MSC) using commercially available Cytodex type 1, type 2, and type 3 microcarriers. In comparison, the Cytodex type 1 microcarriers showed the best results for adherence with over 80% adherent cells after 3 h of incubation, analyzed by the Poisson distribution. Different start cell densities ranging from 1 to 3 x 10(6) cells per 100 cm(2) had only a minor influence on adhesion. The proliferation was examined on Cytodex type 1 microcarriers over a cultivation time of 28 days, which could reveal cell growth and proof of cells recolonizing freshly added microcarriers. Scanning electron microscopy displayed appropriate cell morphology and confirmed cell proliferation. After enzymatic harvest from microcarriers, the osteogenic and chondrogenic differentiation of these cells was induced and shown by relevant histochemistry, such as von Kossa and Alcian blue staining. Totaling the results, we have shown that the three-dimensional expansion of MSC on microcarriers represents a beneficial alternative to the conventional two-dimensional monolayer cultivation method.