【作者】 杨静；

【导师】 郑丽丽；

【作者基本信息】 郑州大学 ， 内科学， 2010， 硕士

【摘要】 背景：动脉粥样硬化(atherosclerosis, AS)是各种心脑血管疾病最重要的危险因素,随着人民生活水平的提高及饮食结构的改变,肥胖以及与肥胖相关的疾病如糖尿病、高血压及高脂血症等的发病率也逐渐增加,并通过多种机制介导着AS的形成与发展。近年来的研究证明,肥胖患者体内过量的脂肪组织分泌的多种细胞因子在AS的发生中发挥着重要的作用。内脏脂肪素(visfatin)是新发现的一种脂肪细胞因子,不仅与多种炎症疾病有关,还参与调节多种炎症因子的合成与分泌,能够促进单核细胞表达内皮细胞粘附分子1(ICAM-1)和内皮细胞表达血管内皮生长因子(VEGF)。已有研究发现内脏脂肪素在颈动脉和冠状动脉粥样硬化斑块中显著表达。提示内脏脂肪素可能作为炎症介质参与了动脉粥样硬化的发生和发展。基质金属蛋白酶1(MMP-1)及基质金属蛋白酶抑制物1(TIMP-1)和ICAM-1在炎症所致的内皮细胞损伤中具有重要的作用。目的：本研究通过观察不同浓度的内脏脂肪素对血管内皮细胞(vein endothelia cell, VEC) MMP-1、TIMP-1及ICAM-1基因表达的影响,在基因表达水平探讨内脏脂肪素损伤血管内皮细胞功能,诱发动脉粥样硬化和斑块破裂的相关机制。研究方法：采用体外培养人脐静脉内皮细胞(HUVEC)株,将传至3-6代时处于对数生长期的细胞作为研究对象。分别用浓度为100ng/ml、200ng/ml、400ng/ml、800ng/ml的内脏脂肪素干预细胞24小时,用Trizol法提取细胞总RNA,采用逆转录-聚合酶链反应(RT-PCR)技术半定量检测不同浓度内脏脂肪素刺激后内皮细胞MMP-1、TIMP-1和ICAM-1mRNA的表达。结果：(1)不同浓度的内脏脂肪素刺激内皮细胞24小时后,内皮细胞表达的MMP-1mRNA显著增多,且呈浓度依赖性。(2)不同浓度的内脏脂肪素刺激内皮细胞24小时后,内皮细胞表达的TIMP-1mRNA呈浓度依赖性下降,MMP-1/TIMP-1比值随内脏脂肪素干预浓度的增高而逐渐升高。(3)不同浓度的内脏脂肪素刺激内皮细胞24小时后,内皮细胞表达的ICAM-1mRNA水平显著升高,且其浓度越高,刺激内皮细胞表达ICAM-1mRNA的作用就越强。结论：(1)内脏脂肪素能够促进体外培养的人血管内皮细胞表达MMP-1,下调TIMP-1的表达,且呈剂量依赖性,MMP-1/TIMP-1的比例失衡,打破了细胞外基质合成与降解的平衡状态。(2)内脏脂肪素促进了体外培养的人血管内皮细胞表达ICAM-1,并随着内脏脂肪素浓度的升高而表达增高,呈现剂量依赖性。(3)内脏脂肪素可能作为炎症因子通过影响人血管内皮细胞MMP-1、TIMP-1及ICAM-1的表达参与了动脉粥样硬化性疾病的发生和发展。更多还原

【Abstract】 Background:As we know that atherosclerosis is the most important risk factor of cardiovascular and cardio-cerebrovascular disease. With the improvement of living standards and changing of eating habits, the incidence of obesity and obesity-related diseases such as diabetes, hypertension and hyperlipemia has gradually increased, and these diseases affect on the formation and development of atherosclerosis through various mechanisms. Recent studies have shown that several cytokines secreted by excess adipose tissue of obesity play an important role in the occurrence of atherosclerosis.Visfatin is the most recently identified adipocytokine(known previously as pre-B cell colony enhancing factor, PBEF).Visfatin has not only associated with a variety of inflammatory diseases,but also can regulate the synthesis and secretion of inflammatory cytokine,stimulate the monoeyte releasing intereellular adhesion moleeule-1 and the endothelium releasing vascular endothelium growth factor. It has been found that visfatin expressed in carotid endarterectomy specimens and coronary artery plaques. This indicates that visfatin is not only an cytokine but also interact with atherosclerosis. MMP-1,TIMP-1 and ICAM-1 play an important role in the inflammation-induced endothelial cell injury.Objective:To investigate the mechanism of artherosclerosis and plaque disurptioninduced by dysfunction of vascular endothelial cell with visfatin in different concentrations, we evaluate the effect of visfatin on the genetic expression of MMP-1,TIMP-1 and ICAM-1 in human vascular endothelium.Method:HUVECs were incubated in vitro and treated with visfatin in 100ng/ml、200ng/ml、400ng/ml、800ng/ml levels for 24 hours respectively.Then total cellular RNA was extracted by Trizol method and the expression of MMP-1,TIMP-1 and ICAM-1 were detected in mRNA levels using RT-PCR method.Results:(1)Visfatin can increase the expression of MMP-1 mRNA significantly in a dose-dependent manner.(2)Visfatin can reduce the expression of TIMP-1 mRNA significantly in a dose-dependent manner,and the ratio of MMP-1/TIMP-1 increased gradually with the increase of visfatin.(3)Visfatin can increase the expression of ICAM-1 mRNA significantly in a dose-dependent manner.Conclusion:(1)The endothelium stimulated with visfatin can increase the expression of MMP-1 mRNA in a dose-dependent manner. And visfatin can down regulate the expression of TIMP-1 mRNA and result in the proportional imbalance of MMP-1/TIMP-1,which break the extracellular matrix synthesis and degradation balance finally.(2)The endothelium stimulated with visfatin can increase the expression of ICAM-1 mRNA in a dose-dependent manner.(3) Visfatin may be involved in the artery atherosclerotic disease through affecting the expression of MMP-1, TIMP-1 and ICAM-1 in human vascular endothelial cells.更多还原