痰瘀同治方基于TGF-β信号通路维持血管稳态治疗动脉粥样硬化的机制研究  

Mechanism of Tanyu Tongzhi Formula in treatment of atherosclerosis by maintaining vascular homeostasis based on TGF-βsignaling pathway

作　　者：崔晓珊 张会雨 陈原原 李亮 高佳明 郝伟[1] 谢成志 刘建勋[1,2] 付建华 郭浩[1] CUI Xiao-shan;ZHANG Hui-yu;CHEN Yuan-yuan;LI Liang;GAO Jia-ming;HAO Wei;XIE Cheng-zhi;LIU Jian-xun;FU Jian-hua;GUO Hao(Institute of Basic Medicine,Xiyuan Hospital,China Academy of Chinese Medical Sciences,Beijing 100091,China;National Clinical Research Center for Traditional Chinese Medicine of Cardiovascular Diseases,Beijing 100091,China)

机构地区：[1]中国中医科学院西苑医院基础医学研究所,北京100091 [2]国家中医心血管病临床医学研究中心,北京100091

出　　处：《中国中药杂志》2024年第23期6429-6438,共10页China Journal of Chinese Materia Medica

基　　金：国家自然科学基金面上项目(82074060);中国中医科学院科技创新工程项目(CI2021A04602);中国中医科学院西苑医院能力提升项目(XYZX0304-02)。

摘　　要：以转化生长因子-β(TGF-β)/SMAD家族成员2/3(Smad2/3)信号通路为切入点,探讨痰瘀同治方治疗小鼠动脉粥样硬化(AS)的可能作用机制及组方配伍意义。8只C57BL/6J小鼠作为正常组,给予普通饲料喂养,35只同品系ApoE^(-/-)小鼠通过高脂饲料喂养8周制备AS模型。造模成功后随机分为模型组、痰瘀同治组(18.2 mg·kg^(-1))、化痰组(10.4mg·kg^(-1))、祛瘀组(7.8 mg·kg^(-1))各8只。除正常组外,其余各组均采用高脂饲料连续8周喂养制备AS模型,之后各组小鼠均灌胃给药8周。采用酶联免疫吸附测定(ELISA)检测总胆固醇(TC)、甘油三酯(TG)、低密度脂蛋白胆固醇(LDL-C)、高密度脂蛋白胆固醇(HDL-C)以及白细胞介素1β(IL-1β)、白细胞介素18(IL-18)水平;苏木素-伊红(HE)染色、油红O染色及Russell-Movat五色染观察小鼠主动脉组织的病理学形态变化,并计算主动脉斑块面积占比、红染脂质面积占比、胶原纤维及弹力纤维占比;免疫荧光法检测基质金属蛋白酶2(MMP2)及组织金属蛋白酶抑制物2(TIMP2)蛋白表达水平;Western blot检测主动脉TGF-β1、TGF-β2、Smad2/3、Smad7的蛋白表达;实时荧光定量PCR法检测小鼠主动脉组织转化生长因子-β受体(TGF-βR)、TGF-β1、Smad2/3、Smad7、细胞黏附分子-1(ICAM-1)、血管细胞黏附分子-1(VCAM-1)mRNA表达量。结果显示,与正常组比较,模型组血浆TC、LDL-C升高,HDL-C显著降低;血浆IL^(-1)β、IL-18显著升高;主动脉斑块面积占比、红染脂质面积占比及胶原纤维占比增加;主动脉弓中MMP2表达显著上调,TIMP2表达显著下调;主动脉组织TGF-βR、TGF-β1、p-Smad2/3蛋白及mRNA表达水平显著升高,Smad7表达水平降低;与模型组比较,痰瘀同治组血浆TC、LDL-C显著降低,HDL-C显著升高;血浆IL-1β、IL-18显著降低;主动脉斑块明显缩小,主动脉斑块病变程度显著减小;主动脉弓中MMP2表达明显下调;ICAM-1、VCAM-1 mRNA表达水平显著降低;主动脉组This study aimed to investigate the potential mechanism and the compatibility significance of Tanyu Tongzhi Formula in treating atherosclerosis(AS)in mice based on the transforming growth factor-β(TGF-β)/Smad2/3 signaling pathway.Eight C57BL/6J mice were as assigned to a normal control group and fed a regular diet,while 35 ApoE^(‒/‒) mice of the same strain were fed a high-fat diet for 8 weeks to establish an AS model.The model mice were randomly divided into a model group,a Tanyu Tongzhi group(18.2 mg·kg^(-1)),a Huatan(phlegm-resolving)group(10.4 mg·kg^(-1)),and a Quyu(blood stasis-resolving)group(7.8 mg·kg^(-1)),with 8 mice in each group.Except for the normal group,all other groups continued to be fed a high-fat diet for 8 weeks to maintain the AS model,and then the mice were treated by gavage for 8 weeks.Plasma levels of total cholesterol(TC),triglycerides(TG),low-density lipoprotein cholesterol(LDL-C),high-density lipoprotein cholesterol(HDL-C),interleukin-1β(IL-1β),and interleukin-18(IL-18)were measured using enzyme-linked immunosorbent assay(ELISA).).Hematoxylin and eosin(HE)staining,oil red O staining,and Russell-Movat pentachrome staining were performed to observe the pathological changes in the aortic tissue.The proportions of aortic plaque area,lipid-stained area,collagen fibers,and elastic fibers were calculated.Immunofluorescence was used to detect the protein expression levels of matrix metalloproteinase-2(MMP2)and tissue inhibitor of metalloproteinases-2(TIMP2).Western blot was used to detect the protein expression levels of TGF-β1,TGF-β2,Smad2/3,and Smad7 in aortic tissue.Real-time fluorescence quantitative PCR(RT-qPCR)was used to measure the mRNA expression levels of TGF-βreceptor(TGF-βR),TGF-β1,Smad2/3,Smad7,intercellular adhesion molecule-1(ICAM-1),and vascular cell adhesion molecule-1(VCAM-1)in aortic tissue.The results showed that compared with the normal control group,the model group had increased plasma TC and LDL-C,significantly decreased HDL-C,and significantly elevated pla

关 键 词：动脉粥样硬化 痰瘀同治方 血管稳态 TGF-Β SMAD2/3 Smad7 

分 类 号：R285.5[医药卫生—中药学]