NLRP3炎症小体促进平滑肌源性泡沫细胞形成参与AS斑块不稳定性的机制研究

Inflammation plays an important role in all phases of atherosclerosis from initiation to its clinical manifestations such as plaque rupture. Activation of NLRP3 inflammasome, which assembles in response to a variety of exogenous and endogenous activators as an initial priming step, and thereby initiates IL-1β processing, is critical to the development of AS plaque instability, but the underlying mechanism is not well known. The formation of lipid laden foam cells, derived predominately from macrophages and vascular smooth muscle cells (VSMC), is closely associated with the development of atherosclerosis and plaque instability.Recent study indicated that at least 50% of the foam cells contributing to the progression of lesions and likely necrosis are smooth muscle cell derived. Therefore, the VSMCs contain a much larger burden of the excess cholesterol in comparison with macrophages in atherosclerosis, as the main origin of foam cells had a more significant effect on AS plaque instability. Study from our laboratory suggested that NLRP3 inflammasome mediated the formation of macrophages-derived foam cells induced by ox-LDL, contributed to the macrophages migration and lipid uptake ability. But the effect of NLRP3 inflammasome on the formation of VSMC-derived foam cells is rarely reported. In this proposed study, we plan to construct transgenic mice with NLRP3 overexpressed in VSMC to test whether NLRP3 inflammasome contributes to VSMC-derived foam cells formation, further promotes the AS plaque instability. In vitro, we will also explore the molecular mechanism of ABCA1 expressions regulated by NLRP3 using cultured VSMC with NLRP3 overexpressed. Eventually, to clarify the mechanistic role of NLRP3 on the formation of VSMC-derived foam cells and atherosclerosis plaque instability, so as to provide insight into the mechanism that links NLRP3 inflammasome to atherosclerosis and plaque instability. The present study will provide evidence for promoting the development of NLRP3inflammasome into a viable therapeutic target in atherosclerosis.

动脉粥样硬化（AS）斑块不稳定性是影响心血管事件发生的病理基础，NLRP3炎症小体活化增加斑块不稳定性，但机制尚待研究。泡沫细胞是AS斑块不稳定性的核心，来源有巨噬细胞和血管平滑肌细胞（VSMC），新的研究提示AS斑块中的泡沫细胞>50%来源于VSMC，改变了传统认识。我们前期发现NLRP3能够促进巨噬细胞源性泡沫细胞形成，但对VSMC源性泡沫细胞形成的影响未见报道。本项目通过构建VSMC过表达NLRP3小鼠模型，从整体动物水平观察NLRP3对VSMC源性泡沫细胞形成及AS斑块不稳定性的影响；并从细胞水平探讨其分子调控机制。验证“NLRP3活化通过调控VSMC的ABCA1表达及功能，影响VSMC胆固醇外流，促进VSMC源性泡沫细胞形成进而增加AS斑块不稳定性”这一科学假说，旨在完善NLRP3促进AS斑块不稳定性的机制，发现潜在干预靶点，为推进NLRP3成为AS炎症靶向药物的开发提供依据。

动脉粥样硬化（AS）斑块不稳定性是影响心血管事件发生的病理基础，炎症机制贯穿动脉粥样硬化的进展，最新的研究提示干预慢性炎症可以在常规标准治疗的基础上进一步降低心血管事件风险。既往研究表明NLRP3炎症小体活化可以促进巨噬细胞来源的泡沫细胞形成，进一步参与动脉粥样硬化的发生及发展，增加斑块不稳定性。泡沫细胞是AS斑块不稳定性的核心，来源有巨噬细胞和血管平滑肌细胞（VSMC），新的研究提示AS斑块中的泡沫细胞>50%来源于VSMC，改变了传统认识。本项目通过构建VSMC过表达NLRP3转基因小鼠模型，发现VSMC过表达NLRP3转基因小鼠与普通对照组小鼠相比，同样予以高脂饮食，其动脉粥样硬化程度明显加重，动脉粥样硬化斑块不稳定性增加，项目组进一步应用NLRP3特异性抑制剂MCC950对动脉粥样硬化造模成功小鼠进行腹腔注射，发现NLRP3特异性抑制剂MCC950可以一定程度上逆转动脉粥样硬化斑块进展，提示NLRP3对VSMC源性泡沫细胞形成及AS斑块不稳定性存在一定影响；细胞水平上，我们通过慢病毒过表达及siRNA干扰等方式，观察过表达或降低NLRP3表达后，对ox-LDL刺激下的平滑肌细胞增殖、泡沫化等存在明显影响，从细胞水平上验证了“NLRP3炎症小体促进VSMC源性泡沫细胞形成 进而增加AS斑块不稳定性”这一科学假说。上述实验结果旨在完善NLRP3促进AS斑块不稳定性的机制，发现 潜在干预靶点，有望推进NLRP3炎症小体成为AS炎症靶向药物的开发。