Vesicle-associated microRNAs are released from blood cells on incubation of blood samples

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• 作者：Verena Kö ; berle ; Bianca Kakoschky ; Ahmed Atef Ibrahim ; Christian Schmithals ; Jan Peveling-Oberhag ; Stefan Zeuzem ; Bernd Kronenberger ; Oliver Waidmann ; Thomas Pleli ; Albrecht Piiper ; ^{piiper@med.uni-frankfurt.de" class="auth_mail" title="E-mail the corresponding author}
• 关键词：Ago2 ; Argonaute 2 ; miRNA ; microRNA ; PBS ; phosphate-buffered saline ; RNase A ; Ribonuclease A ; RT-PCR ; reverse transcriptase polymerase chain reaction
• 刊名：Translational Research
• 出版年：2016
• 出版时间：March 2016
• 年：2016
• 卷：169
• 期：Complete
• 页码：40-46
• 全文大小：364 K

文摘

MicroRNAs (miRNAs) circulating extracellularly in the blood are currently intensively studied as novel disease markers. However, the preanalytical factors influencing the levels of the extracellular miRNAs are still incompletely explored. In particular, it is unknown, whether the incubation of blood samples as occurring in clinical routine can lead to a release of miRNAs from blood cells and thus alter the extracellular miRNA levels before the preparation of serum or plasma from the blood cells. Using a set of marker miRNAs and quantitative RT-PCR, we found that the levels of extracellular miRNA-1, miRNA-16, and miRNA-21 were increased in EDTA and serum collection tubes incubated for 1–3 hours at room temperature and declined thereafter; the levels of the liver-specific miRNA-122 declined monophasically. These events occurred in the absence of significant hemolysis. When the blood was supplemented with Ribonuclease A inhibitor, the levels of miRNA-1, miRNA-16, and miRNA-21 increased substantially during the initial 3 hours of incubation and those of miRNA-122 remained unchanged, indicating that the release of blood cell–derived miRNAs occurred during the initial 3 hours of incubation of the blood tubes, but not at later time points. Separation of 5-hour preincubated blood into vesicle and nonvesicle fractions revealed a selective increase in the portion of vesicle-associated miRNAs. Together, these data indicate that the release of vesicle-associated miRNAs from blood cells can occur in blood samples within the time elapsing in normal clinical practice until their processing without significant hemolysis. This becomes particularly visible on the inhibition of miRNA degradation by Ribonuclease A inhibitor.