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ISSN 1672-9145 Acta Biochim Biophys Sin 2005, 37(5): 349–354 CN 31-1940/Q

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Prolonged Alzheimer-like Tau Hyperphosphorylation Induced by Simultaneous Inhibition of Phosphoinositol-3 Kinase and Protein Kinase C in N2a cells

Guo-Gang XU, Yan-Qiu DENG, Shi-Jie LIU, Hong-Lian LI, and Jian-Zhi WANG*

Department of Pathophysiology, Institute of Neuroscience, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China

Abstract Co-injection of wortmannin (inhibitor of phosphatidylinositol-3 kinase, PI3K) and GF109203X (inhibitor of protein kinase C, PKC) into the rat brain was found to induce spatial memory deficiency and enhance tau hyperphosphorylation in the hippocampus of rat brain. To establish a cell model with durative Alzheimer-like tau hyperphosphorylation in this study, we treated N2a neuroblastoma cells with wortmannin and GF109203X separately and simultaneously, and measured the glycogen synthase kinase 3 (GSK-3) activity by g-³²P-labeling and the level of tau phosphorylation by Western blotting. It was found that the application of wortmannin alone only transitorily increased the activity of GSK-3 (about 1 h) and the level of tau hyperphosphorylation at Ser³⁹⁶/Ser⁴⁰⁴ and Ser¹⁹⁹/Ser²⁰² sites (no longer than 3 h); however, a prolonged and intense activation of GSK-3 (over 12 h) and enhanced tau hyperphosphorylation (about 24 h) were observed when these two selective kinase inhibitors were applied together. We conclude that the simultaneous inhibition of PI3K and PKC can induce GSK-3 overactivation, and further strengthen and prolong the Alzheimer-like tau hyperphosphorylation in N2a cells, suggesting the establishment of a cell model with early pathological events of Alzheimer’s disease.

Key words Alzheimer’s disease (AD); tau; glycogen synthase kinase 3 (GSK-3); protein kinase C (PKC); phosphatidylinositol-3 kinase (PI3K); N2a cells

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Received: January 13, 2005 Accepted: March 11, 2005

This work was supported by grants from the National Natural Science Foundation of China (30430270, 30472030 and 30400068), Science and Technology Committee of China (G1999054007) and National Educational Committee of China (2001-171)

*Corresponding author: Tel, 86-27-83692625; Fax, 86-27-83693883; E-mail, [email protected]