白细胞微粒促进AS中巨噬细胞MI极化的机制及益气活血解毒方的干预研究

Microparticles are membrane-like vesicles shedding of activated or apoptotic cells which act as a biological information carrier to promote a variety of pathological processes. Recent studies have shown a significantly increased expression of leukocyte microparticles (LMPs) in the circulation and plaque of AS patients which promote atherosclerosis progression. M1/ M2 macrophage polarization has recently been considered as "key power" of AS development, but the molecular mechanism of LMPs, “switch” of multiple pathological pathways in AS, promoting M1 / M2 macrophage polarization is unknown. Based on pre-study, Yiqihuoxuejiedu Formula can significantly inhibit macrophage infiltration, reduce IL-6 and vascular remodeling. We propose hypothesis that LMPs induce polarization of macrophages M1, unbalance M1 / M2 macrophages and lead to progresses of AS, and mechanism of polarization mainly involve CD40/CD40L-mediated signaling pathway. In this study, we examined the phenotypic characteristics of circulating LMPs and the ratio of M1 / M2 macrophages in patients with carotid plaques. The effect of LMPs on macrophage polarization and its molecular mechanism were investigated by cell experiment. The molecular mechanism of LMPs promoting macrophages polarization and the therapeutic target of Yiqihuoxuejiedu Formula were elucidated by animal experiments. This study provided a new mechanism and therapeutic method for prevention and treatment of AS. This is an original innovative research.

微粒是细胞活化或凋亡时脱落的膜样囊泡，其作为生物信息载体促进多种病理进程。近期发现，AS患者循环及斑块中，白细胞微粒（LMPs）表达显著增高并促进AS的发展；M1/M2型巨噬细胞极化近来被认为是AS发展的“关键动力”，而LMPs作为AS多个病理环节的启动开关，其是否能促进M1/M2型巨噬细胞极化及分子机制仍是研究的盲点。基于前期基础，益气活血解毒方明显抑制巨噬细胞的浸润、下调IL-6，改善血管重构。我们提出假说：LMPs诱导巨噬细胞M1极化促进AS发展，其机制是CD40/CD40L介导的信号通路。本研究检测临床颈动脉斑块患者循环LMPs的表型特点及巨噬细胞M1/M2的比例；细胞实验探讨LMPs对巨噬细胞极化的影响及其分子机制；动物模型验证该机制，以阐明LMPs诱导巨噬细胞M1极化的分子机制及益气活血解毒方对该机制作用的靶点，为AS防治提供全新的思路和治疗手段。本研究为原始创新性研究。

在白细胞微粒（LMPs）对早期动脉粥样硬化（AS）具有早期诊断价值的基础上，本研究进一步阐明其机制。通过临床试验，观察颈动脉粥样硬化（CAS）患者LMPs、巨噬细胞M1及M2亚型，与颈动脉斑块的相关性；通过细胞实验观察LMPs对巨噬细胞极化的机制；通过AOPE-/-小鼠实验观察LMPs对AS斑块的影响，以揭示LMPs促进巨噬细胞极化加速AS进展的新机制以及益气活血解毒方的药理作用。研究发现：.1.与正常人相比， CAS患者LMPs显著升高，且与CAS病变具有一定正相关性， M1型单核巨噬细胞水平以及M1/M2的比值也明显增加，且与颈动脉内中膜厚度成正相关性，说明早期CAS患者外周血中促炎型和抗炎型单核巨噬细胞存在失衡现象，并且这种失衡很有可能与AS患者斑块形成有关。.2. 细胞和动物实验发现：白细胞微粒通过与巨噬细胞结合，激活CD40-CD40L- NF-κB 通路，促进巨噬细胞向M1极化，导致M1/M2的失衡，促进斑块的不稳定性，促进AS发展。该研究阐明了白细胞微粒促进AS发展的新机制，揭示了白细胞微粒对早期AS具有早期诊断价值的机理，具有重要的科学价值和应用价值。白细胞微粒有望成为AS早期诊断的标志物。.3．益气活血解毒方能明显减少AS斑块内脂质成分，增加胶原成分，稳定LMPs导致的易损性斑块。同时能够降低AS斑块中M1型巨噬细胞含量和NF-κB表达量，增加M2型巨噬细胞含量，改善斑块内炎症环境。该研究结果对于AS的预防和治疗具有重要的实用价值。.本研究按照计划顺利完成，已完成相关内容及目标。主要成果为已发表见刊的中文文章2 篇和4 篇SCI 文章，另有1篇高质量的SCI文章在投。培养硕士研究生2名。相关学术交流4次。支出经费58.05万元，剩余经费7.95万元。

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