【作者】 林勇；

【导师】 汪钟；

【作者基本信息】 中国协和医科大学 ， 心血管药理学， 1998， 博士

【摘要】 白细胞与血小板、内皮细胞及平滑肌细胞的粘附是多种急、慢性心血管疾病发生、发展的重要环节。这些心血管疾病包括动脉粥样硬化、血栓形成、缺血再灌损伤、经皮腔内血管成形术后血管再狭窄等。抑制病理状态下细胞与细胞间的相互粘附能有效地阻止或降低急、慢性炎症引起的组织损伤，预防上述疾病的发生与发展。已知许多因素影响细胞间的粘附，对这些影响因素，特别是药物的研究，不论在基础理论还是在临床实践方面都有十分重要的意义。 正常人或动物的白细胞与血小板的粘附率很低，但急性心肌梗塞病人和动物病理模型的白细胞与血小板的粘附率则显著增加。研究结果表明：白细胞与血小板的激活是导致这两种细胞相互粘附的原因。凝血酶激活的血小板与白细胞的粘附率也显著增加。白细胞与血小板的粘附是钙依赖性的，肝素、硫酸右旋糖苷可抑制心肌梗塞大鼠白细胞与血小板的粘附，而不含硫酸根的右旋糖苷则无抑制作用。提示GMP—140至少部分介导白细胞与血小板的粘附。急性心肌梗塞病人的白细胞CD11a、CD11b的表达明显高于正常人，表明粘附分子CD11a／CD18及CD11b／CD18可能参与白细胞与血小板的粘附。槲皮素在心肌梗塞形成前静脉给药，或在血小板被凝血酶激活之前体外给药均可抑制白细胞与血小板的粘附。阿司匹林也有类似的抑制作用。 本文结果显示，未激活的白细胞可抑制血小板聚集，阿司匹林及槲皮素均能加强白细胞对血小板激活的抑制作用。但当白细胞被PMA激活后，其抑制血小板聚集的作用则消失，如在PMA激活白细胞前加槲皮素与血小板孵育，白细胞对血小板聚集的抑制作用又可恢复，提示槲皮素可抑制PMA对白细胞的激活。 TNF_α激活的内皮细胞与白细胞粘附呈量效和时效关系。与内皮细胞粘附的单核细胞较中性粒细胞略多，且随刺激时间的延长作用更为明显。抗ICAM-1，E-selectin及VCAM-1单克隆抗体抑制白细胞与内皮细胞粘附，提示这些粘附分子介导上述细胞粘附。ELISA法及流式细胞术显示，静息状态的内皮细胞表面有少量的ICAM-1及VCAM-1表达，无E-selectin表达，但RT-PCR法却显示内皮细胞内含有这三种粘附分子的mRNA。TNF_α诱导／增加ICAM-1，E-selectin及VCAM-1的表达也呈量效和时效关系。然而，它们的时效曲线却各具特点。E-selectin起效最快，达峰时间最短，为4～6小时，12小时已基本消退。ICAM-1表达较E-selectin慢，2小时略有增加，12小时达高峰，24小时更多还原

【Abstract】 Adhesion of leukocyte to platelet, endothelial cell （EC） and vascular smooth muscle cell （SMC）, is an essential component of various acute and chronic cardiovascular diseases including atherosclerosis, thrombosis, reperfusion injury and restenosis after percutaneous transluminal coronary angioplasty etc. Inhibition of cells adhesion of pathological status is effective in reducing the tissue injury induced by acute or chronic inflammations and in preventing the genesis and progress of the diseases mentioned above. It is known that several factors influence the cells adhesion. To study these regulating factors, especially drugs, may be of considerable importance for basic theory and clinical application.The adhesion of leukocyte/platelet was very low in healthy volunteers and normal animals. It was increased dramatically in patients with acute myocardial ischaemia （AMI） or experimental pathological animal models. The results indicated that the activation of both leukocyte and platelet played an important role in increasing both cells adhesion in AMI. The leukocyte/platelet adhesion rate was also increased when platelets were stimulated by thrombin in vitro. The adhesion of leukocyte/platelet was Ca²⁺ dependent, heparin or dextran sulfate inhibited both cells binding but not dextran. This indicates that adhesion molecule—GMP-140 might be involved in the mechanism of leukocyte/platelet adhesion. In addition, the expression of CDlla and CD 11b on the surface of leukocyte in AMI patients was significantly increased than normal subjects. It suggests that the adhesion molecules— CD11a/CD18 and CD11b/CD18 may mediate leukocyte/platelet adhesioaQuercetin inhibited the leukocyte/platelet adhesion in AMI animals when administered intravenously before the formation of AMI model or before the platelet stimulated by thrombin. Aspirin had similar effects as quercetin.The results showed that unactivated leukocyte inhibited platelets aggregation. Both quercetin and aspirin enhanced the inhibitory effects of leukocyte on platelet activation. However, the inhibition of platelet aggregation was disappeared when leukocyte was activated by PMA. It is interested to find out that if incubation of quercetin with platelets before activation of leukocyte by PMA, the inhibitory effect of leukocyte on platelet aggregation was restored. It indicates that quercetin has the ability to prevent the activation of leukocyte by PMA.TNF_α increased the adhesion of EC to leukocyte both time- and dose-dependently. More monocytes bound to EC than neutrophils, and it became more significant with the prolongation of stimulation time. Anti-ICAM-1, anti-E-selectin and anti-VCAM-1 monoclonal antibodies blocked the binding of TNF_α stimulated EC to leukocyte, suggesting that the adhesion molecules, ICAM-1, E-selectin and VCAM-1, mediated both cells adhesion.ELISA and flow cytometric analysis revealed that the expression of ICAM-1 and更多还原