Endothelial permeability has been extensively investigated in the context of pathologies such as cancer and also in studies of drug delivery from the circulation. Hypoxia is a critical regulator of endothelial cell (EC) behavior and affects the barrier function of endothelial linings, yet its role has been little studied. This paper reveals the effect of hypoxia on the permeability of an EC monolayer by cellular experiments using a microfluidic device and a conventional cell culture dish. Human umbilical vein endothelial cells (HUVECs) were seeded into one microfluidic channel, creating an EC monolayer on each vertical surface of a collagen gel confined to a central chamber. Oxygen tension was regulated to produce normoxic (21% O2) or hypoxic (3% O2) conditions by the supply of gas mixtures of oxygen, carbon dioxide, and nitrogen at predefined ratios into channels fabricated into the device. Permeability of the EC monolayer quantified by analyzing diffusion of fluorescence-labelled dextrans into the collagen gel increases with barrier function loss by 6 hour hypoxic exposure, showing 11-fold and 4-fold increases for 70 kDa and 10 kDa dextrans, respectively, on average. Consistent with this, subsequent immunofluorescent staining and separate western blot analysis of HUVECs on a culture dish demonstrate loose cell-cell adhesion resulting from internalization of VE-cadherin under hypoxia. Thus, hypoxic stress increases endothelial permeability by altering cell-cell junction integrity.

中文翻译：

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在受控氧张力下内皮单层渗透性。

在诸如癌症的病理情况下以及在从循环中递送药物的研究中，已经广泛研究了内皮渗透性。缺氧是内皮细胞（EC）行为的关键调节器，并影响内皮细胞壁的屏障功能，但其作用尚未得到研究。本文通过使用微流体装置和常规细胞培养皿的细胞实验揭示了缺氧对EC单层渗透性的影响。将人脐静脉内皮细胞（HUVEC）播种到一个微流体通道中，在局限于中央室的胶原蛋白凝胶的每个垂直表面上形成EC单层。通过供应氧气，二氧化碳的混合气体，调节氧气张力以产生常氧（21％O2）或低氧（3％O2）条件。和氮气以预定的比例进入制造到设备中的通道。通过分析荧光标记的葡聚糖在胶原蛋白凝胶中的扩散来量化EC单层的通透性，随着低氧暴露6小时，屏障功能丧失，屏障功能丧失，表明70 kDa和10 kDa葡聚糖的平均分别增加11倍和4倍。与此相一致的是，随后在培养皿上对HUVEC进行的免疫荧光染色和单独的Western印迹分析表明，在缺氧条件下VE-钙黏着蛋白的内在化导致了松散的细胞粘附。因此，低氧应激通过改变细胞-细胞连接完整性来增加内皮通透性。通过分析荧光标记的葡聚糖在胶原蛋白凝胶中的扩散来量化EC单层的通透性，随着低氧暴露6小时，屏障功能丧失，屏障功能丧失，表明70 kDa和10 kDa葡聚糖的平均分别增加11倍和4倍。与此相一致的是，随后在培养皿上对HUVEC进行的免疫荧光染色和单独的Western印迹分析表明，在缺氧条件下VE-钙黏着蛋白的内在化导致了松散的细胞粘附。因此，低氧应激通过改变细胞-细胞连接完整性来增加内皮通透性。通过分析荧光标记的葡聚糖在胶原蛋白凝胶中的扩散来量化EC单层的通透性，随着低氧暴露6小时，屏障功能丧失，屏障功能丧失，表明70 kDa和10 kDa葡聚糖的平均分别增加11倍和4倍。与此相一致的是，随后在培养皿上对HUVEC进行的免疫荧光染色和单独的Western印迹分析表明，在缺氧条件下VE-钙黏着蛋白的内在化导致了松散的细胞粘附。因此，低氧应激通过改变细胞-细胞连接完整性来增加内皮通透性。随后在培养皿上对HUVEC进行的免疫荧光染色和单独的Western印迹分析表明，在缺氧条件下VE-钙黏着蛋白的内在化导致了松散的细胞粘附。因此，低氧应激通过改变细胞-细胞连接完整性来增加内皮通透性。随后在培养皿上对HUVEC进行的免疫荧光染色和单独的Western印迹分析表明，在缺氧条件下VE-钙黏着蛋白的内在化导致了松散的细胞粘附。因此，低氧应激通过改变细胞-细胞连接完整性来增加内皮通透性。