Quantitative microfluidic delivery systems for studying tissue microenvironment and microvascular blood flow regulation in vivo

背景：所有身体组织的营养需求是由血流到全身微血管网络的动态分布提供的。研究体内的血流调节需要专门的、微创的手术准备，以便分离感兴趣的组织，同时保持调节系统本身的正常功能和控制。我们之前已经开发了气体交换室来操纵活鼠骨骼肌中的气体分压。这项工作证明了这些装置的可行性和有效性，并且微循环可以响应周围微环境的变化。我们建议开发新的微流体装置，允许将染料和药理学剂引入骨骼肌，以研究血流调节和物质运输。目的：1)开发一种微流体装置和流体流动系统，能够在体内完整组织表面保持固定的溶质浓度、温度和生理气体条件，同时允许组织内血流的可视化和量化。2)使用染料、荧光探针和血管活性药物验证设备将化合物输送到覆盖组织的功效。3)建立微流控装置和上覆组织的数学质量传递模型，在给定微流控流室内已知条件、组织的物理性质和微循环中的血流的情况下，计算组织内的组织浓度和溶质梯度。4)应用微流控系统和数学模型，利用各种血管活性激动剂、拮抗剂和药物，研究完整骨骼肌的微血管调节机制。科学方法：微流体装置将使用精密激光切割玻璃和软光刻技术制成的模塑塑料来构建。设备内的流量将通过连接到计算机系统的标准微流量阀进行控制。可视化和测量微血管血流将实现使用大鼠骨骼肌的高倍显微镜。计算机模型将用于告知实验结果，并计算通过微流体通道输送的物质将渗透到组织中的深度。影响：与该计划相关的项目将为对心血管系统基本功能感兴趣的年轻科学家和学生提供良好的培训机会。本研究将为研究微血管在体内的功能提供一种新的技术，并为研究活体组织的微环境提供一个平台。这些创新将影响生物医学研究的许多领域，特别是与理解综合微循环有关的领域。

BACKGROUND: The nutritive requirements of all body tissues are provided for by the dynamic distribution of blood flow to microvascular networks throughout the body. Studying blood flow regulation in vivo requires specialized, minimally invasive, surgical preparations in order to isolate a tissue of interest while preserving the normal function and control of the regulatory system itself. We have previously developed gas exchange chambers to manipulate gas partial pressures in skeletal muscle of live rats. This work demonstrated the feasibility and efficacy of these devices and that the microcirculation can respond to changes in the surrounding microenvironment. We propose the development of novel microfluidic devices that allow for the introduction of dyes and pharmacological agents to skeletal muscle for the purposes of studying blood flow regulation and mass transport. OBJECTIVES: 1) Develop a microfluidic device and fluid flow system capable of maintaining a fixed solute concentration, temperature, and physiologic gas conditions, at the surface of an intact tissue in vivo while simultaneously allowing visualization and quantification of blood flow within the tissue. 2) Validate the efficacy of the device to deliver compounds into the overlying tissue using dyes, fluorescent probes, and vasoactive drugs. 3) Create mathematical mass transport models of the microfluidic device and overlying tissue to calculate the tissue concentration and gradients of solutes within the tissue given known conditions within the microfluidic flow chamber, physical properties of the tissue, and blood flow in the microcirculation. 4) Apply the microfluidic system and mathematical model to interrogate microvascular regulatory mechanisms in intact skeletal muscle using various vasoactive agonist, antagonists, and drugs.SCIENTIFIC APPROACH: Microfluidic devices will be constructed using precision laser cut glass and molded plastics made with soft lithography techniques. Flow within the device will be controlled using standard micro-flow valves connected to a computer system. Visualization and measurement of microvascular blood flow will be achieved using high-powered microscopy of rat skeletal muscle. Computer models will be used to inform the experimental results and calculate the depth to which substances delivered via the microfluidic channel will penetrate into the tissue.IMPACT: The projects related to this program will provide excellent training opportunities for young scientists and students interested in the fundamental function of our cardiovascular systems. This research will produce a novel technology for studying microvascular function in vivo as well as a platform for interrogating the microenvironment of living tissue. These innovations will influence many areas in biomedical research, particularly those related to understanding integrated microcirculation.