Characterization and analysis of extracellular vesicle-derived miRNAs from different adipose tissues in mice

Figures

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  Fig. 1. Isolation and characterization of sEVs from different adipose tissues of mice. (A) Schematic diagram of the experimental procedure for the isolation of sEVs. (B) Transmission electron microscopy (TEM) images of sEVs released from iBAT, iWAT, and eWAT. (C and D) Nanoparticle tracking analysis (NTA) and quantification of sEVs derived from iBAT, iWAT, and eWAT (n = 6 each group, from 2 independent experiments). (E) Western blot detection of EV markers ALIX, CD63, and CD9 in sEVs derived from iBAT, iWAT, and eWAT (n = 6 each group, from 2 independent experiments). (F) Western blot analysis of non-vesicular extracellular particles (NVEPs) TGFBI and LDHA in the tissue, cell debris, and their derived sEVs from iBAT, iWAT, and eWAT (n = 6 each group, from 2 independent experiments).

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  Fig. 2. Comparative Analysis of sEV-Derived miRNA Expression Profiles in Different Adipose Tissues. (A) The heatmap illustrates the expression levels of EV-derived miRNAs from eWAT, iWAT, and iBAT, The color scale represents the correlation between color and expression levels, with intensity ranging from low (blue) to high (red). For each tissue type, samples were collected from 6 mice, with each row representing pooled samples from 2 distinct mice, resulting in a total of three composite sample pools. (B) Among the top 30 expressed adipose-derived extracellular vesicle-miRNAs (ADEV-miRNAs), there are 16 miRNAs common to the three types of adipose tissues. (C) The bubble chart of KEGG enrichment analysis for the target genes of the 16 common miRNAs, with p-values indicating the statistical significance of each pathway. (D) GO analysis of the pathways related to the target genes of the 16 common miRNAs, conducted using DAVID Bioinformatics Resources and categorized based on biological processes, cellular components, and molecular functions, with each color representing a different category. (E) Molecular interaction network based on the pathways related to the target genes of the 16…

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  Fig. 3. Comprehensive Analysis of iBAT-Specific EV-derived miRNA and Their Unique Profiles. (A) Venn diagram illustrating a subset of 13 extracellular vesicle (EV)-derived microRNAs (miRNAs) exclusive to iBAT. (B) KEGG enrichment analysis bubble plot for the target genes of 13 iBAT-specific EV-derived miRNAs, with p-values indicating the statistical significance of each pathway. (C) GO analysis of the pathways associated with the target genes of iBAT-specific EV-derived miRNAs, conducted using DAVID Bioinformatics Resources and categorized by biological process, cellular component, and molecular function, with each color representing a different category.(D) Molecular interaction network of the target genes of 13 iBAT-specific EV-derived miRNAs, highlighting key nodes that represent genes with a high degree of connectivity. (E) Graphical representation of the quantitative analysis of HepG2 cell migration, expressed as the percentage of wound closure relative to the initial scratch area at time 0. Scale bars: 200 μm. (F) Assessment of HepG2 cell viability using the Cell Counting Kit-8 (CCK-8) assay (n = 6 each group, from 2 independent experiments). (G) Relative expression levels of genes…

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  Fig. 4. Analysis of iWAT-Specific Enriched miRNAs and Shared miRNAs with iBAT. (A) Venn diagram highlighting a subset of three EV-derived miRNAs exclusively identified in iWAT. (B) KEGG enrichment analysis bubble plot for the target genes of three iWAT-specific EV-derived miRNAs, with p-values indicating the statistical significance of each pathway. (C) GO analysis of the pathways associated with the target genes of iWAT-specific EV-derived miRNAs, conducted using DAVID Bioinformatics Resources and categorized by biological process, cellular component, and molecular function, with each color representing a different category. (D) Venn diagram focusing on a subset of one EV-derived miRNA shared between iBAT and iWAT. (E) KEGG enrichment analysis bubble plot for the target genes of the shared miRNA between iBAT and iWAT, with p-values indicating the statistical significance of each pathway. (F and G) Western blot and quantification analysis of phosphorylated AKT in iWAT-derived SVF cells (n = 6 biological replicates, from three independent experiments).

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  Fig. 5. Analysis of eWAT-Specific EV-derived miRNAs and shared miRNAs between eWAT and iWAT. (A) Venn diagram highlighting a subset of four extracellular vesicle (EV)-derived microRNAs (miRNAs) exclusively identified in eWAT. (B) KEGG enrichment analysis bubble plot for the target genes of four eWAT-specific EV-derived miRNAs, with p-values indicating the statistical significance of each pathway. (C) Gene Ontology (GO) analysis of the pathways associated with the target genes of eWAT-specific EV-derived miRNAs, performed using DAVID Bioinformatics Resources and categorized by biological process, cellular component, and molecular function, with distinct colors representing different categories. (D) Venn diagram illustrating the overlap of 10 EV-derived miRNAs shared between eWAT and iWAT. (E). KEGG enrichment analysis bubble plot for the target genes of 10 EV-derived miRNAs common to both eWAT and iWAT, with p-values indicating the statistical significance of each pathway. (F) GO analysis of the target genes of EV-derived miRNAs shared between eWAT and iWAT, performed using DAVID Bioinformatics Resources and categorized by biological process, cellular component, and molecular function,…