组织型纤溶酶原激活剂及其抑制剂-1在兔深静脉血栓模型股静脉壁中的表达

摘要

目的 观察静脉壁中纤溶相关因子组织型纤溶酶原激活剂(t-PA)、组织型纤溶酶原激活物抑制剂-1(PAI-1)在深静脉血栓(DVT)形成及消退过程中的动态表达变化,揭示t-PA、PAI-1在DVT成和消退中的作用.方法 50只体质量为2.0~2.5 kg的日本大耳白兔随机分为正常对照组(n=4)和模型组(n=46).模型组采用钳夹双侧股静脉+石膏固定双下肢建立兔DVT模型,建模后每24 h行B超监测血栓形成状况.根据建模后预设的时间点及血栓状态的不同,再将模型组分为8个亚组:建模后4 h、8 h、12 h、1 d(无血栓形成)、1 d(有血栓形成)、3 d、5 d、7 d;在各相应的时间点取4只兔处死切取股静脉壁,用SYBR Green I 核酸凝胶染液荧光实时定量PCR及ELISA检测股静脉组织中t-PA、PAI-1 mRNA、蛋白在兔DVT模型建模后的表达.结果 模型组有1只兔因麻醉过量死亡,49只进入实验组.创伤后24 h,经B超监测,模型组血栓形成率为72.73%(24只);血栓栓子随造模后时间延长逐渐缩小机化;从第5天开始经B超监测模型组血栓形成的血管腔有断续血流信号.与对照组相比,t-PA mRNA、蛋白在造模后4、8、12 h呈逐渐下降趋势(P＜0.05),12 h下降到最低点(P＜0.01),在血栓形成后1 d升高至对照组水平(P＞0.05),3、5 d继续升高(P＜0.05),5 d达到高峰(P＜0.01),7 d再次回落至对照组水平(P＞0.05);与t-PA变化相反,PAI-1 mRNA、蛋白在造模后4、8、12 h呈逐渐升高趋势(P＜0.05),12 h达到最高点(P＜0.01),血栓形成后1、3、5 d逐渐下降(P＜0.05),3 d下降至对照组水平(P＞0.05),5 d到最低点(P＜0.01),7 d再次升高至对照组水平(P＞0.05).结论 t-PA、PAI-1参与了DVT形成过程;在血栓形成后,t-PA、PAI-1参与了血栓的溶解、机化再通过程.%Objective This sludy was lo establish rabbil LraumaLic deep vein ihrombosis model, and on ihe basis of model Lo observale ihe expression of L - PA and PAI - 1 of venous wall in ihrombosis and resolution, preliminarily reveal ihe role of I - PA and PAI - 1 in ihe deep vein ihrombosis and resolution. Methods 50 rabbils which is belween 2. 0 ~ 2. 5 kg weighl randomly divided inlo normal conlrol group (re =4) and model group (re =46). The rabbils belong lo model group were established rabbil deep vein ihrombosis model by clamping bilaleral femoral venous + fixing plasler casl of lower limbs, ihe modeling were moniloring ihrombosis condition every 24 h by B ullrasonic. According lo iheLime and ihrombosis after modeling of ihe slale, ihe model group was divided inlo 8 subgroups; posl - establishing 4 li, 8 h, 12 h, 1 d ( no ihrombosis) , Id ( ihrombosis) , 3 d, 5 d, 7 d. In each of ihe lime poinls lake 4 rabbils lo pul lo dealh and harvesl femoral vein wall, wilh SYBR (R) Green I fluorescence real - lime quanlilalive PCR and ELISA dynamic deleclion ihe change of expression aboul I - PA and PAI - 1 mRNA and prolein of femoral vein organizalion in ihe rabbil deep vein ihrombosis afler modeling. Results There was one rabbil of model group dead because of anesthesia, experimental group had 45 raabbils. Posl - Iraumalic 24 h, model group ihrombosis rale was 72. 73% (24) by B - ullrasonography moniloring. Thromboembolus gradually reduced and be organizalion along lime; from ihe 5 day il could be moniloring inlermillenl flow signals in model group ihrombosis vascular by B - ullrasonography. Afler establishing model, ihe I - PA in 4 h, 8 h, 12 h declined ( P < 0. 05 ) , inl d, 3 d, 5 d was become gradually increasing ( P < 0. 05 ) , 7 d is lo ihe control group level ( P > 0. 05 ) , and PAI - 1 expression in 4 h,8 h, 12 h increased ( P < 0. 05 ) , in 1 d, 3 d, 5 d was become gradually declined (P < 0. 05 ) , 7 d is lo the control group level ( P > 0. 05). Conclusion The results demonsIrated lhat I - PA and PAI - 1 involved in the Iraumalic deep vein ihrombosis process; In thrombosis, I - PA and PAI - 1 involved in the dissolution and organization of ihrombi.