Kandabashi T, Shimokawa H, Miyata K, Kunihiro I, Kawano Y, Fukata Y, Higo T, Egashira K, Takahashi S, Kaibuchi K, Takeshita A
Departments of Cardiovascular Medicine, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan.
Circulation. 2000 Mar 21;101(11):1319-23. doi: 10.1161/01.cir.101.11.1319.
We recently demonstrated that the Rho-kinase-mediated pathway plays an important role for coronary artery spasm in our porcine model with interleukin-1beta (IL-1beta). In this study, we examined whether or not Rho-kinase is upregulated at the spastic site and if so, how it induces vascular smooth muscle hypercontraction.
Segments of the left porcine coronary artery were chronically treated from the adventitia with IL-1beta-bound microbeads. Two weeks after the operation, as reported previously, intracoronary serotonin repeatedly induced coronary hypercontractions at the IL-1beta-treated site both in vivo and in vitro, which were markedly inhibited by Y-27632, one of the specific inhibitors of Rho-kinase. Reverse transcription-polymerase chain reaction analysis demonstrated that the expression of Rho-kinase mRNA was significantly increased in the spastic compared with the control segment. Western blot analysis showed that during the serotonin-induced contractions, the extent of phosphorylation of the myosin-binding subunit of myosin phosphatase (MBS), one of the major substrates of Rho-kinase, was significantly greater in the spastic than in the control segment and that the increase in MBS phosphorylations was also markedly inhibited by Y-27632. There was a highly significant correlation between the extent of MBS phosphorylations and that of contractions.
These results indicate that Rho-kinase is upregulated at the spastic site and plays a key role in inducing vascular smooth muscle hypercontraction by inhibiting myosin phosphatase through the phosphorylation of MBS in our porcine model.
我们最近证明，在我们用白细胞介素-1β（IL-1β）构建的猪模型中，Rho激酶介导的信号通路在冠状动脉痉挛中起重要作用。在本研究中，我们检测了Rho激酶在痉挛部位是否上调，如果上调，它又是如何诱导血管平滑肌过度收缩的。
用结合有IL-1β的微珠从外膜对猪左冠状动脉节段进行长期处理。如先前报道，术后两周，冠状动脉内注射血清素在体内和体外均能反复诱导IL-1β处理部位的冠状动脉过度收缩，而这被Rho激酶的特异性抑制剂之一Y-27632显著抑制。逆转录-聚合酶链反应分析表明，与对照节段相比，痉挛部位Rho激酶mRNA的表达显著增加。蛋白质印迹分析显示，在血清素诱导的收缩过程中，Rho激酶的主要底物之一肌球蛋白磷酸酶的肌球蛋白结合亚基（MBS）的磷酸化程度在痉挛节段显著高于对照节段，并且Y-27632也显著抑制了MBS磷酸化的增加。MBS磷酸化程度与收缩程度之间存在高度显著的相关性。
这些结果表明，在我们的猪模型中，Rho激酶在痉挛部位上调，并通过使MBS磷酸化来抑制肌球蛋白磷酸酶，从而在诱导血管平滑肌过度收缩中起关键作用。
