Calpain activation and alpha-spectrin cleavage in rat brain by ethanol.
Calpain activation and alpha-spectrin cleavage in rat brain by ethanol.
复制标题
乙醇对大鼠脑中钙蛋白酶的激活和α-血影蛋白的裂解。
作者：
Yadavalli Rajgopal;M. Vemuri
钙蛋白酶是一种钙活化的半胱氨酸蛋白酶，与神经元变性和死亡有关。在这项研究中，我们利用体内慢性乙醇暴露的实验范式，表征了成年大鼠大脑皮层和小脑中的钙蛋白酶激活。乙醇处理增加了大鼠皮层和小脑的钙蛋白酶活性，但在皮层中增加的程度更高。Western blot分析显示m-calpain水平显著降低，而calpastatin水平不变。通过α -谱蛋白（fodrin）和蛋白激酶c - α (pkc - α)的蛋白水解进一步监测Calpain的活化。蛋白酶特异性谱蛋白分解产物显示calpain产生150和145 kda片段。此外，我们还在小脑中观察到一个120 kda的caspase-3激活片段。乙醇可降低皮质和小脑的pkc - α水平。乙醇处理后，Calpain的激活、α -谱蛋白裂解为Calpain特异性特征片段以及pkc - α蛋白水平的降低，证明了除了caspase-3介导的皮层和小脑细胞死亡外，Calpain也参与其中。考虑到钙蛋白酶在细胞死亡中的作用，本研究中钙蛋白酶活性增加后α -谱蛋白分裂表明钙蛋白酶在乙醇介导的细胞损伤和酒精性神经退行性变中是重要的效应物。
Calpain, a calcium-activated cysteine protease, has been implicated in neuronal degeneration and death. In this study, we have characterized calpain activation in adult rat cerebral cortex and cerebellum, using an experimental paradigm of in vivo chronic ethanol exposure. Ethanol treatment increased the calpain activity in cortex and cerebellum, but to a higher extent in the cortex. Western blot analysis revealed a significant decrease in m-calpain levels while calpastatin levels were unaltered. Calpain activation was further monitored by the proteolysis of alpha-spectrin (fodrin) and protein kinase C-alpha (PKC-alpha). Protease specific spectrin breakdown products revealed calpain generated 150- and 145-kDa fragments. In addition, we also observed a 120-kDa fragment characteristic of caspase-3 activation in the cerebellum. PKC-alpha levels were decreased in the cortex and cerebellum by ethanol. Calpain activation, cleavage of alpha-spectrin into calpain specific signature fragments and decreased PKC-alpha protein levels after ethanol treatment provide the evidence of calpain involvement besides caspase-3-mediated cell death in the cortex and cerebellum. Given the role of calpains in cell death, increased calpain activity followed by alpha-spectrin cleavage in this study suggests that calpains are important effectors in ethanol-mediated cell injury and alcoholic neurodegeneration.