Precise control over the oxygen conditions within the Boyden chamber using a microfabricated insert
Precise control over the oxygen conditions within the Boyden chamber using a microfabricated insert
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DOI：
10.1039/c004856a
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发表时间：
2010-01-01
期刊：
影响因子：
6.1
通讯作者：
Eddington, David T.
中科院分区：
文献类型：
--
作者：
Oppegard, Shawn C.;Blake, Alexander J.;Eddington, David T.
细胞迁移是癌细胞转移的标志，与肿瘤中的低氧密切相关。Boyden小室是一个基于多孔膜的迁移平台，由于其对常见培养容器的适应性和相对易用性，已广泛用于体外迁移和侵袭分析。缺氧室是一种目前可以用来研究细胞对氧气模式的反应的工具。不幸的是，这种方法缺乏空间和时间上的精确度，无法准确地模拟许多生理现象。在这篇文章中，我们提出了一种新开发的微细加工聚二甲基硅氧烷(PDMS)设备，该设备易于适应Boyden小室，并提供对暴露在细胞中的氧合条件的更多控制。该装置在大约20分钟内平衡到1%的氧气，从而展示了研究人员建立短期、持续和间歇性低氧状态的系统的能力。使用对二甲苯-C薄膜涂层来防止环境空气穿透本体PDMS，并发现其改善了平衡时间和终点浓度。以乳腺癌细胞株MDA-MD-231为模型细胞，研究氧浓度对细胞在Boyden小室多孔膜中迁移的影响。与常氧对照相比，持续低氧降低了细胞的迁移，在0%到21%氧气(0 21%IH)之间的间歇低氧循环(IH)也是如此。然而，暴露于5.21%IH的细胞与其他条件相比以及相对于常氧对照组显示出更多的迁移。这里给出的结果表明，该装置可以用于实施博伊登小室进行体外低氧研究的实验，使实验比目前可能进行的更快和更精确。
Cell migration is a hallmark of cancer cell metastasis and is highly correlated with hypoxia in tumors. The Boyden chamber is a porous membrane-based migration platform that has seen a great deal of use for both in vitro migration and invasion assays due to its adaptability to common culture vessels and relative ease of use. The hypoxic chamber is a current tool that can be implemented to investigate the cellular response to oxygen paradigms. Unfortunately, this method lacks the spatial and temporal precision to accurately model a number of physiological phenomena. In this article, we present a newly developed microfabricated polydimethylsiloxane (PDMS) device that easily adapts to the Boyden chamber, and provides more control over the oxygenation conditions exposed to cells. The device equilibrates to 1% oxygen in about 20 min, thus demonstrating the capabilities of a system for researchers to establish both short-term continuous and intermittent hypoxia regimes. A Parylene-C thin-film coating was used to prevent ambient air penetration through the bulk PDMS and was found to yield improved equilibration times and end-point concentrations. MDA-MD-231 cells, an invasive breast cancer line, were used as a model cell type to demonstrate the effect of oxygen concentration on cell migration through the Boyden chamber porous membrane. Continuous hypoxia downregulated migration of cells relative to the normoxic control, as did an intermittent hypoxia regime (IH) cycling between 0% and 21% oxygen (0 21% IH). However, cells exposed to 5 21% IH exhibited increased migration compared to the other conditions, as well as relative to the normoxic control. The results presented here show the device can be utilized for experiments implementing the Boyden chamber for in vitro hypoxic studies, allowing experiments to be conducted faster and with more precision than currently possible.