微流控芯片技术研究血管内皮细胞肾素（前体）受体表达的调控机制及在动脉粥样硬化中的作用

The importance of renin-angiotensin-aldosterone system in cardiovascular diseases is generally accepted. It was demonstrated that angiotensin II wasn’t the only active substrate by both experimental researches and clinical observations. Thus it was of great concern when (pro)renin receptor [(P)RR] was discovered in 2002. It was demonstrated that overexpression of (P)RR could activate cellular signal pathways, and increase cell migration, proliferation, and cause proteinuria, glomerular sclerosis, cardiac fibrosis, and result in hypertension and vascular diastolic dysfunction, independent of Angiotensin II. However, the atherogenic mechanism of (P)RR in vivo has not got yet for the limitation of animal model. It is of great help when the microfluidic chips was invented and applied to the biological field, which made it possible to 3D cell culture and imitation of microvascular environment in vivo. Our study aims to observe changes of (P)RR expression in endothelial cells and its effects on the key factor of cellular signal pathway, G-coupled protein, NF-κB and epithelial apoptosis, intervened by hyperglycemia and different shear stresses through microfluidic chips. Meanwhile, let monocytes roll over the epithelial cells cultured tunnels on chips to observe the effects of (P)RR in the adhesion and migration of monocyte. Try to detect a new target for the prevention and treatment of atherosclerosis.

肾素-血管紧张素-醛固酮系统在心血管疾病中的重要作用已得到广泛认可。基础研究和临床实践均表明，血管紧张素Ⅱ(AngⅡ)不是其唯一的活性物质。新近发现的肾素(前体)受体[(P)RR]及其病理生理作用备受关注。国外及我们的研究已证实(P)RR过度表达能够不依赖于AngⅡ的产生而激活细胞信号通路，介导细胞增殖和炎症反应，导致蛋白尿、心肌纤维化，引起高血压和血管舒张功能障碍。但受动物模型限制，(P)RR致动脉粥样硬化(AS)机制的在体研究尚为空白。微流控芯片技术的发明与应用，使细胞三维立体培养、模拟在体血管微环境成为可能。本研究拟应用微流控芯片的独特优势，观察不同切应力及高糖干预下，内皮细胞(P)RR表达的变化及其对G偶联蛋白、NF-κB 和对内皮细胞凋亡的影响。同时将单核细胞与内皮细胞在同一基质中共培养，观察(P)RR在单核细胞粘附、迁移改变中的作用，为AS防治提供新的靶点。

慢性炎症高尿酸血症是诱导血管老化的重要因素。但目前为止很多炎症因子，包括尿酸的潜在致炎性病变导致的血管内皮改变的机制仍然不清楚。诸多慢性炎症因子中尿酸可能存在激活血管紧张素醛固酮系统，进而导致血管内皮细胞的一系列改变。我们课题组以往的研究发现内皮细胞表达肾素（前体）受体，肾素（前体）受体与配题结合后能够进一步激活血管紧张素醛固酮系统。因此，我们提出假设，某些其他的炎症因子，如尿酸是否能够激活内皮细胞表达的肾素（前体）受体，促进内皮细胞慢性炎症的发生发展，导致动脉硬化进展。为了验证这一假说，以尿酸干预原代体外培养的人脐静脉内皮细胞,分别用免疫印迹免疫荧光及实时定量PCR方法监测PRR的蛋白及mRNA表达情况，同时以微流室共培养内皮细胞及巨噬细胞，观察共培养的单核巨噬细胞的迁移及粘附状况。研究发现尿酸干预内皮细胞后内皮细胞膜表达PRR增高，尿酸刺激血管炎症反应。阻断尿酸的作用后，PRR表达下降，内皮细胞的慢性炎症反应水平降低。同时，微流控芯片技术也验证了尿酸能够通过激活PRR，导致内皮细胞慢性炎症反应，促进单核细胞巨噬化，迁移粘附增强，促进动脉粥样硬化的发生发展。因此，本研究发现动脉硬化的新靶点，为AS的防治提供新理论。

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