人主动脉脂纹超微结构的进一步观察

为进一步探讨脂纹的超微结构,我们在以前工作基础上,用钌红染色做透射电镜检查另外11例主动脉的24对脂纹区及其附近正常区内膜。结果显示脂纹区某些内皮细胞(EC)糖萼明显变薄、有的EC退变甚至脱落、偶见细胞间隙扩张现象,进一步提示EC损伤和通透性增高在动脉粥样硬化(AS)中的作用。脂纹区有钙化小体,提示钙盐沉积可能开始于AS早期脂纹阶段。     

动脉粥样硬化形成和消退中平滑肌细胞定量研究

用网格交点计数法测量了动脉粥样硬化(AS)形成组和消退组及对照组的平滑肌细胞(SMC)胞质内的超做结构。AS形成组内膜或中膜第一层(靠内弹性膜侧)SMC跑质中肌丝减少,而高尔基体、粗面内质网和线粒体增多,AS消退组的肌丝、高尔基体和粗面内质网与对照组相比没有明显差别。结果提示:AS形成组SMC由收缩型向合成型方向转变,可能与SMC增生和转移及与分泌细胞外基质有关;AS消退组SMC由合成型向收缩型转变,可减少细胞外基质的分泌。     

血小板源性生长因子与动脉粥样硬化的关系

PlateletDerivedGrowthFactorandAtherosclerosisChenZhong(DepartmentofPathology,FuwaiHospital.CAMS,Beijing)动脉粥样硬化（AS）病变的一个重要特征是动脉内膜的平滑肌细胞（SMC）增殖。SMC从动脉中膜迁移至内膜并增殖的过程受到众多因素的影响．其中血小板源性生长因子（Plateleterlvedgrowthfactor，PDGF）起着极其重要的作用。现将近年来有关PDGF与AS关系的文献综述如下。IPDGF的来源及分布体外培养的细胞大多数需要在有血清的条件下才能分裂增殖，这是因为血清中含有细胞增殖所需要的特异性物质，血小板释放的PDG…     

动脉粥样硬化狭窄模型的建立及NF-κB、IGF的表达

目的 :建立动脉粥样硬化狭窄 (Atherosclerosis,AS)动物模型并探讨影响AS形成、发展的分子生物学方面的机制。方法 :10只大耳白兔 ,高脂饲料喂饲一周后 ,采用颈动脉为逆行入径 ,将球囊导管顺行插入并随机至一侧髂动脉远端 ,使其内膜损伤 ,(另一侧不损伤 ,作为对照 ) ,高脂喂养 6周后建立髂动脉AS狭窄模型 ,并通过影像学、病理学光镜、电镜及免疫组化分析证实 ,并观察核因子 κB(nuclearfactor κappaB ,NF κB)、胰岛素样生长因子 1(insulin likegrowthfactor 1,IGF 1)的免疫组织化学分析。结果 :模型组动脉造影显示平均狭窄度 6 9 0± 19 12 % ;HE染色示内膜明显增厚至管腔偏心性狭窄 ,增厚内膜主要由泡沫细胞、平滑肌细胞组成 ,内弹力膜分离断裂 ,中膜平滑肌细胞迁移入内膜层 ;电镜示膜结构损伤 ,可见凋亡小体 ;免疫组化示NF κB、IGF 1蛋白表达量均高于对照组 (p <0 0 5或p <0 0 0 5 )。结论 :(1)通过球囊损伤血管内膜加高脂饮食成功建立了兔髂动脉AS狭窄模型 ,多数为偏心、局限性的 4 0 %～ 10 0 %的狭窄 ,模型确切、可靠、是用于经皮冠状动脉腔内成形术 (per cutaneoustransluminalcoronaryangioplasty ,PTCA)的理想动物模型 ;(2 )NF κB及其靶基因IGF 1的激活与AS病变的发生与发展密切相关     

平滑肌细胞与基质在静脉桥内膜增生的时空变化

目的 :研究移植静脉桥内膜增厚过程中平滑肌细胞 （SMC）增殖、表型转化和细胞外基质积聚的时间变化与空间分布。方法 :将兔自体颈浅静脉桥移植到颈动脉 ,在术后 3,8,15 d和 2 ,6个月取血管桥行免疫组化染色。结果 :静脉桥中层 SMC在术后 3d表现受损并开始增殖 ,8d迁移到内层并形成新内膜层 ,SMC表型从正常成熟型向胚胎型转化。术后 15 d,SMC增殖率达高峰 ,2月后迅速下降 ,且绝大部分细胞从胚胎型转变为成熟表型 ,但在术后6个月 ,仍有少量胚胎型 SMC存在。新内膜层自形成后在整个观察期呈进行性增厚。内膜层细胞外基质主要成分 : 型胶原、硫酸肝素、硫酸皮肤素。术后 2月基质大多堆积在内膜深层细胞稀疏区。结论 :静脉桥新内膜的形成起源于表型转化胚胎型 SMC的迁移 ,细胞增殖和基质积聚使新内膜增厚。而胚胎型 SMC在新生内膜的持续存在 ,可能促使基质不断堆积和新内膜层进行性增厚 ,易使移植静脉桥造成狭窄     

动脉粥样硬化狭窄模型的建立及NF-KB、IGF的表达

目的建立动脉粥样硬化狭窄(Atherosclerosis,AS)动物模型并探讨影响AS形成、发展的分子生物学方面的机制.方法10只大耳白兔,高脂饲料喂饲一周后,采用颈动脉为逆行入径,将球囊导管顺行插入并随机至一侧髂动脉远端,使其内膜损伤,(另一侧不损伤,作为对照),高脂喂养6周后建立髂动脉AS狭窄模型,并通过影像学、病理学光镜、电镜及免疫组化分析证实,并观察核因子-kB(nuclear factor-k appaB,NF-kB)、胰岛素样生长因子-1(insulin-like growth factor 1,IGF-1)的免疫组织化学分析.结果模型组动脉造影显示平均狭窄度69.0±19.12%;HE染色示内膜明显增厚至管腔偏心性狭窄,增厚内膜主要由泡沫细胞、平滑肌细胞组成,内弹力膜分离断裂,中膜平滑肌细胞迁移入内膜层;电镜示膜结构损伤,可见凋亡小体;免疫组化示NF-kB、IGF-1蛋白表达量均高于对照组(p＜0.05或p＜0.005).结论(1)通过球囊损伤血管内膜加高脂饮食成功建立了兔髂动脉AS狭窄模型,多数为偏心、局限性的40%～100%的狭窄,模型确切、可靠、是用于经皮冠状动脉腔内成形术(percutaneous transluminal coronary angioplasty,PTCA)的理想动物模型;(2)NF-kB及其靶基因IGF-1的激活与AS病变的发生与发展密切相关.     

癫狂梦醒汤对兔动脉粥样硬化血管基质金属蛋白酶-9表达的影响

目的观察癫狂梦醒汤对兔动脉粥样硬化(AS)血管基质金属蛋白酶-9(MMP-9)表达的影响,以期为癫狂梦醒汤抗AS提供理论依据。方法将32只日本大耳白兔随机分成空白对照组、模型组、癫狂梦醒汤组。采用高脂喂饲制作AS模型。第12周末观察动脉血管内膜厚度,免疫组化法观察MMP-9蛋白表达。结果以高脂喂养成功建立AS模型,模型组血管内膜增厚,MMP-9蛋白表达明显增强;癫狂梦醒汤组血管内膜变薄,MMP-9蛋白表达明显下降(P0.01)。结论癫狂梦醒汤可抑制MMP-9蛋白表达,明显抑制AS发生发展。     

动脉粥样硬化大鼠主动脉内膜增生与树突状细胞分布的关系

目的:探讨动脉粥样硬化(AS)大鼠树突状细胞(DCs)在外周血、动脉壁以及脾脏分布情况及与主动脉内膜增生的关系。方法:经高脂饮食喂养12周形成AS大鼠模型,分别取外周血、动脉壁以及脾脏组织,并制备组织单细胞悬液。应用流式细胞仪检测DCs及T淋巴细胞比例。结果:AS大鼠主动脉壁内膜较正常对照大鼠内膜增生明显(P<0.05),其外周血DCs分布明显减少且主动脉壁内DCs比例明显升高(P<0.001)。T淋巴细胞比例检测发现,AS大鼠较正常大鼠,外周血及主动脉壁内T淋巴细胞比例均升高(P<0.001)。结论:AS大鼠主动脉内膜增生与DCs及T淋巴细胞比例变化有关,DCs在AS大鼠主动脉内膜增生、管壁粥样硬化形成中可能起重要作用。     

血管紧张素Ⅱ1型受体在血管球囊损伤后平滑肌细胞凋亡中的作用

目的探讨血管球囊损伤后平滑肌细胞(SMC)凋亡的机制。方法采用末端脱氧核苷酸转移酶介导的三磷酸脱氧尿嘧啶缺口末端标记法(TUNEL)和免疫组织化学技术检测球囊损伤后血管平滑肌细胞凋亡及血管紧张素Ⅱ1型受体(AT1R)表达的变化。结果球囊损伤后第3天,血管中层AT1R表达比假手术组显著增多(P<0.05),以后无显著改变;损伤后第7天内膜层AT1R为中层的近2倍;至损伤后第28天,内膜层AT1R表达最高;球囊损伤后第3天,血管中层出现凋亡的SMC;损伤后第7天,内膜和中层SMC凋亡率最高,凋亡的SMC主要分布在内膜层;以后逐渐降低,至损伤后第28天,仅内膜层有少量凋亡的SMC,AT1R拮抗剂Irbesartan显著增加SMC凋亡。结论血管球囊损伤后,AT1R上调,血管紧张素Ⅱ通过与AT1R结合抑制VSMC凋亡。     

胰岛素与动脉粥样硬化关系的实验研究

对正常大鼠、糖尿病大鼠、INS治疗的糖尿病大鼠及单纯注射INS的正常大鼠的主动脉进行了形态学的对比观察,并测定了各组大鼠的血糖、血脂、过氧化脂质和血清INS水平。结果显示:糖尿病大鼠的主动脉出现早期动脉粥样硬化(AS)病变,即内膜增厚、内皮下间隙加宽、内弹力板破坏、内膜和中膜浅层平滑肌细胞增生及细胞内脂滴等。INS治疗组大鼠,虽然INS缺乏得到补充,DM状态下的糖、脂质代谢紊乱得到明显纠正,但是主动脉出现类似DM组的病变,且病变程度重于DM组。注射INS的正常大鼠,血清INS水平明显增高,主动脉病变与前两组相比,相对更重。结果表明,INS缺乏的实验性糖尿病大鼠,由于高血糖,高血脂及血小板功能异常等多种因素,可以导致AS的发生;而胰岛素水平过高(绝对或相对的),在DM-AS的发病中,直接或间接的起着重要的促进作用。     

The evolution of early fibromuscular lesions hemodynamically induced in the dog renal artery. I. Light and transmission electron microscopy.

In view of the important roles of arterial intimal fibromuscular lesions as precursors of atherosclerotic plaque and occlusive lesions in arterial reconstructions, a model has been developed for the rapid hemodynamic induction of these lesions by anastomosis of the dog right renal artery to the inferior vena cava. Light and transmission electron microscopic observations were made on the arterial shunt after periods of rapid flow ranging form 10 minutes to 2 hours to identify initial factor(s) and evolutionary mechanisms in the etiology of the lesions. The sequence of events included aberrations in ruthenium red staining of the endothelial luminal membrane at 10 minutes, multilayered thickening of the subendothelial basement membrane (BM) at 15 minutes, and initial reorientation and migration of smooth muscle cells (SMC) into the intima along with the appearance of areas of degeneration of the internal elastic lamina (IEL) at 30 minutes. The endothelial cells were still intact in some areas overlying the SMC migration and IEL degeneration, but they were separating from the surface in other such areas. As subendothelium became exposed, some platelet adherence was noted. By 2 hours, the entire wall reaction was fully developed. Initial observations indicate that in the evolution of this hemodynamically induced lesion visible alteration in the endothelial cells is not prerequisite to degeneration of the underlying IEL and reorientation and migration of medial SMC.     

Identification of intimal subendothelial cells from human aorta in primary culture

Subendothelial cells (SEC) were obtained from the inner intimal layer of adult human aorta by collagenase treatment. SEC were identified in primary culture either as smooth muscle cells by staining with FITC-labeled antisera against human smooth muscle myosin or as macrophages, foam cells and contaminating endothelial cells by their uptake of malondialdehyde treated low density lipoproteins labeled with fluorescent dye 3,3'-dioctadecylindocarbocyanine. Between 1 and 5 days in culture, along with smooth muscle cells (SMC, 38-82%), endothelial cells (0-9%), macrophages and foam cells (2-32%), one more type of cell was found. This cell type resembled SMC in size and shape, but was not stained by antisera to SMC myosin. By ultrastructural criteria these cells were characterized as modulated SMC for they contained prominent rough endoplastic reticulum and Golgi complex together with basement membrane and a large number of plasmalemmal vesicles. Like SMC they reacted with phalloidin and were stained by anti-vimentin but not by anti-desmin monoclonal antibodies. The proportion of such cells varied from 5 to 33% of total cell number and increased in parallel to macrophages and foam cells in vessels with well developed atherosclerotic lesions. We conclude that the applied technique may be used for identification of cultured vascular cells including modulated SMC.     

Phenotypic expression of surface antigens of rabbit aortic smooth muscle cells in culture. Monoclonal antibody, 2P1A2, characteristic of smooth muscle cells present in atherosclerotic plaque, is not correlated with cell proliferation

The expression of smooth muscle cell (SMC) antigens was studied in culture by immunofluorescence and immunoelectron microscopy. As specific SMC markers, we used 2 monoclonal antibodies (MAb), 1PC1 and 2P1A2 which are able to detect atherosclerotic plaques in the rabbit. MAb 1PC1 recognizes an antigen expressed on the cell surface, starting on the 7th day in primary culture after serum activation, and then secreted. On a confluent SMC monolayers this antigen appears outside the cell as an important filamentous network. The kinetics of secretion of this external protein recognized by 1PC1 corresponds to the kinetics of the secretory phenotype described by Chamley-Campbell and Campbell (Atherosclerosis, 40 (1981) 347). 2P1A2 MAb is specific for SMCs exclusively present in the rabbit atherosclerotic plaque. We studied the degree of reactivity of 2P1A2 with SMCs during primary cell culture. This "atherosclerotic" antigen of SMCs recognized by 2P1A2 is expressed in culture conditions by SMCs from rabbit normal media. This antigen appears after 3 days of serum activation, and heparin growth inhibition does not interfere with its expression. 2P1A2 recognized antigen is expressed during all cell cycle phases without amplification. 3 days after fetal calf serum (FCS) stimulation of cells which are in G0/G1, 89% are labelled by 2P1A2, 4 days later G0/G1 positive cells constitute 49%. We conclude that 2P1A2 immunolabelling on the SMC surface reflects an activated state which is not correlated with SMC proliferation.     

Atherosclerotic lesions in the coronary arteries of hyperlipidemic swine. Part 1. Cell increases, divisions, losses and cells of origin in first 90 days on diet

The intima of the proximal portion of the coronary arteries of young swine is normally thickened by accumulations of cells about 90% of which are smooth muscle cells (SMC) and about 10% are of probable monocyte origin. Extracellular components such as collagen and elastic tissue are also present but we have chosen to emphasize their cellular nature by calling the regions of thickened intima, intimal cell masses (ICM). We have previously shown that atherosclerotic lesions produced in the coronary arteries of swine by 90 days of feeding a hyperlipidemic (HL) diet arise almost exclusively in the normally occurring ICM. We are reporting here a study of the pathogenesis of these lesions following killing at 0, 14, 49 and 90 HL diet days with comparisons between ICM in control mash-fed swine and ICM-lesions in the HL swine. We found that in the ICM: lipid accumulation was present by 14 days and increased thereafter; the lipid was mostly in SMC but percentage wise the monocyte-macrophages were involved as much or more, cell division activity was increased 3-4-fold by 49 days, cell numbers in ICM were similar in HL and control swine at 49 days but were about 6-fold greater in the HL swine at 90 days, (now in ICM-lesions), at 90 days, circa 90% of the cells appeared to be of SMC and circa 10% of monocyte origin both in the ICM-lesions of the HL swine and in the normal ICM of the controls. The data suggest but do not prove that early lipid accumulation precedes increased cell divisions especially among the SMC component and this in turn precedes increased numbers of cells in the ICM. Although SMC constitute the major cell component of the ICM-lesion at 90 days, the monocyte-macrophage-like cells also increase in number as a result of the HL diet and constitute a small but definite minor component. One possible explanation for the increased cell division activity is that one of the lipid constituents is acting as mitogen; another possibility is that the effect of a well known mitogen such as platelet-derived growth factor is enhanced by the lipid; another is that the monocytes are being stimulated to produce monocyte-derived growth factor. In any event in the very early stage of atherogenesis in the coronary arteries in these experiments excessive proliferation of resident SMC in the ICM appears to be the predominant feature.     

Separation and characterization of macrophages and smooth muscle cells in rabbit atherosclerotic lesions

Atherosclerotic aortic intimas of cholesterol-fed rabbits were enzymatically dispersed into single cells by collagenase and elastase. And monocyte-macrophages (M phi) were separated from smooth muscle cells (SMC), using the ability of M phi to adhere to a plastic dish firmly even in the enzyme solutions. Round or oval, heavily lipid-laden cells, so-called foam cells (FC), belonged to the M phi fraction. M phi-FC showed very strong activity for non-specific esterase using alpha-naphthyl butyrate, while SMC showed little or no activity. Some of the FC were large and multinucleated (multinucleated giant foam cells). They also showed positive non-specific esterase staining and are thought to be derived from M phi. M phi-FC synthesized various proliferate in the medium and the number decreased gradually within several days. Some SMC were heavily lipid-laden; however, they retained their original spindle shape. SMC lost lipid droplets gradually as they proliferated to confluence. SMC from atherosclerotic lesions showed higher proliferative activity than those from normal-appearing medias of atherosclerotic aortas or control aortas. Almost no M phi-FC were obtained from the intima-medias of grossly normal portions of atherosclerotic aortas and control aortas. The present method will be useful for studying the role of these cells in the pathogenesis of atherosclerosis.     

Endoglin, a TGF-beta receptor-associated protein, is expressed by smooth muscle cells in human atherosclerotic plaques

Endoglin is a transmembrane protein that is found in association with transforming growth factor-beta (TGF-beta) superfamily receptor complexes and has an expression pattern that appears to be restricted primarily to endothelial cells, activated macrophages, trophoblasts, and fibroblasts. Since mutations in endoglin have been shown to be linked to hereditary hemorrhagic telangiectasia type 1, a disease manifested as vascular malformations characterized by excessive layers of vascular smooth muscle cells (VSMC), the expression of endoglin was investigated in VSMC. In vivo, the majority of SMC in human atherosclerotic plaques expressed high levels of endoglin, while endoglin was not detected in SMC from samples of the normal arterial wall. In vitro studies demonstrate that human aortic smooth muscle cells (HASMC) express the L-isoform of endoglin. Like endothelial cells, HASMC express endoglin protein as a dimer on the cell surface that binds TGF-beta1. In vitro, endoglin expression by HASMC is upregulated in response to TGF-beta1, suggesting that the presence of this factor in the atherosclerotic plaque might be responsible for the increased expression of endoglin. The demonstration of increased levels of endoglin in VSMC in human atherosclerotic plaques suggests a role for SMC endoglin in the maintenance of vascular integrity and in the response of the vessel wall to injury.     

经皮腔内血管成形术后形态学改变及组织修复过程的实验研究

本文对34只家兔48支血管的实验性髂动脉粥样硬化性狭窄进行了腔内血管成形术(PTA)后即刻至30天的组织学研究。结果表明,内腆及斑块撕裂是PTA的主要机制。球囊扩张后平滑肌细胞(SMC)是修复过程中的主要细胞成份,薄层附壁血栓对维持动脉壁的完整结构有着重要作用。SMC增生程度与内膜撕裂的深浅有关。暴露的中膜比内皮损伤更易发生血小板粘附和血栓形成。3支血管发生再狭窄,2支由于内腆SMC过度增生。另1支由于血栓机化所致。     

Proteoglycan distribution in the intima and media of the aortas of young and aging rabbits: an ultrastructural study

Aortas from normal healthy rabbits, approx. 3 months old, were examined by light and transmission electron microscopy. The proteoglycan of the extracellular matrix, which was stained by ruthenium red and appeared as granules by transmission electron microscopy, was quantitated morphometrically in the intima and the superficial media. The intima included areas which were thickened and which contained connective tissue, including proteoglycan, and some smooth muscle cells. In the thickened intima there was a greater proportion of extracellular space which was occupied by proteoglycan, and the proteoglycan was present in higher concentration than in the media. In the aortas of rabbits, approx. 2 years old, the extent of intimal thickening and the concentration of proteoglycan increased in the thickened intima but there was no evidence of extracellular lipid deposition. The endothelial basement membrane contained small proteoglycan granules (heparan sulphate) which decreased in concentration in older animals. It is possible that the accumulation of proteoglycan in the thickened intima increases the susceptibility of the intima to accumulate lipid following an additional stimulus, such as hyperlipaemia, in the initial stages of atherosclerosis.