Periodic "flow-stop" perfusion microchannel bioreactors for mammalian and human embryonic stem cell long-term culture
Periodic "flow-stop" perfusion microchannel bioreactors for mammalian and human embryonic stem cell long-term culture
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DOI：
10.1007/s10544-008-9212-5
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发表时间：
2009-02-01
影响因子：
2.8
通讯作者：
Levenberg, Shulamit
中科院分区：
文献类型：
--
作者：
Korin, Natanel;Bransky, Avishay;Levenberg, Shulamit
本研究探讨了使用自动定期“流动停止”灌注系统的长期培养哺乳动物细胞在微通道生物反应器。该方法用于在微通道（高度100 μ m）中长时间（> 7天）培养人包皮成纤维细胞（HFF）和人脐静脉内皮细胞（HUVEC）。设计参数，质量输运和剪切应力的问题进行了理论研究，通过数值模拟。实验监测细胞生长和形态，并测量与恒定灌注微反应器和培养皿中的传统培养物相比增强的生长速率。此外，我们证明了使用该方法在微通道中的HFF饲养细胞上共培养人胚胎干细胞（hESC）的未分化集落。成功的hESC-HFF在微生物反应器中的共培养是由于所开发的方法的两个重要特征实现的-短时间暴露于流动，随后是长的静态孵育期。短脉冲暴露于剪切使得剪切敏感细胞（例如，hESC）以承受培养基更新流。长的静态孵育期可以使分泌的因子（例如，饲养细胞分泌的因子）局部积累。因此，所开发的方法可能适合于微系统中敏感多细胞复合体的长期培养。
The present study examines the use of automated periodic "flow-stop" perfusion systems for long-term culture of mammalian cells in a microchannel bioreactor. The method is used to culture Human Foreskin Fibroblasts (HFF) and Human Umbilical Vein Endothelial Cells (HUVEC) for long periods of time (> 7 d) in a microchannel (height 100 mu m). Design parameters, mass transport and shear stress issues are theoretically examined via numerical simulations. Cell growth and morphology are experimentally monitored and an enhanced growth rate was measured compared to constant perfusion micro-reactors and to traditional culture in Petri dishes. Moreover, we demonstrate the use of the method to co-culture undifferentiated colonies of human Embryonic Stem Cells (hESC) on HFF feeder cells in microchannels. The successful hESC-HFF co-culture in the microbioreactor is achieved due to two vital characteristics of the developed method-short temporal exposure to flow followed by long static incubation periods. The short pulsed exposure to shear enables shear sensitive cells (e.g., hESC) to withstand the medium renewal flow. The long static incubation period may enable secreted factors (e.g., feeder cells secreted factors) to accumulate locally. Thus the developed method may be suitable for long-term culture of sensitive multi-cellular complexes in microsystems.