Mechanism of NFT formation

我们构建了PDGF-β启动子驱动的过表达人4重复tau的小鼠，该启动子含有V337M额颞叶痴呆-17（FTDP-17）突变。表达量约为内源性小鼠tau的5-10%，并且在11月龄时在海马CA2和3区和大脑皮层中可见超磷酸化tau的积累。在这个年龄，Tg小鼠体内的tau不溶于SDS，形成丝状结构，随着年龄的增长，Tg小鼠可以形成NFT。电生理实验观察到海马神经元活动减弱。此外，Tg小鼠在升高加迷宫试验中未表现出习惯化，且在张开臂上停留时间明显延长。因此，明确了NFT的形成降低了海马的神经活动并导致了行为障碍。另一种tau Tg小鼠在CAMK II启动子控制下过表达带有R406W突变的人4重复tau时，小鼠模型也表现出NFT样病理并伴有记忆缺陷，其病理生理特征与AD和R406W患者非常相似。此外，我们在神经元中发现了GSK-3b和JNK的激活，这些神经元积累了过度磷酸化的tau，这表明GSK-3和JNK可能参与了NFT的形成。为了证实GSK-3和JNK参与NFT的形成，我们在COS7细胞中表达了JNK、deltaMEKK、GSK-3b和人tau。Tau在12个磷酸化位点显示磷酸化增加，包括AT100免疫反应位点，这是AD中PHF-tau的特殊抗原。SDS不溶性材料显示抗磷酸化tau免疫反应聚集体。因此，GSK-3 β和JNK对Tau的过度磷酸化参与了NFT的形成。另外，当GSK-3被β淀粉样蛋白激活时，tau TG小鼠导致神经原纤维缠结形成，突触丢失，神经元死亡。

We generated the mouse overexpressing human 4 repeat tau driven by PDGF-β promoter, which contains V337M frontotemporal dementia-17(FTDP-17) mutation. The amount of expression was about 5-10% of endogenous mouse tau, and the accumulation of hyper-phosphorylated tau was seen in the hippocampus CA2 and 3 region and the cerebral cortex at 11 month old. At this age tau in Tg mouse became insoluble for SDS and form filamentous structure, and Tg mouse could form NFT with aging. The decrease in the neuronal activity in the hippocampus was observed in the electrophysiological experiments. In addition, Tg mice didn't show habituation, and significantly stay longer in open arm in elevated plus maze test. Thus, it was clarified that NFT formation reduced neural activity in hippocampus and caused behavioral disorder.In case of the other tau Tg mouse overexpressed human 4 repeat tau with the R406W mutation under control of CAMK II promoter, the mouse model also showed NFT like pathology accompanying with memory deficit, which pathophysiological features are very similar to the patient of AD and R406W. In addition, we could find the activation of GSK-3b and JNK in neurons, which accumulate hyperphosphorylated tau, suggesting that GSK-3 and JNK might be involved in NFT formation.To confirm the involvement of GSK-3 and JNK on NFT formation, we expressed JNK, deltaMEKK, GSK-3b, and human tau in COS7 cells. Tau showed the increased phosphorylation at 12 phosphorylation sites, including AT100 immunoreactive site, which is known as a peculiar antigen of PHF-tau seen in AD. SDS insoluble materials showed anti-phospho-tau immunoreactive aggregates. Thus, an excessive phosphorylation of Tau by GSK-3 β and JNK took part in NFT formation. In addition, when GSK-3 was activated by β amyloid, tau TG mouse resulted in the neurofibrillary tangle formation, the synapse loss, and the neuron death.