Donepezil modulates amyloid precursor protein endocytosis and reduction by up-regulation of SNX33 expression in primary cortical neurons

Yuki Takada-Takatori,Tetsuro Urushidani,Akinori Akaike,Yumiko Mizukawa,Yousuke Nao,Riko Kimata,Yasuhiko Izumi,Katsuharu Tsuchida,Toshiaki Kume,Shota Nakagawa

doi:10.1038/s41598-019-47462-4

Abstract

Donepezil, a therapeutic drug for Alzheimer’s disease, ameliorates cognitive dysfunction through selective inhibition of acetylcholinesterase. However, recent studies have also reported off-target effects of donepezil that likely contribute to its therapeutic effects. In this study, we investigated the (i) role of donepezil in amyloid precursor protein (APP) processing and (ii) involvement of sorting nexin protein 33 (SNX33), a member of the sorting nexin protein family, in this processing. Results showed that donepezil induces an increase in SNX33 expression in primary cortical neurons. The secretion of sAPPα in culture media increased, whereas the expression of full-length APP in the cell lysate remained unchanged. Exposure of cortical cultures to donepezil led to a decrease in amyloid β (Aβ) protein levels in a concentration- and time-dependent manner. This decrease was not affected by concomitant treatment with acetylcholine receptor antagonists. SNX33 knockdown by target-specific morpholino oligos inhibited the effects of donepezil. Donepezil treatment increased cell membrane surface expression of APP in SNX33 expression-dependent manner. These results suggested that donepezil decreases the level of Aβ by increasing SNX33 expression and APP cleavage by α-secretase in cortical neurons.

Highlights

Introduction of morpholino oligosWe delivered 1 mM MOs; Genetools, Philomath, OR, USA) into the cells using the Endo Porter delivery reagent
To examine the mechanism involved in the neuroprotective effects observed by donepezil and galantamine treatment in primary cortical neurons, we initially screened for genes whose expression levels are up-regulated by these two drugs
Among candidate genes identified by microarray analysis of transcripts obtained from primary cortical neurons treated with donepezil or galantamine for 24 h, we focused on the up-regulation of sorting nexin protein 33 (SNX33) expression because the sorting nexin protein (SNX) family is believed to be involved in endomembrane transport of various transmembrane proteins, including APP11

Summary

Introduction

Introduction of morpholino oligosWe delivered 1 mM MOs; Genetools, Philomath, OR, USA) into the cells using the Endo Porter delivery reagent. We examined the effects of donepezil on APP processing in primary cortical neurons. To examine the mechanism involved in the neuroprotective effects observed by donepezil and galantamine treatment in primary cortical neurons, we initially screened for genes whose expression levels are up-regulated by these two drugs.

Results

Conclusion