Analysis of extracellular RNA in cerebrospinal fluid

Julie A Saugstad; Theresa A Lusardi; Kendall R Van Keuren-Jensen; Jay I Phillips; Babett Lind; Christina A Harrington; Trevor J McFarland; Amanda L Courtright; Rebecca A Reiman; Ashish S Yeri; M Yashar S Kalani; P David Adelson; Jorge Arango; John P Nolan; Erika Duggan; Karen Messer; Johnny C Akers; Douglas R Galasko; Joseph F Quinn; Bob S Carter; Fred H Hochberg

doi:10.1080/20013078.2017.1317577

Analysis of extracellular RNA in cerebrospinal fluid

J Extracell Vesicles. 2017 May 24;6(1):1317577. doi: 10.1080/20013078.2017.1317577. eCollection 2017.

Authors

Julie A Saugstad¹, Theresa A Lusardi², Kendall R Van Keuren-Jensen³, Jay I Phillips¹, Babett Lind⁴, Christina A Harrington⁵, Trevor J McFarland⁵, Amanda L Courtright³, Rebecca A Reiman³, Ashish S Yeri³, M Yashar S Kalani⁶, P David Adelson⁷, Jorge Arango⁷, John P Nolan⁸, Erika Duggan⁸, Karen Messer⁹, Johnny C Akers¹⁰, Douglas R Galasko¹¹, Joseph F Quinn¹², Bob S Carter¹⁰, Fred H Hochberg¹⁰

Affiliations

¹ Department of Anesthesiology & Perioperative Medicine, Oregon Health & Science University, Portland, OR, USA.
² Bioinformatics Core, School of Medicine, Oregon Health & Science University, Portland, OR, USA.
³ Neurogenomics, Translational Genomics Research Institute,Phoenix, AZ, USA.
⁴ Department of Neurology, Layton Aging and Alzheimer's Center, Oregon Health & Science University, Portland, OR, USA.
⁵ Integrated Genomics Laboratory, Oregon Health & Science University, Portland, OR, USA.
⁶ Department of Neurosurgery, Radiology, Anatomy and Neurobiology, University of Utah School of Medicine and the Barrow Neurological Institute, Salt Lake City, UT, USA.
⁷ Barrow Neurological Institute at Phoenix Children's Hospital, Department of Child Health, University of Arizona College of Medicine, Phoenix, AZ, USA.
⁸ Scintillon Institute, San Diego, CA, USA.
⁹ Division of Biostatistics and Bioinformatics, University of California, San Diego, CA, USA.
¹⁰ Department of Neurosurgery, University of California, San Diego, CA, USA.
¹¹ Department of Neurosciences, University of California, San Diego, CA, USA.
¹² Department of Neurology, Portland VA Medical Center, Oregon Health & Science University, Portland, OR, USA.

Abstract

We examined the extracellular vesicle (EV) and RNA composition of pooled normal cerebrospinal fluid (CSF) samples and CSF from five major neurological disorders: Alzheimer's disease (AD), Parkinson's disease (PD), low-grade glioma (LGG), glioblastoma multiforme (GBM), and subarachnoid haemorrhage (SAH), representing neurodegenerative disease, cancer, and severe acute brain injury. We evaluated: (I) size and quantity of EVs by nanoparticle tracking analysis (NTA) and vesicle flow cytometry (VFC), (II) RNA yield and purity using four RNA isolation kits, (III) replication of RNA yields within and between laboratories, and (IV) composition of total and EV RNAs by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and RNA sequencing (RNASeq). The CSF contained ~106 EVs/μL by NTA and VFC. Brain tumour and SAH CSF contained more EVs and RNA relative to normal, AD, and PD. RT-qPCR and RNASeq identified disease-related populations of microRNAs and messenger RNAs (mRNAs) relative to normal CSF, in both total and EV fractions. This work presents relevant measures selected to inform the design of subsequent replicative CSF studies. The range of neurological diseases highlights variations in total and EV RNA content due to disease or collection site, revealing critical considerations guiding the selection of appropriate approaches and controls for CSF studies.

Keywords: Extracellular vesicles; cerebrospinal fluid; extracellular RNA; neurological diseases.

Abstract

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