Basic Study
Copyright ©The Author(s) 2020. Published by Baishideng Publishing Group Inc. All rights reserved.
World J Gastroenterol. May 28, 2020; 26(20): 2570-2583
Published online May 28, 2020. doi: 10.3748/wjg.v26.i20.2570
Serum outperforms plasma in small extracellular vesicle microRNA biomarker studies of adenocarcinoma of the esophagus
Karen Chiam, George C Mayne, Tingting Wang, David I Watson, Tanya S Irvine, Tim Bright, Lorelle T Smith, Imogen A Ball, Joanne M Bowen, Dorothy M Keefe, Sarah K Thompson, Damian J Hussey
Karen Chiam, George C Mayne, Tingting Wang, David I Watson, Tanya S Irvine, Tim Bright, Lorelle T Smith, Sarah K Thompson, Damian J Hussey, Discipline of Surgery, College of Medicine and Public Health, Flinders University of South Australia, Adelaide, SA 5042, Australia
Karen Chiam, George C Mayne, Tingting Wang, David I Watson, Tanya S Irvine, Tim Bright, Lorelle T Smith, Damian J Hussey, Flinders Health and Medical Research Institute Cancer Program, College of Medicine and Public Health, Flinders University, Adelaide, SA 5042, Australia
Lorelle T Smith, Discipline of Surgery, The University of Adelaide, Adelaide, SA 5005, Australia
Imogen A Ball, Joanne M Bowen, Adelaide Medical School, The University of Adelaide, Adelaide, SA 5005, Australia
Dorothy M Keefe, Faculty of Health Sciences, The University of Adelaide, Adelaide, SA 5005, Australia
Author contributions: Hussey DJ conceived and supervised the study; Chiam K, Mayne GC, Watson DI, Bowen JM, Keefe DM, Thompson SK and Hussey DJ contributed to study design; Watson DI, Irvine TS, Bright T, Smith LT and White I collected patient blood samples; Wang T processed patient blood samples and performed the laboratory assays; Chiam K, Mayne GC and Hussey DJ analysed the data; Chiam K wrote the first draft of the paper; All authors contributed to revision of the manuscript in its final version.
Supported by National Health and Medical Research Council (NHMRC) Project Funding, No. APP1104281; and NHMRC Centres of Research Excellence (CRE) Grant, No. APP1040947.
Institutional review board statement: Ethical approval was obtained from the Southern Adelaide Clinical Human Research Ethics Committee and the Royal Adelaide Hospital Research Committee.
Conflict-of-interest statement: The authors have no conflict of interest to declare.
Data sharing statement: No additional data are available.
Open-Access: This article is an open-access article that was selected by an in-house editor and fully peer-reviewed by external reviewers. It is distributed in accordance with the Creative Commons Attribution NonCommercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited and the use is non-commercial. See: http://creativecommons.org/licenses/by-nc/4.0/
Corresponding author: Damian J Hussey, PhD, Senior Scientist, Discipline of Surgery, College of Medicine and Public Health, Flinders University of South Australia, Bedford Park, Adelaide, SA 5042, Australia. damian.hussey@flinders.edu.au
Received: January 1, 2020
Peer-review started: January 1, 2020
First decision: January 19, 2020
Revised: March 2, 2020
Accepted: May 13, 2020
Article in press: May 13, 2020
Published online: May 28, 2020
Processing time: 148 Days and 11 Hours
ARTICLE HIGHLIGHTS
Research background

Small extracellular vesicles (sEVs), including exosomes, are shed from tumors into the blood circulation. These circulating sEVs are an excellent source of microRNA (miRNA) biomarkers for cancer research. Blood serum and blood plasma both contain sEVs, however at present there is no consensus on which of these two blood sample types is most useful for biomarker analysis.

Research motivation

Extracellular vesicle preparations derived from serum and plasma are known to be enriched in sEVs, but they also contain significant amounts of non-vesicle associated miRNAs derived from sources such as blood cells and protein-bound miRNAs. These non-vesicles associated miRNAs could interfere with cancer biomarker discovery. Our study was motivated by the need to determine which blood sample contains the least amount of non-vesicle associated miRNAs. This knowledge has the potential to improve cancer biomarker discovery and translation.

Research objectives

We sought to compare the miRNA profiles between serum and plasma sEV preparations to determine their suitability for biomarker studies. We also sought to compare the diagnostic performance of these two sample types using a previously established multi-miRNA biomarker panel for esophageal adenocarcinoma.

Research methods

Matched serum and plasma samples from 10 healthy controls and 10 patients with esophageal adenocarcinoma were used for this study. sEVs were isolated with using ExoquickTM. RNA extracted from the vesicles was profiled using the Taqman Openarray qPCR.

Research results

The overall miRNA content was higher in plasma sEV preparations (480 miRNAs) and contained 97.5% of the miRNAs found in the serum sEV preparations (412 miRNAs). The expression of commonly expressed miRNAs was highly correlated (Spearman’s R = 0.87, P < 0.0001) between the plasma and serum sEV preparations but was consistently higher in the plasma sEV preparations. Specific blood-cell miRNAs (hsa-miR-223-3p, hsa-miR-451a, miR-19b-3p, hsa-miR-17-5p, hsa-miR-30b-5p, hsa-miR-106a-5p, hsa-miR-150-5p and hsa-miR-92a-3p) were expressed at 2.7 to 9.6 fold higher levels in the plasma sEV preparations compared to serum sEV preparations (P < 0.05). In plasma sEV preparations, the percentage of protein-associated miRNAs expressed at relatively higher levels (cycle threshold 20-25) was greater than serum sEV preparations (50% vs 31%). While the percentage of vesicle-associated miRNAs expressed at relatively higher levels was greater in the serum sEV preparations than plasma sEV preparations (70% vs 44%). A 5-miRNA biomarker panel produced a higher cross validated accuracy for discriminating patients with esophageal adenocarcinoma from healthy controls using serum sEV preparations compared with plasma sEV preparations (AUROC 0.80 vs 0.54, P < 0.05).

Research conclusions

Although plasma sEV preparations contained more miRNAs than serum sEV preparations, they also contained more miRNAs from non-vesicle origins.

Research perspectives

Serum appears to be more suitable than plasma for sEV miRNAs biomarkers studies. Future studies on sEV associated cancer biomarkers may benefit from using serum as the sample type for analysis.