MOLECULAR BIOLOGICAL STUDIES ON THE MECHANISM OF CEREBROVASCULAR DEMENTIA

我们利用分子生物学技术建立实验动物模型，研究了脑血管性痴呆的发病机制。所得结果如下：(1)我们证明了脑缺血导致沙鼠和大鼠大脑中选择性易感神经元的基因表达改变。利用沙鼠缺血脑热休克蛋白70 （HSP70）和热休克同源蛋白70 (HSC70) cDNA探针进行原位杂交。在沙鼠中，HSP70 mRNA和HSP70蛋白的丢失先于CA1神经元死亡，这表明转录和翻译紊乱在缺血性神经元死亡中的作用。相反，面对HSP70的合成，大鼠出现神经元死亡。(2)我们还研究了亚致死缺血预处理后诱导缺血耐受的机制。脑内微透析测量的缺血时兴奋性氨基酸谷氨酸释放量未因预处理而改变。然而，HSP70的作用被认为是由于HSP70表达的时间谱与缺血耐受相似，并且由于抗HSP70抗体或槲皮素（一种HSP表达抑制剂）可以抑制耐受性。我们观察到耐受获得性脑中HSP70 mRNA和蛋白的诱导。在缺血耐受大鼠脑中，HSP27主要在神经胶质细胞中被诱导，提示通过神经元-神经胶质相互作用产生神经保护作用。因此，研究结果强烈提示脑缺血诱导神经元和神经胶质细胞的基因表达，从而激活直接和间接的神经保护系统。

We investigated the mechanisms of cerebrovascular dementia using experimental animal models with molecular biological techniques. Results obtained are as follows. (1) We demonstrated that cerebral ischemia gives rise to altered gene expressions in selectively vulnerable neurons in brains of gerbils and rats. We used cDNA probes for heat shock protein-70 (HSP70) and heat shock cognate protein-70 (HSC70) cloned from ischemic gerbil brains and performed in situ hybridization. In gerbils, loss of HSP70 mRNA and HSP70 protein preceded CA1 neuronal death, which suggested the role of transcriptional and translational disturbances in ischemic neuronal death. In contrast, neuronal death occurred in face of HSP70 synthesis in the rat. (2) We also investigated the mechanisms of ischemic tolerance that is induced after preconditioning with sublethal ischemia. The amount of excitotory amino acid glutamate released during ischemia as measured with intracerebral microdialysis was not altered by preconditioning. However, the role of HSP70 was suggested because temporal profile of HSP70 expression was similar to that of ischemic tolerance and because the tolerance was inhibited by treatment with anti-HSP70 antibody or quercetin, an inhibitor of HSP expression. We observed facilitated induction of HSP70 mRNA and protein in the tolerance-acquired brain. In rat brain with ischemic tolerance, HSP27 was induced predominantly in glial cells, suggesting neuroprotection through neuron-glia interaction. Thus, the findings strongly suggest that cerebral ischemia induced gene expressions in neurons and glial cells, thereby activating direct and indirect systems of neuroprotection against injury.