基于微流控芯片的细胞内信号操纵新技术研究

项目名称： 基于微流控芯片的细胞内信号操纵新技术研究

项目编号： No.31471257

项目类型： 面上项目

立项/批准年度： 2015

项目学科： 遗传学与生物信息学、细胞生物学

项目作者： 杜伟

作者单位： 华中科技大学

项目金额： 80万元

中文摘要： 钙信号是细胞采用的一种普遍的调控方式，由于信号分子受体一般同多条信号通路偶联，难以将特定形式的钙信号与某条信号通路对应起来，而人工建立细胞内信号可以避免旁路信号的干扰，有助于更深入揭示钙信号调控机理。目前常用的细胞内信号操纵方法无法精确控制信号在胞内形成特定的空间分布。在我们建立的基于水力门控的细胞刺激微流控芯片基础上，本项目提出了基于微流控生物芯片的细胞内信号操纵新技术研究：利用高度可控的双负压系统快速切换细胞外液体环境，在细胞内形成特定振幅、频率与浓度梯度分布的钙振荡以及不均一激活的Rac1信号，从而建立可形成更加真实、时空特性更好的胞内信号的微流控芯片平台。并进一步结合功能光学成像技术深入研究钙信号以及Rho-GTPase信号在调控细胞迁移、信号小分子的时空变化、关键蛋白的活性中所起的作用。本课题对于阐明细胞内信号转导机理具有重要科学价值，并促进学科交叉与人才培养。

中文关键词： 微流控芯片；光学分子成像；细胞迁移；钙信号；信号转导

英文摘要： Almost all physiological activities are regulated by calcium signaling in cells. However, the details of regulation by calcium signals are still poorly understood. Since many signal molecules and receptors are involved in multiple signaling pathways, it is difficult to study which temporal-spatial change of calcium regulates the specific pathway. Currently, an attractive method for resolving the roles of signaling network components is directly activation of these components in a spatially constrained and rapid manner, which is independent of upstream signaling and avoids the interference of alternative signal pathways. Calcium clamp is the most common method for controlling calcium oscillation in cells. This method suffers from several disadvantages, such as high reagent consumption, incapability of dealing with individual cells, and difficulty in controlling calcium oscillation. Another methods use optical activation to control calcium and key protein signals in cells. However, these methods only can create highly localized activation of signal transduction in focal point area. In contrast, in more physiological setting, a cell processes a shallow gradient of external environmental cue into a similar gradient of intracellular effectors. Thus, it is necessary to develop new methods to generate more natural, spatially distributed artificial signals in individual cells. In this project, we propose a novel microfluidic approach for generating artificial calcium oscillation and Rho-GTPase signaling in cells. By using interface shifting of laminar flows, the frequency, amplitude and distribution of calcium oscillation and the gradient of activated Rac could be controlled in cells. In combination with optical molecular imaging, we further investigate the role of artificially induced calcium and Rac signals in the regulation of cell polarity as well as the distribution of cAMP/cGMP, PIP3 and Rho-GTPase. This project exhibits important scientific value in revealing the mechanism of signal transduction, and provides a theoretical basis and an innovative technological platform for drug screening.

英文关键词： microfluidic chip;optical molecular imaging;cell migration;calcium oscillation;signal transduction