流体剪切应力对晚期内皮祖细胞生物学功能的影响

目的 研究流体剪切应力处理对晚期内皮祖细胞（endothelial progenitor cells,EPCs）体外及体内生物学功能的影响。 方法 密度梯度离心法分离大鼠骨髓单核细胞,应用EGM-2MV进行体外培养。以3~4代的EPCs,即晚期EPCs为靶细胞,对其施以1.2 Pa剪切应力处理。采用EdU标记技术、黏附能力测定实验、改良的Boyden小室、Annexin V/PI、β 半乳糖苷酶检测法、Matrigel法、荧光定量RT PCR等方法分别检测剪切应力对晚期EPCs增殖、黏附、迁移、凋亡、衰老、体外成血管及VEGF mRNA表达等生物学功能的影响。应用大鼠颈动脉损伤模型及细胞原位移植等实验手段检测剪切应力预处理对晚期EPCs修复受损内皮的影响。结果1.2 Pa剪切应力处理可不同程度提高晚期EPCs的增殖、黏附、迁移及成血管能力（P＜0.01）,上调VEGF的基因表达,抑制晚期EPCs的衰老及凋亡（P＜0.01）;移植经剪切应力预处理的晚期EPCs可加速损伤内皮的修复,减缓内膜的增生。结论 流体剪切应力可改善晚期EPCs的功能活性,提高晚期EPCs修复损伤血管内皮的能力, 这为EPCs的临床应用及剪切应力介导的细胞疗法提供了实验依据。     

药物促进冠脉支架置入术后血管再内皮化

经皮冠状动脉球囊扩张和支架置入术是目前治疗冠心病的重要手段。早期的冠脉支架为金属裸支架（BMS）,由于BMS置入术后冠状动脉的再狭窄率较高,药物涂层支架（DES）应运而生。DES通过抑制血管内膜的增生降低支架置入术后再狭窄,但同时也带来了内皮延迟愈合,内皮化延迟是支架内晚期血栓发生的基础,晚期血栓常可带来非常严重的后果。目前已知很多药物可促进血管再内皮化,预防晚期血栓的发生。本文将重点就晚期血栓发生的机制、常见促进再内皮化的药物如粒细胞集落刺激因子、过氧化物酶体增殖物激活受体激动剂、促红细胞生成素、雌激素、他汀的机制及研究现状进行综述。     

G-CSF对动脉粥样硬化兔颈总动脉球囊损伤后内皮修复的影响

目的研究重组人粒细胞集落刺激因子（G-CSF）的骨髓动员作用能否促进动脉粥样硬化性血管内膜损伤后的再内皮化过程,来预防损伤血管再狭窄.方法将35只新西兰雌性大白兔,随机分为正常饲养组（对照组）5只;高脂饲料喂养组30只在喂养10周后随机抽取20只行右颈总动脉囊扩张损伤,术后随机将其分为：G-CSF干预＋损伤组（n=10）,予静脉注射G-CSF50μg/d,连续7d,NS＋损伤组（n=10）,术后静脉注射生理盐水2mL/d,连续7d;第12周处死所有试验动物,取右颈总动脉、胸主动脉行HE染色观察血管粥样硬化病变评价模型制作情况,弹力纤维染色计算内膜（）I、中膜面积（M）比值I/M以评价内膜增生程度;伊文氏蓝染色并计算内皮再生面积（An）、内皮总面积（A）t比值An/At以评价再内皮化率;CD31免疫组化染色检测损伤局部CD31细胞表达的阳性率以评价G-CSF对EPCs动员效果以及EPCs对受损内皮的修复情况.结果 （1）高脂饲料喂养12周后可形成典型的动脉粥样硬化斑块.（2）G-CSF＋损伤组与NS＋损伤组相比I/M值降低,差异有统计学意义（P〈0.01）.（3）An/At明显升高,有统计学意义（P〈0.01）.（4）免疫组织化学染色显示颈动脉内皮CD31细胞表达阳性率G-CSF＋损伤组高于NS＋损伤组,有统计学意义（P〈0.01）.结论 G-CSF可以动员骨髓EPCs到外周血并迁移到血管内膜受损部位,促进再内皮化,抑制内膜增生,有效预防血管损伤后再狭窄的发生。     

白藜芦醇对损伤动脉再内皮化和内膜增生的影响

目的探讨白藜芦醇对大鼠主动脉内膜剥脱后内皮修复、内膜增生的影响及其潜在机制。方法54只雄性SD大鼠随机分为假手术组(n=9)、对照组(n=15)、小剂量[10 mg/(kg.d)]白藜芦醇组(n=15)和大剂量[50mg/(kg.d)]白藜芦醇组(n=15),术前2周至动物处死期间经灌胃器给药;术后1周、2周分别应用Ⅷ因子检测和伊文思蓝染色评价各组损伤血管内皮修复情况;术后2周、4周采用HE染色观察内膜增生情况;采用逆转录聚合酶链反应及免疫组织化学法评价术后1周4、周损伤血管的内皮型一氧化氮合酶mRNA和蛋白水平的表达;体外分离、培养大鼠外周血内皮祖细胞,并采用免疫组织化学法进行鉴定;通过倒置荧光显微镜计数双阳性细胞以评价各组术后1周的外周循环内皮祖细胞数量。结果10 mg/(kg.d)白藜芦醇明显促进术后1周及2周的内皮修复,而50mg/(kg.d)不能;10 mg/(kg.d)白藜芦醇显著抑制术后2周、4周的内膜增生,而50 mg/(kg.d)只能抑制术后4周的内膜增生,且效果不如前者;10 mg/(kg.d)白藜芦醇明显促进损伤血管的内皮型一氧化氮合酶mRNA水平和蛋白水平的表达,而50 mg/(kg.d)组与对照组无明显差异;与对照组相比,10 mg/(kg.d)白藜芦醇显著升高术后1周的外周血内皮祖细胞数量(32.46±6.52比21.58±3.69,P<0.05),而50 mg/(kg.d)白藜芦醇无明显差异(22.48±6.89比21.58±3.69,P>0.05)。结论小剂量白藜芦醇可上调损伤血管内膜的内皮型一氧化氮合酶表达、促进内皮祖细胞动员、加快内皮修复及抑制内膜增生;而大剂量白藜芦醇只能抑制术后4周的内膜增生。     

理阿诺碱对雷帕霉素诱导的内皮生长晕细胞功能抑制的保护作用

目的 探讨理阿诺碱对雷帕霉素诱导的内皮生长晕细胞（EOCs）功能抑制的影响.方法 密度梯度离心法分离脐血单个核细胞,培养并扩增EOCs,免疫组化法、荧光染色法鉴定其内皮细胞特性.分别加雷帕霉素、理阿诺碱、理阿诺碱预处理1h再加雷帕霉素,与EOCs作用24h,CCK8检测细胞增殖能力,Transwell小室检测细胞迁移能力,并在倒置显微镜下检测细胞的黏附能力.结果 雷帕霉素抑制EOCs的增殖、迁移、黏附能力（P＜0.05）.理阿诺碱预处理1h能改善雷帕霉素对EOCs的增殖、迁移、黏附的抑制作用（P＜0.05）,理阿诺碱单独作用对EOCs的增殖、迁移、黏附无影响（P ＞0.05）.结论 雷帕霉素抑制EOCs的增殖、迁移、黏附能力 理阿诺碱可以改善雷帕霉素诱导的对体外培养的EOCs增殖、迁移、黏附的抑制作用.     

Carbon Monoxide Releasing Molecule Accelerates Reendothelialization after Carotid Artery Balloon Injury in Rat

Objective This study was aimed to investigate the effects of carbon monoxide releasing molecule (CORM-2), a novel carbon monoxide carrier, on the reendothelialization of carotid artery in rat endothelial denudation model.
Methods Male rats subjected to carotid artery balloon injury were treated with CORM-2, inactive CORM-2 (iCORM-2) or dimethyl sulfoxide (DMSO). The reendothelialization capacity was evaluated by Evans Blue dye and the immunostaining with anti-CD31 antibody. The number of circulating endothelial progenitor cells (EPCs) was detected by flow cytometry. The proliferation, migration, and adhesion of human umbilical vein endothelial cells (HUVECs) were assessed by using [3H]thymidine, Boyden chamber and human fibronectin respectively. The expressions of protein were detected by using western blot analysis.
Results CORM-2 remarkably accelerated the re-endothelialization 5 d later and inhibited neointima formation 28 d later. In addition, the number of peripheral EPCs significantly increased in CORM-2-treated rats than that in iCORM-2 or DMSO-treated rats after 5 d later. In vitro experiments, CORM-2 significantly enhanced the proliferation, migration and adhesion of HUVECs. The levels of Akt, eNOS phosphorylation, and NO generation in HUVECs were also much higher in CORM-2 treated group. Blocking of PI3K/Akt/eNOS signaling pathway markedly suppressed the enhanced migration and adhesion of HUVECs induced by CORM-2.
Conclusion CORM-2 could promote endothelial repair, and inhibit neointima formation after carotid artery balloon injury, which might be associated with the function changes of HUVECs regulated by PI3K/Akt/eNOS pathway.     

Endothelial progenitor cell therapy in atherosclerosis: A double-edged sword?

Atherosclerosis, an inflammatory process that selectively affects arteries, is highly prevalent in human. Thrombo-occlusive complications of atherosclerosis, including stroke and myocardial infarction, are becoming major causes of morbidity and mortality in the industrialized world. Atherosclerosis develops in response to local endothelial injuries. Endothelial dysfunction and cell loss are prominent features in atherosclerosis. Restoring the endothelial lining to normal is critical for slowing or reversing the progression of atherosclerosis. Increasing data suggest that endothelial progenitor cells (EPCs) play a significant role in reendothelialization of the injured blood vessels. This review focuses on the effects of EPC mobilization and transfusion in the condition of atherosclerosis. The aim of the review is to provide an update on the progress in this research field, highlight the role of EPCs in atherosclerosis and discuss the possible mechanisms and potential risks of progenitor cell-based therapy in atherosclerosis.     

Timing of the vascular actions of estrogens in experimental and human studies: Why protective early,and not when delayed?

Estrogens, and in particular 17β-estradiol (E2), play a pivotal role in sexual development and reproduction and are also implicated in a large number of physiological processes including the cardiovascular system. Although epidemiological studies and Nurses' Health Study suggested, and all animal models of early atheroma clearly demonstrated a vasculoprotective action of both endogenous and exogenous estrogens, the Women's Health Initiative did not confirm the preventive action of estrogens against coronary heart disease (CHD). However, women who initiated hormone therapy closer to menopause tended to have reduced CHD risk compared with increased CHD risk among women more distant from menopause. Thus, it is now mandatory to try to understand the mechanisms that could have influenced the actions of estrogens at various stages of atherosclerosis and/or of life. In this current review, we will summarize our understanding of the potential cellular targets and mechanisms of the vasculoprotective actions of estrogens, as well as of the lack of action of estrogens when administered after a period of hormonal deprivation. The mechanisms of the aggravating role of progestogens such as medroxyprogesterone acetate will be considered. Finally, we will analyze the possibilities to uncouple some beneficial from other undesirable actions following the partial/selective activation of estrogen receptors.     

缺氧预处理外周间充质干细胞对兔血管成形术后再狭窄的影响

目的： 探讨经骨髓动员后外周血分离的间充质干细胞（MSCs）移植对动脉粥样硬化模型兔颈动脉球囊成形术后再狭窄的防治作用及可能机制。方法： 粒细胞集落刺激因子（G-CSF）动员、外周血分离体外扩增获得MSCs,增强型绿色荧光蛋白(EGFP)报告基因标记备用。建立兔动脉粥样硬化狭窄模型,随机分为缺氧预处理MSCs移植组(HP)、非缺氧预处理MSCs移植组（NHP）及对照组（C）,再于球囊成形术后立即耳缘静脉内分别注入标记的缺氧处理、非处理MSCs或培养液。术后7 d、14 d、28 d采集外周血用ELISA法检测血管内皮生长因子（VEGF）水平;术后28 d处死动物留取血管标本通过Weigert弹力纤维染色及免疫组化染色进行形态学分析、归巢MSCs分析、再内皮化程度分析。结果: HP组、NHP组术后各时点VEGF水平均显著高于C组(P<0.01);术后7 d、14 d HP组显著高于NHP组(P<0.05),28 d则差异无显著。术后7 d血管组织中仅HP组、NHP组血管组织中有EGFP（+）细胞分布;术后28 d HP组内膜增殖程度、再狭窄率明显低于NHP组(P<0.05),而该2组均显著低于C组(P<0.01);再内皮化程度HP组明显优于NHP组(P<0.05)、同时该2组又均显著优于C组(P<0.01)。结论： MSCs移植能够抑制模型兔颈动脉球囊成形术后内膜增生、促进损伤血管快速内皮化从而降低再狭窄率,移植经缺氧预处理的MSCs可增强上述作用,可能与缺氧应激诱导MSCs分泌细胞因子功能增强有关。