IBB - Institute for Biotechnology and Bioengineering, Centre for Biological and Chemical Engineering, Instituto Superior Técnico, Av. Rovisco Pais, 1049-001 Lisboa, Portugal.
J Biotechnol. 2010 Apr 15;146(4):194-7. doi: 10.1016/j.jbiotec.2010.02.015. Epub 2010 Feb 25.
Bioreactor systems have been developed as alternatives to standard culture flasks due to their homogeneous nature, easiness of monitoring and increased cell production. Here we investigated the in vitro expansion of bone marrow (BM) mesenchymal stem cells (MSC) in spinner flasks, using gelatin microcarriers (Cultispher S) to support cell adhesion and proliferation. MSC expansion was performed using a low-serum containing medium (2% of fetal bovine serum, FBS). A strategy was defined for the maximization of cell expansion: microcarriers were pre-coated with FBS in order to increase cell seeding efficiency and an adequate feeding regime was established (25% medium exchange everyday). The maximum cell density, 4.2 x 10(5)cells/mL, was obtained at day 8, corresponding to a fold increase in total cell number of 8.4+/-0.8. Expanded MSC retained their differentiation potential into adipogenic and osteogenic lineages, as well as their clonogenic ability. Harvested cells expressed >90% of CD73, CD90 and CD105 markers. These results demonstrated that a microcarrier-based stirred culture system is adequate for human MSC expansion, using a low-serum containing medium, allowing the generation of significant cell numbers for potential applications in regenerative medicine.
生物反应器系统已经被开发出来作为标准培养瓶的替代品，因为它们具有均一的性质、易于监测和提高细胞产量。在这里，我们研究了在旋转瓶中使用明胶微载体（Cultispher S）来支持细胞黏附和增殖，从而进行骨髓（BM）间充质干细胞（MSC）的体外扩增。MSC 的扩增使用含有低血清的培养基（2%胎牛血清，FBS）进行。定义了一种最大化细胞扩增的策略：预先用 FBS 包被微载体以提高细胞接种效率，并建立了适当的喂养方案（每天 25%的培养基更换）。在第 8 天获得了最高细胞密度 4.2×10(5)个/mL，总细胞数增加了 8.4±0.8 倍。扩增的 MSC 保留了向脂肪和成骨谱系分化的潜力，以及其克隆形成能力。收获的细胞表达了>90%的 CD73、CD90 和 CD105 标志物。这些结果表明，使用低血清含量的培养基，基于微载体的搅拌培养系统足以进行人 MSC 的扩增，允许产生大量的细胞，用于再生医学的潜在应用。
