Stability of circulating microRNAs in serum

Abstract

There is a strong body of evidence by several translational studies which demonstrate the potential of circulating miRNAs as a potential biomarker in oncology. However, recent reports documented varying stability of these small RNA molecules in serum samples. The aim of our pilot study was to evaluate the stability of miRNAs in serum in relation to food intake and sample storage. Serum miRNA expression levels of 16 different miRNAs from 8 healthy volunteers were quantified by real-time PCR. 4 samples from each donor were analysed—2 samples (fasting, in the morning and after food intake, at noon) were analysed within 24h and 2 samples (fasting and after food intake, at noon) were stored at -80°C for 14 days and subsequently analysed. Student´s t-test was used to determine significant differences. The detectability of the distinct miRNA as a surrogate for the stability of these small RNA molecules was slightly altered by the storage conditions, but only a miRNA 22-3p, out of the analysed 16 miRNAs, shows significant lower dCq expression (3.821 vs. 4.530; p<0,01) by qPCR dependent on storage conditions (-80°C vs. 4°C). However, miRNA levels were not affected by food intake. The difference between samples taken in the morning (fasting) and at noon (after a normal meal) did not show any significant differences. MiRNAs can be considered to be a relatively stable tool in laboratory diagnostics, but clearly every new assay needs thorough evaluation. The stability of miRNAs documented here in healthy volunteers shows their potential in the search for innovative biomarkers in oncology.

Changes in microRNAs during Storage and Processing of Breast Milk

“…In a study by Kupec et al, serum miRNA was not affected by food intake or sample collection times. However, there was a significant difference in expression according to the storage temperature (−80 vs. 4 °C) [ 28 ]. Glinge et al confirmed that miRNA levels were stable for at least 24 h at room temperature in whole blood but were significantly changed after 72 h [ 29 ].…”

A Glimmer of Hope for Patients with a T3 Transformation Zone: miRNAs Are Potential Biomarkers for Cervical Dysplasia

“…Hsa-miR-205-5p was found to be a key regulator of VEGFA during cancer-related angiogenesis in hepatocellular carcinoma [42]. Since miRNAs are extremely stable even at room temperature and survive long freezing-thawing cycles without notable changes in expression [24], these small molecules have become more and more prominent research targets for human cancer detection and classification.…”

“…Several biomarkers, including p16, ki-67, methylation, and genotyping, have been tested so far, but none were able to close the diagnostic gap in clinical practice [21,22]. On the other hand, miRNAs have become an interesting tool in distinguishing genetically different cellular entities and are a source of potential biomarkers [23][24][25][26]. MicroRNAs are noncoding regulatory RNAs 19-25 nucleotides (nt) in size that are produced by RNA polymerase II and II derived from transcripts of coding or noncoding genes.…”

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“… 18 The overall stability of plasma miRs remains a concern given the small size, circulatory RNase susceptibility, and impact of collection mechanisms. 29 …”