尿毒症血管钙化新机制:内皮微粒致血管平滑肌AEG-1表达激活STAT3/MMP2通路

We have reported the vascular calcification in uremia; the high level of endothelial microparticles (EMPs) in chronic kidney disease; observation of EMPs in the gaps between the activated or injured endothelial cells. However, the relationship of vascular smooth muscle cell (VSMC) calcification and EMPs and its potential mechanism in uremia is unclear. In this study, the activated process of endothelial cells in uremia state is vividly traced on releasing EMPs filled with heterogeneous expression of miRNAs of which targeting gene can be expected by bioinformatics. To identify the new target on VSMC calcification, we imitate uremia state in vivo or in vitro, dynamically monitoring the expression of AEG-1 and calcification in VSMCs stimulated by EMPs in uremia circumstances, achieving the benefits of interdisciplinary subjects. We detect the expression of STAT3/MMP2 signaling and calcification in the stimulated VSMCs with AEG-1 overexpression or silence to understand the molecular signaling pathway in uremia calcification. This study aims to elucidating the role of AEG-1 on calcification by activating STAT3/MMP2 signaling in VSMCs stimulated by EMPs and establishing new therapy approach for vascular calcification in uremia.

申请人报道过尿毒症患者有血管钙化、肾脏病患者有内皮微粒水平升高及观察到了内皮微粒穿越在损伤内皮细胞间隙，但是尿毒症中内皮微粒与平滑肌钙化关系及机制仍不清。根据我们的前期研究及预实验，我们提出科学假说：内皮细胞在尿毒症状态下释放微粒，微粒通过损伤的细胞间隙进入血管平滑肌细胞，调节AEG-1表达，促进血管平滑肌钙化。本项目以内皮微粒和平滑肌细胞为研究对象，对尿毒症状态下内皮细胞释放微粒进行高分辨示踪，用生物信息学方法阐明此状态下内皮微粒miRNAs异质性表达及其靶基因；体外及体内模拟尿毒症内环境，实时、动态监测内皮微粒刺激下平滑肌细胞钙化及AEG-1表达水平，明确AEG-1参与血管钙化新靶点；AEG-1过表达或沉默其表达后检测内皮微粒致平滑肌细胞钙化情况及STAT3/MMP2通路表达水平，从细胞和分子信号转导水平揭示血管钙化病因并为其治疗提供新靶点。

项目背景：慢性肾脏病中内皮微粒水平可以升高。高磷被认为是决定慢性肾脏病血管钙化的重要因素。因此我们想探究高磷状态下释放的内皮微粒在血管钙化中的作用及潜在的机制。.主要研究内容，重要结果及关键数据：我们观察到平滑肌细胞可以摄取高磷诱导的内皮微粒，根据时间的推移摄取的越来越多，到24小时基本吸收完全。我们的结果显示高磷诱导的内皮微粒可以促进血管钙化，或者加速高磷诱导的钙化通过STAT3/BMP2通路。AEG-1活性在高磷刺激的内皮微粒诱导的平滑肌细胞钙化中，在尿毒症大鼠的血管中，或者在尿毒症大鼠注射了高磷诱导的内皮微粒中都是升高的。AEG-1的缺失是抑制血管钙化的，AEG-1的高表达是促进血管钙化的。AEG-1，是miR-153的靶基因，可以被agomiR-153-3p抑制。尾静脉注射agomiR-153-3p适配体至尿毒症大鼠中可以减弱尿毒症血管钙化。.科学意义：我们的研究表面尿毒症高磷状态下的内皮微粒可以诱导或者加强尿毒症状态下的血管钙化，AEG-1可能作为高磷状态下内皮微粒刺激血管钙化的关键调控分子。这个研究结果可能可以为尿毒症血管钙化防治提供新的视角。