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Proc Natl Acad Sci U S A. 2018 Jun 5;115(23):E5334-E5343. doi: 10.1073/pnas.1714397115. Epub 2018 May 18.

Human plasma and serum extracellular small RNA reference profiles and their clinical utility.

Author information

1
Howard Hughes Medical Institute, The Rockefeller University, New York, NY 10065.
2
Laboratory for RNA Molecular Biology, The Rockefeller University, New York, NY 10065.
3
Department of Nephrology, Hadassah-Hebrew University Medical Center, Jerusalem 91120, Israel.
4
Department of Obstetrics and Gynecology, Columbia University Medical Center, New York, NY 10032.
5
Department of General, Visceral, and Pediatric Surgery, University Medical Center Göttingen, 37075 Göttingen, Germany.
6
Department of Pathology and Cell Biology, Columbia University Medical Center, New York, NY 10032.
7
Department of Pediatrics, Columbia University Medical Center, New York, NY 10032.
8
Department of Obstetrics and Gynecology, Columbia University Medical Center, New York, NY 10032; zw2421@cumc.columbia.edu ttuschl@rockefeller.edu.
9
Howard Hughes Medical Institute, The Rockefeller University, New York, NY 10065; zw2421@cumc.columbia.edu ttuschl@rockefeller.edu.

Abstract

Circulating extracellular RNAs (exRNAs) have the potential to serve as biomarkers for a wide range of medical conditions. However, limitations in existing exRNA isolation methods and a lack of knowledge on parameters affecting exRNA variability in human samples may hinder their successful discovery and clinical implementation. Using combinations of denaturants, reducing agents, proteolysis, and revised organic extraction, we developed an automated, high-throughput approach for recovery of exRNAs and exDNA from the same biofluid sample. We applied this method to characterize exRNAs from 312 plasma and serum samples collected from 13 healthy volunteers at 12 time points over a 2-month period. Small RNA cDNA library sequencing identified nearly twofold increased epithelial-, muscle-, and neuroendocrine-cell-specific miRNAs in females, while fasting and hormonal cycle showed little effect. External standardization helped to detect quantitative differences in erythrocyte and platelet-specific miRNA contributions and in miRNA concentrations between biofluids. It also helped to identify a study participant with a unique exRNA phenotype featuring a miRNA signature of up to 20-fold elevated endocrine-cell-specific miRNAs and twofold elevated total miRNA concentrations stable for over 1 year. Collectively, these results demonstrate an efficient and quantitative method to discern exRNA phenotypes and suggest that plasma and serum RNA profiles are stable over months and can be routinely monitored in long-term clinical studies.

KEYWORDS:

biofluid DNA isolation; biofluid RNA isolation; exRNA biomarker; exRNA reference profiling; extracellular nucleic acids

PMID:
29777089
DOI:
10.1073/pnas.1714397115
[Indexed for MEDLINE]
Free PMC Article

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