RNA from stabilized whole blood enables more comprehensive immune gene expression profiling compared to RNA from peripheral blood mononuclear cells

Fleur van der Sijde; Yunlei Li; Rick Schraauwen; Willem de Koning; Casper H J van Eijck; Dana A M Mustafa

doi:10.1371/journal.pone.0235413

RNA from stabilized whole blood enables more comprehensive immune gene expression profiling compared to RNA from peripheral blood mononuclear cells

PLoS One. 2020 Jun 26;15(6):e0235413. doi: 10.1371/journal.pone.0235413. eCollection 2020.

Authors

Fleur van der Sijde¹, Yunlei Li², Rick Schraauwen³, Willem de Koning², Casper H J van Eijck¹, Dana A M Mustafa³

Affiliations

¹ Department of Surgery, Erasmus MC, University Medical Center Rotterdam, Rotterdam, The Netherlands.
² Department of Pathology, Clinical Bioinformatics Unit, Erasmus MC, University Medical Center Rotterdam, Rotterdam, The Netherlands.
³ Department of Pathology, Tumor Immuno-Pathology Laboratory, Erasmus MC, University Medical Center Rotterdam, Rotterdam, The Netherlands.

Abstract

Monitoring changes in the immune profile in blood samples can help identifying changes in tumor biology and therapy responsiveness over time. Immune-related gene expression profiles offer a highly reproducible method to monitor changes of the immune system. However, measuring gene expression profiles in whole blood samples can be complicated because of the high protein and enzyme abundancy that affect the stability and quality of the RNA. Peripheral blood mononuclear cells (PBMCs) are one the most commonly used source for immune cell RNA extraction, though, this method does not reflect all components of the peripheral blood. The aim of this study was to determine the differences in immune-related gene expression between RNA isolated from stabilized whole blood and RNA isolated from PBMCs. Whole blood samples from 12 pancreatic cancer patients were collected before and after chemotherapy (n = 24). Blood samples were collected in both EDTA tubes, and Tempus tubes containing an RNA stabilizer (total n = 48). PBMCs were isolated from EDTA samples using Ficoll and were snap frozen. Subsequently, immune-related gene expression was profiled using the PanCancer Immune Profiling Panel of NanoString technology. Gene expression profiles of PBMCs were compared to that of Tempus tubes using the Advanced Analysis module of nSolver software. Both types of samples provided good quality RNA and gene expression measurements. However, RNA isolated from Tempus tubes resulted in significantly higher gene counts than PBMCs; 107/730 genes were exclusively detected in Tempus samples, while under the detection limit in PBMCs. In addition, 192/730 genes showed significantly higher gene counts in Tempus samples, 157/730 genes showed higher gene counts in PBMCs. Thus, RNA isolated from whole blood stabilizing blood tubes, such as Tempus tubes, enable higher gene counts and more comprehensive measurements of gene expression profiles compared to RNA isolated from PBMCs.

Publication types

Comparative Study
Research Support, Non-U.S. Gov't

MeSH terms

Cohort Studies
Female
Gene Expression Profiling / methods*
Humans
Leukocytes, Mononuclear / immunology*
Leukocytes, Mononuclear / metabolism*
Male
Pancreatic Neoplasms / blood
Pancreatic Neoplasms / genetics
Pancreatic Neoplasms / immunology
RNA / blood*
RNA / genetics*

Substances

RNA

Grants and funding

This study was funded by the Support Casper Foundation. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.