Open-Space Microfluidics as a Tool to Study Signaling Dynamics

IF 5.4 2区 工程技术 Q1 BIOCHEMICAL RESEARCH METHODS
Lab on a Chip Pub Date : 2025-09-18 DOI:10.1039/d5lc00521c
Maude Proulx, Pierre Clapperton-Richard, Alisa Piekny, Laurent Potvin-Trottier, Thomas Gervais
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引用次数: 0

Abstract

The temporal dynamics of cell signaling are a crucial way for cells to regulate their transcriptional targets and consequently may heavily influence cell responses. Improving our understanding of signaling dynamics is important for drug treatments targeting specific signaling pathways. However, studying signaling dynamics requires multiplexed, time-sensitive experiments. Here, we use an open-space microfluidic device, the microfluidic display, which enables liquid delivery from above a surface, forming defined and stable confinement zones without enclosing samples into a chip. A device with rapid reagent switching (<7 seconds) and 6 independent confinement areas is first designed. Using this platform, we study the Notch pathway in engineered C2C12 cells to display constitutively active Notch receptors upon which we force highly controlled time-dependent modulation patterns by delivering time-varying doses of the Notch inhibitor DAPT. We replicate previous findings on Notch activation with our methodology by confirming the Notch-regulated gene Hes1 is upregulated for short Notch activation pulses, while Hey1 required sustained activation. We confirm a previously observed regime switch from Hes1 to Hey1 dominance between 2h and 3h of activation. Finally, by varying signal pulses while keeping dose constant in six independent experiments performed simultaneously, we further show the upregulation of the Hes1 gene for multiple short pulses, while Hey1 activation depends on duty cycle length. These results highlight microfluidic displays as a valuable tool for systems biology, enabling multiplexed, high temporal resolution stimulation of signaling pathways.
开放空间微流体作为研究信号动力学的工具
细胞信号的时间动态是细胞调节其转录靶点的重要途径,因此可能严重影响细胞反应。提高我们对信号动力学的理解对于针对特定信号通路的药物治疗是重要的。然而,研究信号动力学需要多路复用、时间敏感的实验。在这里,我们使用一个开放空间的微流控装置,微流控显示器,它使液体从表面上方输送,形成明确和稳定的约束区,而不将样品封闭到芯片中。首先设计了具有快速试剂切换(<;7秒)和6个独立约束区的装置。利用这个平台,我们研究了工程C2C12细胞中的Notch通路,以显示组成活性的Notch受体,我们通过递送时变剂量的Notch抑制剂DAPT,在其上施加高度控制的时间依赖性调节模式。通过证实Notch调控基因Hes1在短Notch激活脉冲下上调,而Hey1需要持续激活,我们重复了之前关于Notch激活的研究结果。我们证实了先前观察到的在激活2h和3h之间从Hes1到Hey1主导的状态转换。最后,通过在保持剂量恒定的情况下改变信号脉冲,我们在同时进行的六个独立实验中进一步证明了多个短脉冲下Hes1基因的上调,而Hey1的激活取决于占空比长度。这些结果突出了微流控显示器作为系统生物学的一种有价值的工具,能够实现信号通路的多路复用、高时间分辨率刺激。
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来源期刊
Lab on a Chip
Lab on a Chip 工程技术-化学综合
CiteScore
11.10
自引率
8.20%
发文量
434
审稿时长
2.6 months
期刊介绍: Lab on a Chip is the premiere journal that publishes cutting-edge research in the field of miniaturization. By their very nature, microfluidic/nanofluidic/miniaturized systems are at the intersection of disciplines, spanning fundamental research to high-end application, which is reflected by the broad readership of the journal. Lab on a Chip publishes two types of papers on original research: full-length research papers and communications. Papers should demonstrate innovations, which can come from technical advancements or applications addressing pressing needs in globally important areas. The journal also publishes Comments, Reviews, and Perspectives.
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