Fig. 3
Comparison of miRNA cargo isolated from young and old mouse plasma EVs
(a) Schematic diagram showing timeline of miRNA isolation, quality control checks and qPCR-based array. Plasma EVs from three mice per group were analysed. (b) Total miRNA yield from Y_EVs and O_EVs. Samples were compared by unpaired t-test. (c) Principal components analysis (PCA) of Y_EV (silver) and O_EV (green) miRNA cargo. (d) Cycle threshold (CT) values for spike-in miRNAs UniSP6 (pre-reverse transcription), UniSP2, 4 and 5 representing high, medium and low expressed miRNAs, and UniSP3 inter-plate calibration (IPC). cel-miR-39-3p is also included. (e) Percentage of miRNAs with high (CT ≤ 30), medium (30-34.9), low (35–40) and absent expression out of 752 assayed miRNAs. No significant differences in populations were detected. (f) The most abundant miRNAs in Y_EV and O_EV samples, listed from highest to lowest. (g) Scatter diagram of Y_EV (x axis) vs. O_EV (Y axis) miRNA expression levels normalised using NormFinder. A correlation value is shown, and some highly abundant miRNAs are annotated. (h) Volcano plot showing statistical significance (Y axis) against fold-change (X axis). The Y axis line represents a P value of 0.05 and X axis lines show fold changes of + 2.0 or -2.0. Significantly different miRNAs are annotated. (j) Venn diagram showing overlap of the 50 most abundant miRNAs in Y_EV and O_EV samples. The miRNAs found in only one population are annotated. Target prediction of differentially-regulated miRNAs relevant to macrophages