基于边界效应和驻点流的微流控单细胞捕获与动力学分析

The dynamic analysis of the deformability and mechanical properties as well as the intracellular calcium signaling of single cells play vital roles in cellular physiological and pathological studies, early diagnosis at cellular level, and drug screening of serious diseases. Existing microfluidic platforms for single-cell trapping and dynamic analysis are not able to be adopted for both the measurement of the deformability and mechanical properties, and the dynamic analysis of intracellular calcium signaling of single cells. In this project, a microfluidic array for single-cell trapping and dynamic analysis with high throughput and efficiency is proposed based upon the principle of boundary effect and stagnation point flow. In the proposed microfluidic array, the micropipette aspiration is implemented to investigate the deformability and mechanical properties of single cells under the stimuli of different aspiration pressure differences. An external system for generating, detecting and controlling dynamic biochemical signals is constructed to analyze the intracellular calcium dynamics in different types of single cells in response to the combined effects of aspiration pressures and/or dynamic biochemical signals. Furthermore, the quantitative relationship between the intracellular calcium dynamics and the cellular mechanical properties is also investigated. These studies will provide not only a novel experimental platform for analyzing both the mechanical properties and the intracellular signaling dynamics of single cells, but also some evidences and insights for the regulation of the cellular functions and behaviors by controlling intracellular signaling dynamics of single cells via modulating dynamic stimuli in their extracellular microenvironments.

单细胞变形能力、力学特性以及胞内钙信号的动力学分析对于重大疾病的细胞生理和病理研究、细胞水平的早期诊断、以及药物筛选等都具有十分重要的意义。已有的微流控单细胞捕获与动力学分析平台不能用于既测量单细胞变形能力和力学特性又分析胞内钙信号动力学。本项目结合边界效应和驻点流原理设计和构建能用于单细胞高通量、高效率捕获与动力学分析的微流控阵列；在设计的微流控阵列上实现微管吸吮技术，研究单细胞在不同压差刺激下的细胞变形能力和力学特性；构建用于加载、检测和控制动态生化信号的外围系统，分析不同类型吸吮压力或/和动态生化信号组合刺激下单细胞力学特性与胞内钙信号的动力学响应，进一步探寻钙信号动力学特征与细胞力学特性之间的定量关系。这些工作将不仅为单细胞力学特性与细胞内信号动力学分析提供新的实验平台，而且为进一步研究通过调控动态微环境刺激引发单细胞动力学响应进而影响细胞功能和行为等提供一定的线索和依据。

单细胞变形能力和力学特性、以及钙离子信号的动力学分析对于重大疾病发生发展的细胞生物学机制、细胞层面的早期诊断、以及药物筛选等方面都具有十分重要的意义。已有的微流控单细胞捕获与动力学分析平台不能同时实现单细胞力学特性测量和胞内钙信号动力学分析。本项目将微流控技术与驻点流原理、通道边界效应、惠斯通电桥原理和微管吮吸技术等相结合，优化设计并构建了三种不同的单细胞高通量捕获与动力学分析微流控阵列及外围加载、检测与分析系统；基于构建的微流控实验平台，定量分析了微管吮吸作用与细胞变形、细胞表观粘度、弹性模量等力学性质间的关系；利用数值仿真方法分析了不同微通道内生化信号的传输规律，并实验验证了精确加载浓度随时间动态变化或呈时空梯度分布的生化信号刺激的有效性和准确性；研究了不同种类细胞在吮吸压力、剪切力和动态生化信号单独或不同组合刺激下胞内钙离子信号的动力学规律，并建立相应数学模型进一步分析影响胞内钙离子信号动力系统特性的关键力学因素。这些工作不仅为开展机械力和/或生化刺激下单细胞力学特性与细胞内信号动力学分析提供了新的实验平台，而且为进一步研究通过调控动态微环境刺激引发单细胞动力学响应进而影响细胞功能和行为等提供一定的线索和依据。