Analysis of Argonaute 2-microRNA complexes in ex vivo stored red blood cells.

BACKGROUND

Human enucleated mature red blood cells (RBCs) contain both mature microRNAs (miRNAs) and mRNAs, and we have previously correlated RBC storage lesion processes such as eryptosis, adenosine 5'-triphosphate loss, and RBC indices with differentially expressed miRNAs. Here we have characterized Argonaute 2 (AGO2)-miRNA complexes in stored mature RBCs as a first step toward understanding their role, if any.

STUDY DESIGN AND METHODS

In this report AGO2-bound miRNAs in mature RBCs isolated from RBCs collected from three different healthy donors and stored for 24 hours at 4 to 6°C were identified by anti-AGO2 immunoprecipitation (IP) followed by next-generation sequencing of the RNA isolated from the IP. The data were analyzed by various bioinformatics tools.

RESULTS

The analysis highlighted 28 mature AGO2-bound miRNAs that are common to all three donors, representing 95.6% of the identified miRNAs. Among these, miR-16-5p (20.6%), miR-451a-5p (16.7%), miR-486-5p (12.6%), and miR-92a-3p (12.6%) are the most abundant miRNAs. Functional enrichment analysis for mRNA targets of the 28 common miRNAs identified molecules related to various diseases, biofunctions, and toxicity functions such as cardio-, hepato-, and nephrotoxicity.

CONCLUSION

Overall, these results demonstrate the existence of multiple intracellular AGO2-bound miRNAs in 24-hour-stored RBCs and warrant further experiments to determine whether AGO2-miRNAs are functional in RBCs.