Behavior of HepG2/C3A cell cultures in a microfluidic bioreactor
Behavior of HepG2/C3A cell cultures in a microfluidic bioreactor
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DOI：
10.1016/j.bej.2010.10.007
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发表时间：
2011-01-15
影响因子：
3.9
通讯作者：
Leclerc, Eric
中科院分区：
文献类型：
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作者：
Baudoin, Regis;Griscom, Laurent;Leclerc, Eric
毒性研究中的一个重要问题是开发相关的新的体外测试，它将能够提供体内测试方法的替代。目前在组织工程和微技术领域的发展使得使用微流控生物反应器作为一种增强体外研究的工具成为可能。然而，细胞在复杂环境中的行为和它们对化学物质的反应都需要更好地了解，特别是为了未来对任何新测试的验证。为了表征这种方法的敏感性，我们研究了微流控生物反应器中两个细胞培养参数变化对肝细胞模型的影响：接种细胞密度(0.35×10(6)、0.45×10(6)和0.65×10(6)个/生物反应器)和微流控流速(0、10和25亩L/分钟)。我们还调查了一个环境污染物，模拟了三种氨浓度(0，5和10 mm)。在生物反应器中的增殖与流速和接种细胞密度有关。这导致在培养96h后，在没有氨负荷的情况下，生物反应器3D微环境中的平均值为1.2+/-0.2×10(6)个细胞。细胞代谢率，如葡萄糖和谷氨酰胺消耗或细胞色素P1a解毒，在动态条件下比静态条件下更高。此外，氯化铵浓度的增加反过来又增加了葡萄糖和谷氨酰胺的消耗以及细胞色素P1a的活性。在氯化铵分析中，当细胞浓度为0.35×106个/生物反应器时，细胞浓度为5 mM时，细胞增殖抑制(IC50)为50%。相反，对于较大的细胞密度0.65×106个/生物反应器，在5 mm处检测不到任何影响，表明生物反应器中的浓度和细胞密度依赖。这项研究强调了HepG2/C3A细胞对微流控培养条件的敏感性，并说明了使用微流控生物反应器进行更大规模体外毒性研究的可能性。(C)2010爱思唯尔B.V.保留所有权利。
An important issue in toxicity studies is the development of pertinent new in vitro tests that will be able to provide an alternative to in vivo testing methods. Current developments in the fields of tissue engineering and microtechnology make it possible to propose the use of microfluidic bioreactors as a tool for enhanced in vitro investigations. However, both the cells' behavior in complex environments and their response to chemicals need to be better understood, especially for future validation of any new assay. To characterize the sensitivity of this approach, we investigated the behavior of a liver cell model with respect to variations of two cell culture parameters in a microfluidic bioreactor: inoculated cell density (0.35 x 10(6), 0.45 x 10(6) and 0.65 x 10(6) cells/bioreactor) and microfluidic flow rates (0, 10 and 25 mu L/min). We also investigated an environmental pollutant modeled with three ammonia concentrations (0, 5 and 10 mM). Proliferation in the bioreactor was found to be flow rate and inoculated cell density dependent. This led to a mean value of 1.2 +/- 0.2 x 10(6) cells in the 3D microenvironment of the bioreactor without ammonia loadings after 96 h of cultures. Cell metabolism rates, such as glucose and glutamine consumption or CYP1A detoxification, were found to be higher in dynamic conditions than in static conditions. Furthermore, increased ammonium chloride concentration in turn increased glucose and glutamine consumptions and CYP1A activity. Inhibition of 50% of cell proliferation (IC50) during the ammonium chloride analysis was found at 5 mM when cell concentrations of 0.35 x 106 cells/bioreactor were inoculated. In contrast, no effect could be detected at 5 mM for larger cell densities of 0.65 x 106 cells/bioreactor, demonstrating concentration and cell density dependence in the bioreactors. This study highlighted the sensitivity of the HepG2/C3A cells to microfluidic culture conditions and illustrated the potential for larger in vitro toxicity studies using microfluidic bioreactors. (C) 2010 Elsevier B.V. All rights reserved.