Microvessels-on-a-chip have enabled in vitro studies to closely simulate in vivo microvessel environment. However, assessing microvessel permeability, a functional measure of microvascular exchange, has not been attainable in nonpermeable microfluidic platforms. This study developed a new approach that enables permeability coefficients (Ps) to be quantified in microvessels developed in nonpermeable chip platforms by integrating avidin/biotin technology. Microvessels were developed on biotinylated fibronectin-coated microfluidic channels. Solute transport was assessed by perfusing microvessels with fluorescence-labeled avidin. Avidin molecules that crossed endothelium were captured by substrate biotin and recorded with real-time confocal images. The Ps was derived from the rate of avidin/biotin accumulation at the substrate relative to solute concentration difference across microvessel wall. Avidin tracers with different physiochemical properties were used to characterize the barrier properties of the microvessel wall. The measured baseline Ps and inflammatory mediator-induced increases in Ps and EC [Ca²⁺]_i resembled those observed in intact microvessels. Importantly, the spatial accumulation of avidin/biotin at substrate defines the transport pathways. Glycocalyx layer is well-formed on endothelium and its degradation increased transcellular transport without affecting EC junctions. This study demonstrated that in vitro microvessels developed in this simply designed microfluidics structurally possess in vivo-like glycocalyx layer and EC junctions and functionally recapitulate basal barrier properties and stimuli-induced responses observed in intact microvessels. This new approach overcomes the limitations of nonpermeable microfluidics and provides an easily executed highly reproducible in vitro microvessel model with in vivo microvessel functionality, suitable for a wide range of applications in blood and vascular research and drug development.

中文翻译：

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利用亲和素/生物素相互作用量化微血管芯片的渗透性

芯片上的微血管使体外研究能够密切模拟体内微血管环境。然而，评估微血管渗透性，微血管交换的功能测量，在不可渗透的微流体平台中尚未实现。本研究开发了一种新方法，通过集成抗生物素蛋白/生物素技术，可以量化在非渗透性芯片平台上开发的微血管中的渗透系数 (Ps)。微血管是在生物素化的纤连蛋白涂层微流体通道上开发的。通过用荧光标记的抗生物素蛋白灌注微血管来评估溶质转运。穿过内皮细胞的抗生物素蛋白分子被底物生物素捕获并用实时共聚焦图像记录。Ps 源自亲和素/生物素在底物上的积累速率，相对于微血管壁上的溶质浓度差异。具有不同理化性质的亲和素示踪剂用于表征微血管壁的屏障性质。测量的基线 Ps 和炎症介质引起的 Ps 和 EC 增加 [Ca²⁺ ]_我类似于在完整微血管中观察到的那些。重要的是，亲和素/生物素在底物上的空间积累决定了运输途径。糖萼层在内皮上形成良好，其降解增加了跨细胞转运而不影响 EC 连接。这项研究表明，在这种简单设计的微流体中开发的体外微血管在结构上具有体内样糖萼层和 EC 连接，并在功能上概括了在完整微血管中观察到的基础屏障特性和刺激诱导反应。这种新方法克服了非渗透性微流体的局限性，提供了一种易于执行、高度可重复的体外微血管模型，具有体内微血管功能，适用于血液和血管研究以及药物开发中的广泛应用。