The amyloid plaque proteome in early onset Alzheimer's disease and Down syndrome.

Amyloid plaques contain many proteins in addition to beta amyloid (Aß). Previous studies examining plaque-associated proteins have shown these additional proteins are important; they provide insight into the factors that drive amyloid plaque development and are potential biomarkers or therapeutic targets for Alzheimer's disease (AD). The aim of this study was to comprehensively identify proteins that are enriched in amyloid plaques using unbiased proteomics in two subtypes of early onset AD: sporadic early onset AD (EOAD) and Down Syndrome (DS) with AD. We focused our study on early onset AD as the drivers of the more aggressive pathology development in these cases is unknown and it is unclear whether amyloid-plaque enriched proteins differ between subtypes of early onset AD. Amyloid plaques and neighbouring non-plaque tissue were microdissected from human brain sections using laser capture microdissection and label-free LC-MS was used to quantify the proteins present. 48 proteins were consistently enriched in amyloid plaques in EOAD and DS. Many of these proteins were more significantly enriched in amyloid plaques than Aß. The most enriched proteins in amyloid plaques in both EOAD and DS were: COL25A1, SMOC1, MDK, NTN1, OLFML3 and HTRA1. Endosomal/lysosomal proteins were particularly highly enriched in amyloid plaques. Fluorescent immunohistochemistry was used to validate the enrichment of four proteins in amyloid plaques (moesin, ezrin, ARL8B and SMOC1) and to compare the amount of total Aß, Aß40, Aß42, phosphorylated Aß, pyroglutamate Aß species and oligomeric species in EOAD and DS. These studies showed that phosphorylated Aß, pyroglutamate Aß species and SMOC1 were significantly higher in DS plaques, while oligomers were significantly higher in EOAD. Overall, we observed that amyloid plaques in EOAD and DS largely contained the same proteins, however the amount of enrichment of some proteins was different in EOAD and DS. Our study highlights the significant enrichment of many proteins in amyloid plaques, many of which may be potential therapeutic targets and/or biomarkers for AD.

Isolation of Extracellular RNA from Bile.

The study of extracellular RNA has been recently reported as a tool for biomarker discovery. Extracellular vesicles can be isolated from different types of body fluids which contain protein, mRNA, and noncoding RNA. Extracellular RNA isolated from bile could be a useful tool for analyzing biliary tract diseases or cancer. Herein, we describe protocols based on modifications of commercially available kits for the collection, processing, and isolation of extracellular RNA from bile.