清心解瘀方调控NLRP3炎症小体抑制巨噬细胞焦亡稳定动脉粥样硬化易损斑块的机制研究

Pyroptosis is a newly discovered and established type of programmed cell death with proinflammatory property. Pyroptosis of macrophages plays an important role in the development, rupture and immune inflammatory response of atherosclerotic vulnerable plaque. NLRP3 inflammasome is an important regulator of pyroptosis. Qingxin Jieyu Formula is developed from Yugeng Tongyu Decoction of academician Keji Chen according to the ‘blood-stasis and toxin causing catastrophe’ theory. In our previous studies, Qingxin Jieyu Formula showed reliable anti-inflammatory effect in patients with coronary artery disease. However, the mechanism of its anti-inflammatory effect remains unclear. Therefore, we put forward the hypothesis that Qingxin Jieyu Formula can inhibit pyroptosis of macrophages and decrease release of inflammatory cytokines IL-1β and IL-18 via regulating the NLRP3 inflammasome, thereby stabilizing atherosclerotic vulnerable plaque. This project aims to explore mechanism of plaque stabilizing and inflammation inhibiting effect of Qingxin Jieyu Formula by observing its effects on vulnerable plaque in ApoE-/- mice, pyroptosis of macrophages and activation of NLRP3 inflammasome through both in-vivo and in-vitro study. Results of this project may provide traditional Chinese medicine (TCM) with a novel method in the treatment of atherosclerosis, present more scientific evidence for the ‘blood-stasis and toxin’ theory, and finally improve efficient of TCM in treating atherosclerosis.

细胞焦亡是新近发现并被证实的一种促炎症性程序性细胞死亡方式，巨噬细胞焦亡在动脉粥样硬化（AS）易损斑块的形成、破裂及免疫炎症反应中扮演着重要角色，NLRP3炎症小体是介导细胞焦亡的重要调节器。清心解瘀方是在“瘀毒致变”理论指导下，由陈可冀院士经验方愈梗通瘀汤精简化裁而来。课题组前期研究发现，清心解瘀方在冠心病治疗中具有良好的抗炎作用，但其机制尚不清楚。因此提出假说：清心解瘀方可能通过调控NLRP3炎症小体，抑制巨噬细胞焦亡的发生，减少炎症因子IL-1β、IL-18的释放，从而稳定AS易损斑块。本项目拟体内与体外研究相结合，探索清心解瘀方对ApoE-/-小鼠易损斑块的干预效应及其与巨噬细胞焦亡和NLRP3炎症小体的关系，从整体-细胞-分子水平探索其稳定AS易损斑块的作用及抗炎免疫机制，以期为AS的中医药干预提供新的思路，丰富“瘀毒”理论科学内涵，进一步提高AS性心血管病的中医防治效果。

动脉粥样硬化（AS）是发生于动脉血管壁的慢性炎症性疾病，细胞焦亡是一种新型促炎症性程序性细胞死亡方式。研究表明，巨噬细胞焦亡在AS易损斑块的形成、破裂及免疫炎症反应中扮演着重要角色。本研究在冠心病“瘀毒”病因病机理论指导下，以NLRP3炎症小体介导的巨噬细胞焦亡为切入点，围绕“清心解瘀方可通过抑制NLRP3炎症小体的活化、调控巨噬细胞焦亡的发生、减少炎症因子IL-1β和IL-18的释放，从而发挥抗炎、稳定AS易损斑块的作用”这一科学假说开展了三部分研究：通过动物实验揭示清心解瘀方调控巨噬细胞焦亡、稳定AS易损斑块的效应特点和作用规律；通过细胞实验阐明清心解瘀方抑制NLRP3炎症小体活化、调控巨噬细胞焦亡的效应特点和分子机制；通过网络药理学研究探索清心解瘀方抑制NLRP3炎症小体活化的潜在生物学机制。研究表明：⑴ 清心解瘀方可改善ApoE-/-小鼠脂质代谢和炎症反应，减少主动脉斑块内巨噬细胞浸润和脂质沉积，稳定AS易损斑块，其机制可能与抑制NLRP3炎症小体活化介导的巨噬细胞焦亡和炎症级联反应有关。⑵ 清心解瘀方可调控LPS联合ATP诱导的J774A.1巨噬细胞焦亡，其机制与抑制NLRP3炎症小体活化有关。⑶ 清心解瘀方可从生物网络调控水平抑制NLRP3炎症小体活化，具有多成分、多靶点、多途径整合调节的效应特点。项目从动物、组织、细胞、分子和生物网络等多个层面解析了活血解毒代表方药——清心解瘀方调控巨噬细胞焦亡、稳定AS易损斑块的作用机制，从细胞焦亡新视角揭示了其稳定AS易损斑块的效应机制，丰富了冠心病“瘀毒”病因病机理论的科学内涵，为活血解毒防治ASCVD的临床转化应用提供了新的科学依据。