A rapid and efficient method for expansion of human mesenchymal stem cells
A rapid and efficient method for expansion of human mesenchymal stem cells
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DOI：
10.1089/ten.2005.0513
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发表时间：
2007-01-01
影响因子：
--
通讯作者：
De Bruijn, Joost D.
中科院分区：
文献类型：
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作者：
Both, Sanne K.;Van der Muijsenberg, Adrie J. C.;De Bruijn, Joost D.
在过去的十年中，人们对人类间充质干细胞（hMSC）在骨组织工程中的应用产生了浓厚的兴趣。 HMSCs 获得相对容易，并可在培养物中快速扩增，但出于临床目的，通常需要大量细胞，并且成本应保持在最低水平。因此，快速有效的培养方案将是有益的。在这项研究中，我们研究了不同培养基成分对来自 19 名供体的骨髓源性 hMSC 的扩增和成骨分化的影响。我们还研究了低接种密度和地塞米松对 hMSC 扩增及其体外和体内成骨分化能力的影响。以100个细胞/cm(2)密度接种的HMSCs的生长速率明显高于5000个细胞/cm(2)密度，添加地塞米松进一步提高了生长速率。根据 CD29、CD44、CD105 和 CD166 的阳性染色，对扩增的 hMSC 进行体外表征。通过流式细胞仪碱性磷酸酶染色评估扩增的 hMSC 的体外成骨潜力。通过将细胞接种在陶瓷支架中，然后皮下植入裸鼠，并在植入6周后对从头骨形成进行组织病理学评估，评估hMSC的体内成骨潜力。来自所有供体的扩增 hMSC 表现出相似的成骨潜力，与培养条件无关。基于这些结果，我们开发了一种有效的 hMSC 培养方法，将细胞以 100 个细胞/cm(2) 接种到含有地塞米松的 α 最低必需培养基中。
During the past decade, there has been much interest in the use of human mesenchymal stem cells (hMSCs) in bone tissue engineering. HMSCs can be obtained relatively easily and expanded rapidly in culture, but for clinical purposes large numbers are often needed and the cost should be kept to a minimum. A rapid and efficient culturing protocol would therefore be beneficial. In this study, we examined the effect of different medium compositions on the expansion and osteogenic differentiation of bone marrow-derived hMSCs from 19 donors. We also investigated the effect of low seeding density and dexamethasone on both hMSCs expansion and their in vitro and in vivo osteogenic differentiation capacity. HMSCs seeded at a density of 100 cells/cm(2) had a significantly higher growth rate than at 5000 cell/cm(2), which was further improved by the addition of dexamethasone. Expanded hMSCs were characterized in vitro on the basis of positive staining for CD29, CD44, CD105, and CD166. The in vitro osteogenic potential of expanded hMSCs was assessed by flow cytometric staining for alkaline phosphatase. In vivo bone-forming potential of the hMSCs was assessed by seeding the cells in ceramic scaffolds, followed by subcutaneous implantation in nude mice and histopathologic assessment of de novo bone formation after 6-week implantation. Expanded hMSCs from all donors displayed similar osteogenic potential independent of the culture conditions. On the basis of these results we have developed an efficient method to culture hMSCs by seeding the cells at 100 cells/cm(2) in an alpha-minimal essential medium-based medium containing dexamethasone.