Suspension Culture of Human Pluripotent Stem Cells in Controlled, Stirred Bioreactors
Suspension Culture of Human Pluripotent Stem Cells in Controlled, Stirred Bioreactors
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DOI：
10.1089/ten.tec.2011.0717
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发表时间：
2012-10-01
影响因子：
3
通讯作者：
Zweigerdt, Robert
中科院分区：
文献类型：
--
作者：
Olmer, Ruth;Lange, Andreas;Zweigerdt, Robert
多能干细胞及其衍生物的治疗和工业应用需要在限定条件下产生大量细胞。为此，我们将人多能干细胞（hPSC;包括人诱导多能干细胞[hiPS]和人胚胎干细胞[hESC]）的单细胞接种悬浮培养物转移到搅拌罐生物反应器中。这些系统广泛应用于生物制药行业，可以直接放大并详细在线监控关键工艺参数。为了确保最低培养基消耗，但同时对工艺监测所必需的所有探针进行功能整合，即pO（2）和pH，在由四个独立控制容器组成的“微型反应器平台”中以100 mL培养体积进行实验。通过建立严格控制细胞接种和聚集体形成的规定参数，在7天内的单次工艺运行中生成了高达2 x 108个hiPSC/100 mL。多能性标记物的表达和细胞在体外分化成所有三个胚层的衍生物的能力得以维持，强调了这种新方法的实际效用。所提供的数据提供了人类iPS和ES细胞可规模化大规模扩增的关键步骤，从而使干细胞研究能够转化为相关大型动物模型中的（前）临床应用，以及用于药物开发和验证的有价值的体外试验。
Therapeutic and industrial applications of pluripotent stem cells and their derivatives require large cell quantities generated in defined conditions. To this end, we have translated single cell-inoculated suspension cultures of human pluripotent stem cells (hPSCs; including human induced pluripotent stem cells [hiPS] and human embryonic stem cells [hESC]) to stirred tank bioreactors. These systems that are widely used in biopharmaceutical industry allow straightforward scale up and detailed online monitoring of key process parameters. To ensure minimum medium consumption, but in parallel functional integration of all probes mandatory for process monitoring, that is, for pO(2) and pH, experiments were performed in 100 mL culture volume in a "mini reactor platform'' consisting of four independently controlled vessels. By establishing defined parameters for tightly controlled cell inoculation and aggregate formation up to 2 x 10(8) hiPSCs/100 mL were generated in a single process run in 7 days. Expression of pluripotency markers and ability of cells to differentiate into derivates of all three germ layers in vitro was maintained, underlining practical utility of this new process. The presented data provide key steps toward scalable mass expansion of human iPS and ES cells thereby enabling translation of stem cell research to (pre)clinical application in relevant large animal models and valuable in vitro assays for drug development and validation as well.