Production of reactive oxygen species in endothelial cells under different pulsatile shear stresses and glucose concentrations
Production of reactive oxygen species in endothelial cells under different pulsatile shear stresses and glucose concentrations
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DOI：
10.1039/c0lc00651c
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发表时间：
2011-01-01
期刊：
影响因子：
6.1
通讯作者：
Luo, K. Q.
中科院分区：
文献类型：
--
作者：
Chin, L. K.;Yu, J. Q.;Luo, K. Q.
本研究开发了一种血流动力学芯片实验室系统。该系统有两个独特的特点：(1)它由一个微流体网络组成，其中包含一系列内皮细胞播种点，可以在多种条件下进行测试；(2)微通道中培养基的流速和频率由无脉动泵控制，以模拟不同生理条件下血管中的血液流动情况。研究的生理条件为：(1)正常剪切应力15达因cm（-2）、正常心率70 bpm的静息状态；(2)高剪切应力30达因cm（-2）、快速心率140 bpm的穷尽运动状态；(3)恒定高剪切应力30达因cm（-2）。研究了两种化学条件（10 mM和20 mM葡萄糖）来模拟糖尿病患者的高血糖状况。使用开发的血流动力学芯片实验室系统，通过评估两个参数，研究了不同剪切应力单独或与不同葡萄糖浓度联合对内皮细胞的影响。一种是通过荧光探针H(2)DCFDA测定细胞内活性氧（ROS）水平。另一个是用荧光染料MitoTracker Green FM显示的线粒体形态。结果表明，在剧烈运动60分钟后，ROS水平升高了近4倍。我们发现，流体的脉动性是导致细胞中ROS产生的决定因素，因为在恒定剪切应力条件下几乎没有检测到ROS的增加。同样，与静态条件下相比，在正常或高脉冲剪切应力下，将10 mM葡萄糖施加于细胞时，检测到的ROS水平要高得多。这些结果表明，有必要使用脉动剪切应力来表示血流的生理状况，并证明在未来的剪切应力相关研究中，使用这种新开发的血液动力学Labon-a-chip系统比传统的非脉动系统有优势。
A hemodynamic Lab-on-a-chip system was developed in this study. This system has two unique features: (1) it consists of a microfluidic network with an array of endothelial cell seeding sites for testing them under multiple conditions, and (2) the flow rate and the frequency of the culture medium in the microchannel are controlled by a pulsation free pump to mimic the flow profile of the blood in the blood vessel under different physiological conditions. The investigated physiological conditions were: (1) the resting condition in a normal shear stress of 15 dyne cm(-2) with a normal heart rate of 70 bpm, (2) an exhaustive exercise condition with a high shear stress of 30 dyne cm(-2) and a fast heart rate of 140 bpm, and (3) a constant high shear stress of 30 dyne cm(-2). Two chemical conditions were investigated (10 mM and 20 mM glucose) to mimic hyperglycemic conditions in diabetes patients. The effects of various shear stresses either alone or in combination with different glucose concentrations on endothelial cells were examined using the developed hemodynamic Lab-on-a-chip system by assessing two parameters. One is the intracellular level of reactive oxygen species (ROS) determined by a fluorescent probe, H(2)DCFDA. Another is the mitochondrial morphology revealed with a fluorescent dye, MitoTracker Green FM. The results showed that ROS level was elevated nearly 4-fold after 60 min of exhaustive exercise. We found that the pulsatile nature of the fluid was the determination factor for causing ROS generation in the cells as almost no increase of ROS was detected in the constant shear stress condition. Similarly, much higher level of ROS was detected when 10 mM glucose was applied to the cells under normal or high pulsatile shear stresses compared with under a static condition. These results suggest that it is necessary to use pulsatile shear stress to represent the physiological conditions of the blood flow, and demonstrate the advantage of utilizing this newly developed hemodynamic Labon-a-chip system over the conventional non-pulsatile system in the future shear stress related studies.