和厚朴酚通过Wnt/β-连环蛋白通路抗动脉粥样硬化  

The effect of honokiol on the forming process of atherosclerosis by regulating Wnt/β-catenin signaling pathway

作　　者：詹萍[1] 熊尚全[1] 柴大军[2] 蔡宏达[3] ZHAN Ping;XIONG Shang-quan;CHAI Da-jun;CAI Hong-da(Department of Cadiology,the First Affiliated People’s Hospital,Fujian Traditional Medical University,Fuzhou,Fujian 350004,China;Department of Cardiology,The First Affiliated Hospital of Fujian Medical University;Department of Anesthesiology,The First Affiliated Hospital of Fujian Medical University)

机构地区：[1]福建中医药大学附属人民医院心血管内科,福建福州350004 [2]福建医科大学附属第一医院心血管内科 [3]福建医科大学附属第一医院麻醉科

出　　处：《中华高血压杂志》2022年第2期153-160,共8页Chinese Journal of Hypertension

基　　金：福建省自然科学基金项目(2018J01328);福建省卫健委医学创新课题(2019-CX-25)。

摘　　要：目的体内外观察和厚朴酚(HNK)对动脉粥样硬化(AS)模型Wnt/β-连环蛋白(β-catenin)信号通路及动脉硬化的影响;探讨和厚朴酚调控Wnt/β-catenin信号通路对AS形成过程影响的可能机制。方法动物实验:8只SD大鼠作为正常对照组,24只高脂喂养SD大鼠随机分为AS组(高脂喂养+维生素D_(2)注射建立AS模型),AS+HNK组[建立模型后HNK腹腔注射5 mg/(kg·d),2次/d,连续8周]和AS+HNK-PNU组[AS+HNK组的基础上加用Wnt/β-catenin拮抗剂PNU-74654100 mg/(kg·d),连续8周],每组8只。检测各组血脂水平,HE染色图像分析测定胸主动脉中膜面积及血管壁厚度,免疫印迹法检测大鼠胸主动脉组织Wnt/β-catenin的蛋白表达。细胞实验:培养人脐静脉内皮细胞(HUVEC)及人主动脉平滑肌细胞(HA-VSMC)分为对照组、AS模型组[使用20 mg/L氧化型低密度脂蛋白(ox-LDL)干预48 h诱导建立细胞AS模型],AS+HNK组(AS+HNK 32μmol/L干预48 h)、AS+HNK-PNU组(AS+HNK 32μmol/L+PNU-7465450μmol/L干预48 h);分别采用实时荧光定量聚合酶链反应(RT-PCR)法和Western blot法检测细胞Wnt3a和β-catenin mRNA及蛋白质表达。采用Transwell小室实验检测HA-VSMC迁移能力。结果动物实验:与对照组相比,AS组血清三酰甘油、总胆固醇、低密度脂蛋白胆固醇(LDL-C)水平升高(均P<0.05),与AS组比较,AS+HNK组和AS+HNK-PNU组血脂水平降低(P<0.05)。与对照组比较,AS组大鼠胸主动脉中膜面积及血管壁厚度增加(均P<0.01);胸主动脉组织Wnt/β-catenin蛋白表达水平增加(P<0.05);和厚朴酚可降低AS模型大鼠胸主动脉中膜面积及血管壁厚度和Wnt/β-catenin蛋白表达水平(P<0.05),联合使用PNU-74654此效应下降。细胞实验:两种细胞株模型组Wnt3a及β-catenin mRNA和蛋白表达水平高于对照组;AS+HNK组的Wnt3a、β-catenin蛋白表达与mRNA相对表达量低于AS+HNK-PNU组及AS组(均P<0.05)。AS组HA-VSMC迁移能力高于对照组,AS+HNK组与AS+HNK-PNU组HA-VSMC迁移能力低于AS组(P<0.Objective To explore the effects and the possible mechanisms of honokiol(HNK)on Wnt andβ-catenin signaling pathway and arteriosclerosis in both vitro and vivo arteriosclerosis model.Methods In animal experiments,eight Sprague Dawley(SD)rats were used as control group,and twenty-four high-fat-chow-fed SD rats(to establish atherosclerosis rats model)were randomly divided into 3 groups:AS group(atherosclerosis model induced by high fat diet and vitamin D_(2) injected),AS+HNK group[intervention with honokiol 5 mg/(kg·d)for eight weeks]and AS+HNK-PNU group[intervention with honokiol 5 mg/(kg·d)plus PNU-74654100 mg/(kg·d)for eight weeks].Blood lipid levels were detected.Media cross-sectional area and wall thickness of thoracic aorta were measured by HE staining.Wnt andβ-catenin protein levels were detected by Western blot.For in vitro experiments,human umbilical vein endothelial cells(HUVEC)and human aortic vascular smooth muscle cells(HA-VSMC)were cultured,and both were divided into four groups:normal control group,AS group[induced by 20 mg/L oxidized low density lipoprotein(ox-LDL)for 48 hours],AS+HNK group[intervention with honokiol 32μmol/L for 48 hours]and AS+HNK-PNU group[intervention with honokiol 32μmol/L plus 50μmol/L PNU-74654 for 48 hours].Real time fluorescence quantitative polymerase chain reaction(RT-PCR)and Western blot were used to detect the mRNA and protein level of Wnt andβ-catenin in cells after intervention.Transwell assay was used to detect cell migration.Results Compared with control group,triglyceride,total cholesterol and low-density lipoprotein cholesterol(LDL-C)levels increased in AS group.Blood lipid levels decreased in AS+HNK group and AS+HNK-PNU group compared with AS group.Compared with control group,media cross-sectional area and wall thickness,Wnt andβ-catenin protein levels of thoracic aorta increased in AS group.Honokiol reduced media cross-sectional area,wall thickness and Wnt andβ-catenin protein levels of thoracic aorta in AS model,which can be partially blocked by PNU-7

关 键 词：和厚朴酚 Wnt蛋白 Β-连环蛋白 内皮细胞 血管平滑肌细胞 迁移 大鼠 动脉粥样硬化 

分 类 号：R285[医药卫生—中药学]