Proteomic Characterization of Postmortem Amyloid Plaques Isolated by Laser Capture Microdissection

Lujian Liao,Dongmei Cheng,Jian Wang,Duc M Duong,Tatyana G Losik,Marla Gearing,Howard D Rees,James J Lah,Allan I Levey,Junmin Peng

doi:10.1074/jbc.m403672200

Lujian Liao, Dongmei Cheng + Show 8 more

Open Access

https://doi.org/10.1074/jbc.m403672200

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Abstract

The presence of amyloid plaques in the brain is one of the pathological hallmarks of Alzheimer's disease (AD). We report here a comprehensive proteomic analysis of senile plaques from postmortem AD brain tissues. Senile plaques labeled with thioflavin-S were procured by laser capture microdissection, and their protein components were analyzed by liquid chromatography coupled with tandem mass spectrometry. We identified a total of 488 proteins co-isolated with the plaques, and we found multiple phosphorylation sites on the neurofilament intermediate chain, implicating the complexity and diversity of cellular processes involved in the plaque formation. More significantly, we identified 26 proteins enriched in the plaques of two AD cases by quantitative comparison with surrounding non-plaque tissues. The localization of several proteins in the plaques was further confirmed by the approach of immunohistochemistry. In addition to previously identified plaque constituents, we discovered novel association of dynein heavy chain with the plaques in human postmortem brain and in a double transgenic AD mouse model, suggesting that neuronal transport may play a role in neuritic degeneration. Overall, our results revealed for the first time the sub-proteome of amyloid plaques that is important for further studies on disease biomarker identification and molecular mechanisms of AD pathogenesis.

Highlights

The presence of amyloid plaques in the brain is one of the pathological hallmarks of Alzheimer’s disease (AD)
Senile plaques labeled with thioflavin-S were procured by laser capture microdissection, and their protein components were analyzed by liquid chromatography coupled with tandem mass spectrometry
The abbreviations used are: AD, Alzheimer’s disease; LC-MS/MS, liquid chromatography coupled with tandem mass spectrometry; LCM, laser capture microdissection; A␤, amyloid ␤-peptide; APP, A␤ precursor protein

Summary

Introduction

The presence of amyloid plaques in the brain is one of the pathological hallmarks of Alzheimer’s disease (AD). Senile plaques labeled with thioflavin-S were procured by laser capture microdissection, and their protein components were analyzed by liquid chromatography coupled with tandem mass spectrometry. Our results revealed for the first time the sub-proteome of amyloid plaques that is important for further studies on disease biomarker identification and molecular mechanisms of AD pathogenesis. The abbreviations used are: AD, Alzheimer’s disease; LC-MS/MS, liquid chromatography coupled with tandem mass spectrometry; LCM, laser capture microdissection; A␤, amyloid ␤-peptide; APP, A␤ precursor protein. The first major breakthrough in understanding the molecular pathogenesis of AD came from the biochemical purification of amyloid ␤-peptide (A␤) from senile plaques, as described by Glenner and Wong [6], and the subsequent sequencing and identification of the A␤ precursor protein (APP) gene. Mass spectrometry has greatly simplified the analysis of post-translational modifications [21, 22]

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