- 作者：MiMi P. Macias ; Am ; a M. Gonzales ; Ashley L. Siniard ; Aaron W. Walker ; Jason J. Corneveaux ; Matthew J. Huentelman ; Marwan N. Sabbagh ; Boris Decourt
- 关键词：Alzheimer's ; BE(2)-M17 ; Amyloid-beta ; In vitro ; Secretase ; TNF
- 刊名：Journal of Neuroscience Methods
- 出版年：15 February, 2014
- 年：2014
- 卷：223
- 期：Complete
- 页码：114-122
- 全文大小：1561 K
Background
A hallmark pathologic feature of Alzheimer's disease (AD) is accumulation of neuritic senile plaques in the brain parenchyma. Neurotoxic plaque cores are composed predominantly of amyloid-尾 (A尾) peptides of 40 and 42 amino acids in length, formed by sequential cleavage of amyloid precursor protein (APP) by 尾-, and 纬-secretases. There is a great interest in approaches to modulate A尾 peptide production and develop therapeutic interventions to reduce A尾 levels to halt or slow the progression of neurodegeneration.New method
We characterized and present the BE(2)-M17 human neuroblastoma cell line as a novel in vitro model of the APP-cleavage cascade to support future (1) functional studies of molecular regulators in A尾 production, and (2) high-throughput screening assays of new pharmacotherapeutics.
Results
In BE(2)-M17 cells, both RNA (i.e., RT-PCR, RNA sequencing) and protein analyses (i.e., Western blots, ELISA), show endogenous expression of critical components of the amyloidogenic pathway, APP-cleavage intermediates CTF83 and CTF99, and final cleavage products A尾40 and A尾42. We further report effects of retinoic acid-mediated differentiation on morphology and gene expression in this cell line.
Comparison with existing method(s)
In contrast to primary isolates or other cell lines reported in current literature, BE(2)-M17 not only sustains baseline expression of the full contingent of APP-processing components, but also remains stably adherent during culture, facilitating experimental manipulations.
Conclusions
Our evidence supports the use of BE(2)-M17 as a novel, human, cell-based model of the APP processing pathway that offers a potential streamlined approach to dissect molecular functions of endogenous regulatory pathways, and perform mechanistic studies to identify modulators of A尾 production.