A xeno-free microcarrier-based stirred culture system for the scalable expansion of human mesenchymal stem/stromal cells isolated from bone marrow and adipose tissue
A xeno-free microcarrier-based stirred culture system for the scalable expansion of human mesenchymal stem/stromal cells isolated from bone marrow and adipose tissue
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DOI：
10.1002/biot.201400586
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发表时间：
2015-08-01
影响因子：
4.7
通讯作者：
Cabral, Joaquim M. S.
中科院分区：
文献类型：
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作者：
Carmelo, Joana G.;Fernandes-Platzgummer, Ana;Cabral, Joaquim M. S.
人间充质干细胞/基质细胞（MSC）是基于细胞的疗法的有前途的候选者，并且在具有明确定义的无异种成分的可扩展生物反应器中开发基于微载体的培养物代表了这些细胞的临床规模生产的重要里程碑。在这项工作中，我们优化了我们以前开发的基于无异种微载体的系统，用于从骨髓分离的人MSC（BM MSC）和脂肪源性干细胞/基质细胞（ASC）的可扩展扩增。通过在转瓶中的初始细胞接种/培养阶段调整搅拌/补料方案，我们能够最大化细胞扩增速率和最终细胞产量。在培养7天和8天时，BM MSC（0.60 +/- 0.04天（-1））和ASC（0.9 +/- 0.1天（-1））培养物的最大细胞密度分别为3.6 x 10（5）和1.9 x 10（5）个细胞/mL。即用型微载体Synthemax（R）II和Enhanced Attachment（R）支持BM MSC的相同扩增性能，将这些有效替代品转化为我们的无异种可扩展培养系统中使用的预涂塑料微载体。重要的是，扩增的MSC保持了其免疫表型和多向分化潜能。此外，分泌组分析表明，引发效应的搅拌培养条件对细胞因子的生产MSC。该培养系统产生了相当大的最终细胞密度，其可以按比例放大到受控的大规模生物反应器，从而允许用于临床环境的更有效、安全和具有成本效益的MSC生产。
Human mesenchymal stem/stromal cells (MSC) are promising candidates for cell-based therapies and the development of microcarrier-based cultures in scalable bioreactors with well-defined xenogeneic-free components represent important milestones towards the clinical-scale production of these cells. In this work, we optimized our previously developed xeno-free microcarrier-based system for the scalable expansion of human MSC isolated from bone marrow (BM MSC) and adipose-derived stem/stromal cells (ASC). By adapting the agitation/feeding protocol at the initial cell seeding/cultivation stage in spinner flasks, we were able to maximize cell expansion rate and final cell yield. Maximal cell densities of 3.6 x 10(5) and 1.9 x 10(5) cells/mL were obtained for BM MSC (0.60 +/- 0.04 day(-1)) and ASC (0.9 +/- 0.1 day(-1)) cultures, upon seven and eight days of cultivation, respectively. Ready-to-use microcarriers Synthemax (R) II and Enhanced Attachment (R) supported identical expansion performance of BM MSC, turning those effective alternatives to the precoated plastic microcarriers used in our xeno-free scalable culture system. Importantly, expanded MSC maintained their immunophenotype and multilineage differentiation potential. Moreover, secretome analysis suggested a priming effect of stirred culture conditions on cytokine production by MSC. This culture system yielded considerable final cell densities that can be scaled-up to controlled large-scale bioreactors allowing a more efficient, safe and cost-effective MSC production for clinical settings.