Possible role of protein kinase-dependent smooth muscle contractile mechanism in the occurrence of delayed spasm after subarachnoid hemorrhage

我们已经报道了以下实验证据，表明PKC（蛋白激酶C）激活参与蛛网膜下腔出血后慢性血管痉挛的发生。[1]犬基底动脉（BA）具有PKC系统，即使在无Ca^<2+>的培养基中也能充分发挥作用，并且在体内或体外条件下，通过应用PKC抑制剂（如H-7和staurosporine），可以逆转暴露于“双出血”的BA的狭窄或张力增强。[2]与非痉挛动脉相比，痉挛动脉中磷脂酰胆碱和磷脂酰乙醇胺的转化刺激了PKC内在激活剂1,2-二酰基甘油的含量显著增加。[3]与非痉挛动脉相比，痉挛动脉在第4天和第7天，胞浆PKC活性下降了40-45%，而膜PKC活性没有显著变化。20 kDa肌球蛋白轻链磷酸化的程度在痉挛和非痉挛BA之间没有变化。第14天，随着血管痉挛的缓解，动脉胞浆PKC活性恢复到正常控制水平。Western blot分析PKC亚型发现，痉挛动脉中PKCα和PKCε表达下调，PKCζ表达下调，提示PKCα和PKCε在慢性血管痉挛过程中被激活。4]主动平滑肌收缩的增强，可能是由PKC激活引起的，在慢性血管痉挛的发生中起作用，这比动脉壁被动物理性质的改变（动脉壁的可有可无性降低）更重要。[5]此外，氧化二酰基甘油诱导正常和剥皮BA持续收缩。在pCa=5.5下，用棕榈酰-花生四烯基二酰基甘油-过氧化氢（5x10 ^<-5>M）培养剥皮的基底动脉显示PKC a和e的显著下调。众所周知，蛋白的下调是显示其先前激活的直接证据，氧化二酰基甘油诱导的收缩被认为是由PKC a或ε的激活介导的。[6]最后，在实验性蛛网膜下腔出血脑组织中发现PAG=OH和PLG-OH作为氧化DAG。上述研究表明，PKC的激活，尤其是在平滑肌收缩的不依赖钙的机制中发挥核心作用的e型PKC激活，可能参与了蛛网膜下腔出血后慢性血管痉挛的发生

We have already reported the following experimental evidence suggesting that PKC (protein kinase C) activation participates in the occurrence of chronic vasospasm following subarachnoid hemorrhage. 1] The canine basilar artery (BA) possesses a PKC system which fully functions even in the Ca^<2+>-free medium, and that the narrowing or the augmented tonus of the BA exposed to 'two-hemorrhage' can be reversed by application of PKC inhibitors such as H-7 and staurosporine either in vivo or in vitro condition. 2] The content of 1,2-diacylglycerol, an intrinsic PKC activator, is significantly increased through the stimulated turnover of phosphatidylcholine and phosphatidylethanolamine in the spastic BA in comparison with that in the nonspastic BA.3] In spastic arteries at Days 4 and 7, the cytosolic PKC activity showed a decrease of 40-45% with no significant changes in membrane PKC activity as compared with nonspastic control arteries. The extent of 20 kDa myosin light chain phosphorylation … More was unchanged between the spastic and nonspastic BA.On Day 14, the cytosolic PKC activity of arteries returned toward the normal control level with the remission of vasospasm. Western blot analysis of the PKC isoforms revealed that the amounts of PKCα and PKCε but not PKCζ were down-regulated in spastic arteries, suggesting the activation of PKCα and PKCε in the course of chronic vasospasm. 4] The augmentation of active smooth muscle contraction, possibly induced by PKC activation, plays a role in the occurence of chronic vasospasm which is more important than the altered passive physical properties of the arterial wall (decreased dispensability of arterial wall). 5] In addition, it is revealed that oxidized diacylglycerol induces sustained contraction of normal and skinned BA.Under the pCa=5.5, an incubation of skinned basilar artery with palmitoyl-arachidonyl diacylglycerol-hydroperoxide (5xl0^<-5>M) shows a remarkable down-regulation of PKC a and e. As it is known that the down-regulation of the protein is the direct evidence to show its preceding activation, the oxidized-diacylglycerol induced contraction is thought to be mediated by an activation of PKC a orε. 6] Finally, PAG=OH and PLG-OH as an oxidized DAG were found in the brain tissue subjected to experimental subarachnoid hemorrhage. The above line of investigations render it likely that PKC activation, especially type e which is though to play a central role in the calciun-independent mechanism of smooth muscle contraction, participates in the occurrence of chronic vasospasm following subarachnoid hemorrhage Less