■心外膜脂肪组织,外泌体,miRNAs在动脉粥样硬化中具有重要的活性。本研究的目的是建立冠状动脉粥样硬化患者心外膜脂肪组织来源的外泌体的miRNA表达谱。
■从有和没有冠状动脉疾病的患者获得心外膜脂肪组织的活检(CAD,n=12和NCAD,n=12)在选择性心脏直视手术中。将组织与DMEM-F12孵育24小时。外泌体被分离,然后纳米粒子跟踪分析,透射电子显微镜,进行免疫印迹以确认外泌体的存在。对外来体中的总RNA进行高通量测序以鉴定差异表达的miRNA。进行微小RNA靶基因预测,通过基因本体论(GO)分析目标基因,京都基因和基因组百科全书(KEGG),和mirPath来标识函数。进行逆转录定量PCR以确认差异表达的miRNA。
■鉴定了53个独特的miRNA(调整后p<0.05,变化倍数>2),在冠心病患者中,32个miRNAs上调,21个miRNAs下调.逆转录定量PCR验证了7种miRNA的结果,包括miR-141-3p,miR-183-5p,miR-200a-5p,miR-205-5p,miR-429,miR-382-5p和miR-485-3p,最后两个被下调。通过mirPath的GO和KEGG分析表明,这些差异表达的miRNA在细胞存活中富集,凋亡,扩散,和差异化。
■冠状动脉疾病患者与没有冠状动脉疾病的患者相比,表现出不同的心外膜脂肪组织外泌体miRNA表达。该结果为进一步研究动脉粥样硬化的机制提供了线索。
UNASSIGNED: 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.
UNASSIGNED: 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.
UNASSIGNED: 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.
UNASSIGNED: 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.