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）具有一个PKC系统，即使在无Ca^2+的介质中也能充分发挥其功能，并且在体内或体外条件下应用PKC抑制剂如H-7和星形孢菌素（staurosporine）可逆转“二次出血”引起的BA狭窄或张力增强。2]与非痉挛性BA相比，痉挛性BA中的磷脂酰胆碱和磷脂酰乙醇胺的刺激周转显著增加了内源性PKC激活剂1，2-二酰甘油的含量。[3]在痉挛性动脉中，与非痉挛性对照动脉相比，在第4和第7天，胞浆PKC活性显示出40-45%的下降，而膜PKC活性没有显著变化。20 kDa肌球蛋白轻链磷酸化程度 关于我们 在第14天，随着血管痉挛的缓解，动脉细胞浆PKC活性恢复到正常对照水平。Western blot分析显示，在痉挛血管中PKCα和PKCε表达下调，而PKC β表达无明显变化，提示PKCα和PKCε在慢性血管痉挛过程中激活。4]可能由PKC激活引起的主动平滑肌收缩的增强在慢性血管痉挛的发生中起作用，这比动脉壁的被动物理性质改变（动脉壁的可分配性降低）更重要。5]在pCa=5.5时，用棕榈酰-花生四烯酰二酰基甘油-氢过氧化物（5 × 10 - 4 M）孵育带皮基底动脉<-5>，显示PKC α和e的显著下调。由于已知蛋白质的下调是显示其先前激活的直接证据，因此认为氧化二酰甘油诱导的收缩是由PKC α或ε的激活介导的。6]最后，PAG=OH和PLG-OH作为氧化DAG被发现在实验性蛛网膜下腔出血的脑组织中。上述研究表明，PKC激活，特别是在平滑肌收缩的钙非依赖性机制中发挥核心作用的e型PKC激活，可能参与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