Attempt to develop animal model for the formation of paired helical filaments in the brains of Alzheimer's disease patients via sustained inactivation of integrin-linked kinase in mouse brain.

整合素连接激酶（ILK）是一种粘着斑丝氨酸/苏氨酸蛋白激酶，在整合素和生长因子信号通路中具有重要作用。最近，我们证明ILK失活导致异常tau磷酸化，这与阿尔茨海默病患者脑中成对螺旋丝中的一些异常磷酸化位点相同，通过培养的N1 E-115细胞中GSK-3β的持续激活。在这项研究中，我们研究了ILK的持续失活是否会导致异常的tau蛋白磷酸化，并在小鼠脑中产生成对的螺旋丝。ILK的激酶缺陷型突变体（DN-ILK）在小鼠脑内的转染和表达导致ILK活性的抑制和活性形式GSK-3β的增加，并在转染后3 wk诱导海马中异常的tau磷酸化。然而，由DN-ILK诱导的内源性ILK活性的抑制不被保留，并且通过DN-ILK转染后12周内源性ILK蛋白的显著增加而恢复到基础水平。这种代偿机制导致GSK-3β的活性形式恢复到正常水平，并导致异常tau磷酸化水平降低。这些结果强烈表明，ILK通过抑制GSK-3β活性在体内保护免于异常tau磷酸化。虽然我们不能在阿尔茨海默病患者的大脑中建立成对螺旋丝的小鼠模型，但我们的结果表明，激活ILK活性的药物可能对阿尔茨海默病早期患者的治疗有效。ILK在脑内失活后的代偿机制有待进一步研究。

Integrin-linked kinase (ILK) is a focal adhesion serin/threonine protein kinase with an important role in integrin and growth factor signaling pathways. Recently, we demonstrated that ILK inactivation results in aberrant tau phosphorylation, which is identical to some of the aberrant phosphorylation sites in paired helical filaments in the brains of patients with Alzheimer's disease, via sustained activation of GSK-3β in cultured N1E-115 cells. In this study, we examined whether sustained inactivation of ILK leads to aberrant tau phosphorylation and produces paired helical filaments in mouse brain. Transfection and expression of a kinase-deficient mutant of ILK (DN-ILK), which behaves as a dominant negative, to mouse brain in vivo lead to inhibition of ILK activity and increase in active form of GSK-3β, and then induce aberrant tau phosphorylation in the hippocampus 3 wk after the transfection. However, the inhibition of endogenous ILK activity induced by DN-ILK is not retained and recovered to the basal level via a marked increase of endogenous ILK protein 12 wk after DN-ILK transfection. This compensatory mechanism leads the active form of GSK-3β to normal level and results in a decrease in the levels of aberrant tau phosphorylation. These results strongly suggest that ILK protects against aberrant tau phosphorylation via inhibition of GSK-3β activity in vivo. Although we could not create the mouse model for paired helical filaments in the brains of patients with Alzheimer's disease, our results imply that medical drugs to activate ILK activity might be effective in the therapy of patients with the early stage of Alzheimer's disease. Further studies are required to elucidate the compensatory mechanism after ILK inactivation in the brain.