猪冠状动脉侧支循环生成的免疫组化研究

Immunohistochemical analysis of coronary collateral circulation development in pigs

目的:探讨免疫组化法分析猪冠状动脉侧支循环生成的可行性,为冠心病康复的机制研究奠定方法学基础。方法:健康成年小型猪45只,建立可控性心肌缺血动物模型,分别设立假手术组、缺血组和运动组。缺血组予以单纯冠状动脉水囊充气诱发的短暂缺血,运动组除行缺血刺激外再给予平板有氧训练,共持续8周。实验终点时取冠状动脉钝缘支支配区心肌组织行心肌毛细血管密度测定:①常规HE染色后镜检计数;②免疫组化分析Ⅷ因子相关抗原标记并镜检计数。对两种方法检测毛细血管密度的重复性进行检验。结果:HE染色与免疫组化均显示运动组心肌缺血区毛细血管密度明显高于单纯缺血组(P<0.01)及假手术组(P<0.01),缺血组明显高于假手术组(P<0.01)。免疫组化检测的毛细血管密度显著高于HE检测(P<0.05)。两种方法的结果有较好的重复性(P>0.05),均与冠状动脉侧支血流量(CCBF)正相关,但免疫组化检测结果与CCBF相关系数显著高于HE(P<0.05)。结论:免疫组化可以准确有效地量化分析猪冠状动脉侧支血管状态。

Objective：To study the feasibility of applying immunohistochemical analysis in coronary collateral circulation development in mini pigs, which will be a methodological foundation in mechanism of cardiac rehabilitation. Method： An animal model of controllable myocardial ischemia was established in 45 mini pigs. The animals were equally divided into four groups, which were the sham-operated group （SO）, sedentary ischemic group （SI） and exercise training group （ET）. Repeated transient myocardial ischemia was induced by inflation of balloon constrictor or aerobic exercise training for 8 weeks. The myocardium attributed to the oblique marginal artery was finally sampled at the end point of the study for measurement of capillary density （CD）. The methods of the measurement were H.E. staining and immunohistochemical assay labeled by Factor Vm-RA. The reproductivity of the two methods for CD analysis was studied. Result： Both methods demonstrated significant increase of myocardial CD in ET group, and it was significantly higher than SI（P〈0.01） and SO（P〈0.01）. Furthermore, the myocardial CD in SI was also higher than SO （P〈0.01）. The CD analyzed by immunohistochemistry was higher than HE （P〈0.05）. The reproducibility between HE staining and immunohistochemical assay was excellent（P〉0.05）. Coronary collateral blood flow （CCBF） was positively correlated with CD analyzed by both methods. The correlation coefficient between immunohistochemical assay and CCBF was significantly higher than H.E. staining （P 〈0.05）. Conclusion： Immunohistochemical analysis is an accurate and reliable method for quantitative analysis of the coronary collateral circulation development in mini pig.