Wang Tzu-Wei, Wu Hsi-Chin, Wang Hsin-Yen, Lin Feng-Huei, Sun Jui-Sheng
Institute of Biomedical Engineering, College of Medicine and College of Engineering, National Taiwan University, Taipei, Taiwan.
J Biomed Mater Res A. 2009 Mar 15;88(4):935-46. doi: 10.1002/jbm.a.31914.
The aim of this study was to examine the feasibility of expanding and regulating mesenchymal stem cells (MSCs) from isolated adult human bone marrow mononuclear cells, seeded on gelatin-hyaluronic acid biomatrices, and then to quantitatively compare the gene expression in three different culture systems. Individual and interactive effects of model system parameters on construct structure, function, and molecular properties were evaluated. The results showed that these adult human MSCs even at old age not only expressed primitive mesenchymal cell markers but also maintained a high level of colony-forming efficiency and were capable of differentiating into osteoblasts, chondrocytes, and adipocytes upon appropriate inductions. After 21 days of culture, we found that the osteoblastic and chondrocytic lineage gene expression were earlier and higher expressed in spinner flask bioreactor culture group when compared with the static culture and rotating wall vessel reactor culture. The osteogenic lineage proteins type I collagen, alkaline phosphatase, and osteocalcin were strongly stained in histological sections of spinner flask bioreactor culture, whereas these were less detected in the other two groups, especially in rotating wall vessel reactor culture. As for the markers associated with the chondrogenic lineage differentiation proteins, type II collagen was apparently expressed in spinner flask culture group, while the expression of proteoglycans (aggreacan, decorin) in three culture conditions took the lead of each other. We conclude that the spinner flask bioreactor with appropriate induction medium reported in this study may be used to rapidly expand adult MSCs and is likely to possess better induction results toward osteoblastic and chondrocytic lineages.
本研究的目的是检验从分离的成人骨髓单个核细胞中扩增和调控间充质干细胞（MSCs）的可行性，将其接种在明胶-透明质酸生物基质上，然后定量比较三种不同培养系统中的基因表达。评估了模型系统参数对构建体结构、功能和分子特性的个体和交互作用。结果表明，这些成人MSCs即使在老龄时不仅表达原始间充质细胞标志物，还保持高水平的集落形成效率，并且在适当诱导后能够分化为成骨细胞、软骨细胞和脂肪细胞。培养21天后，我们发现与静态培养和旋转壁式生物反应器培养相比，转瓶生物反应器培养组中成骨和软骨谱系基因表达更早且更高。在转瓶生物反应器培养的组织学切片中，成骨谱系蛋白I型胶原蛋白、碱性磷酸酶和骨钙素被强烈染色，而在其他两组中检测到的较少，尤其是在旋转壁式生物反应器培养中。至于与软骨形成谱系分化蛋白相关的标志物，II型胶原蛋白在转瓶培养组中明显表达，而蛋白聚糖（聚集蛋白聚糖、核心蛋白聚糖）在三种培养条件下的表达相互领先。我们得出结论，本研究报道的带有适当诱导培养基的转瓶生物反应器可用于快速扩增成人MSCs，并且可能对成骨和软骨谱系具有更好的诱导效果。
