Department of Medicine, University of Vermont, Burlington, VT, USA.
J Alzheimers Dis. 2012;29(2):449-57. doi: 10.3233/JAD-2011-111337.
Enzymatic cleavage of amyloid-β protein precursor (AβPP) produces amyloid-β (Aβ) peptides which form the insoluble cortical plaques characteristic of Alzheimer's disease (AD). AβPP is post-transcriptionally processed into three major isoforms with differential cellular and tissue expression patterns. Changes in AβPP isoform expression may be indicative of disease pathogenesis in AD, but accurately measuring AβPP gene isoforms has been difficult to standardize, reproduce, and interpret. In light of this, we developed a set of isoform specific absolute quantification real time PCR standards that allow for quantification of transcript copy numbers for total AβPP and all three major isoforms (AβPP695, AβPP751, and AβPP770) in addition to glyceraldehyde-3-dehydrogenase (GAPDH) and examined expression patterns in superior frontal gyrus (SFG) and cerebellar samples from patients with (n = 12) and without AD (n = 10). Both total AβPP and AβPP695 transcripts were significantly decreased in SFG of patients with AD compared to control (p = 0.037 and p = 0.034, respectively). AβPP751 and AβPP770 transcripts numbers were not significantly different between AD and control (p > 0.15). There was trend for decreased percentage AβPP695 (p = 0.051) and increased percentage AβPP770 (p = 0.013) expression in SFG of patients with AD. GAPDH transcripts levels were also decreased significantly in the SFG of patients with AD compared to control (p = 0.005). Decreasing total AβPP and AβPP695 copy number was associated with increased plaque burden and decreased cognitive function. In this study we describe a simple procedure for measuring AβPP isoform transcripts by real-time PCR and confirm previous studies showing altered AβPP isoform expression patterns in AD.
淀粉样前体蛋白（AβPP）经酶切产生淀粉样β（Aβ）肽，这些肽形成阿尔茨海默病（AD）皮质斑块的不溶性特征。AβPP 经转录后加工为三种主要的同工型，具有不同的细胞和组织表达模式。AβPP 同工型表达的变化可能是 AD 发病机制的指标，但准确测量 AβPP 基因同工型一直难以标准化、重现和解释。有鉴于此，我们开发了一套同工型特异性绝对定量实时 PCR 标准，可用于定量总 AβPP 和所有三种主要同工型（AβPP695、AβPP751 和 AβPP770）以及甘油醛-3-磷酸脱氢酶（GAPDH）的转录本拷贝数，并检查了来自 AD 患者（n = 12）和非 AD 患者（n = 10）的额上回（SFG）和小脑样本中的表达模式。与对照组相比，AD 患者 SFG 中的总 AβPP 和 AβPP695 转录本均显著降低（p = 0.037 和 p = 0.034）。AD 和对照组之间 AβPP751 和 AβPP770 转录本数量无显著差异（p > 0.15）。AD 患者 SFG 中 AβPP695 表达百分比降低（p = 0.051）和 AβPP770 表达百分比增加（p = 0.013）的趋势。与对照组相比，AD 患者 SFG 中的 GAPDH 转录本水平也显著降低（p = 0.005）。总 AβPP 和 AβPP695 拷贝数的减少与斑块负荷的增加和认知功能的下降有关。在这项研究中，我们描述了一种通过实时 PCR 测量 AβPP 同工型转录本的简单程序，并证实了之前的研究表明 AD 中 AβPP 同工型表达模式发生改变。
