Microfluidic System for Applying Shear flow to Endothelial Cells on Culture Insert With Collagen Vitrigel Membrane

2021 ◽  
pp. 130675
Author(s):  
Yuya Morimoto ◽  
Shogo Nagata ◽  
Miki Matsumoto ◽  
Keisuke Sugahara ◽  
Shigenori Miura ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Shear Flow ◽  
Microfluidic System ◽  
Collagen Vitrigel

scholarly journals Estimation of nitric oxide generation by endothelial cells EA.hy926 under flow-induced mechanical stress in a microfluidic system

2020 ◽  
pp. 71-77
Author(s):  
А.А. Московцев ◽  
А.Н. Мыльникова ◽  
Д.В. Колесов ◽  
А.А. Микрюкова ◽  
Д.М. Зайченко ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Mechanical Stress ◽  
Microfluidic System ◽  
Hydrodynamic Characteristics ◽  
No Production ◽  
Hydrodynamic Conditions ◽  
Cellular Mechanisms ◽  
Vascular Walls

Эндотелиальные клетки, выстилающие стенки сосудов, преобразовывают деформацию собственных структур, вызванную током крови, в химические сигналы, одним из которых является важный регулятор просвета сосуда - оксид азота (NO). К настоящему моменту накоплен большой объём данных о клеточных механизмах активации продукции NO, однако сведений о динамике генерации оксида азота эндотелиальными клетками в зависимости от гидродинамических условий недостаточно. В этой связи разработка микрофлюидных систем in vitro, имитирующих кровеносное русло, и изучение в них эндотелия в сложных гидродинамических условиях является актуальной задачей. В данной работе для создания контролируемых гидродинамических условий для монослоя эндотелиоцитоподобных клеток EA.hy926 была спроектирована и разработана микрофлюидная система, имитирующая линейные участки микрососудистого русла. Методом непрямого определения содержания оксида азота (II) NO с использованием флуоресцентного зонда 4,5-диаминофлуоресцеина DAF-2 впервые получены данные об увеличении продукции NO клетками EA.hy926 при механическом стрессе, создаваемом потоком ростовой среды. Представлены расчетные гидродинамические характеристики микрофлюидной системы, а также методика измерения продукции NO. Возможность исследования функциональной активности эндотелия позволяет использовать разработанную микрофлюидную модельную систему как для изучения клеточно-автономных регуляторных свойств эндотелия при действии ряда вазоактивных фармакологических препаратов и других методов воздействия на эндотелий, так и при моделируемой дисфункции эндотелия. Endothelial cells lining vascular walls transform the flow-induced deformation of their own structures into chemical signals, one of which, nitric oxide (NO), is an important regulator of the vascular lumen diameter. By present, a large amount of data on cellular mechanisms for activation of NO production has been accumulated. However, there is insufficient information on changes in endothelial NO generation under different hydrodynamic conditions. Therefore, development of microfluidic systems that model blood vessels in vitro and using them to study the endothelium under complex hydrodynamic conditions are relevant tasks. In this study, a microfluidic system was developed to create controlled hydrodynamic conditions for a monolayer of endotheliocyte-like cells EAhy.926. This system simulates linear sections of the microvasculature. By indirect measurement of NO (II) content with a fluorescent 4,5-diaminofluorescein (DAF-2) probe, we showed an increase in the NO production by EAhy.926 cells under mechanical stress generated by the medium flow. The article presents the method for measuring NO production and the calculated hydrodynamic characteristics of the microfluidic system. The results showed that the developed microfluidic model system is promising for studying cell-autonomous regulatory properties of the endothelium both under the action of vasoactive agents and in simulated endothelial dysfunction.

scholarly journals Photoconversion and chromatographic microfluidic system reveals differential cellular phenotypes of adhesion velocity versus persistence in shear flow

Lab on a Chip ◽  
2020 ◽  
Vol 20 (4) ◽  
pp. 806-822 ◽  
Author(s):  
Katherine G. Birmingham ◽  
Isaac E. Robinson ◽  
Erin E. Edwards ◽  
Susan N. Thomas
Keyword(s):  
Shear Flow ◽  
Cancer Cell ◽  
Cell Sorting ◽  
Metastatic Cancer ◽  
Microfluidic System ◽  
Velocity Versus ◽  
Metastatic Cancer Cell ◽  
Cellular Phenotypes ◽  
Cell Dissemination

Engineered cell sorting, photoconversion platform to interrogate mechanisms of metastatic cancer cell dissemination.

scholarly journals Shear flow increases S-nitrosylation of proteins in endothelial cells

2009 ◽  
Vol 83 (3) ◽  
pp. 536-546 ◽  
Author(s):  
Bin Huang ◽  
Shih Chung Chen ◽  
Danny Ling Wang
Keyword(s):  
Endothelial Cells ◽  
Shear Flow

scholarly journals Integrin VLA-4 enhances sialyl-Lewisx/a-negative melanoma adhesion to and extravasation through the endothelium under low flow conditions

2008 ◽  
Vol 295 (3) ◽  
pp. C701-C707 ◽  
Author(s):  
Shile Liang ◽  
Cheng Dong
Keyword(s):  
Endothelial Cells ◽  
Shear Flow ◽  
Tumor Cells ◽  
Melanoma Cell ◽  
Melanoma Cells ◽  
Host Cells ◽  
High Shear ◽  
Sialyl Lewis ◽  
Adhesive Interactions ◽  
Low Shear

During their passage through the circulatory system, tumor cells undergo extensive interactions with various host cells including endothelial cells. The capacity of tumor cells to form metastasis is related to their ability to interact with and extravasate through endothelial cell layers, which involves multiple adhesive interactions between tumor cells and endothelium (EC). Thus it is essential to identify the adhesive receptors on the endothelial and melanoma surface that mediate those specific adhesive interactions. P-selectin and E-selectin have been reported as adhesion molecules that mediate the cell-cell interaction of endothelial cells and melanoma cells. However, not all melanoma cells express ligands for selectins. In this study, we elucidated the molecular constituents involved in the endothelial adhesion and extravasation of sialyl-Lewisx/a-negative melanoma cell lines under flow in the presence and absence of polymorphonuclear neutrophils (PMNs). Results show the interactions of α4β1(VLA-4) on sialyl-Lewisx/a-negative melanoma cells and vascular adhesion molecule (VCAM-1) on inflamed EC supported melanoma adhesion to and subsequent extravasation through the EC in low shear flow. These findings provide clear evidence for a direct role of the VLA-4/VCAM-1 pathway in melanoma cell adhesion to and extravasation through the vascular endothelium in a shear flow. PMNs facilitated melanoma cell extravasation under both low and high shear conditions via the involvement of distinct molecular mechanisms. In the low shear regime, β2-integrins were sufficient to enhance melanoma cell extravasation, whereas in the high shear regime, selectin ligands and β2-integrins on PMNs were necessary for facilitating the melanoma extravasation process.

scholarly journals Shear flow-driven actin re-organization induces ICAM-1 nanoclustering on endothelial cells that impact T-cell migration

2020 ◽  
Author(s):  
Izabela K. Piechocka ◽  
Sarah Keary ◽  
Alberto Sosa-Costa ◽  
Lukas Lau ◽  
Nitin Mohan ◽  
...  
Keyword(s):  
T Cells ◽  
Endothelial Cells ◽  
Cell Migration ◽  
T Cell ◽  
Shear Flow ◽  
Membrane Receptors ◽  
Mechanical Forces ◽  
Shear Forces ◽  
T Cell Migration ◽  
The Impact

ABSTRACTThe leukocyte specific β2-integrin LFA-1, and its ligand ICAM-1 expressed on endothelial cells (ECs), are involved in the arrest, adhesion and transendothelial migration of leukocytes. Although the role of mechanical forces on LFA-1 activation is well established, the impact of forces on its major ligand ICAM-1, has received less attention. Using a parallel-plate flow chamber combined with confocal and super-resolution microscopy, we show that prolonged shear-flow induces a global translocation of ICAM-1 on ECs upstream of flow direction. Interestingly, shear-forces promoted ICAM-1 nanoclustering prior to LFA-1 engagement. This spatial nanoscale organization was driven by actin cytoskeleton re-arrangements induced by shear-force. We further assessed the impact of prolonged shear-stress EC stimulation on T cell migration. T cells adhered to mechanically pre-stimulated ECs developed a more pro-migratory phenotype, migrated faster and exhibited shorter EC interactions than when adhered to non-mechanically stimulated ECs. Together, our results indicate that shear-forces increase the number of ICAM-1/LFA-1 bonds due to ICAM-1 nanoclustering, strengthening adhesion and thereby reducing actin retrograde flow of T-cells, leading to their increased migration speed. Our data also underscores the importance of mechanical forces regulating the spatial organization of cell membrane receptors and their contribution to adhesion regulation, regardless of integrin activation.Summary statementWe show that shear forces promote ICAM-1 spatial re-arrangement and actin-dependent nanoclustering on ECs prior to integrin engagement. This mechanism might be important for firm leukocyte adhesion and migration during the immune response.

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