TLR4/PKC/MAKPs信号途径介导抵抗素在川崎病冠状动脉损伤中的作用及机制研究

Kawasaki disease(KD) is the most common cause of vasculitis affecting children, and the leading cause of acquired heart disease in the world.The treatment of KD patients with intravenous immunoglobulin(IVIG) greatly reduces the incidence of coronary artery damage; however, the incidence of coronary artery lesions(CALs) continues to be 25% even in appropriately treated patients.Therefor further investigating the pathogenesis of CALs in KD patients will allow to choose therapeutic intervention for children with KD.Resistin has emerged as a new molecule to stand at the nodal point of signaling pathways to link metabolic disorders and in?ammation.Fortunately,Our previous study provided the ?rst evidence that incresed levels of serum resistin were associated with the development of coronary aneurysm in children with KD but the underlying molecular mechanisms are not yet known.Accordingly, in the present study, we tested the hypothesis that the recently identified marker of inflammation, resistin, might lead to vascular damage in the LCWE-induced coronary arteritis model of KD directly or indirectly;thus, suggesting a pivotal role for resistin in the pathophysiology of CALs.Furthermore, we sought to investigate whether the intracellular signals elicited by resistin are mediated through PKC and MAPKs signaling channel and initiated by resistin binding to the toll-like receptor 4(TLR4) in human coronary artery endothelial cell(HCAECs). Controlling the expression of resistin may represent a novel therapeutic strategy for treatment of KD.

川崎病(KD)的病理特征是全身弥漫性血管炎症，已成为引起小儿后天性心脏病最常见原因之一。经静脉注射免疫球蛋白（IVIG）治疗后，仍有约25%的患者发生冠状动脉损伤（CALs）。因此对其CALs发生机制的深入探讨将有重要理论价值和临床意义。抵抗素是新近发现的炎性因子，其分子位于联结代谢性疾病和炎性疾病信号途径的结点上。我们前期实验首次发现KD患儿血清抵抗素水平的升高与其冠脉瘤的形成有关，但其分子机制尚不明确。故本课题将构建LCWE诱发的KD动物冠脉炎模型，阐明抵抗素是否直接地或间接地造成了其血管损伤，进而表明其在冠脉损伤发生发展中的重要作用；进一步我们将通过人冠脉内皮细胞(HCAECs)的培养与干预,验证重组抵抗素是否通过结合细胞膜toll样受体4(TLR4),从而激活了细胞内信号途径PCK/MAPKs来参与血管损伤过程。控制抵抗素的表达将有望成为伴冠脉损伤的KD患儿新的治疗策略。

目前的结果验证了我们目前的猜测：①与健康对照组比较，血清抵抗素水平在KD患儿中明显升高。与KD合并冠状动脉扩张的患儿比较，血清抵抗素在KD伴有冠状动脉瘤形成的患儿中明显升高。也检测川崎病急性期患儿血清中与抵抗素相关细胞因子TNF-α、IL-1β表达水平。川崎病伴有冠状动脉损伤的患儿血清中TNF-α表达水平和IL-1β表达水平显著高于川崎病无冠状动脉损伤的患儿及健康对照组。②抵抗素可见在受累冠状动脉血管壁分布。注射LCWE后小鼠的冠状动脉周围免疫细胞炎症浸润过程与抵抗素产生的过程一致。抵抗素敲除可降低小鼠炎症细胞因子表达水平及冠状动脉炎发生率。③抵抗素可明显刺激HEACEs的MMP-9mRNA转录和蛋白的分泌，抵抗素能够下调TIMP-1mRNA转录和蛋白的表达，提示抵抗素对冠脉瘤的作用可能部分通过对TIMP-1的抑制作用实现。抵抗素至少部分通过使ERK1/2, P38 MAPK 和JNK信号蛋白磷酸化,从而导致了HEACEs合成MMP-9蛋白。④抵抗素诱导的IL-1β表达依赖于与TLR4受体相关的信号通路，然而抵抗素诱导的TNF-α表达不依赖于与TLR4受体相关的信号通路。抑制经典及非经典NF-κB信号途径，均完全抑制了抵抗素诱导的TNF-α表达，然而只有抑制了经典NF-κB信号途径，才抑制了抵抗素刺激的IL-1β的表达。抵抗素通过激活TLR4/经典NF-κB信号通路刺激IL-1β的表达，而抵抗素通过协同抵抗素/CAP-1相关的信号通路与NF-κB信号通路诱导TNF-α表达，但不依赖于TLR4受体。抵抗素可能为冠状动脉炎严重程度评估的一个有价值的生物标志物，抵抗素可能成为川崎病致冠状动脉损伤治疗的新靶点。