代谢综合征血管内皮损伤炎性机制及柴芪汤调节作用机制

Metabolic syndrome (MS) is to blame for most of acute cardio-/cerebrovascular event. Many studies have found the close relationship among an inflammatory state, endothelial injury and the development of insulin resistance and atherosclerosis. As a key inflammatory mediator, NF-κB is an attractive target for suppressing inflammatory signaling pathway. In our clinical practice, Chaiqi decoction shows efficacy in treatment of MS. We later discovered in our laboratory study that regulation of the expression of the relevant inflammatory cytokines may be the therapeutic mechanism involved. The present study is aimed to investigate the role of inflammatory mechanism in MS and the therapeutic mechanism of Chaiqi decoction from the standpoint of pathology, cell and molecular biology, and genetics. We design to dynamically observe plasma levels of TNF-α, IL-6, IL-1β, MMP-9, VCAM-1, ICAM-1 and NF-κB, expression of VCAM-1, ICAM-1 and NF-κB of abdominal aorta in JCR: Lp-cp and oral-feeding rats before and after treatment of Chaiqi decoction, and expression of VCAM-1, ICAM-1 and NF-κB in vascular endothelial cells (VECs) under different environments. The involved assays include enzyme-linked immunosorbent assay (ELISA), Western Blot, real time-PCR (RT-PCR)，immunohistochemistry, and immunocytochemistry. Through the study, we hope to further understand the mechanism contributing to development of atherosclerosis in MS and validate the method of regulation of the Spleen and Stomach in treatment of MS.

代谢综合征（MS）是导致心脑血管疾病恶性事件发生的主要原因。炎性反应是胰岛素抵抗和动脉粥样硬化的共性机制，既往工作证实柴芪汤治疗MS有效，对相关炎性因子具有调节作用。本课题使用喂养型和JCR：Lp-cp大鼠MS模型、及人脐静脉内皮细胞损伤模型，以内皮损伤炎性反应体系-NF-κB多信号转导途径为切入点，采用ELISA、Western Blot、RT-PCR、免疫组织和细胞化学等方法，动态观察两种大鼠模型血浆中TNF-α、IL-6、IL-1β、MMP-9、VCAM-1、ICAM-1、NF-κB的表达，腹主动脉血管中VCAM-1、ICAM-1、NF-κB的表达，及不同环境下血管内皮细胞中VCAM-1、ICAM-1、NF-κB的表达，从整体、组织、细胞、蛋白、基因水平揭示MS的发生发展机制，证实柴芪汤的干预机制。为健脾疏肝调理脾胃法临床运用提供依据，进一步阐释MS向心脑血管疾病发生发展的内在机制。

本课题使用高盐高糖高脂饲料喂养SD大鼠建立MS模型，第8周末模型组大鼠血糖、血脂升高，出现胰岛素抵抗，评价造模成功。预防组和治疗组较模型组均可降低大鼠血清血糖、血脂、胰岛素、TNF-α、IL-6、ICAM-1、VCAM-1及腹主动脉NF-κB p65、ICAM-1、VCAM-1的表达，腹主动脉血管病理形态学表现优于模型组；但治疗效果不如早期干预效果理想，表现在预防组在大鼠血清血糖、血脂、IRI、TNF-α、IL-6、ICAM-1、VCAM-1及腹主动脉血管病理形态学表现等方面优于治疗组，这也证实课题组前期设想，在疾病发展的早期给予疏肝健脾的治疗干预，可改善代谢性疾病的演变。MS是一个慢性代谢性疾病，选择4、8、12、16周的动态取材可以直接动态观察疾病的发展和演变，更深入了解中药在不同时间节点的作用效果。对于慢性代谢性疾病而言，中医药的早期干预效果不容忽视。使用APOE-小鼠作为MS自发性模型，第12周末模型组小鼠血糖、血脂升高，出现胰岛素抵抗，评价造模成功。中药组较模型组可降低APOE-小鼠血清血糖、血脂、胰岛素、TNF-α、IL-6、ICAM-1、VCAM-1及腹主动脉NF-κB p65、ICAM-1、VCAM-1的表达，腹主动脉血管病理形态学表现优于模型组；说明具有健脾疏肝、调理脾胃作用的柴芪汤对后天喂养型和先天自发型MS模型均有改善作用，对MS炎性信号通路尤其是NF-κB多信号转导途径有调节作用。细胞实验中培养人脐静脉内皮细胞，经过TNF-α处理后ICAM-1和VCAM-1的表达上升，柴芪汤作用细胞后ICAM-1和VCAM-1的表达下调，并呈现出随着柴芪汤浓度的升高而抑制作用增强的趋势。提示柴芪汤在一定程度上对血管炎性过程有良性调节作用。本实验从整体、组织、细胞、蛋白、基因水平揭示MS的发生发展机制，证实了柴芪汤的作用机理。为健脾疏肝、调理脾胃法临床运用治疗MS提供实验依据，为把调节炎性信号通路作为防治MS及延缓MS向动脉粥样硬化发生发展的关键提供理论支撑。