Activation of α7 nicotinic receptor affects APP processing by regulating secretase activity in SH-EP1-α7 nAChR-hAPP695 cells.
Multiple lines of evidence have implicated that nicotinic acetylcholine receptor (nAChR) may be an important therapeutic target for the treatment of Alzheimer's disease (AD). Although there are reports suggesting a link between alpha7 nAChR subtype and AD, there has been little report on the mechanism. The present study investigates whether and how α7 nAChR activation affects APP695 processing in SH-EP1 cell model. Cell line co-expressing α7 nAChR gene and human amyloid precursor protein 695 (hAPP695) gene were constructed by stable transfection. Expression of β-amyloid, α-form of secreted APP (αAPPs) and APP1695 was measured by ELISA, western blotting and real-time PCR respectively. Additionally, α, β, and γ-secretase activities were also analyzed in constructed SH-EP1-α7 nAChR-hAPP695 cell line. The results showed that SH-EP1-α7 nAChR-hAPP695 cell line, expressing both hAPP695 gene and α7 nAChR subtype gene, was constructed successfully. The secreted Aβ was decreased and αAPPs was significantly increased by non-selective nAChR agonist nicotine (10 μM) and specific α7 nAChR agonist GTS-21 (1 μM), and APP expression was not affected. Furthermore, specific α7 nAChR antagonist methyllycaconitine (MLA) reversed the alterations induced by activation of α7 nAChR. CTF-α was increased and CTF-γ was decreased when treated with nicotine (10 μM). In addition, the results of enymatic activity analysis showed that nicotine (1μM) and GTS-21 (0.1, 1 μM) decreased γ-secretase activity, but has no effects on α-secretase activity and β-secretase activity. Our findings demonstrate that, through regulating γ-secretase activity, α7 nAChR activation reduces APP processing in amyloidogenic pathway, and at the same time enhances APP processing in non-amyloidogenic pathway. The constructed SH-EP1-α7 nAChR-hAPP695 cell line might be useful for screening specific nAChR agonists against AD.
Nie
Hui Zhen
HZ
School of Pharmacy, Shanghai Jiao Tong University, Shanghai 200240, China.
Shi
Sha
S
Lukas
Ronald J
RJ
Zhao
Wen Juan
WJ
Sun
Yong Ning
YN
Yin
Ming
M
D053829:Amyloid Precursor Protein Secretases / Q000378:metabolism* / Q000502:physiology; D016564:Amyloid beta-Protein Precursor / Q000037:antagonists & inhibitors / Q000378:metabolism* / Q000502:physiology; D000818:Animals; D002460:Cell Line; D015536:Down-Regulation / Q000235:genetics; D004789:Enzyme Activation / Q000502:physiology; D006801:Humans; D009474:Neurons / Q000201:enzymology / Q000378:metabolism*; D012323:RNA Processing, Post-Transcriptional / Q000235:genetics / Q000502:physiology; D011978:Receptors, Nicotinic / Q000235:genetics / Q000378:metabolism* / Q000502:physiology; D015398:Signal Transduction / Q000235:genetics / Q000502:physiology; D015854:Up-Regulation / Q000235:genetics; D064569:alpha7 Nicotinic Acetylcholine Receptor
D016564:Amyloid beta-Protein Precursor; C579167:Chrna7 protein, human; D011978:Receptors, Nicotinic; D064569:alpha7 Nicotinic Acetylcholine Receptor; D053829:Amyloid Precursor Protein Secretases