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Purification and microRNA Profiling of Exosomes Derived from Blood and Culture Media
Published on: June 14, 2013
MicroRNA Expression Profiles of Epicardial Adipose Tissue-Derived Exosomes in Patients with Coronary Atherosclerosis
Jinxing Liu1, Ang Gao1, Yan Liu1
1Department of Cardiology, Beijing Anzhen Hospital, Capital Medical University, Beijing Institute of Heart Lung and Blood Vessel Diseases, Beijing Key Laboratory of Precision Medicine of Coronary Atherosclerotic Disease, Clinical center for coronary heart disease, Capital Medical University, 100029 Beijing, China.
Background And Aims:
Epicardial adipose tissue, exosomes, and miRNAs have important activities in atherosclerosis. The purpose of this study was to establish miRNA expression profiles of epicardial adipose tissue-derived exosomes in patients with coronary atherosclerosis.
Methods:
Biopsies of epicardial adipose tissue were obtained from patients with and without coronary artery disease (CAD, n = 12 and NCAD, n = 12) during elective open-heart surgeries. Tissue was incubated with DMEM-F12 for 24 hours. Exosomes were isolated, then nanoparticle tracking analysis, transmission electron microscopy, and immunoblotting were performed to confirm the existence of exosomes. Total RNA in exosomes was subjected to high-throughput sequencing to identify differentially expressed miRNAs. MicroRNA target gene prediction was performed, and target genes were analyzed by Gene Ontology (GO), the Kyoto Encyclopedia of Genes and Genomes (KEGG), and mirPath to identify function. Reverse transcription quantitative PCR was performed to confirm the differentially expressed miRNAs.
Results:
Fifty-three unique miRNAs were identified (adjusted p 0.05, fold of change 2), among which 32 miRNAs were upregulated and 21 miRNAs were downregulated in coronary artery disease patients. Reverse transcription quantitative PCR validated the results for seven miRNAs including miR-141-3p, miR-183-5p, miR-200a-5p, miR-205-5p, miR-429, miR-382-5p and miR-485-3p, with the last two downregulated. GO and KEGG analysis by mirPath indicated that these differentially expressed miRNAs were enriched in cell survival, apoptosis, proliferation, and differentiation.
Conclusions:
Coronary artery disease patients showed differential epicardial adipose tissue exosomal miRNA expression compared with patients without coronary artery disease. The results provide clues for further studies of mechanisms of atherosclerosis.
