血管内皮靶向微泡对兔心肌缺血再灌注损伤的超声分子显像

Ultrasound Molecular Imaging of Vascnlar Endothelial Targeted Microbubbles on Myocardial Ischemiu Repertusion Injury in Rabbit

目的:构建针对血管内皮细胞损伤的靶向微泡,实现对兔心肌缺血再灌注损伤的靶向显像。方法:构建血管内皮细胞间黏附分子-1(ICAM-1)靶向阳离子微泡(TCMB)和非靶向普通阳离子微泡(CMB),肿瘤坏死因子α(TNF-α)活化培养至70%融合的原代人脐静脉内皮细胞(HUVEC)/脐静脉,CMB和TCMB分别以2.0dynes/cm^2的剪切应力通过平行板流动腔连接的细胞培养皿/脐静脉,比较CMB和TCMB对HUVEC/脐静脉的靶向粘附能力。20只新西兰大耳兔随机分为CMB组和TCMB组,每组各10只,参照相关文献方法制备心肌缺血再灌注损伤模型,耳缘静脉注射1ml浓度均为2×10~8个/ml的CMB(CMB组)和FITC二抗标记的TCMB(TCMB组),比较CMB和TCMB对兔心肌缺血再灌注损伤的靶向显像能力。结果:CMB与TCMB的粒径差异无统计学意义(2.90±1.90μm vs 3.10±2.10μm,P>0.05),CMB表面电位高于TCMB(+31.30±3.90mV vs+22.00±2.40mV,P<0.01)。与活化血管内皮细胞粘附的TCMB数量是CMB的19.4倍,与脐静脉粘附的TCMB声强度是CMB的3倍,差异均有统计学意义(P<0.01)。TCMB可在兔心肌缺血再灌注损伤区域的冠脉内定向聚集,实现靶向超声分子显像,而CMB无明显靶向显像功能;TCMB组兔损伤区域心肌声强比是CMB组的1.86倍,差异有统计学意义(P<0.01)。结论:ICAM-1靶向微泡能与损伤的血管内皮细胞特异性粘附,实现对心肌缺血再灌注损伤的靶向超声分子显像。

Objective：To construct the vascular endothelium-targeted microbubbles for the target ultrasound imaging of myocardial ischemia reperfusion injury in rabbits. Method.. To construct the vascular endothelium-targeted microbubble （TCMB） by connecting the intercellular adhesion molecule-1 （ICAM-1） antibody to the surface of the cationic microbubble using biotin-streptavidin system. The non-targeted cationic microbubble （CMB） was served as the control. Primary human umbilical vein endothelial cells （HUVECs, 70M confluence） and umbilical vein were activated with 50 ng/ml tumor necrosis faetor-a （TNF-a）. CMBs and TCMBs were passed across the HUVECs and umbilical vein at a shear stress of 2 dynes/cm^2 to compare the targeted adhesion ability of the two microbubbles. Twenty rabbits were constructed myocardial isehemia reperfusion injury animal model and randomly divided into two groups： CMB group and TCMB group, n= 10 per group. One milliliter of CMBs and FITC-IgG labeled TCMBs were infused through the ear vein to compare the target imaging ability of the two for myocardial isehemia reperfusion injury in rabbits. Results.. The average diameters of the CMB and TCMB were no statistic difference （2. 90±1.90μm vs 3.10±2.10μm, P〉0.05） and the zeta potential of the CMB was higher than that of TCMB （＋31.30±3.90mV vs ＋22. 00±2.40mV, P〈0. 01）. The amount of TCMB targeted adhesion to vascular endothelial cells and umbilical vein was 19.4- fold and 3-fold to CMB, respectively（P〈0.01）. TCMB can accumulate in the coronary arteries of myocardial ischemia reperfu- sion injury area for targeted ultrasound molecular imaging, while CMB has no obvious target imaging function. The acoustic intensity ratio of TCMB in the injury area was 1.86 fold of CMB （P〈0.01）. Conclusion： ICAM-1 targeted microbubble can specifically adhere and aggregate with injury vascular endothelial cells, which can be used for targeted ultrasonic molecular imaging of myocardial ischemia reperfusion injury.