Development of a scalable suspension culture for cardiac differentiation from human pluripotent stem cells.
Development of a scalable suspension culture for cardiac differentiation from human pluripotent stem cells.
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DOI：
10.1016/j.scr.2015.08.002
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发表时间：
2015-09
影响因子：
1.2
通讯作者：
Couture LA
中科院分区：
文献类型：
--
作者：
Chen VC;Ye J;Shukla P;Hua G;Chen D;Lin Z;Liu JC;Chai J;Gold J;Wu J;Hsu D;Couture LA
为了满足临床前和临床研究对大量 hPSC 衍生心肌细胞 (CM) 的需求，用于 CM 生产的强大且可扩展的分化系统至关重要。利用我们之前建立的 hPSC 聚集体悬浮培养系统，我们开发了一种无基质、可扩展且符合 GMP 的工艺，通过用小分子调节 Wnt 通路来指导 hPSC 在悬浮培养中分化为 CM。通过优化关键工艺参数，包括：细胞聚集体大小、小分子浓度、诱导时间和搅拌速率，我们能够始终如一地将 hPSC 分化为 >90% CM 纯度，在最大 1L 旋转烧瓶规模下，平均产量为 1.5 至 2×109 CM/L。悬浮培养产生的 CM 显示出典型的遗传、形态和电生理学心肌细胞特征。该悬浮培养系统允许在连续悬浮培养中从 hPSC 扩增到 CM 分化的无缝过渡。它不仅为大规模 CM 生产提供了一种具有成本效益和劳动力效益的可扩展工艺，而且还为细胞制造自动化提供了生物反应器原型，这将加速 hPSC 研究向治疗应用的进展。
To meet the need of a large quantity of hPSC-derived cardiomyocytes (CM) for pre-clinical and clinical studies, a robust and scalable differentiation system for CM production is essential. With an hPSC aggregate suspension culture system we established previously, we developed a matrix-free, scalable, and GMP-compliant process for directing hPSC differentiation to CM in suspension culture by modulating Wnt pathways with small molecules. By optimizing critical process parameters including: cell aggregate size, small molecule concentrations, induction timing, and agitation rate, we were able to consistently differentiate hPSCs to >90% CM purity with an average yield of 1.5 to 2×109 CM/L at scales up to 1L spinner flasks. CM generated from the suspension culture displayed typical genetic, morphological, and electrophysiological cardiac cell characteristics. This suspension culture system allows seamless transition from hPSC expansion to CM differentiation in a continuous suspension culture. It not only provides a cost and labor effective scalable process for large scale CM production, but also provides a bioreactor prototype for automation of cell manufacturing, which will accelerate the advance of hPSCs research towards therapeutic applications.