Synthetic 3D Model of the Carotid Artery to Study Exercise-Induced Changes in Endothelial Gene Expression

总结 大约十分之九的脑血管病发作是由于动脉粥样硬化。此外，内皮 功能障碍目前被认为是动脉粥样硬化的第一个病理生理步骤。内皮细胞 体内平衡和基因表达通过剪切应力高度调节，剪切应力与血液直接相关。 流量变化。有氧运动（AX）与改善心血管（CV）健康有关。然而，在这方面， 只有~50%的AX的有益效果是通过对传统CV风险因素的改善来解释的（例如， 高血压、高胆固醇血症、肥胖症）。AX的其余~50%的有益效果是 未知此外，传统控制的AX不提供个性化的药物，这可能导致 对于大量的无反应者。因此，本提案的主要目的是开发3D 使用3D生物打印技术模拟活体的人颈动脉合成模型 个性化AX诱导的血流模式和内皮剪切应力，并确定基因 表达/转录和分子变化。基于先前 报告和我们的初步数据，我们假设颈动脉的3D合成模型将响应于 运动诱导的血流模式与正常颈动脉相同。此外，我们假设内皮细胞 在相似的血流模式和剪切应力下培养的细胞将增加 动脉粥样硬化保护性mRNA/蛋白（例如，eNOS、PGI 2和SOD）和结构mRNA/蛋白质（例如， 肌动蛋白、硫酸肝素蛋白聚糖[糖萼]和α-肌动蛋白束应力纤维），以及前 动脉粥样硬化和促炎mRNA/蛋白质表达（例如，ICAM-1、VCAM-1和ET-1）在类似的 强度依赖的方式。首先，我们将确定生物力学特性（例如，血管扩张性和 顺应性）的颈动脉在体内在静息条件下和在3 AX强度在健康，年轻 受试者、中风患者和年龄匹配的对照。然后，受试者将接受磁共振检查 成像（MRI）研究以确定确切的形状（例如，长度和轮廓）， 这些图像将用于通过3D生物打印来构建3D合成模型。3D合成模型将模拟 更多的解剖学和血液动力学条件，这将允许更多的生理体外实验。 其次，我们将在3D合成模型上接种的内皮培养细胞中执行几种流动模式。 流动模式将与体内研究期间观察到的模式相似。在应用不同的流量后， 模式，细胞将被收集和处理，以确定特定基因转录因子的变化， 蛋白质表达通过表征不同强度AX期间的血流模式并确定 在这些相同的血流条件下，内皮细胞的基因表达/转录和分子变化 模式，使用“反向”翻译方法，我们将解释心血管疾病的机制。 与AX作为个体化药物相关的保护因素，尤其是在卒中幸存者中。

Summary Approximately nine out of 10 cerebrovascular attacks are due to atherosclerosis. Additionally, endothelial dysfunction is currently accepted as the first pathophysiological step toward atherosclerosis. Endothelial cell homeostasis and gene expression is highly regulated via shear stress, which is directly associated with blood flow changes. Aerobic exercise (AX) has been associated with improved cardiovascular (CV) health. However, only ~50% of the beneficial effects of AX are explained via improvements on traditional CV risk factors (e.g., hypertension, hypercholesterolemia, obesity). The remainder ~50% of the beneficial effects of AX are unknown. Moreover, traditionally controlled AX does not provide personalized medicine, which could account for a high number of non-responders. Therefore, the main purpose of this proposal is to develop a 3D synthetic model of the human carotid artery using 3D bio-printing technology to simulate in vivo personalized AX-induced blood flow patterns and endothelial shear stress and to determine gene expression/transcription and molecular changes in endothelial cultured cells in vitro. Based on previous reports and our preliminary data we hypothesize that a 3D synthetic model of the carotid artery will respond to exercise-induced blood flow patterns as a normal carotid artery. In addition, we hypothesize that endothelial cultured cells under similar blood flow patterns and shear stress will increase the expression of atherosclerosis-protective mRNA/proteins (e.g., eNOS, PGI2, and SOD) and structural mRNA/proteins (e.g., actin, heparin sulfate proteoglycan [glycocalyx], and α-actinin-bundled stress fibers), and a decrease of pro- atherosclerosis and pro-inflammatory mRNA/protein expression (e.g., ICAM-1, VCAM-1, and ET-1) in a similar intensity-dependent manner. First, we will determine biomechanical properties (e.g., vessel distensibility and compliance) of the carotid artery in vivo during resting conditions and at 3 AX intensities in healthy, young subjects, patients with stroke, and age-matched controls. Then, subjects will undergo a magnetic resonance imaging (MRI) study to determine the exact shape (e.g., length and contour) of same tested carotid artery and the images will be used to build a 3D synthetic model via 3D bio-printing. The 3D synthetic model will mimic more anatomical and hemodynamic conditions, which will allow for more physiological in vitro experiments. Secondly, we will perform several flow patterns in endothelial cultured cells seeded on the 3D synthetic model. Flow patterns will be similar to those patterns observed during in vivo studies. After applying the different flow patterns, cells will be collected and processed to determine changes in specific gene transcription factors and protein expression. By characterizing blood flow patterns during different intensities of AX and determining the gene expression/transcription and molecular changes in endothelial cells under these same blood flow patterns, using a ‘reverse’ translation approach, we will explain the mechanisms of the cardiovascular protective factors associated with AX as personalized medicine, especially in stroke survivors.