fgl2在急性心肌缺血再灌注冠脉微循环障碍中的作用及机制研究

Reperfusion therapy in acute myocardial infarction is a double-edged sword that it improves myocardial ischemia as well as causes cardiac microvascular injury, therefore may give rise to no-reflow or slow flow. To unravel the mechanism is an urgent and unsettled clinical problem. Recently, we observed in rats with acute myocardial ischemia/reperfusion (AMI/R) that a novel prothrombinase- - - - fgl2 was highly expressed in cardiac microvascular endothelial cells (CMVECs), and it may mediate the coronary microthrombosis in situ which leads to microcirculatory disturbance of AMI/R. We plan to perform this study in vitro and vivo by the following two techniques in which fgl2 siRNA and the anti-NPG-12 antibody (Patent Application Number: 201110307207.X) will be used to down-regulate fgl2 expression and to inhibit fgl2 activity respectively. Thus by decreasing the formation of thrombin through a possible novel coagulation pathway which is independent of the classical one, we expect three biological effects in cardiac microcirculation. Less microthrombosis in situ due to the less transformation from fibrinogen to fibrin, less hemocytic adhersion to CMVECs due to the downregulation of chemokine and adhesion molecule expression through the inhibiton of PAR-1 activation on endothelial cells, and less platelet aggregation due to the inhibition of platelet activation. In this study, the effects of fgl2 downregulation and inhibition in AMI/R rats will be investigated by histopathological, immunological and molecular biological techniques and the results may address the role of fgl2 in microcirculatory disturbance of AMI/R and provide a novel target for biological therapy in future.

急性心肌梗死再灌注治疗是一把双刃剑,既可改善心肌缺血,也可引起心脏微血管损伤,导致无复流或慢血流的发生,其发生机制是目前临床迫切需要解决的问题。我们近期在AMI/R大鼠模型中发现,一种新的凝血酶原酶fgl2异常高表达于心脏微血管内皮细胞，介导冠脉原位微血栓形成,其可能是导致冠脉微循环障碍重要原因。本研究拟从离体和在体两方面,通过siRNA下调fgl2表达及抗体(专利申请号:201110307207.X)抑制其活性,经由非经典凝血途径负向调控凝血酶生成。由此达到:减少其对纤维蛋白原激活,降低心脏原位微血栓的形成；减少其对内皮细胞PAR-1激活,降低趋化因子和黏附分子的表达，改善微循环中血细胞黏附；减少其对血小板活化,降低血小板聚集。利用组织病理学、免疫学和分子生物学技术进行检测评估,以明确fgl2介导AMI/R微循环障碍的作用和机制，为冠脉微循环障碍的治疗提供新的干预靶点和生物学手段。

急性心肌梗死再灌注治疗是一把双刃剑,既可改善心肌缺血,也可引起心脏微血管损伤,导致无复流或慢血流的发生,其发生机制是目前临床迫切需要解决的问题。我们近期在AMI/R大鼠模型中发现,一种新的凝血酶原酶fgl2异常高表达于心脏微血管内皮细胞，介导冠脉原位微血栓形成,其可能是导致冠脉微循环障碍重要原因。. 本研究从离体和在体两方面，利用组织病理学、免疫学和分子生物学等技术，通过基因沉默下调fgl2的表达以及Anti-NPG-12（专利申请号:201110307207.X）抑制 fgl2的活性，负向调控凝血酶，抑制冠脉原位微血栓形成，从而改善冠脉微循环障碍。. 目前我们已经成功完成NPG-12肽段的筛选和anti-NPG-12抗体的制备，并根据沉默效率最高的大鼠fgl2 siRNA，合成对应的人fgl2 siRNA，验证制备抗体的特异性，并通过该抗体靶向抑制fgl2的促凝活性，经由非经典凝血途径负向负向调控凝血酶的形成；此外，我们成功引进并繁殖fgl2基因敲除小鼠，建立心肌缺血再灌注模型，证实fgl2 表达缺失可以抑制心脏凝血酶和心脏纤维蛋白的沉积，下调PAR-1 的活化及其下游生物学效应，有效改善实验性冠脉无复流面积、炎性细胞浸润和心脏功能，进一步揭示fgl2介导AMI/R微循环障碍的作用和机制，从而为冠脉微循环障碍的治疗提供新的干预靶点和生物学手段。