通心络对IL-1β介导的小型猪冠状动脉内膜增殖及5-HT诱发的冠状动脉痉挛的抑制作用

Inhibitory Effect and Acting Mechanism of Tongxinluo on IL-1β-mediated Coronary Intimal Hyperplasia and 5-hydroxytryptamine-Induced Coronary Vasospasm in Small Swine

目的在体观察通心络抑制猪冠状动脉痉挛的作用及可能机制。方法选取16头小型雄性家猪,麻醉后进行左侧开胸手术,选择心脏左前降支及回旋支近端管腔外径大小近似的两处节段进行仔细分离,在血管外膜包裹吸附白细胞介素-1β(IL-1β,2.5μg)的纸巾,构建冠状动脉痉挛模型,2周后采用冠状动脉造影观察经导管冠脉内给予5-羟色胺(5-HT,10μg/kg)诱发冠状动脉痉挛情况。将成功诱发冠状动脉痉挛的12头小型猪随机分成两组,对照组6只(12个血管段),单纯喂养专用饲料;通心络组6只(12个血管段),喂养专用饲料和通心络〔1g/(kg.d)〕,连续喂养4周后停药1周,进行第2次造影并以同样方法进行诱发痉挛实验。造影结束后处死动物,截取包裹IL-1β的血管段,进行病理学检查,并采用RT-PCR方法测定各组手术处理血管段(Ras hom ologue,Rho)激酶mRNA表达,采用Western blot方法测定其底物肌球蛋白结合亚基磷酸化(myosin-binding subunit phosphorylation,MBS-P)的表达。结果冠状动脉造影显示,12头小型猪冠状动脉局部血管发生不同程度的管腔狭窄(20%～30%),5-HT能诱发其痉挛。第2次造影显示,对照组11血管段5-HT诱发痉挛阳性,1血管段5-HT诱发痉挛阴性。通心络组2血管段5-HT诱发痉挛阳性,10血管段5-HT诱发痉挛阴性。病理学检查,两组均可见不同程度的巨噬细胞聚集现象,通心络组与对照组比较管腔狭窄程度及内膜增殖明显减轻。对照组Rho激酶mRNA的表达及肌球蛋白磷酸酶(MLCP)的MBP-S水平明显上调,通心络组Rho激酶mRNA表达〔(71.5±2.4)%〕被抑制,与对照组〔(98.2±7.7)%〕比较差异有统计学意义(P<0.05),通心络组MBS-P的灰度值(16633±1390)明显降低,与对照组(25818±4745)比较,差异有统计学意义(P<0.05)。结论通心络具有明显抑制IL-1β介导的冠状动脉内膜增殖及5-HT诱发的冠状动脉痉挛的作用;抑制IL-1β包裹血管段细胞内Rho激酶mRNA表达,减少MBS-P表达可能是其作用机制之一。

Objective To observe the inhibitory effect of Tongxinluo （TXL） on coronary vaso spasm in small swine in vivo, and to investigate its possible acting mechanism. Methods The model of coronary atherosclerosis in 16 male small swines was established by left thoracotomy after anesthesia, isolated the sections of left anterio-descending branch and proximal end of rotator branch with similar outer diameter, and encapsulated them with papertowel holding 2.5 μg interleukin-1 β. Two weeks later, the condition of coronary vasospasm induced by catheter intra-coronary injection of 5-hydroxytryptamine （5-HT, 10 μg/kg） was observed through coronary artery contrast examination. The 12 swines with successfully formed coronary vaso spasm were randomly divided into 2 groups, the TXL group and the control group. They were fed with special diet, but TXL 1 g/（kg · d） was administered additionally to the TXL group for 4 weeks. The observation on coronary vasospasm was repeated ! week after discontinuation of TXL treatment, then the animals were sacrificed, their vascular sections enclosed with IL-1β was taken to conduet the pathologic examination and to deteet the expressions of Rho kinase mRNA and its substrate myosin-binding subunit phosphorylation （MBS-P） by RT-PCR and Western blot method. Results Coronary artery contrast showed that local coronary stenosis occurred in the 12 model swines to different extents （20% -30% ） , and vascular spasm on them could be induced by 5-HT. At the time of repeating examination, 11 vascular sections in the control group still maintain their positive spasm reaction to 5-HT, but only 2 in the TXL group did so, the reaction turned to negative in 1 and 10 in the two groups respectively. Pathological examination showed that different degrees of macrophage aggregation could be found in both groups. The degree of lumen stricture and endometrial hyperplasia in the TXL group was obviously attenuated than those in the control group. The expressions of Rho kinase mRNA and MBS-P in the control group were up-regulated obviously. As compared with those in the control group, they were inhibited significantly in the TXL group, as （71.5 ± 2. 4） vs （98.2 ± 7.7）% and 16 633 ± 1 390 vs 25 818 ± 4 745, respectively （ all P 〈 0.05 ）. Conclusion TXL could obviously inhibit the coronary intimal hyperplasia mediated by IL-1β and coronary vasospasm induced by 5-HT, one of its mechanisms is possibly the inhibition on the intracellular Rho kinase mRNA expression in the IL-1β enclosed vascular section to decrease the level of MBS-P.