Platelets confound the measurement of extracellular miRNA in archived plasma

• Type of Material: Article
• Authors: Adam Mitchell, Emory University; Warren D. Gray, Emory University; Salim S. Hayek, Emory University; Yi-An Ko, Emory University; Sheena Thomas, Emory University; Kim Rooney, Emory University; Mosaab Awad, Emory University; John Roback, Emory University; Arshed Quyyumi, Emory University; Charles Searles Jr, Emory University
• Language: English
• Date: 2016-09-13
• Publisher: Nature Publishing Group
• Publication Version: Final Published Version
• Copyright Statement: © The Author(s) 2016.
• License: Creative Commons Attribution 4.0 International
• Final Published Version (URL): http://dx.doi.org/10.1038/srep32651
• Title of Journal or Parent Work: Scientific Reports
• ISSN: 2045-2322
• Volume: 6
• Start Page: 32651
• End Page: 32651
• Grant/Funding Information: Research reported in this publication was supported by the National Heart, Lung, and Blood Institute of the National Institutes of Health (R01HL109559 (CDS), NIH R01HL124879 (CDS), T32HL007745 (WDG)). This work was supported by NIH T32HL069769 (JR); NIH VA Merit I01BX000704 (CDS); AHA 15SFDRN25220002 (CDS).
• Keywords: Science & Technology; Multidisciplinary Sciences; MEMBRANE INTEGRITY; Predictive markers; CANCER; CIRCULATING MICRORNAS; HEART-FAILURE; MICROPARTICLES; ARTERY-DISEASE; STANDARDIZATION; Diagnostic markers
• Research Categories: Health Sciences, General; Health Sciences, Pathology

Abstract

Extracellular miRNAs are detectable in biofluids and represent a novel class of disease biomarker. Although many studies have utilized archived plasma for miRNA biomarker discovery, the effects of processing and storage have not been rigorously studied. Previous reports have suggested plasma samples are commonly contaminated by platelets, significantly confounding the measurement of extracellular miRNA, which was thought to be easily addressed by additional post-thaw plasma processing. In a case-control study of archived plasma, we noted a significant correlation between miRNA levels and platelet counts despite post-thaw processing. We thus examined the effects of a single freeze/thaw cycle on microparticles (MPs) and miRNA levels, and show that a single freeze/thaw cycle of plasma dramatically increases the number of platelet-derived MPs, contaminates the extracellular miRNA pool, and profoundly affects the levels of miRNAs detected. The measurement of extracellular miRNAs in archived samples is critically dependent on the removal of residual platelets prior to freezing plasma samples. Many previous clinical studies of extracellular miRNA in archived plasma should be interpreted with caution and future studies should avoid the effects of platelet contamination.